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Editorial Board

Drozdstoj Stoyanov (Editor-in-chief)Damianka Getova-Spassova(Managing Editor)Ivan Kindekov (Scientific secretary)Boyan Lozanov

Dobrin Svinarov

Grigor Velev

Janet Grudeva-Popova

Margarita Kamenova

Mihail Boyanov

Nadka Bojadjieva

International Advisory Board

Andrew Miles – London, U.K.Ashok Agraval – Clivelandq Phio, USAJuan E. Mezzich – New York, USAJan Kiselovic – Bratislava, SlovakiaKenneth William Fulford – Oxford, U.K.Miroljub Popovic – Murcia, SpainSamuel Refetoff – Chicagp, Illinois, USAStanley B. Prusiner – Nobel Laureate, San Francisco, USA

BULGARIAN MEDICINE ISSN 1314-3387

Редакционна колегия

Дроздстой Стоянов (главен редактор)

Дамянка Гетова-Спасова(изпълнителен редактор)

Иван Киндеков(научен секретар)

Боян Лозанов

Добрин Свинаров

Григор Велев

Жанет Грудева-Попова

Маргарита Каменова

Михаил Боянов

Надка Бояджиева

Международен редакционен съвет

Андрю Майлс – Лондон, Великобритания

Ашок Агравал – Кливланд, САЩХуан Месич – Ню Йорк, САЩ

Ян Киселович – Братислава, Словакия

Кенет Уилиям Фулфорд – Оксфорд, Великобритания

Миролюб Попович – Мурсия, Испания

Самуел Рефетоф – Чикаго, САЩ

Стенли Прузиър – Нобелов лауреат, Сан Франциско, САЩ

Съдържание

А. Каканакова Категориални срещу дименсионални модели

на класификация. Континуум на психозите........... 4

Петя В. Иванова, Ивица ДимовКростолк между различни сигнални пътища

и витамин Д и витамин Д3 рецептор -

ВДР в процесът на онкогенеза в кожа.................. 11

Д. Гетова, Д. Димитрова, М. ТополовСравнително изследване на телесни масти, някои

телесни обиколки и нивата на триглицериди

и холестерол при различни групи пациенти,

включени в НИРДИАБО проекта

от Пловдивска област .............................................25

В. Попова, З. Въжев, Ж. Пешев, Ж. Грудева-Попова, А. БаталовАнализ на експресията на серумните нива на тумор

некрозисфактора алфа и TNF при пациенти с остър

коронарен синдром във връзка с наличието

или отсъствието на възпалително ставно

заболяване, тип заболяване на ставите

и активност на заболяването..................................32

Ангел М. ДжамбовПроучвания върху шума в околната среда

и здравето в България (1970 – 2017)....................41

М. Христамян, Й. Стоилова, П. Печалова, В. Христамян Анализ на познанието за епидемиологията

и клиниката на бисфосфонатно-асоциираната

остеонекроза на челюстите.....................................46

Обзори

Оригинални статии

Бъ

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ска м

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Българска медицина се реферира в международната база данни Index Copernicus International.

Content

Kakanakova A.Categorical vs dimensional models of classification

continuum of psychoses. ........................................... 4

Petya V. Ivanova, Ivica Dimov Crosstalk between different signal pathways and

Vitamin D and Vitamin D3 Receptor - VDR in the

process of oncogenesis in skin................................. 11

Getova D., Dimitrova D., Topolov M.

Comparison of the fat mass, some body circumfer-

ences (waist, hip and thigh) and the levels of choles-

terol and triglycerides in different groups of patients

included in NIRDIABO project from Plovdiv.............25

Popova V., Vazhеv Z., Peshev G., Grudeva-Popova Zh., Batalov A.

Analysis of the expression of the Tumor Necrosis

Factor alpha and TNF serum levels in patients with

acute coronary syndrome in relation to the presence

or absence of inflammatory joint disease, type of joint

disease and disease activity .....................................32

Dzhambov A. M.

Environmental noise and health research

in Bulgaria (1970 – 2017) ........................................41

Hristamyan M., Stoilova Y., Pechalova P., Hristamyan V.

Analysis of the knowledge of the epidemiology

and clinic of bisphosphonate-related

osteonecrosis of the jaws..........................................46

Reviews

Original articles

Bu

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Bulgarian medicine is included inIndex Copernicus International database.

4 Bulgarian medicine vol. 7 № 2/2017

Категориални срещу дименсионални модели на класификация. Континуум на психозите

А. Каканакова,МУ Пловдив, Катедра Психиатрия и медицинска психология

Categorical vs dimensional models of classification. Continuum of psychoses

Kakanakova A., Medical University Plovdiv, Department of Psychiatry and Medical psychology

Reviews

РЕЗЮМЕ:

Настояща практика за диагностика впсихиатрията е използване на синдромнобазирани системи на класификация напсихиатричните разстройства като петотоиздание на ДСМ (diagnostic and statisticalmanual of mental disorders) и десетотоиздание на МКБ (международна класифи-кация на болестите), в които на практикасе ползват симптоми и прояви за описател-но категоризиране на диагностични едини-ци. Основна цел на класификацията в пси-хиатрията е подобряване на лечението ипрофилактиката.

Използваната в днешни дни нозологич-на класификация в Европа и САЩ е кате-гориална и е наследник на дихотомнитекласификации на Крепелин и Блойлер.Дихотомният подход между шизофрения иразстройства на настроението има редицаограничения. В противовес на категориал-ния подход, възниква дименсионален,който описва по-добре комплексността ихетерогенността на разстройствата.Симптоми, общи за различни разстрой-

ABSTRACT

Contemporary practices of diagnostics inPsychiatry use syndrome based systems ofclassification of the psychiatric disorders asDSM (diagnostic and statistical manual ofmental disorders) 5th edition and ICD (inter-national classification of diseases) 10th edi-tion, which use symptoms and signs as a toolof descriptive categorization. The aim of suchclassifications is improvement of therapy andprevention.

These classification systems, used inEurope and the USA are categorical and suc-ceed Kraepelin and Bleuler’s dichotomousclassifications. The dichotomous approach,separating schizophrenia and mood disorders,is subject to numerous restrictions. A dimen-sional approach is created versus the categor-ical, which is better in covering the complexityand heterogeneity of the disorders.Symptoms common to different disordersproduce clinical dimensions.

In dimensional classifications the existenceof a continuum from normal to abnormal isassumed. Such concept stands for the exis-

Bulgarian medicine vol. 7 № 2/2017 5

INTRoDuCTIoN

Contemporary practices of diagnostics inPsychiatry use syndrome based systems ofclassification of the psychiatric disorders asDSM (diagnostic and statistical manual ofmental disorders) 5th edition and ICD (inter-national classification of diseases) 10th edi-tion, which use symptoms and signs as a toolof descriptive categorization.

Deciding on a specific diagnose has fewtasks, the main of them being reaching anexpectance of a predictable therapeutic effectand a prognosis of the course of the disorder,and even working towards better prevention.

Do we need a classification? Classificationis a process of simplifying complex phenome-na by grouping them in categories, accordingto predefined criteria. The aim of such classi-fications in psychiatry is improvement of ther-apy and prevention.

CLASSIFICATIoNS IN PSyChIATRy ThRouGh ThE AGES

The first review of psychiatric classificationssince ancient times has been done by CarlMenninger and his associates. They state thatthe first description of a psychiatric diseasehas been given as early as 3000 BC - princePtahhotep suffering of senile dementia. InSumer and Egyptian texts melancholic andhysteria syndromes have been describedabout 2600 BC. In Eber papyruses (1500 BC)senile dementia and alcoholism has beendescribed. In Ayurveda, 1400 BC, in India aclassification of psychiatric maladies has beenincluded.

Introducing the concept of psychiatric dis-orders in medicine is usually attributed toHippocrates (460–370 BC). He describesacute psychiatric states with or without fever,chronic psychiatric without fever (melan-choly), hysteria and Skit’s disease (probablytransvestism).

Mental retardation and dementia has beendescribed by the French renaissance healerFelix Platter (1536–1614).

Philippe Pinel (1745–1826), French physi-cian, simplifies the preceding complex diagnos-tic system to: melancholia, mania (insanity),dementia, and idiotism. He states that all psy-chiatric disorders are functional (without fever,inflammation, hemorrhage and anatomicaldamage) and so is a form of neurosis.

CoNCEPT oF ThE uNITARy PSyChoSIS

The concept of the unitary psychosis is firstproposed by the Belgian psychiatrist JosephGuislain (1797–1860). In his book „Traité DesPhrénopathies ou Doctrine Nouvelle desMaladies Mentales” in 1833 he describes acomplex system of classification of psychiatricdisorders [2]: there are four consecutivestages in the course of a psychiatric state: 1)mental activity exaltation, 2) brain structuralchanges, 3) brain structural damages, 4)mental exhaustion [3]. He claims that thebasis of every psychiatric state is the „phré-nalgie” or „mental suffering”[3]. Later thepsychiatric disorder would develop in sevensuccessive stages: mania, insanity, stupidity,epilepsy, hallucinations, confusion, anddementia [18].

ства, дават клинични дименсии.При дименсионалните класификационни

системи се приема наличието на континуумот норма до абнормност. Смята са, че кон-цепцията за съществуване на континуум/дименсионален подход, всъщност е в под-крепа на хипотезата за съществуване наедно заболяване. Концепцията за единнатапсихоза първи предлага белгийският пси-хиатър Joseph Guislain. През 20 век тезиидеи възражда и Klaus Conrad.

Ключови думи: категориален/димен-сионален подход, континуум на психозите

tence of a unitary disorder. The concept of theunitary psychosis first is proposed by theBelgian psychiatrist Joseph Guislain. In the20the century this ideas are resumed byKlaus Conrad.

Key words: categorical vs dimensionalapproach, continuum, psychosis


Guislain’s ideas have been adopted by theGerman psychiatrist Ernst Albrecht von Zeller(1804–1877). He states that different psychi-atric disorders are in fact separate stages of asingle morbid process [4]. Zeller followed theidea that soul and character exist as a spiritualunity, according to the contemporary believesin Germany, based on Naturphilosophie andanthropology, and it is the spiritual self that isaffected [15], and both psychological andmoral causes combine to produce the psychi-atric disorder [4]. The developed psychiatricstate is universal and goes through four con-secutive states: melancholia, mania, para-noia, dementia [4].

In the 19th century all psychiatric disordershave been widely accepted as a sign of under-lying somatic pathology, so according specificdamages have been searched for. First Morel(1809–1873) used the course of the diseaseas a basis for a classification and describesdementia praecox.

One of Zeller associates, WilhelmGriesinger (1817–1868), grasps the conceptof the unitary psychosis, even though he didnot follow Zeller’s ideas of the spiritual entity.Griesenger is one of the somaticist and one ofthe founders of materialist psychiatry [3]. Inhis opinion, psychiatric state is developedwhen „psychic reflex action” is either dimin-ished: melancholia or accelerated: mania[15]. As Zeller, he believes that the singleform of psychic state is melancholia, whichmight pass to mania and finally dementia [4].In his work „Mental Pathology andTherapeutics” in 1861, he proposes a simpli-fied classification of psychiatric disorders: onewith predominantly emotional disturbancesand other with predominantly intellectual andvolitional disturbances [32]. According to himthese two categories are two forms but alsotwo consecutive phases of a single morbidprocess [32]. He also states that „psychiatricdisorders are disorders of the brain”, it is justthat at that time there were not enough dataon brain damages so psychiatric disorders arelabeled functional or disorders with stillunknown somatic origin.

One of the most passionate supporters ofthe theory of the unitary psychosis is the 19thcentury German psychiatrist HeinrichNeumann (1814–88) [23]. In his „Lehrbuch

der Psychiatrie” in 1859 he evaluates everyclassification attempt in psychiatry as artificial[3] and states that there is only one type ofpsychiatric disorder, which he names‘„Irresein” and one should define stages, notforms as: insanity (Wahnsinn), confusion(Verwirrheit) and dementia (Blödsinn) [23].Neumann goes further and states that notonly psychiatric conditions exist as continuumdegrees, but there is a continuum of healthand illness and all conditions in-between [3].

One of Neumann’s critics is his coevalKahlbaum, who in 1863 publishes his book„Die Gruppierung der psychischenKrankheiten” (Classification of PsychiatricDisorders) [16]. Kahlbaum separates fourtypes of psychiatric disorders (vesania): vesa-nia acuta, vesania typica, vesania progressiva,vesania catatonica [5]. In his opinion themajor issues with declining any classificationof psychiatric disorders would be the futuredevelopment of the therapeutic approachesand studying the psychopathology in general[15]. He introduces the idea of (1) transientsymptom complex instead of an underlyingdisease, (2) differentiation between organicand functional mental disorder and (3) theimportance of the age of the patient at theonset of the disorder.

In the end of the 19th century, Kraepelin(1856–1926) unites manic and depressivestates as manic-depressive psychosis, whichdiffers from the chronic dementia praecox,named schizophrenia by Bleuler, becausethere are periods of remission. He also sepa-rates paranoia and dementia praecox, deliri-um and dementia and introduces the idea ofpsychogenic neuroses and psychopathies. Hisapproach is based on combining clinical fea-tures, unlike Bleuler (1857–1939) who takespsychopathological processes as a basis.

August Hoch (1868– 1919) is the first tocriticize the consideration of schizophrenia asa disorder. He differentiates disorder, syn-drome (symptom complex) and single symp-tom, and thinks that in fact dementia praecoxis characterized by the chaotic symptoms andthat precisely changes in the symptoms arelongitudinally specific to psychotic states [21],which are supported by other research in thefield in Europe [9].

Freud (1856–1939) introduces the concept


of neurosis/ psychoneurosis (anxiety, anxious-hysteric (phobia), obsessive compulsive, hys-teria).

In 20th century the idea of the unitarypsychosis has been revived numeroustimes, predominantly in attempt to declineKraepelin’s dichotomy [4].

Klaus Conrad (1905–1961), Germanneuropsychiatrist, is convinced that there isonly one endogenic psychosis, upon hisresearch on the manifestation of schizo-phrenia in the offspring of patients withaffective states. [3]. Based on his clinicalpractice, he postulates that such symptomsas hallucinations, delusions, catatonic statewhich are usually attributed to schizophre-nia, might appear in patients with maniaand depression as well and eventually leadto stabile personality changes [4]. His ideasare close to 19th century concept of theunitary psychosis [3; 4].

ThE DIChoToMouS APPRoACh

Although significant research takes placein the last hundred years, we still use basiccategories, proposed by Kraepelin andBleuler (organic disorders, affective disor-ders, schizophrenia) and Freud (neurosisand personality disorders).

Kraepelin is the first to propose thedichotomous approach, defining manic-depressive disorder and dementia praecox,based on the course of the disorder [24].

Bleuler proposes the nameSchizophrenia, aiming to emphasize thatpatients with the chronic course of the dis-order are not „demented”, and it is not nec-essary for it to have an early onset [6]. Heconsiders „schizophrenias” as a spectrum ofpsychoses possessing specific symptoms,with a schism in behavior and emotions.

According to Bleuler affective disordersare not that specific, so recognizing themshould follow exclusion of possible schizo-phrenic state.

Later Bonhoeffer’s works shows that thesame disorder can present with differentpsychopathologic symptoms, where theconflict between etiologic and syndromepsychiatric classification origins.

Another theory, that schizophrenia leads

to dementia and in manic-depressive disor-der there is possible remission, is dis-claimed as well. As early as 1909, one ofKraepelin’s associates makes a researchamong 468 dementia praecox cases inKraepelin’s hospital in Munich. In 29.8%there was remission, which supposedly wasdue to misdiagnosis. This early data is con-firmed by contemporary research on courseand development of schizophrenia in Huberet al. [19], Ciompi and Müller [9], Bleuler[7], Marneros et al. [25], Möller et al. [29]works.

It is expected that in the course of theaffective psychoses there is remission, butBumke objects the opposite [8]: above15% takes chronic course and in lot of thecases there are residual affective signsbetween episodes.

Now across Europe and the USA a noso-logical classification is used which appearsto be e successor of Kraepelin’s andBleuler’s dichotomy [1].

Separate disorders appear to be differentsigns and symptoms combinations. That ishow we differentiate Bipolar disorder andSchizophrenia. Such approaches have cer-tain advantages – high reliability and defi-ciencies – low validity.

The dichotomous approach has a numberof drawbacks. In reality lot of the patientssuffer from states that cover as affectivesymptoms and schizophrenic symptoms aswell. This naturally leads to the introducingof a separate category – Schizoaffective dis-order.

As counterbalance of the categoricalapproach comes the dimensional approach[10; 27; 28; 33], which surpasses the cate-gorical in describing complexness and het-erogeneity of the disorders. Symptoms com-mon to different disorders produce separatedimensions...

For instance psychotic, anxious symp-toms, impaired impulse control are observedin a number of disorders and in terms of thedimensional approach, new understandingand overcoming of the disorders are possi-ble. In fact human biology in general is com-plex and heterogenic, so dimensionalapproach is much more fit [22].


CoNTEMPoRARy CLASSIFICATIoN SySTEMS

The first predecessor of the DSM is pub-lished by APA (American PsychiatricAssociation) in 1844, as a statistical classifica-tion of institutionalized psychiatric inpatients.After World War Two, up to now there are fiveeditions published and some Text revisions.The first edition of DSM is published in 1952.

International Classification of Diseases(ICD) is the main alternative of DSM and isthe official nosological system used in Europeand the USA as well, because it considers notonly psychiatric conditions. DSM is widelyused in the USA as for psychiatric disordersand ICD in Europe and the rest of the world.

The last two editions of the DSM, DSM-IV-TR и DSM-V postulate that diagnoses have tobe based on presence or absence of specificsigns and symptoms, caused by different bio-logical factors. Diagnoses are categorical, anddefined.

Ever since 1974, the use of the term disor-der in DSM-IV-TR (diagnostic and statisticalmanual of mental disorders IV edition - textrevision), instead of syndrome or disease,supposes that these conditions are too com-plex to be defined as syndromes, and there isyet not enough data on etiology and patho-genesis, to be described as diseases. Thiscomes on J. Wakefields idea of psychiatric dis-orders as impairing dysfunctions. His conceptis based on quantitative data on manifestationof number of specific criteria out of the maxi-mal, which are signs and symptoms, evaluat-ed by structured clinical interview [35].

In DSM V there is an attempt of introducingdimensional approach but only as an alterna-tive of the widely accepted categorical one,addressing personality disorders.

DSM is a categorical classification system.In such classifications there might be morethan one diagnostic units, or comorbidity.Normal and abnormal are defined specifically.

In dimensional classifications there is con-tinuum of normal to abnormal and differentdegree of manifestation of the same symptomin patients with the same diagnosis.Accordingly there are less nosological units,and lesser comorbidity.

Dimensional models suppose predisposition.

Every patient should be evaluated with a specificscore on different dimensions (as personalitymodel of Eisenk and the dimensions: psychoti-cism, neuroticism, introvert/extrovert) [17].

Unfortunately the dimensional approach isnot fit for the everyday clinical practice,because it does not ease the decision on aspecific therapeutic plan [17].

NEW APPRoAChES

In the last decades researchers worktowards proposing alternative classificationmodels as the prototype model – the nosolog-ical units are more general compared to thetypical DSM and ICD categories, based onexclusionary criteria [36]; cluster model –grouping categories based on common signsand symptoms [36].

„Research Domain Criteria (RDoC)” projectis an initiative of the American Mental HealthInstitute, which opts for a biologically validat-ed classification of psychiatric disorders,based on new research methods in genetics,neurosciences, behavioral sciences. It isestablished in 2008, with predominantlyresearch goals and is based on the dimension-al approach concerning behavioral and neuro-biological parameters. In order to clarifyunderlying causes of the psychiatric disorders,defining and connecting the major biologicaland behavioral models in normal and abnor-mal state is needed. This would lead to thedefinition of valid and reliable phenotypes,with measurable characteristics.

Contemporary work on the ideas in„Research Domain Criteria (RDoC)” model, isthe Project on Translational Validity, which isbased on the strong belief that all data con-cerning psychopathology, usually derived bystructured clinical interviews, need to be vali-dated by comparing it to neuroimagingresults. Juxtaposition of clinical psychopathol-ogy data, from rating scales, and functionalneuroimaging results in real time is needed[14].

CoNCLuSIoNS

Nowadays the existence of SchizoaffectiveDisorder as a diagnostic unit in opposition ofKraepelin’s dichotomy supports the concept ofthe unitary psychosis, which even if it is out of


date is updated and allows the existence of thedifferent psychoses as degrees of a continuum[30]. A list of authors as Crow [11, 12] andTaylor [34] state that psychotic disorders asSchizophrenia (Sch), Schizoaffective disorder(SchA) and Bipolar disorder (BPD), are in facta continuum based on severity. Or they arecontinuum of severity, not a continuum ofdiagnoses. Psychotic features are just symp-toms, not a diagnosis. It is accepted that theconcept of a continuum/ dimensionalapproach, comes to support the hypothesis ofthe existence of a single disorder, with widespectrum of severity of symptomatology.

Lake and associates [25] emphasize on theproblem: does it really matter if the diagnosisis Sch/ SchA/ Psychotic BPD and if the exclu-sion of „neuroses” from DSM - III, is similar toeventually excluding well known diagnoses asSchizophrenia and Schizoaffective disorder,and finally uniting them with affective disor-ders in a spectrum of psychotic disorders,when there is not enough units. Psychotic fea-tures are criterial for Schizophrenia accordingto Bleuler, Schneider and the DSM, but onlycome to show the extent of severity of BPD,which is scientifically validated diagnostic unit.

Introducing the diagnostic unitSchizoaffective disorder creates the so impor-tant connection between Schizophrenia andAffective disorders in time when those twocategories were mutually excluding whichallows researchers to concentrate on compar-ing those categories.

The problem is that there is a continuum ofthe psychopathology from affective to schizo-phrenic symptoms. Mundt [31] in his review,says that Schneiders first rank symptoms,Huber’s basic symptoms, negative symptomsof Andreasen and Olsen, delusions ofChapman and psychophysiological reactionsof Hyman are non-specific. He concludes thatif the symptoms are non-specific, then onecannot consider single disorder but a spec-trum of the idiopathic psychoses. In 1995,Crow says: „Perhaps we should grasp thenettle and admit that the conditions wedescribe are in fact a continuum. There are nodefining signs, needed for the existence of aspecific diagnostic units...”, „and if there areno diagnostic units, we cannot separate a partof the psychopathological spectrum, we needto consider the whole range.” [13].

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36. Стоянов Д., 2016 Учебник по психиатрия застуденти по медицина под редакцията напроф. д-р В.Акабалиев, д.м.н., IV издание, 17.Психиатрична нозология: Класификация иНоменклатура, 81-84

Адрес за кореспонденция:

Д-Р АНДРИАНА КАКАНАКоВА

Катедра Психиатрия и Медицинска психология

Бул. В.Априлов, 15 АПловдив 4000

[email protected]

Corresponding author:

DR. ANDRIANA KAKANAKoVA

Department of Psychiatry andMedical PsychologyBul. V. Aprilov, 15 А

Plovdiv 4000andriana_kakanakova

@hotmail.com


Кростолк между различни сигнални пътищаи витамин Д и витамин Д3 рецептор - ВДР в процесът на онкогенеза в кожа

Петя В. Иванова, Ивица ДимовКатедра Химия и Биохимия, Фармацевтичен Факултет, Медицински Университет Пловдив

Crosstalk between different signal pathwaysand Vitamin D and Vitamin D3 Receptor -

VDR in the process of oncogenesis in skin.

Petya V. Ivanova, Ivica DimovDepartment Chemistry and Biochemistry, Pharmaceutical Faculty,

Medical University of Plovdiv

РЕЗЮМЕ:

ВДР (Витамин Д Рецетор, VDR) протеи-нът се експресира в почти всички нормал-ни човешки клетъчни типове и тъкани, исъщо в ракови клетъчни линии и тумори.Увеличената ВДР експресия се асоциира спо-високи нива на диференциация, липсана лимфни метастази, и добра прогноза вдебелочревен рак, с по-ниска туморна сте-пен, късно развитие на на метастази в лим-фните възли, и по-дълга преживяемост безрецидиви при рак на млечната жлеза, и сподобрена обща преживяемост при про-статен, немалък белодробен рак и мела-ном. Беше доказано че, метаболитите наВитамиин Д инхибират пролиферацията ииндуцират диференциация на меланомни-те клетки експресиращи ВДР. Реду -цираните нива или липсата на ВДР се асо-циират с меланомната прогресия (мелано-генезата може да супресира експресиятана рецепторът (лош прогностичен маркер),водещо до намалено оцеляване на мела-номните пациенти. Витамиин Д супресираключов туморен път в развитието на БЦК(BCCs) - Hedgehog сигналният път.

ABSTRACT

VDR protein is expressed in almost all nor-mal human cell types and tissues, and also incancer cell lines and tumors of several origins.Remarkably, elevated VDR expression is asso-ciated with high tumor differentiation,absence of node involvement, and good prog-nosis in colon cancer, with lower tumor grade,late development of lymph node metastases,and longer disease-free survival in breast can-cer, and with improved overall survival inprostate and non-small cell lung cancer andmelanoma. It was shown that the me¬tabo-lites of vitamin D inhibit proliferation andinduce differentiation of melanoma cellsexpressing VDR. Reduced level or absence ofVDR is associated with melanoma pro¬gres-sion (melanogenesis can suppress theexpression of the receptor (bad prognosticmarker), resulting in deteriorated survival ofmelanoma patients. vitamin D suppresses akey tumour pathway in BCCs development -Hedgehog signalling pathway. 1a,25(OH)2D3also inhibits the growth of SCCs in vivo as wellas in vitro, inhibiting Wnt/β-catenin and inter-feres in ERK1/2 signaling, connected with


MELANoMA

Nowadays the most effective treatment formelanoma is surgical removal of the neoplasticlesions. Therefore, early detection of melano-ma in patient increases the chances for suc-cessful curing of this disease. The significantmortality is caused by the ineffectiveness ofstandard treatment methods, particularlydur¬ing the stage of metastasis. Notableadvance in the treatment of this type of cancerwas the introduction of targeted therapy withinhibitors for tyrosine kinase receptor KIT.Recent developments, including BRAF-inhibi-tors, as well as drugs targeting the MAPK path-way, bring new perspectives to the field, espe-cially when used as a combined immunothera-py [4].

The RAS/RAF/MEK/ERK pathway has beenreported to be activated in over 80% of allcutaneous melanomas, making it the focus ofmany scientific studies in the melanoma field.Discoveries of mutations and aberrant expre-ssion of components in this cascade, in parti-cular, BRAF and NRAS render a deeper under-

standing of the mechanisms responsible foroncogenesis and provide new therapeutic stra-tegies for this deadly disease. Mutations in B-raf are found in 67% of melanomas [16], 50% according Ascierto PA et al., 2012, activa-ting mutations in the TERT promoter wererecently identified in up to 71% of cutaneusmelanoma [7].

In fact, approximately 50% of melanomasharbor activating BRAF mutations. Among theBRAF mutations observed in melanoma, over90% are at codon 600, and among these, over90% are a single nucleotide mutation resultingin substitution of glutamic acid for valine(BRAFV600E: nucleotide 1799 T>A; codonGTG>GAG). The second most common muta-tion is BRAFV600K substituting lysine for vali-ne, that represents 5-6 % (GTG>AAG), follo-wed by BRAFV600R (GTG >AGG), an infre-quent two-nucleotide variation of the predomi-nant mutation, BRAF V600′ E2′ (GTG>GAA),and BRAF V600D (GTG>GAT). The prevalenceof BRAFV600K has been reported as higher insome populations. In melanoma, BRAF muta-tion is most common in patients whose tumors

1a,25(OH)2D3 инхибира също растежът наСЦК (SCC) in vivo както и in vitro, инхиби-райки Wnt/β-катенин и повлиявайкиERK1/2 сигналът, свързан с развитието намеланом. Въпреки че изследванията, invitro и в животински модели, предполагатче vitamin D може да предотврати разви-тието на БЦК и СЦК, допълнителни изслед-вания са необходими за да се оцени ефек-тът от топикалното или системно (орално)използване на Витамин D3 в профилакти-ката на ракът в лицево-челюстната област(кожен рак) при хора. Използването на1,25(OH)2D3 при ракови пациенти се огра-ничава от хиперкалцемичният ефект навитаминът в терапевтични дози, довело доразвитието на няколко аналози, притежа-ващи антитуморна активност, но проявява-щи по-слабо калцемично действие, вклю-чително и при пациенти с панкреатиченрак, орган смятан за нетаргетен за дей-ствието на витаминът.

Ключови думи: 1a,25(OH)2D3, ВДР,БЦК, СЦК, меланом, Кожен рак

development of melanoma. Although the invitro and animal studies suggested that vita-min D may prevent development of BCCs andSCCs, additional studies on humans are need-ed to assess the suitability of topical or oralvitamin D3 supplementation in chemopreven-tion of head and neck skin cancers.Administration of 1,25(OH)2D3 to cancerpatients is restricted by its hypercalcemiceffects at the therapeutic doses, enforcing thedevelopment of several analogs that maintainthe antitumoral properties but have less cal-cemic actions, including pancreatic cancerpatients, organ which is thought not to be tar-get for vitamin D action.

Keywords: 1α,25(OH)2D3, VDR, BCC,SCC, melanoma, Skin cancer


arise on skin without chronic sun-induceddamage, whereas BRAF mutations are rare inmelanomas arising from mucosal and acralsites [1].

Several RAF mutations have been implica-ted in the induction of genomic instability, dri-ving the proliferation of cancer cells with thehighest frequency in melanoma. For instance,mutated BRAF signals as a monomer, indepen-dent of upstream growth stimuli. The most fre-quent BRAF mutation, BRAFV600E, causesconstitutive activation of the kinase as well asinsensitivity to negative feedback mecha-nisms. BRAFV600E has been implicated in dif-ferent mechanisms of melanoma progression,and principally the activation of the downstre-am MEK/ERK pathway, evasion of senescenceand apoptosis, unchecked replicative potential,angiogenesis (through MEK-dependent activa-tion of HIF-1a and VEGF), tissue invasion andmetastasis (via upregulation of several prote-ins involved in migration, integrin signaling,cell contractility, tumor - and microenviron-ment-derived interleukin-8), as well as theevasion of immune response [1].

In melanoma, BRAF mutation is most com-mon in patients whose tumors arise on skinwithout chronic sun-induced damage, whereasBRAF mutations are rare in melanomas arisingfrom mucosal and acral sites [1].

No clear differences in prognosis (time fromprimary diagnosis to distant metastasis) werenoted between BRAF-mutated versus wild-type melanomas. Features of the antecedentprimary melanoma significantly associatedwith a BRAF mutation (P<0.05) were thesuperficial spreading and nodular histopatholo-gical subtypes, the presence of mitoses, thepresence of a single or occult primary melano-ma, a truncal location and age at diagnosis ofthe primary tumor (≤50 years) [1].

Mechanistically, mutated BRAF and NRASexert most of the oncogenic effects throughthe activation of the MAPK pathway, whichboth drive the uncontrolled growth of melano-ma cells and regulate the cell survival. In asubsequent section, clinical efficacy of targetedsmall-molecule inhibitors is highlighted. BRAF-targeted therapies (e.g., vemurafenib, dabra-fenib) have showed impressive results in sys-temic therapy for melanoma harboring activa-ting BRAF V600E mutations (most frequent

BRAF mutation [1]. MEK inhibitors show limitedactivity in phase I trials, and inhibitors directlytargeting mutated NRAS, to date, have notbeen realized. Furthermore, the emergingmechanisms underlying both intrinsic andacquired drug resistance as well as approachesto prevent or abrogate the onset of therapeuticescape are addressed [25]. Nevertheless, see-king for new drugs is still required, since duringthe treatment many tumours acquire resistan-ce to the therapies [20].

Studies conducted in the 70’s showed, thatvitamin D can stimulate the activity of tyrosi-nase, the principal enzyme involved in melaninsynthesis in cultured mel¬anoma cell line.Following studies confirmed the presence ofthe vitamin D receptor (VDR) in the primarymelanoma tissue. Having in mind its antiproli-ferative properties, vitamin D and its analogu-es are potential candidates for melanoma tre-atment. In fact, it was shown that the metabo-lites of vitamin D inhibit proliferation and indu-ce differentiation of melanoma cells expressingVDR. Interestingly, it seems that not transfor-med melanocytes and keratinocytes are pro-tected by 1,25(OH)2D3 and its analogues.There is also evidence, that 1α,25(OH)2D3promotes survival of the cells and inhibits thetumour invasion and angiogenesis. In theGupta’s study, keratinocytes after the treat-ment with 1a,25(OH)2D3 and exposure to UVradiation were characterized by the increasedsurvival and reduced amount of DNA damage.Similar protective effect was observed in mela-noma cells. Furthermore, VDR was shown totake part in activation of DNA repair mecha-nism. Thus, it seems that vitamin D is animportant, physiologically relevant factor pre-venting UV-induced carcinogenesis. It is notsurprising, that a number of recent studiespointed out that the development of melano-ma can be linked with vitamin D deficiency ordefects in vitamin D signalling pathway. Inhumans, VDR gene has multiple splicing vari-ants, which are likely to affect protein expre-ssion and activity of the VDR. In addition, over1000 polymorphic sites have been detected inVDR gene, some of which have been correla-ted with increased risk of melanoma occurren-ce, its aggressiveness and prognosis, as wellas with other pathologies [20].

Reduced level or absence of VDR is associa-


ted with melanoma progression (melanogene-sis can suppress the expression of the recep-tor), resulting in deteriorated survival of mela-noma patients. Recent studies also providedevidence that vitamin D and its analogues maybe effective in melanoma treatment, especiallyrecently tested low-calcemic analogues suchas 20-OH D3 or analogues with pregnenolone-like side chain (21-hydroxypregnacalciferol)[20].

ERK1/2 SIGNALING PAThWAy

Study on breast carcinoma showed thatcalcitriol inhibits Src tyrosine kinase in Ras/ Rafkinase cascade, thereby leading to subsequentdecrease in activity of ERK 1/ 2 and MAP kina-se, this in turn reduces the formation of cyclin/cdk proteins, thereby leading to inhibition ofcell proliferation and alteration of differentiati-on in G1 and other phases of the cell cycle(MAP kinase plays an important role in cell pro-liferation and differentiation). Bernardi et al.have also investigated the action of calcitriol onVEGF-induced TDEC (tumor-derived endothe-lial cells) proliferation and found the vitamin toreduce phospho-ERK 1/ 2 and phospho-Aktlevels in them. Such observation shows thatcalcitriol interferes with both the proliferation -transduction pathways induced by RTKs, the-reby inhibiting angiogenesis [3].

The prognostic role of VDR expression or itsrelationship with PIK3CA or KRAS mutation isuncertain. Among 619 colorectal cancers intwo prospective cohort studies, 233 (38%)tumors showed VDR overexpression by immu-nohistochemistry, which significantly associa-ted with KRAS mutation (odds ratio, 1.55;95% confidence interval, 1.11-2.16) andPIK3CA mutation (odds ratio, 2.17; 95% con-fidence interval, 1.36-3.47). These data sup-port potential interactions between the VDR,RAS-MAPK and PI3K-AKT pathways, and pos-sible influence by KRAS or PIK3CA mutation ontherapy or chemoprevention targeting VDR[13].

Gilad LA et al. [6] previously demonstratedthat 17β-estradiol (E2) regulates the transcrip-tion and expression of the vitamin D receptor(VDR) in rat colonocytes and duodenocytes invivo. E2 has been shown to increase the num-ber of VDR in the osteoblastlike cell line ROS

17/2·8, an increase associated with enhancedresponsiveness of the cells to 1,25(OH)2D3.Increased VDR expression following E2 treat-ment has also been noted in other tissues andcell types, such as the uterus, liver, and humanbreast cancer cells. They compared E2-asso-ciated signaling activity in HT29 colon cancercells, a non-classical E2-target, with that inMCF-7 breast cancer cells, the natural targetsof the hormone. E2 did not affect proliferationof HT29 cells, but enhanced proliferation ofMCF-7 cells. Vitamin D inhibited proliferation ofboth cell lines and the combined treatmentinduced potentiation of vitamin D activity. E2upregulated VDR transcription and proteinexpression concomitantly with ERK 1/2 phosp-horylation in both cell lines. PD98059, a speci-fic mitogen-activated protein kinase (MAPK)inhibitor, prevented E2-mediated activation ofERK 1/2, with concomitant inhibition of VDRexpression. Rapid activation of mitogen-activa-ted protein kinase (MAPK) by E2 in ROS 17/2·8cells has provided the first evidence of MAPKactivation by E2 through phosphorylation, indi-cating the involvement of putative plasmamembrane receptors. Rapid effects exerted byE2 on growth-factor related signaling pathwa-ys have also been demonstrated in neuronalcells, suggesting a potential mechanism bywhich E2 might affect the expression of geneswith promoters that do not contain strictly est-rogen-responsive elements but are responsiveto factors acting through other response ele-ments, such as activation protein-1 (AP-1) andserum response elements [6].

ICI182780 (ER-specific inhibitor) inhibitedVDR expression in HT29 and MCF-7 cell lines.A conjugate of E2 and bovine serum albuminupregulated phosphorylation of ERK 1/2 andconcomitantly enhanced VDR expression in asimilar fashion as the nonconjugated hormo-ne. Expression of ERα and ERβ was detected inMCF-7 and HT29 cell lines respectively, whichlocalized to the nuclei, cytosol and caveolarmembrane rather than non-caveolar membra-ne. Disruption of lipid rafts/caveolae by deple-ting cellular cholesterol with the cholesterol-binding reagent β-methylcyclodextrin blockedERK 1/2 phosphorylation concomitantly withVDR upregulation. The tyrosine phosphorylati-on inhibitor suramin and src kinase inhibitorPP2 inhibited both ERK 1/2 phosphorylation


and VDR expression. E2 induced phosphoryla-tion of Raf and Jun in a time-dependent man-ner. The Ras/Raf dependent inhibitor of tran-sactivation sulindac sulfide also blocked E2effects. The specific c-Jun phosphorylationinhibitor SP600125 dose dependently inhibitedc-Jun phosphorylation and VDR expression.The MAPK/ERK kinase inhibitor PD 98059downregulated both c-Jun phosphorylation andVDR expression indicating that upstream anddownstream events in the signaling cascadeare all related to the control of VDR expression,through AP-1 binding site on the VDR promo-tor. Taken together, the authors’ data suggestthat E2 binds to receptors (ERa or ERβ with dif-ferent tissue distribution) compartmentalizedto membranal caveolar domains in HT29 andMCF-7 cells, tyrosine phosphorylation activitytriggers the Ras-ERK pathway, which ultimate-ly can activate transcription factors such as c-Jun or c-Fos to bind specific sequences withinthe VDR gene (such as the AP-1 binding site)and finally induce upregulation of transcriptionand expression of the VDR gene [6].

For better understanding the functionalinteractions between VDR and the ERK signa-ling pathway, Narayanan R et al. [17] soughtto determine whether 1,25-D activates ERK inthe osteoblastic cell lines, MG-63 (humanosteosarcoma) and MC3T3-E1 (osteoblasticcell line from a C57BL/6 mouse calvaria withhigh alkaline phosphatase (ALP) activity in theresting state), and to evaluate the effects ofERK on VDR activity. The authors found that1,25-D rapidly induced ERK activity and thatthis activation persisted at 24 h in both celllines. Surprisingly, the effects of ERK activationon VDR activity in the two cell lines were verydifferent. Overexpression of Raf-1 (an upstre-am activator of ERK) reduced VDR activity inMC3T3-E1 cells, but stimulated activity in MG-63 cells. Similarly, inhibition of ERK by the MEKinhibitor U0126 stimulated VDR activity inMC3T3-E1 cells. However, it inhibited VDR acti-vity in MG-63 cells as well as in HeLa cells, acervical carcinoma cell line commonly utilizedto study the functions of nuclear receptors.Despite this difference, U0126 treatmentenhanced expression of VDR in both cell lines.Although previous studies in keratinocytessuggested a potential for differential coactiva-tor usage as a function of differentiation, and

coactivators are phosphoproteins, more highlydifferentiated MC3T3-E1 cells responded simi-larly to the undifferentiated cells.Supplementation with either SRC-1 orDRIP205, two VDR coactivators, had no effecton the response to the ERK pathway [17].

The primary effect of U0126 in MC3T3-E1cells appears to be enhancement of nuclearlocalization and DNA binding. The increase inDNA binding in MC3T3-E1 cells may, in part, bedue to increased nuclear levels of VDR, but thebinding is not strictly proportional to VDRexpression as 1,25-D treatment alone was lesseffective than U0126 in increasing DNA bindingdespite the higher levels of VDR. The lack ofcorrelation between nuclear VDR levels andDNA binding suggested that the VDR heterodi-mer partner, RXR, might be limiting. An analy-sis of RXR isoform expression levels revealedthat MC3T3-E1 cells had lower levels of RXRβand RXRγ than did HeLa or MG-63 cells, bothof which are cancer cell lines, suggesting thatin MC3T3-E1 cells the VDR may be moredependent upon RXRα.Previous studies haveshown that RXRα is a substrate for ERK andthere is evidence that phosphorylation of thissite reduces the effectiveness of RXRα as aVDR partner. Ras-transformed keratinocytesare resistant to 1,25-D-mediated differentiati-on, and this resistance is caused by the phosp-horylation of Ser260 of RXR. Mutation of thisserine to alanine eliminated the resistance.Interestingly, the response of TR, which alsoforms heterodimers with RXR, was not diffe-rentially affected by ERK signaling in the twocell lines. Consistent with the correlation of thelevels of RXR isoforms, elevated expression ofRXRγ eliminated the U0126 stimulation of VDRactivity. Thus, although coactivator functionmay be modified by ERK signaling, the domi-nant determinant of the effect of ERK signalingon VDR function is its RXR partner. In cells inwhich RXRα predominates, the elevation ofERK signaling by 1,25-D likely blunts the VDRtranscriptional responses whereas in cells con-taining higher levels of the other isoforms, theenhanced ERK signaling stimulates VDR activi-ty. The RXR isoform-dependent response indi-cates that depending upon the cellular milieu,treatment with 1,25-D will activate ERK withlittle activation of the transcriptional activity ofVDR (RXRα dominant cells) whereas in other


cells (RXRβ or RXRγ dominant), 1,25-D poten-tiates VDR activity both through its action as aligand as well as through activation of ERK.ERK activity is often associated with cellgrowth, whereas 1,25-D acting through VDRoften inhibits growth and induces differentiati-on, although 1,25-D stimulates proliferation insome cell types. Thus, the relative abundanceof RXR may play a role in determining theextent to which 1,25-D alters proliferation. incells in which RXRa is the dominant VDR par-tner, activation of ERK by 1,25-D reduces theactivity of VDR, whereas in cells utilizing RXRβor RXRγ, the activation of ERK enhances theactivity of VDR [17].

IN SKIN

Inflammation, elicited in the skin followingtissue damage or pathogen invasion, maybecome chronic with deleterious consequen-ces. Tumor necrosis factor (TNF) is a keymediator of cutaneous inflammation and thekeratinocyte an important protagonist of skinimmunity. Calcitriol (v.D3, 1,25(OH)2D3) andits analogs attenuate epidermal inflammationand inhibit the hyperproliferation of keratinocy-tes associated with the inflammatory disorder,psoriasis. Since activation of extracellular sig-nal-regulated kinase (ERK) promotes keratino-cyte proliferation and mediates epidermalinflammation, the effect of calcitriol on ERKactivation in HaCaT keratinocytes exposed tothe ubiquitous inflammatory cytokine TNF wasstudied. Ziv E. et al.established that TNF acti-vated ERK in an EGFR and Src dependent andan EGFR and Src independent modes. EGFRdependent activation resulted in the upregula-tion of the transcription factor, c-Fos, while theEGFR independent activation mode was of ashorter duration, did not affect c-Fos expressi-on but induced IL-8 mRNA expression.Calcitriol, enhanced TNF-induced EGFR-Srcdependent ERK activation and tyrosine phosp-horylation of the EGFR, but abolished theEGFR-Src independent ERK activation. Theseeffects were mirrored by enhancement of c-Fos and inhibition of IL-8 induction by TNF.Treatment with calcitriol increased the rate ofthe de-phosphorylation of activated ERK, acco-unting for the inhibition of EGFR-Src indepen-dent ERK activation by TNF. It is possible that

effects on the ERK cascade contribute to theeffects of calcitriol and its synthetic analogs oncutaneous inflammation and keratinocyte pro-liferation [30].

Cystatin A, a cysteine proteinase inhibitor,belongs to the cystatin superfamily, is a corni-fied cell envelope constituent and a differentia-tion marker of keratinocytes. In the epidermiscystatin A is expressed in the upper spinous togranular cell layers. 1,25(OH)2D3 suppressedNHK proliferation in a dose-dependent mannerwith the maximal effect at 1×10-7 M. It alsostimulated cystatin A promoter activity and itsexpression with similar dose effects. Theincreased cystatin A was detected by 24 h andthe effect was accompanied by the suppressi-on of ERK activity. The increased cystatin Awas detected by 24 h and the effect wasaccompanied by the suppression of ERK activi-ty. Cystatin A promoter activity was not affec-ted by cotransfection of vitamin D3 receptor orretinoid X receptor. Further analyses disclosedthat the 12-o-tetradecanoylphorbol-13-aceta-te (TPA)-responsive element (TRE), T2 (−272to −278), in cystatin A promoter is critical forthe regulation by 1,25(OH)2D3. Transfectionof the dominant-negative form of ERK adeno-virus (Ad-dnERK) increased cystatin A promo-ter activity and its expression, which was mar-kedly augmented by 1,25(OH)2D3 treatment.Transfection of the dominant-active form ofRaf-1 (Ad-daRaf-1) or MEK1 (Ad-daMEK1)inhibited 1,25(OH)2D3-dependent cystatin Apromoter activity and its expression.Consistent with these results, the MEK1 inhibi-tor, PD98059, further augmented1,25(OH)2D3-induced cystatin A promoteractivity and its expression. This study demon-strated that the 1,25(OH)2D3-responsive ele-ment in the cystatin A gene is identical to theTRE, T2 (−272 to −278), and that the suppre-ssion of Raf-1/MEK1/ERK1,2 signaling path-way increases cystatin A expression of NHK[11]. Additionally, cystatin A is positively regu-lated via the Ras/MEKK1/MKK7/JNK signaltransduction pathway. In contrast, transfectionof dominant negative forms of MKK3, MKK4, orp38 did not affect cystatin A promoter activity[22]. The AP-1 proteins, c-Jun and c-Fos, orby c-Jun and JunD, regulate cystatin A promo-ter activity probably via PKCα activation [21].Loss-of-function mutations in the gene for pro-


tease inhibitor cystatin A (CSTA) result in skinfragility due to impaired cell-cell adhesion:autosomal-recessive exfoliative ichthyosis oracral peeling skin syndrome [8] and is theunderlying genetic cause of exfoliative ichthyo-sis [15]. Electron microscopy of skin biopsiesfrom affected individuals revealed that thelevel of detachment occurs in the basal andlower suprabasal layers. Immunostaining aga-inst keratin 14 and E-cadherin showed a wide-ning of intercellular spaces and major disorga-nization of keratinocytes in the lower supraba-sal and basal layers [2]. siRNA knockdown ofCSTA resulted in cytoplasmic localization ofDsg2 (desmoglein 2), perturbed cytokeratin14 staining and reduced levels of desmoplakinin response to mechanical stretching. CSTA isderegulated in several skin cancers, includingsquamous cell carcinomas (SCC) and loss offunction mutations lead to recessive skin fragi-lity disorders [8].

We could not find data concerning CystatinD expression in human epidermis (keratinocy-tes), influencing E-cadherin expression andEMT. Cystatin M/E (CST6) is another memberof cystatin family, a nonredundant, epithelium-specific protease inhibitor with a presumed rolein epidermal differentiation and tumor suppre-ssion. Zeeuwen PL et al., have previouslyreported that cystatin M/E deficiency in Cst6(-/-) mice causes neonatal lethality because ofexcessive transepidermal water loss. Theabsence of cystatin M/E in adult animals cau-sed scarring alopecia. Biochemical evidencesuggests that cystatin M/E controls the activityof legumain, cathepsin L, cathepsin V, andtransglutaminase-3. The authors concludedthat a tightly regulated balance between cat-hepsin L and cystatin M/E is essential for tissueintegrity in epidermis, hair follicles, and cornealepithelium [28].

Furthermore, Verdier-Sevrain S et al. havedemonstrated that keratinocytes express bothestrogen receptor ER-a and ER-β. At physiolo-gical concentrations, estradiol up-regulates thelevel of ER-a receptors in keratinocytes andinduces proliferation in neonatal keratinocyte[24]. E2 increased protein and mRNA levels ofcyclin D2, and resultantly enhanced assemblyand kinase activities of cyclin D2-cyclin-depen-dent kinases 4 or 6 complexesin human kera-tinocytes. 17β-estradiol (E2) enhanced cyclin

D2 promoter activity, via CREB phosphorylati-on by protein kinase A, dependent on cAMP.These effects of E2 may be mediated via cellsurface GPR30 (G-protein-coupled receptor)[11]. It is known that in human keratinocytesERK1/2 induces keratinocytes proliferation,and increases expression of cyclin B1, but notthat of cyclin D [5].

The ER-a agonist, TGF β1 dependentlyincreases fibroblast migration and keratinocyteproliferation. By contrast, the ER β agonistdoes not affect cell migration and increaseskeratinocyte proliferation in a TGF β1 indepen-dent manner. Accordingly, in vivo experimentshave shown that targeting ER β, but not ER αleads to accelerated re epithelialization in miceand rats. Furthermore, ER α has been provento be responsible for impaired wound healingin male mice. Perzelova V et al. demonstratedthat the pharmacological activation of ER β,but not that of ER α, in HaCaT keratinocytes,expressed both ER α and –β, led to an increasein cell proliferation and keratin 19 expression,characteristic which exemplifies the stem celllike character of keratinocytes. In relation tomarkers, their previous research has demon-strated that the ER β agonist does not induceSox 2 expression, a character¬istic of stemlike properties, in keratin 19 positive cells [19].

Typical of the type II nuclear receptors,1,25(OH)2D3-induced VDR activation inhibitedproliferation and promoted differentiation ofkeratinocytes, though VDR/RXRα heterodi-mers, which can transactivate keratinocyticgenes independent of 1,25(OH)2D3 binding[10].

RXRa, the primary isoform in skin and hairfollicles, has stronger expression levels compa-red to RARs, and RXRa/RARγ heterodimerappears to be the major retinoid transducingelement in epidermal biology. RXRa/RARγheterodimerization is also critical in the deve-lopment and formation of epidermal lamellargranules by repression of target genes, asRARγ agonists promote lamellar granuledefects in murine skin. Similarly,RXRa/PPARβ/δ heterodimers are equallyimportant to stratum corneum homeostasisthrough activation of gene transcription [10].

Mice with an epidermal-specific ablation ofRXRα (RXRαep−/−) presented epidermalhyperplasia, keratinocyte hyperproliferation


and aberrant terminal differentiation, alopecia,dermal cysts and a cutaneous inflammatoryresponse [10,15]. Likewise, mice lacking bothRXRα and RXRβ in keratinocytes (RXRaβep−/−)developed a chronic dermatitis similar tohuman AD (Atopic dermatitis) patients, eleva-ted serum IgE/IgG and cytokine productionassociated with Th2-type response.Importantly, thymic stromal lymphopoietin(TSLP) was strongly upregulated from thebasal keratinocytes, potentially influencing thesystemic AD phenotype in these mice. To fur-ther support the hypothesis that loss of RXRαcontributed to a derepressive mechanism ongene expression leading to inflammatory res-ponses, expression of RXRα has been reportedto decrease in human psoriatic lesions, withlevels in progressive disease further reducedcompared to stable stages [10].

The transcription factor KLF4 is required forproper EPB formation in mice and dysregula-ted KLF4 activity is shown to be oncogenic.KLF4 regulates expression of RXRa and KLF4-induced malignant transformation is sensitiveto retinoids in vitro. Importantly, rexinoid app-lication drastically prevented formation of SCCin a KLF4-activated transgenic mouse line.These results suggest existence of a crosstalkbetween RXR and KLF4 signaling, where KLF4-mediated expression of RXR contributes totumor suppressor activity within the epider-mis. RXRαep−/− mice, selectively lacking RXRαin epidermal keratinocytes, subjected to a7,12-dimethylbenz(a)anthracene (DMBA)/12-O-tetradecanolyphoral-13-acetate (TPA) indu-ced two-step chemical carcinogenic protocoldeveloped higher numbers of epidermaltumors compared to control mice. TheRXRαep−/− papillomas progressed towardsSCC in a murine model, further validating therole of RXRa as a cutaneous tumor suppressor[10].

Type II NR-mediated signaling via RXRdimerization is essential to melanocyte biology.RXRa and β expression has been detected inB16 and S91 murine melanoma cells, withRXRβ being the predominant isoform.Interestingly, loss of melanocytic RXRα expre-ssion was seen in human primary and metas-tatic melanoma compared to benign nevi, indi-cating the importance of this signaling path-way to the differentiation of these cell types.

RARβ expression was also downregulated inmelanoma and its RA-induced expression waslinked to growth inhibition and differentiationin these cells. Sequential occupation of the RAresponse element located on the Rarβ promo-ter by RXR/RXR and RXR/RAR combinations isbelieved to be a molecular switch responsiblefor Rarβ transcriptional activation in thesecells. Since ligand activation of RXR heterodi-meric partners regulates transcriptional activi-ty of target genes involved with differentiationand growth arrest, it is possible that combina-torial activation of both dimer partners mayassist in melanoma therapy [10].

Interestingly, in a two-step carcinogenesismodel, RXRaep−/− mice developed a highernumber of dermal melanocytic growths (nevi)compared to control mice, implicating contri-bution of keratinocyte-derived factors in mela-nomagenesis. Only nevi from RXRα mutantmice progressed to melanoma-like tumors,suggesting that RXRa-mediated distinct non-cell autonomous actions suppressed nevi for-mation and melanoma progression in mice[10].

Similarly, VDR−/− mice undergoing identicaltreatments also developed higher numbers ofmelanocytic lesions, indicating RXRα/VDRheterodimerization may be the causative fac-tors, at least in part, in these non-cell autono-mous events. Finally, the loss of keratinocyticRXRα alongside an activated-CDK4 mutationenhanced the metastatic transformation ofcutaneous melanoma after chemical carcino-genesis. Loss of epidermal RXRα was also seenin human melanoma progression and couldpotentially be utilized as a therapeutic biomar-ker [10].

NoN-MELANoMA SKIN CANCERS

Similarly as in case of melanomas, thereare number of studies concerning the role ofvitamin D in basal cell carcinoma (BCC) andsquamous cell carcinoma (SCC). Just like kera-tinocytes, BCC cells also express VDR, and fur-thermore, peripheral cells forming BCC tumo-urs show even higher expression of VDR thanneighbouring, un¬affected epidermal cells. Itwas demonstrated that vitamin D sup¬pressesa key tumour pathway in BCCs development -Hedgehog signalling pathway. It was shown


that VDR-knockout mice, after the exposure toa carcinogen, were more prone to BCCs skintumours development than the wild type ani-mals. Another studies on mice showed thattopical application of vitamin D3 reduces BCCcell proliferation and also inhibits theHedgehog signalling pathway, both in vitro andin vivo. 1α,25(OH)2D3 also inhibits the growthof SCCs in vivo as well as in vitro. Similar stu-dies on animals showed that mice lacking VDRand exposed to high and prolonged doses ofUVB are predisposed to SCC tumour formati-on. In addition, as in BCCs, topically applied1a,25(OH)2D3 inhibits formation of chemical-ly induced tumour in a dose-dependent man-ner [20].

What is more, 20(OH)D3, 20,22(OH)D3and 20,23(OH)D3, novel vitamin D33 analogu-es produced by P450scc, show pro-differentia-tion, anti-proliferative and anticancer properti-es. Although the in vitro and animal studiessuggested that vitamin D may prevent deve-lopment of BCCs and SCCs, additional studieson humans are needed to assess the suitabilityof topical or oral vitamin D3 supplementa¬tionin chemoprevention of non-melanoma skincancers [20].

hEDGEhoG SIGNALING PAThWAy

In humans, the hedgehog (Hh) genes werefound to be an essential developmental signa-ling pathway in maintaining tissue polarity andstem cell population. Inactivation or hyperac-tivation of this pathway can cause serioushealth problems, including developmentaldefects such as holoprosencephaly, and diffe-rent forms of cancer including Basal CellCarcinomas (BCCs), medulloblastomas, leuke-mia, gastrointestinal, prostate, ovarian, breastand lung cancers. The receptor for Hhs is atransmembrane protein called Patched (PTC,Ptch). In the absence of Hh ligands, PTC isbound to another transmembrane protein,smoothened (SMO), and functions as an inhi-bitor of SMO. The binding of Hh ligands to PTCreleases SMO from the inhibitory effect of PTCand allows SMO to transduce signals leading tothe activation of transcription factor, calledglioma associated (Gli), and the expression ofgenes involved in regulating embryonic andpostnatal development, and the transformati-

on of cancer- and metastasis-initiating cells[4].

In the absence of Hh ligands (A), Ptch1receptor at the base of the primary cilium inhi-bits the function of SMO by preventing its entryinto the cilium. Gli3 and, to a lesser extentGli2, are converted to C-terminally truncatedrepressor forms (Gli-R) and translocate intothe nucleus, where they inhibit the transcripti-on of Hh target genes. Formation of Gli-R ispromoted by sequential phosphorylation of fulllength Gli by a kinase cascade that includesPKA (Protein Kinase A), GSK-3β (GlycogenSynthase Kinase 3β), and CK1 (Casein Kinase1), which creates binding sites for the adapterprotein β-TrCP (β-transducin repeat-containingprotein). The Gli/β-TrCP complex is ubiquitina-ted by Cul1-based E3 ligase, which results inpartial Gli degradation by the proteasome andformation of Gli-R. In addition to partial degra-dation, full-length Gli may be completely deg-raded by the proteasome through SPOP(Speckle-type POZ Protein)-mediated Cul3-based E3 ligase ubiquitination. Upon Hh ligandbinding (B), Ptch is displaced from the cilium,becomes internalized in endosomes, and deg-raded. SMO relocates from intracellular vesic-les to the cilium. Active SMO promotes a sig-naling cascade that leads to translocation ofactivated forms of Gli (Gli-A) into the nucleus,where they induce the transcription of Hh tar-get genes, such as Gli1, Ptch1, and HHIP (Hh-interacting protein). HHIP competes with bin-ding of the Hh ligands, while the GPI-linkedGas1 (Growth Arrest-Specific protein 1) andthe Ig/Fn repeat-containing surface proteinsCdo and Boc (brother of Cdo) act as corecep-tors of Hh [12,18].

Recent studies indicate a cross-talk betwe-en vitamin D3 and Hh signaling mediated by atleast two mechanisms. First, PTC has beenshown to stimulate the secretion of a vitaminD3-related compound, which is likely respon-sible for the inhibitory action of PTC on SMO.Second, 1α,25(OH)2D3 can down regulate theexpression of some members of the Hh path-way genes, including PTC, SMO and Gli in anepidermal explants culture system, suggestinga direct regulation by 1a,25(OH)2D3. Theseresults are in agreement with the increasedexpression of Shh in the keratinocytes of the


VDR-null animal and hyperactivation of the Hhpathway, predisposing the skin to the develop-ment of both malignant and benign epidermalneoplasms. More interestingly, Uhmann et al.demonstrated that 1a,25(OH)2D3 was capableof inhibiting Hh signaling at the level of SMO inthe absence of VDR. Similar conclusion wasobtained by Tang et al. who studied murinebasal cell carcinomas (BCC) in vitro and invivo, and found that the effect of1a,25(OH)2D3 induced Gli expression is likelyindependent of VDR. The results providestrong evidence of the non-genomic and rapidnon-VDR action of 1a,25(OH)2D3 on cellgrowth and differentiation mediated by Hh/Glisignaling pathway [4].

In the absence of the Hh ligands, Hh recep-tor (PTC) binds SMO, the key Hh signal tran-sducer, via a vitamin D3-related compound.Under this condition, Gli molecules are proces-sed into repressor forms (Gli R), which turn offthe expression of Hh-inducible genes. In thepresence of Hh, Hh binds to PTC. The bindingcauses the release of SMO from T. After SMOis freed from the PTC inhibition, it undergoesconformational changes to activate Gli to formactive Gli (Gli A) in the cytosol. Cytosolic Gli Ais then transported into the nuclei to act as atranscription factor to induce gene expression.Vitamin D may affect Hh signaling pathway viatwo potential mechanisms. PTC can stimulatethe secretion of a vitamin D3-related compo-und, which is responsible for the inhibitoryaction of PTC on SMO in the absence of Hh.This results in a down-regulation of Hh signa-ling and increased expression of p21 and p27.Right panel depicts that the active form of vita-min D3, 1a,25(OH)2D3, can down-regulate theexpression of PTC, SMO and Gli proteins, thatin turn decreases Hh signaling, leading toenhanced p21 and p27 expression and otheractions, including rapid nongenomic responses[4].

WNT/β-CATENIN SIGNALING PAThWAy

Expression of E-cadherin and β-catenin(proteins forming intracellular junctions) arealso decreased in skin malignant tumours,including basal cell carcinoma, squamous cell

carcinoma and melanoma . One of the studiesrevealed, that 4-day incubation of humankeratinocytes with 1a,25(OH)2D3 caused theassembly of adherens junctions, upregulatingE-cadherin expression, but not of desmoso-mes. The same study demonstrated that1a,25(OH)2D3 may induce formation of intra-cellular junctions by protein kinase C (PKC)activation. Thus, it is speculated that1a,25(OH)2D3-induction of cell–cell junctionsformation may be a novel, promisingmecha¬nism of the anti-neoplastic and anti-proliferative cancer treatment [20].

Moreover, protein kinase C inhibitors blockE-cadherin, P-cadherin, a-catenin, and vinculintranslocation to cell-cell contacts and theassembly of adherens junctions promoted by1,25(OH)2D3 in cultured human keratinocytes[24].

Many epidermal genes induced by WNT/β-catenin contain VDR response elements andwere activated independently of TCF/LEF,implying that it is part of a TCF/LEF-indepen-dent aspect of WNT signaling. Likewise, deple-tion of follicular keratinocyte populations inVDR-null mice was linked to aberration of thecanonical WNT pathway [9].

Wnt/β-catenin is an evolutionarily conser-ved signaling pathway that plays an essentialrole in a diverse array of biologic processes,including organogenesis, tissue homeostasisand, in some instances, pathogenesis of disea-ses, including cancers. Earlier studies indicatethat 1a,25(OH)2D3 and its analogs are able topromote the differentiation of colon cancercells by inhibiting Wnt/β-catenin signalingpathway mediated by VDR competing withtranscription factor TCF-4 for β-catenin bin-ding. The molecular mechanism of the inducti-on by 1a,25(OH)2D3 was further studied inLS180 colon cancer cells using chromatinimmunoprecipitation-seq and gene expressionanalyses. It was found that VDR and RXR co-localized to VDRE sites in a ligand-dependentmanner near a set of genes that included c-FOS and c-MYC. The expression of both c-FOSand c-MYC was modulated by 1α,25(OH)2D3.At the c-FOS gene, both VDR/RXR andTCF4/β-catenin bound to a single distal enhan-cer located 24kb upstream of the transcriptio-nal start site. At the c-MYC locus, binding wasat a cluster of sites between - 139 and -165 kb


and at a site located -335 kb upstream (down-regulation), where both VDR and β-catenin acti-vation was interlinked to basal and1a,25(OH)2D3-inducible activities. In addition,1a,25(OH)2D3 is known to regulate two genesencoding two extracellular Wnt inhibitors, DICK-KOPF-1(DKK-1) and DICKKOPF-4 (DKK-4), inopposite directions; 1a,25(OH)2D3 up-regulatesDKK-1 which acts as a tumor suppressor inhuman colon cancer cells, whereas1α,25(OH)2D3 down-regulates DKK-4, anoncogenic protein and a target of the Wnt/β-catenin pathway. Taken together, these datareveal complex modes of action in the regulati-on of target genes 1α,25(OH)2D3 [4].

NoN-GENoMIC ACTIoN oF VITAMIN D3

The genomic actions of 1a,25(OH)2D3 aremediated through its binding to vitamin Dreceptor (VDR), a member of the nuclearreceptor superfamily, to modulate the expre-ssion of various genes. The liganded VDRforms a heterodimer with retinoid X receptor(RXR) and binds to vitamin D response ele-ment (VDRE) to modulate the gene expressi-on. Since VDR is expressed in almost all tissu-es, 1a,25(OH)2D3 has been found to exertvarious anti-cancer actions, including anti-pro-liferation, anti-inflammation, pro-differentiati-on, pro-apoptosis and antiangiogenesis in atissue- and cell-specific manner. Recent evi-dence indicates that 25(OH)D3 is a naturalligand for VDR, and is capable for causing bio-logicaleffects without converting to1a,25(OH)2D3 [4].

So-called, non-genomic mechanism of rapidvitamin D response has been described recen-tly. This mechanism does not di¬rectly affectgene expression or require additional proteinsynthesis. Rapid vitamin D response wasshown to mod¬ulate intracellular calciumlevels, affects activity of sev¬eral intracellularsignalling pathways, through activation ofselected phosphate kinases and phosphatases.These activities take minutes and occur in thecytoplasm of the cell rather than in the nucle-us. Potential mechanism of non-genomic res-ponse involves interaction of vitamin D to1a,25(OH)2D3 membrane–associated rapidresponse steroid-binding protein (1,25 D-

MARRSBP), also known as the protein-disulfi-deisomerase-associated 3 (PDIA3) or endop-lasmic reticulum stress protein 57 (ERp57).PDIA3 activates phospholipase C in a G prote-in-coupled process and results in production ofinositol trisphosphate (IP3) and diacylglycerol.These two cellular messengers mediate therapid release of calcium from the cellular sto-res [26].

Furthermore, an alternative ligand-bindingpocket in the VDR has been identified by mole-cular docking using a receptor conformationalensemble model generated from x-ray crystalstructure of the VDR ligand bindingdomain.This structural flexibility has been pro-posed as a base of rapid and nongenomic bio-logical responses induced by 1a,25(OH)2D3and some of its analogs. The non-genomicrapid (diect) response has been shown inseveral biological systems, that may be media-ted through a functional VDR in some systemsor may be independent of VDR in other sys-tems. To support the involvement of VDR inthe rapid non-genomic actions of vitamin D, ithas been reported that VDR is present incaveolae-enriched plasma membranes andbinds 1a,25(OH)2D3 with high affinity in vivoand in vitro [4].

The existence of non-classical membraneVDR has been found to be related to the rapidactions, including activation of protein kinase Cand protein phosphatase PPIc [4].

The actions have been shown to result insubsequent ion channel activity modulation.Furthermore, it has been known for some timethat 1a,25(OH)2D3 reduces UV induced DNAdamage in the form of cyclobutane pyrimidinedimers (CPD) in human keratinocytes in cultu-re, and in mouse and human skin. The photo-protection by 1a,25(OH)2D3 against oxidativeinsults is thought to be mediated by a non-genomic signaling mechanism, because1a,25(OH)3 lumisterol 3, which has almost notransactivating activity, reduces UV-inducedDNA damage, apoptosis and immunosuppre-ssion with similar potency as1a,25(OH)2D3[4].

In the earlier section under HedgehogHh/Gli signaling pathway, it was mentioned thework by Uhmann et al. who demonstrated that1a,25(OH)2D3 was capable of inhibiting Hhsignaling at the level of SMO in the absence of


VDR, and by Tang et al. who also found that1a,25(OH)2D3-induced Gli expression in muri-ne BCC cells was independent of VDR. Thus, inaddition to the report by Wali et al, these twostudies provide strong evidence of the non-genomic and rapid non-VDR action of1a,25(OH)2D3 on cell growth and differentiati-on [4].

Tang et al. who studied murine basal cellcarcinomas (BCC) in vitro and in vivo, andfound that the effect of 1a,25(OH)2D3 on Gliexpression is likely independent of VDR. Theresults provide strong evidence of the non-genomic action of 1a,25(OH)2D3 on cellgrowth and differentiation mediated by Hh/Glisignaling pathway [4].

Rapid activation of extracellular signal-regu-lated kinases, ERK1/ERK2 in NB4 promyelocy-tic leukemia cells can be induced not only by1,25-D, but also by analogs that are unable toactivate VDR, suggesting the possibility of aseparate receptor. Antibodies to a membraneprotein block the ability of 1,25-D to inducerapid calcium uptake and activation of PKC incartilage cells. VDR-/- osteoblasts take up cal-cium and activate PKC similar to the wild-typeosteoblasts, implicating proteins other thanVDR in these actions. In contrast, Gniadeckihas described activation of ERK through 1,25-D-induced activation of Raf as a result of inte-ractions between VDR and the adaptor proteinShc. VDR-null osteoblasts do not exhibit ionchannel responses in response to 1,25-D andErben et al. have reported that deletion of theVDR DNA binding domain also eliminates non-genomic responses. Thus some of the rapidactions of 1,25-D may be dependent uponVDR, whereas others are not [17].

SuMMARy AND FuTuRE DIRECTIoNS

The reversal of EMT through the use of VDRhas been found to play critical roles in the con-trol of tumor invasion, metastasis, and drugresistance in PDAC (pancreatic ductal adeno-carcinoma). Downregulation of VDR could tri-gger EMT by several factors, including cytoki-nes and cellular signaling pathways such as β-catenin, FoxM1, and CSCs. More importantly,specific natural compounds could partiallyreverse the EMT phenotype to mesenchymal-

to-epithelial transition (MET), resulting in thereversal of drug resistance. Therefore, targe-ting the VDR pathway with nontoxic naturalagents could be a novel potential therapeuticstrategy for the treatment of metastatic PDAC.However, the molecular mechanisms of EMTare very complicated and are not yet fully elu-cidated. Therefore, further investigation isnecessary to explore the mechanisms underly-ing EMT progression in PDAC. Future researchwill surely focus on uncovering the molecularsimilarities and differences among the EMTprograms. EMT research in the next few yearspromises to be exciting, as new mouse modelsand molecular probes are identified to addressthe important, still-unanswered questions[29].

Cell fate and phenotype are strictly regula-ted by extracellular signals. 1,25(OH)2D3 andEMT-TFs have opposite effects on epithelial cellphenotype and they antagonize each other(Figure 3 - part I). 1,25(OH)2D3 induces epit-helial differentiation while it inhibits the expre-ssion of several EMT inducers. Conversely,expression of keyEMT-TFs in epithelial cellspromotes the acquisition of a mesenchymalphenotype, which in the case of SNAIL1 andSNAIL2 is associated with VDR gene repressi-on and the blockade of 1,25(OH)2D3 action onepithelial differentiation. Thus, a double nega-tive feedback loop operates between1,25(OH)2D3 and EMT inducers that may con-tribute to the complete acquisition of the phe-notype dictated by the extracellular cues.Theloop may first amplify the signal and later sta-bilize cell fate once the process is completed.Hence, the balance between1,25(OH)2D3/VDR and SNAIL family of tran-scription factors determines cell fate, and itsimbalance may explain the reversibility of theEMT process. Of note, the transition betweenepithelial and mesenchymal phenotypes is alsogoverned by similar double negative feedbackloops among EMT-TFs and certain microRNAs,such as the ZEB/miR-200 and theSNAIL1/miR-34 regulatory circuits [14].

The implication of EMT in cancer progressi-on and organ fibrosis and the inhibitory effectof 1,25(OH)2D3 on EMT have opened the pos-sibility of a therapeutic use of VDR agonistsagainst these diseases. However, the downre-gulation of VDR expression found in several


types of cancer, frequently associated withadvanced stages of the disease, limits the app-licability of vitamin D compounds to preventionin highrisk populations and treatment in pati-ents at early stages of tumor progression [14].In addition, EMT is a transient event duringtumorigenesis, and it has been proposed thatthe reverse process (MET) in the metastaticsites was postulated to be part of the processof metastatic tumor formation [27,14]. Theselines of evidence have led to controversy aboutanticancer therapeutic strategies designed toinhibit EMT, as they may favor the formation ofmetastases, and suggest that these therapiesmay be limited to patients diagnosed at earlystages of the disease to prevent invasion anddissemination. Nevertheless, vitamin D com-pounds as inhibitors of EMT may be interestingtherapeutic agents for fibrosis-associated pat-hologies, in which the EMT process is notreverted and the mesenchymal phenotype ismaintained during disease progression [14].

1a,25(OH)2D3 is a potent anti-canceragent, affecting cancer cells in cultures and

tumor progression in animal models by avariety of mechanisms, including but not limi-ted to anti-proliferation, anti-angiogenesis,pro-apoptosis, pro-differentiation and anti-inflammation (Figure 2-part I). The major hur-dle is the hypercalcemic side effect induced byadministering a high dose of 1a,25(OH)2D3,that is necessary to exert the anti-tumoreffects of vitamin D in humans. The potentialof using less calcemic analogs of vitamin Dwith much higher potency than 1a,25(OH)2D3for treating cancers still exists (Seocalcitol waswith positive opinion from EMA for treatmentof hepatocellular carcinoma and recommendedfrom the producer also for treatment of pan-creatic cancer , showen cardiotoxic activities),especially in combination with other anti-can-cer agents or immunomodulatory drugs. Theassociation between decreased sun exposu-re/vitamin D deficiency and the risk of chronicdiseases, including many types of cancer, indi-cates that maintaining adequate vitamin Dnutrition should be a paramount priority formen and women of all ages [4].

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ПЕтЯ В. ИВАНоВА

Катедра Химия и Биохимия,Фармацевтичен Факултет,

Медицински Университет Пловдив,Бул. Васил Априлов 15А, 4002

[email protected]


PETyA V. IVANoVA

Department Chemistry andBiochemistry, PharmaceuticalFaculty, Medical University of

Plovdiv, 15A Vasil Aprilov Blvd,4002 Plovdiv, Bulgaria.

[email protected]


Original articles

Сравнително изследване на телесни масти,някои телесни обиколки и нивата на триглицериди и холестерол при различни групи пациенти, включени в НИРДИАБо проекта от Пловдивска област

Д. Гетова1, Д. Димитрова1,2, М. Тополов1,3

1Лаборатория по невропсихофармакология, Технологичен център за спешнамедицина Пловдив;

2Катедра по фармакология и клинична фармакология, Медицински факултет,Медицински университет Пловдив;

3Катедра по фармакология и лекарствена токсикология, армацевтичен факултет,Медицински университет Пловдив, България.

Comparison of the fat mass, some body circumferences (waist, hip and thigh)

and the levels of cholesterol and triglyceridesin different groups of patients included

in NIRDIABo project from Plovdiv

Getova D.1, Dimitrova D.1,2, Topolov M.1,3

1Laboratory of Neuropsychopharmacology, High Technological Center for Emergency Medicine Plovdiv;

2Departmet of Pharmacology and Clinical Pharmacology, Medical Faculty, Medical University Plovdiv;

3Department of Pharmacology and Drug Toxicology, Faculty of Pharmacy, Medical University Plovdiv, Bulgaria.

РЕЗЮМЕ:

Въведение: Цел на изследването е дасе сравнят някои телесни обиколки (талия,ханш и бедро), телесните масти и ниватана триглицериди и холестерол при различ-

ABSTRACT

Introduction: The aim of the study was tocompare some body circumference meas-ures, fat mass and triglycerides and choles-terol levels in different age groups in NIRDIA-


ни групи пациенти, включени в НИРДИАБОпроекта от пловдивска област. Метод: Визследването са включени доброволци сBMI над 25, разделени на 5 възрастовигрупи: a) 20-29 г.; б) 30-39 г.; в) 40-49 г.;г) 50-59 г.; д) 60-69 г. (n=5-9). Вземанетона кръвни проби е единствената слабоболезнена процедура. Първите 2 месецапациентите са на ниско калорична диета иприем на пробиотик. Определени са ниватана триглицериди и общ холестерол, както ина HDL and LDL холестерола при първатавизита. Резултати: Измерените телесниобиколки показват завишени стойности всредната възрастова група, телеснитемасти са също по-високи в тази група.Липидният профил показва високи нива натриглицериди, холестерол (HDL и LDL) всъщата група. Заключение: Клиничнотопроучване показва важността на измерва-нето на тези телесни обиколки, сравнява-нето им с телесните масти и нивата на три-глицериди и холестерол, особено на HDL иLDL холестерола в различните възрастовигрупи. Основна рискова група са хората насредна възраст – 40-49 години.Използването на пробиотик и подходящанискокалорична диета може да помогне запонижаването им, да подобри стила наживот и да го удължи.

Ключови думи: телесни масти, телесниобиколки, липиден профил

BO project from Plovdiv region. Method:Volunteers with BMI over 25 were included(male and female), divided in 5 age groups:a) 20-29 years; b) 30-39 years; c) 40-49years; d) 50-59 years; e) 60-69 years (n=5-9). There was one mild invasive procedure –taking blood samples from patients. The levelsof triglycerides, cholesterol (incl. total fraction,HDL and LDL) were measured at first visit.Results: The body circumferences wereincreased in the middle age group, the fatmass also was higher in the same group. Thelipid profile showed higher levels of triglyc-erides, cholesterol (HDL and LDL) in the samegroup. Conclusion: The present studyreveals the importance of measuring somebody circumferences that have to be com-pared with the fat mass levels, the levels oftriglycerides and cholesterol, with focus onHDL and LDL, in different age groups. Themain group at risk with higher parameters andlevels is the middle aged patients – 40-49years. The use of probiotics with an appropri-ate low caloric diet may help them to decreaseit and to improve the life style.

Key words: fat mass, body circumfer-ences, lipid profile

INTRoDuCTIoN

The tendency to develop diabetes mellitustype 2 is mainly due to the rise of overweightand obesity in population [10]. The obesityhas increased globally over the last fewdecades and its association with insulin resist-ance has affected the ability to reduce popu-lation morbidity and mortality. Traditionally,adipose tissue in the visceral fat depot hasbeen considered one of the major culprits inthe development of insulin resistance.However, there is growing evidence support-ing the role of subcutaneous abdominal adi-pose tissue in the development of insulinresistance. There are significant differences inthe functional characteristics of subcutaneousabdominal vs. intra-abdominal vs. gluteo-

femoral fat depot [11]. Therapeutic lifestyle changes continue to

be the most important intervention in clinicalpractice to improve adipose tissue functionand to avoid development of insulin resist-ance.

The ratio between the body circumferenceat the waist and the hips (WHR) is unique tohumans. The waist-hip ratio is an accurateanthropometric parameter for predictinghypertension in overweight and obesepatients [6]. High WHR doubles the risk ofvascular diseases. BMI and fat mass are notthe only and strongest body predictors of„idiopathic” diseases, but also WHR should beconsidered as such [9]. The WHR measuresalso the distribution of body fat and it has


been a focus of extensive research. The distri-bution of body fat provides information aboutage, health and fertility [2]. Usually fat massis the marker for some physiological andpathological conditions, like fat infiltration ofcells [13].

Cholesterol plays a vital role in cell biology.Cholesterol is the precursor for all steroid hor-mones, including gluco- and mineral- corti-coids, sex hormones, vitamin D. All these reg-ulate carbohydrate, sodium, reproductive andbone homeostasis, respectively [4]. Diet-increased serum cholesterol levels do notincrease the low-density lipoprotein/high-den-sity lipoprotein (LDL/HDL cholesterol ratio).Increased levels of LDL cholesterol reduceHDL cholesterol levels and are consideredindependent risk factors for arterial diseases.[8].

The aim of our study was to compare somebody circumference measures as waist, hipand thigh, the fat mass and the levels of cho-lesterol and triglycerides in different agegroups of volunteers that took part in theproject „NIRDIABO”. All volunteers were fromthe Plovdiv region and they participated in thefirst year of the project.

METhoDS

The project aimed to examine the quality oflife and the effects of some non-pharmacolog-ical interventions on volunteers with obesityand high BMI. Both male and female volun-teers with BMI over 25 were included in thisstudy, and were divided in the followinggroups: a) 20-29 years of age; b) 30-39 yearsof age; c) 40-49 years of age; d) 50-59 yearsof age; e) 60-69 years of age. The number ofpatients in each age group was relativelysmall (n=5-9).

All requirements to perform human studywere done, as it was described in our previouspaper [5]. There weren’t any harmful proce-dures on the patients excluding the weak dis-comfort when blood samples were taken.

The including as well as excluding factorsand criteria and the non-pharmacologicalintervention of the study was also performedas it was cited in previous paper [5].

We used „TANITA C 300 BC-420 MA bodycomposition analyzer” apparatus (USA) to

make the following measurements: age, sex,body weight, BMI and fat mass. The circum-ference measures (waist, hip and tight) weredone using simple linear measure in centime-ters.

The levels of cholesterol, triglyceride, HDLand LDL were done at Clinical LaboratoryDepartment of the Medical University – Sofia.The estimations were done using the widelyaccepted method on the system Roche -Hitachi, Cobas 6000 (Japan), according to theinstructions of the producer.

Statistics: all observed values were calcu-lated and the mean for each group was esti-mated. Because of the small number ofpatients in each age group the statistical sig-nificance was not determined.

RESuLTS

The average body measures of circumfer-ences of the waist, hip and thigh in centime-ters (cm) are shown on Fig. 1. There is a ten-dency, not statistically significant, for allmeasures to increase over the years in thefirst 3 age groups, but in the two last groupsthere is a tendency of decreasing in all circum-ferences measured.

The average fat mass was measures bio-electrically on the first and the second visits.There is interesting tendency in the first 2group, more pronounced in the youngergroup, to decrease the fat mass after 2months low caloric diet, performed by thepatients. The third group do not show anychanges between first and second visit. Theolder 2 groups even show slight increasing ofthe fat mass (Fig. 2).

The basal levels of the cholesterol and thetriglycerides were estimated on the first visit.The 1st and the 2nd group of patients were withnormal range levels of cholesterol and triglyc-erides. The 3rd, 4th and 5th groups of patientswere with higher levels of cholesterol (Fig. 3).

The levels of HDL and LDL in all groupsstudied were in the normal range, but thelevels of LDL wwere higher in the last 3groups. Evidently the ration HDL/LDL ismoved towards up due to increased levels ofLDL (Fig. 4).


Fig. 1. Comparison between some body circumference measures (waist, hip andthigh) in the examined age groups.

Fig. 2. The average fat mass measured bioelectrically during first and second visitsin all age groups of patients.


Fig. 3. Basal levels of cholesterol and triglycerides in the different groups ofpatients.

Fig. 4. Basal levels of HDL and LDL cholesterols in different groups of patients.


DISCuSSIoN

Recent research suggests that distributionof body fat is the most important indicator forcardiovascular risk [3]. In particular, increasedvisceral fat has been shown by them as animportant risk factor for diabetes type 2 andother metabolic disorders. An indicator of vis-ceral fat is not only fat mass, but also thewaist circumference is valid marker as moresensitive to the distribution of body fat thanother markers, as BMI. Other authors [1]showed that some other indices of abdominalobesity should be considered – hip and thighcircumferences, which gave better informa-tion about metabolic risk factors. They believethat ethic and racial variation among popula-tion from different regions might need differ-ent cut-off points or use of different anthropo-metric measurements to diagnose obesity andmetabolic disorders.

In our study all circumferences measured –waist, hip and thigh showed increasing valuesin cm with age (from 20 to 49 years), but theydecreased after 50 – 69 years of age.Evidently our findings are in accordance withthose in the literature and the main risk groupis in the 3rd one – 40-49 years, probably dueto hormonal changes in human body duringthis period of life. We assume that our data isvalid for the Eastern European population andshowed the European tendency for increasingthe body fat due to more caloric diet tradition-ally used.

It has been established that abdominalobesity assessed by waist circumference pre-dicts obesity-related health risk [7]. It hasbeen shown that waist circumference and hipor thigh circumferences have independent andopposite effects on metabolic health risk. Theysuggest that waist circumference is positivelyassociated with health risk, but hip and thighcircumferences are negatively associated withhealth. Our data do not support such view,because in our study all circumferences meas-ured have linear relationship.

Other believe, that a large hip or thigh cir-cumferences, or both, which are due probablyto a greater lean mass, have a protectiveeffect on health [1]. Our data shows that fatmass measured bioelectrical is slightlydecreased after 2 months of low caloric diet

performance by patients only in 20-39 yearsof age, do not change in older age groups.Probably those measures have no significantinfluence on metabolic changes associatedwith obesity or diabetes type 2.

It is well known that the levels of triglyc-erides and cholesterol, as well as its HDL andLDL forms play a significant role in obesepatients or diabetic type 2 patients. In ourstudy triglycerides and cholesterol increasedafter 40 years of age and the levels of HDLand LDL cholesterol also.

Clinical and epidemiologic studies haveconstantly shown that there is an inverse rela-tionship between the HDL cholesterol concen-tration and vascular risk [12]. HDL cholesterolhas been proposed to have several anti-ather-osclerotic properties, including the ability tomediate macrophage cholesterol efflux,antioxidant capacity, anti-inflammatory prop-erties, nitric-oxide promoting activity and anability to transport proteins with their ownintrinsic biological activity. HDL particles arecritical acceptors of cholesterol from lipidmacrophages and thereby play an importantrole in the maintenance of cholesterol balancein the arterial wall. On the other hand LDLcholesterol is an important target in clinicalstudies and decreased levels below1.8 moll/Lis of great interest. High LDL cholesterol isprerequisite for formation of atheroscleroticplaque. In our study LDL cholesterol levels arehigher in aged groups – 50-69 years of age,i.e. the bad cholesterol shifts the rationHDL/LDL towards itself.

CoNCLuSIoN

The present study reveal the importance ofmeasure some body circumferences, compareit with fat mass and the levels of triglyceridesand cholesterol, especially HDL and LDL cho-lesterols, in different age groups. The maingroup at risk with higher measures and levelsis the middle aged patients – 40-49 years. Theuse of probiotics with appropriate low caloricdiet may help them to decrease it and toimprove life style and prolong life time.

Acknowledgement: Thanks for laborato-ry tests of the staff of Department of ClinicalLaboratory, Medical University Sofia.


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3. Bozeman SR, Hoaglin DC, Burton TM, PashosCL, Ben-Joseph RH, Hollenbeak CS. Predictingwaist circumference from body mass index.BMC Med Res Methodol, 2012, 12: 115-123.

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5. Getova D, Dimitrova D, Topolov M. Comparisonof body weight, BMI, glucose level, blood pres-sure and heart rate in different groups ofpatients included in NIRDIABO project fromPlovdiv. Bulgarian Medicine, 2017, 7(1): 23-29.

6. Gonzalez-Jimenez E, Montero-Alonso MA,Schmidt-RioValle J. Waist-hip ration as a predic-tor of arterial hypertension risk in chidren andadolescents. Nutr. Hosp, 2013: 28(6): 1993-6.Doi: 10:3305/nutr hosp.v28in06.6653.

7. Janssen I, Katzmarzyk PT, Ross R. Waist circum-ference and not body mass index explains obe-sity related health risk. The American JournalClinical Nutrition, 2004, 79: 379-84.

8. Kapourchali FR, Surendiran G, Goulet A,Moghadasian MH. The role of dietary choles-terol in lipoprotein metabolism and relatedmetabolic abnormalities: a mini review. Crit.Rev. Food Sci, Nutr., 2016, 56(14): 2408-15.

9. Mondelli M, Aretini A, Ginanneschi F, Greco G,Mattioli S. Waist circumference and waist-to-hipration in carpal tunnel syndrome: a case-con-trol study. J. Neurol. Sci, 2014, 338(1-2): 207-13.

10. Nicolls SJ, Brown A. Targeting obesity, diabetesand the metabolic syndrome: are we trying toclose the barn door after the horse bolted?Exprt. Rev. Cardiovasc. Ther., 2014, 12(3):279-90. Doi: 10.1586/1479072.2014.894362.

11. Patel P, Abate N. Body fat distribution andinsulin resistance. Nutrients, 2013, 5(6): 2019-27.

12. Ono K. Current concept of reverse cholesteroltransport and novel strategy for atheroprotec-tion. Journal of Cariodlogy, 2012, 60: 339-343.

13. Vetrovska R, Vilikus Z, Klaschka J, Stranska Z,Svacine S, Svobodova S, Matoulek M. Doesimpedance measure a functional state of thebody fat? Physiol. Res, 2014, suppl. 2: S309-320.


Проф. Д-р ДАМЯНКА ГЕтоВА

Лаборатория по невропсихофармакология,Високотехнологичен център

за спешна медицина, Пловдив. E-mail: [email protected]


Prof. Dr. DAMIANKA GEToVA

Laboratory ofNeuropsychopharmacology, High

Technological Center forEmergency Medicine, Plovdiv.

E-mail: [email protected]


Анализ на експресията на серумните нивана тумор некрозисфактора алфа и TNF при пациенти с остър коронарен синдромвъв връзка с наличието или отсъствието на възпалително ставно заболяване, тип заболяване на ставите и активност на заболяването

В. Попова1, З. Въжев2, Ж. Пешев3, Ж. Грудева-Попова4, А. Баталов1

1 МУ - Пловдив, МФ - Катедра по пропедевтика на вътрешните болести2МУ - Пловдив, МФ - Катедра по сърдечна и съдова хирургия, Клиника по кар-диохирургия

3МУ - Пловдив, МФ, Катедра по обща и клинична патология4МУ - Пловдив, МФ, Катедра по клинична онкология

Analysis of the expression of the TumorNecrosis Factor alpha and TNF serum levels

in patients with acute coronary syndrome in relation to the presence or absence

of inflammatory joint disease, type of joint disease and disease activity

Popova V.1, Vazhеv Z.2, Peshev G.3, Grudeva-Popova Zh.4, Batalov A.1

1MU - Plovdiv, Medical Facility, Department of Propedeutics of Internal Diseases2MU - Plovdiv, Faculty of Medicine, Department of Cardiac and Vascular Surgery

3MU - Plovdiv, Medical Faculty, Department of General and Clinical Pathology4MU - Plovdiv, Medical Faculty, Department of Clinical Oncology

РЕЗЮМЕ:

Повишени нива на Туморния НекрозенФактор алфа (TNF-a) и TNFa серум се асо-циират с наличието на възпалителни став-ни и сърдечни заболявания. Повечето отсъществуващите научни проучвания саизследвали тази връзка поотделно, в рам-

ABSTRACT

Elevated levels of the Tumour NecrosisFactor alpha (TNF-α) and TNFα serum have asignificant association with inflammatoryjoint disease and cardiac disease. However,most of the existing research has examinedthis relationship separately, within inflamma-


INTRoDuCTIoN

Rheumatoid (RA) and psoriatic arthritis(PsR) are chronic inflammatory autoimmunediseases that are associated with higher car-diovascular risk and accelerated atherosclero-

sis. They influence both traditional risk factorsas well as specific factors such as comorbidity,inflammation level, treatment of the diseaseand inflammatory activity. TNF-a is an impor-tant cytokine with a central role in inflamma-

ките на възпалителни ставни заболяванияи в рамките на сърдечни заболявания.Настоящото проучване има за цел да обе-дини тези две области като проследяванивата на експресия на TNFa и TFNa серумпри пациенти с остър коронарен синдром(ACS), разделени на две групи: със и безвъзпалително ставно заболяване. Също сеизледва потенциална асоциация с тип став-но заболяване (RA срещу PsA) и ниво наболестна активнот, измерено чрез DAS28при пациенти с RA и ASDAS при пациенти сPsA.

Изледвана е извадка от 95 пациентите состър коронарен синдром (ОКС). От тях, 46са с възпалително ставно заболяване (ВСЗ)и 49 без ВСЗ. TFN-a експресия е исмереначрез имунохистохимичен анализ на мате-риал, съдържащ атероматозна плака отоперация на сърдечен байпас или другаманипулация без риск за пациента. TNFaсерумните нива се измерват с протоколELISA Kit за тумор некрозис фактор ELISA(TNF) АА 77-233, човешки, сандвич ELISA,чувствителност 1.0 pg / mL ABIN411361-BOSTER.

Резултатите показват значително по-високи нива на експресия на TNFa и TNFaсерум (48 часа) при ОКС пациенти с ВСЗ всравнение с ОКС групата без ВСЗ. Не сеустановява статистически значима разликапри TNFa серум на 24 часа. В групата безВСЗ има значителен спад в серумното нивона TNFa на 48 часа; докато в групата с ВСЗне се наблюдава значително понижение.Установява се, че TNF-a експресията исерумното ниво на TNFa не са свързани свида на възпалителното ставно заболяване(RA спрямо PsA), нито с нивото на актив-ност на заболяването.

Ключови думи: възпалително ставнозаболяване, остър коронарен синдром,TNF-α, TNFα серум, ревматоиден, артрит,псориатичен артрит

tory joint disease and within cardiac disease.The present study aimed to put together bothareas by comparing the levels of TNF-αexpression and TFNα serum in patients withacute coronary syndrome (ACS), divided intotwo groups: with and without inflammatoryjoint disease. Associations with type of jointdisease (RA vs. PsA) and inflammatory jointdisease activity (as measured by DAS28 inpatients with RA and ASDAS in patients withPsA) were also explored.

The sample included 95 ninety-fivepatients with ACS. Among then, 46 hadinflammatory joint disease and 49 did not.TFNα was measured through immunohisto-chemical analysis of material containingatheromatous plaque from cardiac bypasssurgery or other manipulation without a riskfor the patient. TNFa serum levels weremeasured with the following protocol TumorNecrosis Factor ELISA Kit (TNF) AA 77-233Human, Sandwich ELISA, sensitivity 1.0 pg /mL ABIN411361- BOSTER.

TNFα expression and TNFα serum levels(48hrs) were significantly higher in ACSpatients with inflammatory joint disease thanin those without inflammatory join disease.There was no significant difference regardingTNFα serum levels (24hrs). In the group with-out inflammatory joint disease, there was asignificant drop in TNFα serum level at 48 hrs;whereas in the group with inflammatory jointdisease, no significant decrease wasobserved. It is observed that TNFα expres-sion and TNFα serum levels were not associ-ated with type of inflammatory joint disease(RA vs. PsA), nor with level of disease activity.

Key words: inflammatory joint disease,acute coronary syndrome, TNFα, TNFα serum,rheumatoid arthritis, psoriatic arthritis


tory joint disease, atherogenesis and acutecoronary syndromes. Its production may berelated to the level of activity and the extentof systemic and local inflammation. The nor-mal heart does not express and does not pro-duce TNF-a, but in heart failure or other car-diac diseases, the heart produces extremelevels of TNF-a [1, 2].

Atherosclerosis is characterized by hyper-plasia, neointimal and local inflammatory res-ponse. TNFa mediates proinflammatory, proli-ferative, cytostatic and cytotoxic effects invarious cell types, including endothelial cellsand vascular smooth muscle cells (VSMCs)[3]. In addition, the myocardium may synthe-size TNF-a de novo which causes TNF-a serumlevel to rise extremely in response [4-7].Scientific research about these processes isbeneficial to therapeutic approaches utilizingcytokine inhibition and can help achieve con-trol and risk reduction in acute events.

However, most of the existing research hasexamined this relationship separately, withininflammatory joint disease and within cardiacdisease. The present study aimed to puttogether both by comparing the levels of TNFαexpression and TFNα serum in patients withacute coronary syndrome (ACS), divided intotwo groups: with and without inflammatoryjoint disease. Associations with type of jointdisease (RA vs. PsA) and inflammatory jointdisease activity (as measured by DAS28 inpatients with RA and ASDAS in patients withPsA) were also explored.

MATERIAL AND METhoDSPatients

The sample was drawn from cardiac surgi-cal patients at a Bulgarian medical institution.Ninety five patients with acute coronary syn-drome (ACS) (mean age 69.59 ± 7.22), ofwhom 46 (mean age 68.74 ± 7) with inflam-matory joint disease and 49 (mean age 70.39± 7.40 ) without inflammatory joint diseasewere included in the study. Among the 46patients with inflammatory joint disease, 23had rheumatoid arthritis (RA - mean age67.39 ± 7.42) and 23 were with psoriaticarthritis (PsA - mean age 70.09 ± 6.37). Table1 contains stratified demographic information.There was no significant difference between

the groups in terms of average age, p > .05for all comparisons. The samples were pred-ominantly male (77.5% without joint disease;80% with joint disease; 65% RA; 95.6%PsA). There was no significant sex differencebetween the groups with joint disease andwithout joint disease, p = .730; however,there was a significant sex difference betweenthe RA and PsA groups as the male sex waspredominant in the PsA group, p =.009. Table1 contains stratified demographic information.

The procedures were conducted in accor-dance with the WMA Declaration of Helsinki.All patients provided written informed consentprior to the investigation.

Immunohistochemical analysis was perfor-med on material containing atheromatousplaque from cardiac bypass surgery or othermanipulation without a risk for the patient.The immunohistochemical analysis was per-formed according to the IHH protocol usingthe following kits to detect the presence ofTNF alpha: anti-Tumor Necrosis Factor antibo-dy (TNF) - (AA 180-230) -tnf-alpha-Rabbit,Polyclonal, IgG, 1: 100 Biosystems andPolymer Visualization System - CRF™ Anti-Polyvalent HRP Polymer (DAB) Stain Kit. Forthe color intensity, the following numericcodes were used: 0 - absence; 1 - weak; 2 -moderate; 3 - strong. Strong intensity iscounted as this of a positive control, and asabsent based on a negative control. For amore accurate assessment of the color reacti-on, all materials were prepared with a negati-ve control.

TNFa serum levels were tested at 24 and48 hours after onset of ACS with protocolTumor Necrosis Factor ELISA Kit (TNF) AA 77-233 Human, Sandwich ELISA, sensitivity 1.0pg / mL ABIN411361- BOSTER. Their relation-ship to the presence of absence of inflamma-tory joint disease was sought.

Inflammatory joint disease activity in pati-ents with RA was established with DAS28, andin the PsA patients with ASDAS. DAS28 valueswere used to determine categories of diseaseactivity as follows: remission – DAS28 ≤ 2.6;low disease activity – 2.6 < DAS28 ≤ 3.2;moderate disease activity – 3.2 < DAS28 ≤5.1; high disease activity – DAS28 > 5.1.Accordingly, ASDAS levels were defined as:


remission – ASDAS < 1.3; low disease activity– 1.3 ≤ ASDAS ≤ 2.1; medium disease activi-ty – 2.1 ≤ ASDAS ≤ 3.4; high disease activity– ASDAS > 3.5.

AnalysisSPSS software package version 24 (SPSS,

Chicago, Illinois, USA) was used for statisticalanalysis. Normally distributed continuous vari-ables are described in mean and standarddeviation (SD), dichotomous variables areexpressed in number and percentage, and forordinal variables, the median and range areprovided. The dependent variables of interestto this investigation (TNFa, TNFa serum24hrs; and TNFa serum 48hrs) were exami-ned for normality through Kolmogorov-Smirnov goodness-of-fit test. The resultsrevealed that all three of them were not nor-mally distributed, p < .001 for all three vari-ables. This determined the use of non-para-metric tests, including Mann-Whitney U testfor between group comparisons andSpearman Rho correlation for exploring asso-ciations. Additionally, chi-square test was usedfor nominal variables. All statistical results areinterpreted at level of significance, alpha (a) =.05.

RESuLTS

First are presented the results of the bet-ween-groups comparison of ACS patients withand without inflammatory joint disease inrelation to the values of TFNα and TFNα serum(24 and 48 hrs). The descriptive statistics forthese variables are given in Table 1. As men-tioned earlier, all three of them were not nor-mally distributed and for this reason the com-parison was performed with Mann-Whitney Utest for two independent samples. The resultsshowed statistically significant differences ontwo of the three parameters, TNFa and TNFαserum (48hrs). For TNFa, the difference washighly significant, p < .001. The group withoutinflammatory joint disease had a lower medi-an value of 1 (range 0-2), whereas the groupwith inflammatory joint disease had a medianvalue of 2 (range 0-3). A similar trend wasfound for TNFα serum (48 hrs), where the dif-ference was highly significant, p < .001. Thegroup without inflammatory joint disease hada significantly lower mean value of 5.01 (±

3.31) as compared to the group with inflam-matory joint disease, mean of 9.98 (± 4.57).No significant difference was found betweenthe two groups regarding TNFα serum at 24hours, p =.690.

Since the between-group comparisonsrevealed lack of significant differences regar-ding TFN α serum (24 hrs) and a highly signi-ficant difference for TFN α serum (48 hrs), itwas considered important to follow it up withwithin-group comparisons between the twoserum levels. Wilcoxon Signed Ranks test wasused to explore potential internal differencesin TFN α serum levels between 24 and 48 hrs.The results revealed significant decrease inTFN α serum levels for the ACS group withoutinflammatory joint disease, p < .001. Themean value dropped from 9.48 to 5.01.However, for the ACS group with inflammato-ry joint disease no significant reduction in TFNα serum levels was found, p = . 288, with aneven slight increase in mean value, from 9.19to 9.62. The within-group trends of TFN αserum levels between 24 and 48 hrs are illust-rated on Figure 1.

Additionally, the internal relationship bet-ween TNFα and TNFα serum (24 and 48 hrs)was also explored through correlation analysisemploying Spearman Rho test. Two correlati-ons were examined within each ACS patientgroup: 1) between TNFa and TNFα serum(24hrs) and 2) between TNFα and TNFαserum (48hrs), amounting to a total of fourcorrelation analyses. In both groups, no signi-ficant association between TNFa expressionand TNFa serum were found; p > .05 in allfour correlations as shown in Table 2.

At the next level, a comparative analysiswas performed within the group of ACS pati-ents with inflammatory joint disease involvingthe two subgroups of RA and PsA patients.The Mann-Whitney U test was used to explorepotential differences associated with the typeof joint disease. None of the three compari-sons showed statistical significance. Themedian value for TNFα was 2 in both RA andPsA patients. Although the range was different(0-2 for RA) and (0-3 for PsA), only one pati-ent in the PsA group had level 3 of TNFαexpression. The Mann-Whitney U test showedlack of significant difference, p = .616. Themean values of TNFa serum (24hrs) were


Table 1: Demographic features of the participants in the study and dependent vari-ables (TNFa and TNFa serum at 24 and 48 hours

TNFa = Tumor Necrosis Factor measured on an ordinal scale of 0 to 3, where 0 = absenceand 3 = high expression; TNFa serum is measured in ng/ml and is a continuous variable.

Figure 1: Lineplot of groupmeans for TFNα serum levelsbetween 24 and48 hrs


slightly higher in the PsA group (10.35 vs.9.19), but the difference was not statisticallysignificant, p = .974. A similar trend wasobserved regarding TNFa serum (48hrs),where the PsA group had a slightly highermean of 10.34 vs. 9.62 of the RA group, withno statistical significance, p = .953.

Within the RA and PsA groups of patients,a potential relationship between TNFa andTNFa serum (24 and 48 hrs) and disease acti-vity level was explored through SpearmanRho correlation. Disease activity within the RAsubgroup was established on the basis ofDAS28, following the standard values andranges for each level. There were no patientsin remission (DAS28 ≤ 2.6) and no patientswith low disease activity ( 2.6 < DAS28 ≤3.2). Nine patients (42.9%) had moderatedisease activity (3.2 < DAS28 ≤ 5.1) and 12(51.1%) had high disease activity ( DAS28 >5.1).

For the PsA patients, disease activity wasdetermined on the basis of ASDAS, again inaccordance with the established referencevalues. There were no patients in remission(ASDAS < 1.3) and no patients with lowdisease activity (ASDAS 1.3 – 2.1). Five pati-ents showed moderate disease activity(ASDAS 2.1 -3.4) and 16 patients had highdisease activity (ASDAS > 3.5). The descripti-ve statistics for TNF-a TNF-a serum levels at24 and 48 of the stratified samples are shownin Table 3.

Correlation analysis was also performedwithin the RA and PsA groups between disea-se activity level (DAS28 and ASDAS), TNFαexpression and TNFa serum (24 and 48 hrs).All correlation coefficients were low with levelof significance exceeding .05 (Table 4).

CoNCLuSIoNS

The study provides information about diffe-rences and similarities in TNF-a expressionand TNFa serum levels (24hrs and 48hrs) inpatients with ACS in relation to presence andabsence of inflammatory joint disease, typeof inflammatory joint disease (RA vs. PsA) andlevels of disease activity. The results show sig-nificantly elevated levels of TNF-a expressionin patients with ACS and inflammatory jointdisease as compared to patients with ACS, butwithout inflammatory joint disease.Extrapolating from this finding, it can be con-cluded that the presence of inflammatory jointdisease in patients with ACS adds up to theproduction of TNF-a and further elevates itsexpression. However, when patients of twodifferent types of inflammatory joint diseasewere compared, RA vs. PsA, the expression ofTNF-a did not reveal significant differences,which leads to the conclusion that the type ofjoint disease does not have association withthe levels of TNF-a expression.

Table 2: Spearman correlation results for TNFα and TNFα serum (24 and 48 hrs) withinboth groups

Note: None of the correlations are significant, p < .05.


Regarding serum levels at 24hrs after theonset of the coronary syndrome, the studyfound no significant differences between pati-ents with ACS with and without inflammatoryjoint disease. However, a significant differencewas observed in relation to the TNFα serumlevels at 48hrs as the group without jointdisease showed significantly lower levels. Thepost hoc analysis revealed a significant drop inTNFα serum level at 48 hrs as compared to

the level at 24hrs within the group of patientswith ACS, but no inflammatory joint disease.This suggests that the presence of a systema-tic inflammatory joint disease leads to induc-tion of cytokine synthesis after 48 hours. Thisconclusion is valid for both the RA and PsApatients based on the lack of significant diffe-rences between them regarding TNFα serumlevel at 24 and 48 hrs. Inflammatory jointdisease leads to induction of cytokine synthe-

Table 3: Stratification of RA and PsA groups according to disease activity

Note: DAS28 used for determining disease activity in RA patients; ASDAS in PsA patients

Note: None of the correlations are significant, p < .05.

Table 4: Correlation analysis between disease level TNFα expression and TNFaserum


sis regardless of the type of joint disease. The study also sought to explore possible

associations between TNF-a expression andTNF-α serum levels at 24 and 48 hours wit-hin the groups of ACS patients with and wit-hout inflammatory joint disease. No signifi-cant relationship was found suggesting thatTNF-α expression and TNF-α serum levelsserve as independent measures.

In relation to disease activity, the pati-

ents with RA and PsA fell into two categoriesof moderate and high disease activity asmeasured by DAS28 and ASDAS, respecti-vely. No significant relationship was foundbetween disease activity level, TNF-aexpression and TNF-α serum levels. Thismaybe due to the limited variation in disea-se activity parameters as the patients inboth groups were between moderate andhigh levels.

TNF X PLAQUE TNF X CIKATRIX INFARKT

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13. Lampugnani MG, Resnati M, Raiteri M, Pigott R,Pisacane A, Houen G, Ruco LP, Dejana E. Anovel endothelial-specific membrane protein isa marker of cell-cell contacts. J Cell Biol.1992;118(6):1511–22

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Д-р В. ПоПоВА

4001 Пловдив - ул. „София” 64УМБАЛ „Каспела” –

Клиника по ревматологияE-mail: [email protected]

Teл: + 359 879 221 128


Dr. V. PoPoVA

4001 Plovdiv - 64 Sofia StreetUMHAL „Kasella” -

Clinic of RheumatologyE-mail: [email protected]

Tel: + 359 879 221 128

Проучвания върху шума в околната среда и здравето в България (1970 – 2017)

Ангел М. ДжамбовКатедра „Хигиена и екомедицина”, Факултет по обществено здраве, Медицински университет – Пловдив

Environmental noise and health research in Bulgaria (1970 – 2017)

Angel M. DzhambovDepartment of Hygiene and Ecomedicine, Faculty of Public Health,

Medical University of Plovdiv

РЕЗЮМЕ:

Шумът в околната среда е повсеместно раз-пространен замърсител и е свързан с редицаздравни резултати, от лошо психично здраве досърдечно-съдови заболявания. Това е свързаносъс значителни социално-икономически загуби.Шумът е проникнал във всички сфери на живота,от транспорта до дейностите през свободнотовреме, а неблагоприятното социално-икономи-ческо положение в България допълнителноусложнява проблема. В настоящата работа авто-рът цели да направи кратък обзор на българска-та литература по темата, с намерението да уста-нови съществуващите пропуски и да подчертаенеобходимостта от развитието на тази научнаобласт. Обзорът е базиран на електронни и неав-томатизирани търсения, без времеви ограниче-ния, на актуални проучвания, публикувани набългарски или английски. Включени бяха собст-вените публикации на автора заедно с такива наспециалисти по комунална хигиена, които бяхапомолени да прегледат архивите си. Резултатитепоказаха, че докато в по-заможните държави санатрупани значителни данни, тази област на тър-сене е все още в зародиш в България. Въпрекитова, вдъхва увереност фактът, че проучваниятавърху шума в страната набират скорост и са всепо-признати от международната научнаобщност.

Ключови думи: Раздразнение от шума; ССЗ;Психично здраве; Шумова експозиция;Транспортен шум

ABSTRACT

Environmental noise is a ubiquitous pollu-tant, and it is associated with a plethora ofhealth outcomes, ranging from mental illhealth to cardiovascular disease. This trans-lates into considerable socioeconomic loses.Noise has penetrated all aspect of modern life,from transportation to leisure time activities,and the unfavorable socioeconomic situationin Bulgaria further compounds the problem. Inthe current paper, the author aims to brieflyreview the Bulgarian literature on the subjectmatter, with the intention to identify existinggaps and emphasize the necessity to advancethe field. The review was based on electronicand manual searches, with no time restric-tions, of relevant studies published inBulgarian and English. Author’s own publica-tions were included along with those of spe-cialists in environmental hygiene who wereasked to search their archives. Resultsshowed that while substantial data has beenaccumulated in more affluent countries, thisfield of inquiry is still in its infancy in Bulgaria.Still, it is reassuring that noise research in thecountry is gaining momentum and is increas-ingly acknowledged by the international scien-tific community.

Keywords: Noise annoyance; CVD;Mental health; Noise exposure; Traffic noise



INTRoDuCTIoN

Environmental noise is a ubiquitous pollu-tant, with road traffic being the major noisesource in urban settlements (15). In 2012,around 37.8% of Europeans living in majoragglomerations were exposed to road trafficnoise > 55 dB(A), while this proportion wasmore than double, 84.44%, in Bulgaria (19).Houthuijs et al. recently estimated that 46%of Europeans were expose to ≥ 55 dB(A) day-evening-night noise level, and 32%, to ≥ 50dB(A) at night (18). Median daily and nightti-me noise levels in the EU were 54.5 dB(A) and46.5 dB(A), respectively (18).

In the report „Burden of disease from envi-ronmental noise”, WHO experts calculatedthat at least one million healthy life years arelost in Europe every year due to road trafficnoise; of those, 61 000 disability-adjusted life-years (DALYs) are due to ischemic heartdisease, 45 000 DALYs – to cognitive impair-ment in children, 903 000 DALYs – to sleepdisturbance, 22 000 DALYS – to tinnitus, and587 000 – to noise annoyance (23). Accordingto Swinburn et al., nationwide reduction ofnoise levels with 5 dB(A) would result in 1.4%and 1.8% lower prevalence of hypertensionand ischemic heart disease, respectively,which would save around $ 3.9 billion in theUS (20). In Europe, road traffic noise is cul-pable for 1.1 million cases of hypertension, 52600 hospitalizations, and 12 200 cases of pre-mature mortality from ischemic heart diseaseand stroke (18).

Evidently, environmental noise has a subs-tantial impact on public health. Its control,however, remains challenging on a globalscale, let alone in Bulgaria. Noise has penet-rated all aspect of modern life, from transpor-tation to leisure time activities, and the unfa-vorable socioeconomic situation in Bulgariafurther compounds the problem. In the cur-rent paper, the author aims to briefly reviewthe Bulgarian literature on the subject matter,with the intention to identify existing knowled-ge gaps and emphasize the necessity toadvance the field. The review was based onelectronic and manual searches, with no timerestrictions, of relevant studies published inBulgarian and English. Author’s own publicati-ons were included along with those of specia-lists in environmental hygiene who wereasked to search their archives.

MENTAL WELL-BEING AND PSyChoSoCIAL REACTIoNS To NoISE

Evidence of the psychosocial effects ofenvironmental noise in Bulgaria dates back tothe 1970-ties. For instance, Radneva-Dimitrova conducted a socio-acoustic surveyof the resident of 1 030 apartments in thecapital Sofia and indicated that 94.6% ofthose living close to a highway construed traf-fic noise as a major nuisance (29). In a follow-up study, she investigated general noise anno-yance comparing people exposed to 68 – 78dB(A) and 51 – 60 dB(A) (30). Another studyfocused on noise penetrating residential buil-dings and its association with restoration andsleep (26).

Several studies were conducted in the cityof Pazardzhik. In 2000, Gatseva et al. lookedat the association between traffic noise expo-sure and self-reported health status of 100adults and found some indication for highernoise annoyance, sleep disturbance andneurological complaints among participantsliving in noisier neighborhoods (16). In additi-on to adults, Gatseva and Turnovska sampled285 children from Pazardzhik and found hig-her irritability, more complaints and difficultiesconcentrating in children living on streets withhigher traffic intensity (17).

In 2011, Turnovska et al. sampled 400residents of the town of Smolyan exposed to67.4 – 70.6 dB(A); 60 – 70% reported high,and 25%, very high noise annoyance (21).The National Centre for Public Health andAnalyses reported results of a pilot study con-ducted in 2013 in the capital Sofia and alsofocused on citizens living in a noisy area (≈ 73dB(A)) (25). Noise was measured objectivelyand it was linked to participants’ perception ofthe acoustic environment in the neighborho-od. High traffic intensity and noise exposurewere negatively associated with participants’residential satisfaction; 32% reported highnoise annoyance, and more than half reportedsleep disturbance due to transportation noise.

Two multi-item questionnaires for theassessment of noise sensitivity were validatedin Bulgarian to herald the use of this psycho-metric construct in future noise research (8,11). It was confirmed that noise sensitivitywas an important correlate of annoyance,


predicting it above and beyond noise exposure(8, 11). Another study indicated that percei-ved traffic intensity on the residential streetwas a better predictor of noise annoyancethan the actual number of motor vehicles andmeasured noise level (14).

Dzhambov and Dimitrova examined theexposure-response relationship between roadtraffic noise exposure and noise annoyance inPlovdiv (2). They extracted noise data fromthe strategic noise map of the city for 513 par-ticipants who also filled a socio-acoustic ques-tionnaire. The empirically derived relationshipin this sample resembled the polynomialrecommended by the WHO for calculation ofthe proportion of highly noise annoyed peoplein different noise bands. Burden of diseasecalculations following the WHO methodologyshowed an annual loss of 1 188 DALYs inPlovdiv, which translated into around € 14 mil-lion; on a national scale, these estimates were6 400 DALYs and € 77 million (2). The socioe-conomic burden of traffic noise-attributedsleep disturbance was even greater – 15 468DALYs and € 185 million (4).

In the recent years, the interest in mentalhealth as a potential outcome of environmen-tal noise exposure was renewed. In 2013,Dzhambov and Dimitrova sampled 182 resi-dents in one of the neighborhoods of Plovdivwhere noise levels were around 72 dB(A) (1).The authors assessed participants’ perceivedacoustic environment at home and looked atthe association between displaced aggressionand noise. Hearing noises above the perceivednormal threshold more often, exposure tocontinuous noise (vs intermittent), and highernoise sensitivity were independently associa-ted with higher aggression (1).

More recently, the same authors investiga-ted different pathways linking road trafficnoise to general mental health in a sample of399 youth (3). That study extended the inqui-ry to the social sphere and integrated environ-mental and social aspects of the noise –health relationship. Results showed that hig-her noise exposure was associated with worsemental health only indirectly through noiseannoyance, which was associated with lowerneighborhood restorative quality, in turn withless social cohesion and physical activity, andconsequently with worse mental health (3).

CARDIoVASCuLAR AND METABoLIC ouTCoMES

There is limited evidence linking trafficnoise to cardiovascular and metabolic outco-mes in Bulgaria. To name a few examples,Tzenova et al. surveyed 1 062 residents of 52buildings in Sofia and measured their noiseexposure and arterial blood pressure (31).The study indicated higher prevalence ofhypertension in those exposed to > 60 dB(A)compared to < 60 dB(A) (36.97% vs 28.41%,p < 0.01). A similar trend was observed forischemic heart disease (17.38% vs 10.56%, p< 0.01) (31). Differences in systolic and dias-tolic blood pressure, measured on a continu-ous scale in mmHg, between residents inquiet and noisy areas were also statisticallysignificant and more pronounced in hyperten-sive individuals (27). These findings were con-firmed in a sample of 84 normotensive medi-cal students (28). In an ecological study of115 819 participants living in the city ofBurgas, Turnovska et al. found that thoseliving near the airport and exposed to > 50dB(A) had higher prevalence of circulatorydiseases and myocardial infarction (22). Ofnote, the aforementioned studies were carriedout some 10-15 years ago and suffered met-hodological shortcomings, such as exposuremisclassification and lack of adequate controlfor confounding factors.

More recently, data were collected for 513residents of Plovdiv to study the long-termrelationship between road traffic noise and theprevalence of cardiovascular disease, diabe-tes, and obesity in healthy adults (24). In con-trast to previous research, this study accoun-ted for a wide range of confounding factors,e.g. sociodemographic variables, co-exposureto air pollutants, other contextual factors.Results for CVD were inconclusive, as therewere indications that higher noise exposurewas associated with higher odds for hyperten-sion and ischemic heart disease, but the effectestimates were non-significant (5, 10).According to the authors, these null findingscould be due to the smallish sample size. Still,burden of disease calculations for the year2012 showed that 2.9% of all myocardialinfarction cases in Bulgarian urban areas couldbe attributed to road traffic noise. Those addi-


tional cases were valued at around € 11.6 mil-lion per annum (6). As regards metabolic out-comes, participants exposed to > 70 dB(A) athome had higher odds for diabetes (7) andobesity (9).

In order to expand the scope of previousresearch, which was mostly concerned withthe effects of noise in the general population,Dzhambov et al. investigated the impact ofroad traffic noise on several cardiovascularand metabolic indicators in patients with pre-existing cardiovascular disease (24). In a seri-es of studies of patients from the PlovdivProvince, they observed higher blood pressu-re, lower glomerular filtration rate, and lessfavorable lipid profile in some patients expo-sed to higher residential noise (12, 13). Theauthors identified potentially vulnerable subg-roups in which the effect was both substantiveand statistically significant. These findings

added to the limited body of evidence on thesubject and suggested the clinical relevance ofnoise pollution.

CoNCLuSIoN

Environmental noise is a conspicuous thre-at for human health and well-being. To per-suade stakeholders to take action and mitiga-te noise exposure, the current body of eviden-ce needs to be expanded further. While subs-tantial data has been accumulated in moreaffluent countries, this field of inquiry is still inits infancy in Bulgaria. Still, it is reassuringthat the current review found a positive trendin the recent years. Noise research in Bulgariais gaining momentum and is increasingly ack-nowledged by the international scientific com-munity.

REFERENCES

1. Dzhambov A, Dimitrova D. Neighborhood noisepollution as a determinant of displaced aggres-sion: a pilot study. Noise Health.2014;16(69):95-101

2. Dzhambov A, Dimitrova D. Road traffic noise andannoyance: exposure-response relationship andburden of disease calculations in Bulgaria. Scr SciMed. 2015;47(2):22-30

3. Dzhambov A, Tilov B, Markevych I, Dimitrova D.Residential road traffic noise and general mentalhealth in youth: The role of noise annoyance,neighborhood restorative quality, physical activi-ty, and social cohesion as potential mediators.Env Int. 2017;109:1-9.

4. Dzhambov AM, Dimitrova DD, Mihaylova-AlakidiVK. Burden of Sleep Disturbance Due to TrafficNoise in Bulgaria. Fol Med (Plovdiv). 2015;57(3-4):264-9.

5. Dzhambov AM, Dimitrova DD. Associationbetween Noise Pollution and Prevalent IschemicHeart Disease. Fol Med (Plovdiv).2016;58(4):273-281.

6. Dzhambov AM, Dimitrova DD. Evaluation of thesocial and economic burden of road traffic noise-attributed myocardial infarction in Bulgarianurban population. Arh Hig Rada Toksikol.2015;66(1):15-21

7. Dzhambov AM, Dimitrova DD. Exposures to roadtraffic, noise, and air pollution as risk factors fortype 2 diabetes: A feasibility study in Bulgaria.Noise Health. 2016;18(82):133-42.

8. Dzhambov AM, Dimitrova DD. Psychometric

properties of the Bulgarian translation of NoiseSensitivity Scale Short Form (NSS-SF):Implementation in the field of noise control.Noise Health. 2014;16:361-7.

9. Dzhambov AM, Dimitrova DD. Road traffic noiseexposure association with self-reported bodymass index. Noise Contr Eng J. 2015;63(6):572-81.

10. Dzhambov AM, Dimitrova DD. Road trafficnoise-related hypertension and ethnicity as aneffect modifier. In: Łucjan C, Gérard D (Editors),Traffic Noise: Exposure, Health Effects andMitigation. New York: Nova Science Publishers,2017, pp. 41-58. ISBN: 978-1-53612-550-4.

11. Dzhambov AM, Dimitrova DD. Validating a shortBulgarian version of a psychometric instrumentfor multidimensional noise sensitivity assess-ment. Fol Med (Plovdiv). 2014;56(2):116-25

12. Dzhambov AM, Tokmakova M, Gatseva P,Zdravkov N, Gencheva D, Ivanova N, KarastanevK, Vladeva S, Donchev A, Dermendzhiev S.Community noise exposure and its effect onblood pressure and renal function in patients withhypertension and cardiovascular disease. FolMed, in press.

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24. Джамбов АМ. Шумовото замърсяване от авто-мобилен трафик – рисков фактор за някоисърдечно-съдови и метаболитни заболявания

сред пълнолетното население [дисертация].Пловдив: Медицински университет –Пловдив; 2017.

25. Иванова П, Израел М, Петкова Хр, ДимитроваМ, Димитрова Д. Оценка на въздействието нашума върху населението в район с установенанаднормена шумова експозиция. Юбилейнаконференция с международно участие:Общественоздравна политика и практика.Плевен, 2015.

26. Радев Ат, Раднева-Димитрова Р, Радева-Александрова Н. Проникващи шумове вжилищните сгради и въздействието им върхупочивката и съня на населението. В: СНМД -Дружество по Хигиена. Резюмета на доклади-те, изнесени на III Национален конгрес похигиена. София, 13 - 15 X, 1977, стр. 34.

27. Раднева Р, Торбова С, Ценова Б, Цветков Д,Боев М. Акустични характеристики на жилищ-ната среда и епидемиология на артериалнатахипертония. Хигиена и здравеопазване.2001(44);1:14-19.

28. Раднева Р, Ценова Б, Торбова С, Цветков Д,Боев М, Попова К. 24-часово мониториране наартериалното налягане и пулсовата честотапри лица, живеещи в шумни градски зони.Хигиена и здравеопазване. 2001;44(1):19-22.

29. Раднева-Димитрова Р. Проучване на субек-тивната оценка на въздействието на шумавърху отдиха и възстановяването на човеш-кия организъм. В: СНМД - Дружество поХигиена. Резюмета на докладите, изнесени наIII Национален конгрес по хигиена. София, 13- 15 X, 1977, стр. 34.

30. Раднева-Димитрова Р. Шумът в жилището истепента на общата обезпокоеност на обита-телите. В: СНМД - Дружество по Хигиена.Резюмета на докладите, изнесени на IIIНационален конгрес по хигиена. София, 13 -15 X, 1977, стр. 34.

31. Ценова Б, Раднева Р, Цветков Д, Вуков М.Характеристики на шумовото въздействие всъвременната жизнена среда и техните здрав-ни аспекти. Хигиена и здравеопазване.2000;5/6:29-33.


Ас. д-р АНГЕЛ М. ДЖАМБоВ

Катедра „Хигиена и екомедицина”,Факултет по обществено здраве,

Медицински университет – Пловдив,бул. „Васил Априлов” 15-А,

4002 Пловдив, БългарияE-mail: [email protected]


Asst. Prof. ANGEL M.DzhAMBoV, MD

Department of Hygiene andEcomedicine, Faculty of PublicHealth, Medical University ofPlovdiv, 15-A Vassil Aprilov

Blvd., 4002 Plovdiv, BulgariaE-mail: [email protected]


Анализ на познанието за епидемиологията и клиниката на бисфосфонатно-асоциираната остеонекроза на челюстите

М. Христамян1, Й. Стоилова1, П. Печалова2, В. Христамян3,4

1Катедра по Епидемиология и МБС, Медицински университет Пловдив, България; 2Катедра по Орална хирургия, Медицински университет Пловдив, България;3Клиника по лицево-челюстна хирургия , УМБАЛ „Свети Георги”, Пловдив,България;

4Катедра по Лицево-челюстна хирувгия, Медицински университет Пловдив,България

Analysis of the knowledge of the epidemiology and clinic of bisphosphonate-

related osteonecrosis of the jaws

Hristamyan M.1, Stoilova Y.1, Pechalova P.2, Hristamyan V.3,4

1Department of Epidemiology and Disaster Medicine, Medical University, Plovdiv, Bulgaria;

2Department of Oral surgery, Medical University , Plovdiv, Bulgaria;3Clinic of Maxillofacial Surgery, University Hospital „Sveti Georgi”, Plovdiv, Bulgaria;

4Department of Maxillofacial surgery, Medical University , Plovdiv, Bulgaria.

РЕЗЮМЕ:

Бисфосфонатно асоциираната остео-некроза на челюстите (БАОНЧ) е състоя-ние, описано сред работещите във фабри-ки за кибритени клечки през 19 век подимето „фосфорна челюст“, но докладваноофициално за първи път едва през 2003година. БАОНЧ е страничен ефект от при-ложението на бисфосфонати (БФ) – групамедикаменти, повлияващи костния мета-болизъм. Развитието на състоянието е при-чина за сериозни увреждания и диском-форт на част от увеличаващия се бройпациенти, лекувани с БФ, особено приинтравенозна апликация. Съществуватмножество дилеми и спорове на различнигрупи специалисти относно определение-то, епидемиологията и рисковите фактори,

ABSTRACT

Bisphosphonate related osteonecrosis ofthe jaws (BRONJ) is a condition, firstdescribed in the 19th Century among workersin match factories as "Phossy Jaw", but offi-cially reported in 2003. BRONJ is a side effectfrom the use of bisphosphonates (BP), agroup of drugs that affect bone metabolism.The development of the condition is the causeof serious impairment and discomfort of partof the increasing number of patients treatedwith BР, especially by intravenous application.There are several dilemmas and controversiesamong different groups of experts on the def-inition, epidemiology and risk factors, mecha-nisms of development, classification, clinicalmanifestations and approaches to treatmentand prevention. That’s why, in the literature,


INTRoDuCTIoN

Bisphosphonates (BP) are a group ofdrugs, synthesized in the late 1960s, thataffect the cycle of bone metabolism by sup-pressing the processes of absorption. Theyare used to treat diseases that feature highbone resorption - multiple myeloma, osteo-lytic bone metastases, osteoporosis, Paget'sdisease , fibrous dysplasia , McCune-Albright syndrome, hypocalcaemia of malig-nancy and are most commonly administe-red by intravenous infusion(IV).

Bisphosphonates are classified in twogroups based on their chemical structure(24): • Aminobisphosphonates contain nitrogen

in the side atomic chain of their molecule,with a much stronger effect

• Non-aminobisphosphonates - metaboli-zed by osteoclasts to inactivate non-hyd-roxylysine adenosine triphosphate-analo-gues that have a direct cytotoxic effectwhich lead to apoptosis (40).Although they have a good safety profile,

these drugs can also cause some seriouscomplications after oral and especially long-

term intravenous use. Treatment with bisp-hosphonates may result in short or long-term side effects such as renal failure, upperintestinal tract disorders, esophageal can-cer, acute early phase reaction, uveitis andother ocular inflammatiory diseases ,arthralgia, fever, muscle pain , hypocalcae-mia, bone turnover suppression, risk of aty-pical femoral fractures. In recent years,reports of the occurrence of cases of bisp-hosphonate-related osteonecrosis of the jaw(BRONJ) have increased significantly (31).

hISToRy

Pure phosphorus was first obtained fromanimal bone tissue in the early 19th century.The compound immediately found use inthe matchmaking industry in the form ofwhite phosphorus (yellow phosphorus).

The infamous epidemic of a disease cal-led "phossy jaw" began around 1858 andcontinued until 1906, after which onlyoccurred sporadic cases. The epidemic waslinked to the "yellow phosphorus", keyingredient in the matches. Depending onthe duration of exposure, many workers

механизма на възникване, класификация-та, клиничните изяви и подходите за лече-ние и превенция. Това говори, че се касаеза изключително сложен и многофакторенпроцес, изискващ внимателно мониторира-не и индивидуален подход към всеки паци-ент. Тъй като състоянието се счита за необ-ратимо, усилията трябва бъдат насочениосновно към неговата превенцията, кактопреди, така и след началото на БФ терапия.От ключово значение е увеличаването наинформираността по проблема на денталнилекари, онколози и пациенти и готовносттаим да работят заедно като екип с цел свеж-дане до минимум риска от развитие на товаусложнение.

Ключови думи: Бисфосфонати, Бис -фос фонатно асоциираната остеонекроза начелюстите, епидемиология, рискови факто-ри, превенция

BRONJ is described as a complicated processrequiring careful monitoring and individualapproach to each patient. As the condition isconsidered to be irreversible, efforts should bedirected primarily to its prevention, bothbefore and after the onset of BP therapy. It iscrucial to increase the awareness of dentalpractitioners, oncologists and patients andtheir willingness to work together as a team tominimize the risk of developing this complica-tion.

Key words: Bisphosphonates, Bis -phosphonate Related Osteonecrosis of theJaws, Epidemiology, Risk Factors, Prevention


exposed to heated fumes, containingphosphorus, developed a condition charac-terized by a painful bone exposure in theoral cavity, whereas the people, working inthe office environment didn’t have that con-dition. After finding effective and relativelysafer substitutes, the European countriessigned an agreement (the BerneConvention, 1906), obliging the participantsnot to produce or import matches contai-ning white (yellow) phosphorus (23).

Yellow phosphorus has a simple chemicalstructure (P4O10) and in a medium rich inoxygen and carbon dioxide environment,such as the oral cavity, and in the presenceof amino acids (lysine), the compound canform bisphosphonates almost identical toalendronate (Fosamax, NovartisPharmaceuticals , East Hanover, NJ) andpamidronate (Aredia, NovartisPharmaceuticals). A century later, the causeof the "phossy jaw" disease was establis-hed, and it was verified that the 19th-cen-tury epidemic actually represented cases ofbisphosphonate-related osteonecrosis of thejaws, long before the pharmaceuticalindustry began to market and sell thesedrug. The contemporary cases of BRONJrepresent a second "phossys jaw" epidemic(23).

More recent data on BRONJ dates back to2002, when 9 months after the intravenousbisphosphonate zoledronic acid was appro-ved by marketing regulators, the FDA recei-ved reports of nine cancer patients whowere treated with zoledronic acid and unex-pectedly developed a condition characteri-zed by exposed jawbone (15). The first offi-cial report was published in 2003 by Marx,who reported 36 cases of "painful exposureof the lower and upper jaw bone in patientsundergoing bisphosphonate treatment withpamidronate and zoledronate (25). Thatsame year Migliorati reported five cases(27), Carter and Gross - four (12), Wang -three (39). In 2004, Ruggierro and co. pub-lished 63 cases of osteonecrosis of the jaw-bone in patients treated with bisphosphona-

tes (35). In 2004, Novartis, the manufactu-rer of Aredia and Zometa, reported to heal-thcare professionals about additional war-nings to these products, regarding the riskof development of osteonecrosis of the jaws(19). In 2005, the manufacturer officiallydeclared 475 cases of BRONJ and the pre-cautions were changed for all members ofthe bisphosphonate group, including oralbisphosphonates.

Information about BRONJ continues to bepublished in the scientific literature, butunfortunately, the number of cases both inBulgaria and globally remains unknown.

The first practical guidelines on how toensure safe dental treatment for patients onbisphosphonate therapy are published in theJournal of Clinical Oncology Practice inJanuary 2006 (34).

NoMENCLATuRE AND DEFINITIoN

In Bulgaria, the most common acronym,used for describing the complication isBAONJ (BP associated osteonecrosis of thejaws). Various abbreviations are available inthe English-language literature - BRONJ,BRON, BON, BAONJ, ONJ. The recognitionof jaw necrosis as a complication associatedwith the use of other drugs, excluding BP,provoked the American Association of Oraland Maxillofacial Surgery (AAOMS) torecommend the term "medication-associa-ted osteonecrosis of the jaw" (MRONJ) (6).

Two working definitions for BRONJ areproposed. The first one is presented by aspecial task force convened by the AmericanSociety for Bone and Mineral Research defi-nes BRONJ as the presence of exposed bonein the maxillofacial region that does not healwithin eight weeks after identification by ahealth care professional. (21) In the wor-ldwide and in the Bulgarian literature , thesecond defitition , provided my AAMOS, andupgraded in their last position paper from2014 (6), is adopted: „necrosis of the jawbone, related or unrelated to dental proce-dures, persisting for more than 6 to 8


weeks, refractory to conservative treat-ment, in patients having no history of priorradiotherapy in the affected area, treatedintravenously with amino-containing bisp-hosphonates for at least one year, or orallyfor a much longer period, for a generaldisease causing bone resorption.” (25). Thisdefinition is also adopted by an internationaltask force on osteonecrosis of the jaw (20).Some authors consider this definition to beinaccurate. Fedele et al. believe that usingthe traditional definition could result in 25%of patients remaining undiagnosed (16).

EPIDEMIoLoGy AND RISK FACToRS

The epidemiology of BRONJ still remainsunclear due to the inconsistency and limita-tions of available studies, including the lackof specific ICD (International Classificationof Diseases) code, unrealistic and reducedreporting in drug surveillance systems, therecent introduction of preventive measures,short-term monitoring and lack of cumulati-ve rates of morbidity in the long term. It issurprising that BRONJ was not found in theinitial testing of BF and the first cases wereofficially reported and published in 2003(11).

Most authors present statistical studieson patients with multiple myeloma, breastcancer, prostate cancer and lung carcinoma,as these are the largest groups of patientstreated with bisphosphonates. Sook-BinWoo et al. quote a study of 1203 cases, inwhich 7 % of the 904 patients with multiplemyeloma and 4 % of the 299 patients withlung carcinoma developed osteonecrosis ofthe jaws, related to the usage of BF (40).Kohl et al. report a wide incidence of BRONJfrom 0 to 27.5% , with/at an average inci-dence of 7% (22). In one of the latest revi-ews on the topic, Rugani et al. conducted asearch in PubMed and found a total inciden-ce of 2.09% in breast cancer patients, 3.8%in prostate cancer patients and 5.16% inmultiple myeloma patients (33).

Very little information can be found aboutthe incidence of BRONJ in patients withosteoporosis. Solomon et al. found a morbi-dity rate of 0.028% to 4.3% in the literatu-re, and in their own cohort studies, the inci-dence was 0.02% (95% CI 0.004% -0.11%) (37).

Risk factors related to bisphosphonates

Type of BF: There is no absolute unanimi-ty in the literature on which BF causes morefrequent development of BRONJ, but scien-tific evidence supports the prevailing viewthat Zoledronic acid has highest risk-poten-tial (6, 9, 11, 26).

Route of administration: The risk is hig-her with intravenous BP (6, 9, 11, 26) thanwith oral BP, but this factor may be closelyrelated to their widespread use in patientswith malignant neoplasms that receive sig-nificantly higher overall doses and their tre-atment usually has longer duration (11).According to the literature, the lower risk fororal BP intake is due to poor intestinalabsorption (0.64%) (11).

Administration schedule: The frequencyof administration is also recognized as a riskfactor by many oncologists who now pre-scribe intravenous zoledronate every 3 or 6months, which is different from the manu-facturer's recommendation for 3 weeks toone month. This reduction in the frequencyof administration has led to a decrease inthe number and severity of the cases (26).

Dose: It is well known that for almost anypharmaceutical product, increased dosingresults in a greater effect, and respectively,to a greater and more significant severity ofthe complications. Available data shows ahigher risk of developing osteonecrosis ofthe jaw by increasing the total dose of BPadministered intravenously monthly to can-cer and hematology patients (11).


Duration of treatment: The incidence ofdevelopment of BRONJ is higher with longertreatment duration, especially when theduration of therapy exceeds four years. Thisperiod may be reduced by the presence ofcertain concomitant diseases, chronic gluco-corticosteroids, or co-administration of BFwith angiogenesis inhibitors (9).

Risk potential: Alendronate and zoledro-nate have the highest potency and causethe majority of the cases of BRONJ (26).

Risk factors associated with the dento-alveolar system

Anatomical comorbidity

The alveolar process, both in the lowerand upper jaw, is the most vulnerable part.Typically, cases of spontaneously occurringnecrosis are within alveolar bone (26).

Mandible / Maxilla: Most of the studiesbased on a large number of cases show atendency for BACCP to appear more fre-quently in the lower jaw than in the upperjaw (2: 1) (26). Rarely, both jaws are affec-ted. (4.5%). (6).

Torus: The mandibular torus, palatinetorus and bone exostosis are places, thatare more easily locally traumatization in thecourse of day-to-day activities,and are oftenaffected. (11, 26).

Dental diseases and procedures

Dental procedures: According to the lite-rature, the prevalence of bisphosphonate-associated osteonecrosis of the jaws asso-ciated with previous dental procedures ispredominant compared to the so-calledspontaneous BRONJ. Woo Sook-Bin and call.found that between 33% and 86% of thecases were manifested after various dentalprocedures (21). Dental extraction (6, 9,

11, 26) has the leading role - initiator in upto 62% of the cases (26).

Hyper oclusion / Prosthetics: Excessivechewing forces initiate cases of BRONJ, dueto the higher velocity and remodeling ratesin the alveolar bones (26).It is assumed thatthe mechanism of development of necrosisafter putting dentures is analogous (6, 11).

Periodontal diseases: Active periodontalinflammation is associated with the deve-lopment of osteonecrosis of the jaw in pati-ents receiving BP (6, 9, 11, 26). This is dueto further increased bone remodeling stimu-lated by the inflammatory process.

Surgical interventions in the alveolarbone: Surgical manipulations, other thantooth extraction - dental implants, endodon-tic, periodontal and peri-implant surgery,also increase the rate of bone remodelingand can therefore lead to the appearance ofBRONJ. (6, 9, 11,26).

other risk factors

Many authors discuss the role of additio-nal to the BP intake therapy, such as che-motherapy (9, 26), corticosteroids (6, 9, 11,26), antiangiotensive agents (6, 9, 11), aswell as the influence of age and gender (9,11), geographic location (9), smoking, alco-hol, obesity (6, 9, 11, 26), genetic factors(6, 9, 11), type of tumor (6, 9, 11, 26) andaccompanying diseases - hypocalcemia,hyperparathyroidism and disturbances inbone mineralization (11), diabetes, anemia(6, 26).

However, about 25% of the BAONC casesare spontaneous (26).

Abu-Id et al have proposed a predictivescale for risk of development of BRONJ:1. At high risk: patients with malignancy

receiving intravenous BF (Zoledronate orPamidronate) and / or with a history ofchemotherapy, radiotherapy or ongoingexogenous steroid use.


2. At low risk: patients receiving oral BF wit-hout a history of chemotherapy, radiothe-rapy or ongoing exogenous use of stero-ids (mostly patients with non-steroid-induced osteoporosis) (1).

PAThoPhySIoLoGy

Although the first cases of BRONJ werereported more than a decade ago, the pat-hophysiology of this disease has not beenfully elucidated. A source of debate amongclinicians and researchers is the potentialmechanisms underlying BRONJ (36). Theleading hypotheses for explaining the uni-que location in the jaws include the role ofremodeling or excessive suppression ofbone resorption, inhibition of blood flow,permanent microtrauma, suppression ofinnate or acquired immunity, deficiency ofvitamin D, BP toxicity on soft tissues andinfection / inflammation. None of thesehypotheses can explain all the cases.

CLASIFFICATIoN

In 2006, Ruggiero et al. proposed an ONJclassification comprising three stages :stage 1 = bone exposure but without signsor symptoms of infection; stage 2 = boneexposure/necrosis with clinical evidence ofinfection; stage 3 = the above manifestati-ons and also alterations such as pathologicalfractures, extraoral fistulas or osteolysisextending to the inferior mandibular margin(34).

In 2007 the AAOMS adopted this classifi-cation, though in addition to the group ofpatients with BRONJ (with its three stages),it included another group of patients com-prising individuals at risk. These patientswere defined as subjects without evidentexposed or necrotic bone or symptoms, butwho have been treated with oral or intrave-nous BPs (4).

In the year 2009, the AAOMS added astage 0 to its classification, involving altera-tions (pain, tooth mobility, fistulas, radio-graphic changes, etc.) that may have been

due to treatment with BPs, but withoutexposed bone. The risk of progressiontowards more advanced stages of the disea-se was not known at that time (5).

In 2014, AAOMS updated its classification(6):

At risk category - No apparent necroticbone in patients who have been treated witheither oral or IV bisphosphonates

Stage 0 - No clinical evidence of necroticbone, but non-specific clinical findings,radiographic changes and symptoms

Stage 1 - Exposed and necrotic bone, orfistulae that probes to bone, in patients whoare asymptomatic and have no evidence ofinfection

Stage 2 - Exposed and necrotic bone, orfistulae that probes to bone, associated withinfection as evidenced by pain and erythe-ma in the region of the exposed bone withor without purulent drainage

Stage 3 Exposed and necrotic bone or afistula that probes to bone in patients withpain, infection, and one or more of the fol-lowing: exposed and necrotic bone exten-ding beyond the region of alveolar bone,(i.e., inferior border and ramus in the man-dible, maxillary sinus and zygoma in themaxilla) resulting in pathologic fracture,extra-oral fistula, oral antral/oral nasal com-munication, or osteolysis extending to theinferior border of the mandible of sinus floor

n 2016, Bakardjiev, as a result of his cli-nical observations, proposed the formationof sequesters as an addition to the AAOMSStage III criteria (8).

CLINICAL MANIFESTATIoN

Clinically, intraoral lesions at BRONJ looklike spaces of yellow-white hard bones, witha soft or thickened board. Their dimensionsmay vary from a few millimeters to a fewcentimeters. These areas of exposed andnecrotic bone may remain asymptomatic forweeks, months, or even years, and show notendency for spontaneous recovery anddiminishing, which is a hallmark of BRONJ


(3) Clinical manifestation occurs in thedevelopment of inflammation in surroun-ding soft tissues. The main symptom is aprotracted, dull, irritating pain in the jaw,which is suppressed by analgesics, but doesnot disappear completely. Usually, mobilityof teeth is observed, which can lead to theirloss, periosteum, gingivitis. The surroundingmucous membrane is hyperemised, painful,in some places distorted and covered bynecrotic scarring (3). The presence of extra-and intraoral fistulas is typical. As the pro-cess progresses, the lesions grow and enga-ge vast spaces in the oral cavity, leading toeating, speech and oral hygiene difficulties.Cases of pathological fractures of the jawbone have been observed.

In the literature are described somecases of a subclinical form of BRONJ, whichoccurs without bone exposure. The clinicalmanifestations of this form of BRONJ inclu-des unexplained jaw pain, fistula formation,edema, mobility of the teeth, and pathologi-cal fractures (30).

PARACLINICAL TESTS

Radiological /X-ray findings in patientswith BRONJ include osteosclerosis (mostcommonly) , osteolysis, dense woven bone,thickened lamina, subperiosteal bone depo-sition, and inability to post-operative remo-deling.

In the early stages of disease develop-ment, X-ray changes may be absent (40).Some authors recommend computedtomography (10). Chiandussi et al. believethat late changes in the jaws in bisphospho-nate-associated osteonecrosis are visualizedby X-ray, and CT scanning or nuclear mag-netic resonance is better for earlier detecti-on of lesions (13). Other authors recom-mend scintigraphy of the jaw with Tc-99mmethylene diphosphonate (28).

Microbiological testing most commonlyfinds Actinomyces druses. In a 10-yearreview of the literature on microbiologicaldata, Hinson et al. report presence of acti-

nomyces in 68.8% of the cases, followed byStreptococci, isolated in 54.7% of the orallesions reported in the review (17).

Histopathological analysis shows theabsence of a finding comparable to the bisp-hosphonate-treated disease, but the pre-sence of a necrotic bone surrounded by bac-teria that do not enter it (32).

TREATMENT

Despite the significant number of publica-tions on the subject, a particularly vagueaspect, and certainly one of the most con-troversial, is the treatment of BRONJ. Thereis still no prospective evidence-based studywith long-term follow-up to guide treat-ment.

In particular, it is not known whether theaffected persons should receive symptoma-tic therapy with pain and infection controland possibly minimally invasive debris of thesurface of the necrotic bone, or resectivesurgical methods should be used. The sub-ject is complicated because some patientshave minimal symptoms and can thereforebenefit from minimally invasive therapy. Incontrast, there are others, with a painful,advanced disease that does not respond tosymptomatic therapy and require moreaggressive intervention.

With respect to surgical treatment, thereis still controversy among researchers onthe type of surgery and the time of perfor-ming it. One group of authors (25) recom-mend superficial curettage and prolongedconservative treatment. Another group ofauthors (2) recommend hyperbaric oxyge-nation, superficial debridement, laser treat-ment, and in advanced cases – aggressiveresection of the diseased bone and soft tis-sue. Hoff, et al (18), reported healing pro-cess in 23% of the patients treated withconservative therapy only. Some authorscompletely remove the diseased bone,covering the site with mucosa, and reportgood results in 85% to 100% of the patients(38). Similar results were reported by anot-


her author (29), who identified the bounda-ries of the diseased bone using fluorescentlight. This technology has been used inBulgaria from Bakardjiev for two years. Theanalysis of a number of publications showsthat in the early stages of the disease(AAOMS 1 and 2) most authors administermedication therapy and conservative treat-ment, and in stage 3 - resection and/orsequestrectomy are performed. The methodproposed by Otto S et al. (29), reported byAssaf AT et al. (7), uses fluorescent light forvisualization of vital and non-vital bone, is anew and promising method for demarcationwhen performing resection of the jaw

Eventually, regardless of the stage of thedisease, areas of necrotic bone that are thesource of chronic soft tissue irritation andsequestration should be removed or recon-stituted so that the soft tissue healing isoptimized.

In Bulgaria, Pechalova et al. (31), bysummarizing the information found in theliterature, provided a algorithm for treat-ment of patients, suspected to have BRONJ:1. A careful clinical examination to find the

location and volume of exposed necroticbone

2. Imaging diagnostics of the affected jaw -X-rays, CT scans, MRI, bone scintigraphy

3. Providing material for histopathologicalanalysis to exclude the presence of a sys-tematic

process in the jaw (in case of multiple mye-loma, a metastasis in case of oncologicaldiseases or a primary neoplasm)

4. Providing material for microbiologicalexamination with emphasis on fungal orother

pathological bacterial infections290 A Textbook of Advanced Oral and

Maxillofacial Surgery5. Treatment- conservative and surgical

There is no consensus on the need fordiscontinuation of bisphosphonate treat-ment after diagnosing osteonecrosis of thejaw. Marx suggests that behavior in terms of

bisphosphonate therapy in cancer patientsshall be discussed with the oncologist on thepurpose of determining the benefit to riskratio, in view of the long half-life (10 years)of bisphosphonates(24). Dunstan et al. sug-gest that bisphosphonate therapy be dis-continued (14). Expert group guidelines arealso divergent (9). The decision should betaken individually on a case-by-case basis,taking into account the expected benefitsand harm to the individual patient.

PREVENTIoN

To date, there is no evidence-based the-rapeutic strategy with regard to BACCP andthe condition is considered irreversible, sothe focus of the medical community's focusis on its prevention capabilities (9, 11 ,40).

Prior to initiation of bisphosphonate therapyThe American Association of Clinical

Oncology (ASCO), AAOMS, the InternationalTask Force on Osteonecrosis of the Jaw andthe European Medicines Agency (EMA)recommend, that all patients planning anti-resorptive or angiogenesis inhibitor therapyshould undergo a full dental examinationprior to beginning the therapy. Preventivedental activities include extraction of thenon-prospective teeth, endodontic treat-ment and retreatment, periapical surgery,the main purpose of which is that the pati-ent does not require dental manipulation aslong as possible (20).

Pechalova et al. recommend the followingplan for behavior (31):1. A thorough clinical examination of the

dentition and oral cavity, panoramicradiograph and, by the discretion of thedoctor, targeted periapical radiographs asa mandatory required minimum.

2. Conducting dental treatment aimed at eli-mination of infection and the need forinvasive procedures in the short- andmid-term futureAll invasive dental procedures should be

performed at least one month before initia-


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ting bisphosphonate therapy in order toallow sufficient time for recovery of the jaw-bone (31).

After the start of bisphosphonate therapyAfter the start of bisphosphonate therapy

patients are subject to preventive check-upsat every four months, with radiographic exa-mination, which should seek vigilantly for thepresence of osteolysis, osteosclerosis, andexpansion of periodontium and involvementof furcations. If additional treatment is requi-red, non-invasive dental procedures are pre-ferred (31).

Oral hygiene should be carefully monito-red. Patients should be encouraged to quitsmoking (9).

Patients should be asked about any plan-ned dental procedures prior to administrationof each dose of an anti-resorptive agent and

should be reminded to avoid invasive dentalprocedures (tooth extraction, implants, andany other procedures involving manipulationof the jawbone or perineum ) while beingtreated with these types of drugs (24)

CoNCLuSIoN

BRONJ is a complication, associated withBP intake, that seriously degrades the qualityof life of many patients. As the condition isconsidered to be irreversible, and treatmentoptions are still a complex and controversialsubject among the clinicians, our efforts sho-uld be directed primarily to its prevention,both before and after the onset of BP thera-py. It is crucial to increase the awareness ofdental practitioners, oncologists and patientsand their willingness to work together as ateam to minimize the risk of developing thiscomplication.


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ХРИСтАМЯН М.

Катедра по епидемиология и медицина при бедствия,Медицински университет,

Пловдив, България; e-mail: [email protected]


hRISTAMyAN M.

Department of Epidemiologyand Disaster Medicine, MedicalUniversity , Plovdiv, Bulgaria;e-mail: [email protected]

bulgarian medicine issn 1314-3387basa.bg/en/images/files/br_02_2017.pdf · МУ Пловдив,...

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