Prevalence of Coeliac Disease in Adult Saudi Patients with

Symptoms of Irritable Bowel Syndrome; pilot study.

Shendy M. Shendy* and Nihal Al-Assaly** , Naema I. El-Ashry**.

*Tropical, Hepatology and gastroenterology department, **Clinical biochemistry dep, Theodor Bilharz research Institute.

-Accepted for publication in Journal of Arab Society for Medical Research (JASMR), 29- 12- 2006

Abstract:

Few recent studies have found higher prevalence of coeliac disease among patients with diagnosis of irritable bowel syndrome (IBS) than general population (3-11% vs. 0.2-0.6%). Similar studies showed that coeliac disease is as common in Middle Eastern countries as in Europe; in both the general population and at-risk groups. The aim of this work is to estimate the prevalence and the potential clinical consequences of coeliac disease testing in adult Saudi patients with IBS. Materials and methods: This is a prospective pilot study including 320 Arab patients with features compatible with IBS as defined by Rome III criteria without any other co-morbidity. The age of patients ranged between18-70 years. All patients were subjected to good history taking, clinical examination, and some investigations if needed such as stool, urine, CBC, liver enzymes, kidney function tests, ECG, electrolytes, H pylori serology, upper and lower endoscopy when indicated. Those diagnosed as having persistent criteria of IBS were tested for coeliac disease by IgA and IgG anti-gliadin antibodies, anti endomysial antibodies (EMA) IgA and anti-TG2 (IgA and IgG). Upper endoscopy and duodenal biopsies were done and gluten free diet was implemented for only those with positive serological test. The same tests were repeated after period of about 6 months. Results: Anti-gliadin antibodies were found positive in 15/320(4.69%) patients (14 with IgA and 13 IgG), EMA IgA in 13/320 (4.06%), anti-TG2 IgA in 12/320 (3.76%) and anti- TG2 IgG in 13/320 (4.06%). Abdominal pain, diarrhea, dyspepsia, postprandial distress, epigastric pain, distension and chronic diarrhea were significantly higher and more common in combinations in those with positive serology in comparison to serologically negative patients (P < 0.05). Haemoglobin level, serum iron, albumin and calcium were found to be significantly lower in those with positive serology in comparison to serologically negative patients (P < 0.05). All these parameters improved significantly after gluten free died (GFD) for about 6 months (P< 0.05). Only 11 patients (74.44% of those with positive serology and 3.49% of total patients) were diagnosed by biopsies as compatible with coeliac disease of which, two patients have family history of coeliac disease in first degree relatives. After gluten free died (GFD) for about 6 months, seroconversion to negative tests occurred in 6 patients for AGA-IgA, 4 for AGA- IgG, 3 for EMA IgA, 5 for Anti-TG2 IgA and 5 for Anti-TG2 IgG. Also, the grade of histopathology showed complete healing in 4 patients and improvement to lower grades in 4 patients after GFD. Worsening occurred in one case and still 7 cases showed the same grade of the disease. Conclusion: It is concluded from this study that minimally symptomatic coeliac disease can easily be mistaken for IBS. The presence of many persistent gastrointestinal symptoms in addition to the lower serum levels of some nutritional parameters must alert the physicians to screen for coeliac disease. Any serological test can be used for the screening but this must be confirmed by tissue diagnosis which is the gold standard for diagnosis. Finally, screening for coeliac disease among patients with IBS must be considered to offer better prognosis to these patients simply by gluten free diet.

Introduction:

Irritable bowel syndrome (IBS) is a highly prevalent, multi-symptomatic, gastrointestinal motility disorder that has a wide clinical spectrum. This disease is associated with gastrointestinal dysmotility and/or visceral hypersensitivity.

Although the recent trend has been to consider IBS a condition that can be diagnosed based on symptom criteria, as opposed to regarding it as a 'diagnosis of exclusion' only to be made after extensive testing, a limited screen for other diseases is recommended in patients with symptoms suggestive of IBS.[2,3,4] The yield of haematology, chemistry and thyroid function tests, sigmoidoscopy or colonoscopy, and microbiological stool studies in patients with 'suspected' IBS ('s'-IBS) is modest.[2] Recent studies suggest that the prevalence of coeliac disease, a gluten-sensitive enteropathy characterized by intestinal villous blunting and malabsorption,[5] is 3-11%[6,7,8] among patients diagnosed with IBS, compared with 0.02-0.65%[6,7,8,9] in the general population. Coeliac disease may be present in patients with IBS-like symptoms with diarrhoea or constipation predominance, or alternating bowel habit.[6,7]

While a classic presentation of coeliac disease consists of steatorrhea and weight loss, the presentation can be more subtle, including symptoms characteristic of IBS (abdominal discomfort, altered defecation, bloating, gas), and diagnosis can be delayed. [10,11] 24-37% of coeliac disease patients were initially diagnosed with IBS.[10,11,12] Whereas IBS is a chronic condition with no cure and with limited therapeutic options, adherence to a gluten-free diet may improve quality of life and prevent long-term complications in coeliac disease.[5,12-19]

Recent studies showed that coeliac disease is as common in Middle Eastern countries as in Europe; in both the general population and at-risk groups, e.g. patients with irritable bowel syndrome or type 1 diabetes. Also, these studies showed that presentation with non-specific symptoms or no symptoms is as common in the Middle East as in Europe. Frequent exposure to wheat protein may lead to immune tolerance, leading to milder symptoms that may be misdiagnosed as irritable bowel syndrome or unexplained gastrointestinal disorders (20).

Symptom overlap and comorbidity between IBS and other gastrointestinal motility disorders (eg, chronic constipation, functional dyspepsia, gastroesophageal reflux disease), with gastrointestinal disorders that are not related to motility (eg, celiac disease, lactose intolerance), and with somatic conditions (eg, fibromyalgia, chronic fatigue syndrome), are frequent. The clinical associations and pathophysiologic links between IBS and these disorders continue to be explored (22).

Measurement of IgA antibody to human recombinant tissue transglutaminase (TTG) is recommended for initial testing for CD. Although as accurate as TTG, measurement of IgA antibody to endomysium (EMA) is observer dependent and therefore more subject to interpretation error. Because of the inferior accuracy of the antigliadin antibody tests (AGA), the use of AGA IgA and AGA IgG tests is no longer recommended for detecting CD. In individuals with known selective IgA deficiency and symptoms suggestive of CD, testing with TTG IgG is recommended. It is recommended that confirmation of the diagnosis of CD require an intestinal biopsy in all cases. Because the histologic changes in CD may be patchy, it is recommended that multiple biopsy specimens be obtained from the second or more distal part of the duodenum. There is good evidence that villous atrophy (Marsh type 3) is a characteristic histopathological feature of CD (34). The presence of infiltrative changes with crypt hyperplasia (Marsh type 2) on intestinal biopsy is compatible with CD but with less clear evidence. The presence of infiltrative changes alone (Marsh type 1) on intestinal biopsy is not specific for CD in children. Concomitant positive serological tests for CD (TTG or EMA) increases the likelihood such an individual has CD. In circumstances where the diagnosis is uncertain additional strategies can be considered, including determination of the HLA type, repeat biopsy or a trial of treatment with a gluten-free diet (GFD) and repeat serology and biopsy. The diagnosis of CD is considered definitive when there is complete symptom resolution after treatment with a strict GFD in a previously symptomatic individual with characteristic histologic changes on small intestinal

biopsy. A positive serological test that reverts to negative after treatment with a strict GFD in such cases is further supportive evidence for the diagnosis of CD (33).

The detection of high titres of antigliadin antibody (AGA), antireticulin antibody (ARA) or antiendomysium (EMA) antibody at the time of diagnosis, and the subsequent decrease in titres after removal of dietary gluten, add specificity to the histologic findings of coeliac disease (31). The identification of tissue transglutaminase (tTG 2) as the main antigen of EmAs (28 ) allows a new diagnostic approach to CD. A large number of ELISA methods, mainly based on commercially available guinea pig tTG, have been produced. Anti-tTG antibodies are highly sensitive and specific for the diagnosis of CD (29,30 ). Enzyme-linked immunosorbent assay (ELISA) tests for IgA anti-tTG antibodies are now widely available and are easier to perform, less observer-dependent, and less costly than the immunofluorescence assay used to detect IgA endomysial antibodies. The diagnostic accuracy of IgA anti-tTG immunoassays has been improved further by the use of human tTG in place of the nonhuman tTG preparations used in earlier immunoassay kits.

In one study of 288 patients with significant complaints and physical signs attributable to the lower GIT in Eastern Province of the Kingdom of Saudi Arabia; 128 patients (44.5%), sigmoidoscopy and rectal and/or colonic biopsies did not reveal any pathological abnormalities. These patients were considered to have various disorders such as irritable bowel syndrome or parasitic infestation. Eighty-one patients (28%) were found to have mild to moderate non-specific colitis or proctitis. In another 49 patients (17%) the diagnosis of schistosomiasis mansoni was made. Ulcerative colitis and colorectal carcinoma were detected in only 11 (4%) and 4 (1.5%) patients respectively. In the remaining 15 patients (5%), other lower GIT diseases were found (21). It has been suggested that patients with IBS should be tested for coeliac disease.[2,6,7]

Our aims were to estimate the prevalence and the potential clinical consequences of coeliac disease testing in adult Saudi patients with IBS.

Materials and methods:

This is a prospective pilot study evaluating adult patients attending GIT outpatient clinic in Riyadh city in SA. Those patients with symptoms suggestive of IBS and those also diagnosed after exclusion as IBS were included in the study. The study included 320 patients. All patients were Arabs. They were 184 males and 136 females. Their age ranged from 18-67 years. They were chronic patients attending the clinic for more than 6 months complaining of abdominal pain with features compatible with IBS as defined by Rome III criteria.

Exclusion criteria:1- Those with age below 18 years or above 70 years2- Presence of red flag signs or alarm features as severe unrelenting diarrhoea, nocturnal

symptoms, unintentional weight loss, haematochezia, a family history of organic gastrointestinal diseases such as IBD, celiac sprue or malignancy

3- Symptoms or signs of coeliac disease or those already on gluten free diet being diagnosed already as coeliac disease.

4- Those with other medical illnesses or on chronic medications for organic disease.5- Those who did not agree to continue follow up and giving consent to study.6- Females during pregnancy or breast feeding

All patients were subjected to good history taking, clinical examination, and some investigations if needed such as stool, urine, CBC, liver enzymes, kidney function tests, ECG,

electrolytes, H pylori serology, upper and lower endoscopy when indicated. Those with mixed or atypical presentation were investigated properly for other diagnoses and then managed accordingly. Those diagnosed as having persistent criteria of IBS were included in the study. As serological tests and small bowel biopsy remain the cornerstones of diagnosis of coeliac disease (23), these patients were subjected to:

1- Serum IgA and IgG anti-gliadin antibodies (AGA), Anti endomysial antibodies (EMA) IgA (The test result is reported simply as positive or negative, since even low titers of serum IgA endomysial antibodies are specific for CD) and Anti-TG2 (IgA and IgG).

2- Upper endoscopy for those with positive serology and histopathology of duodenal biopsy.

3- Gluten free diet for those with positive serology. 4- All the above tests were repeated after period of 6-12 months.

During endoscopy of patients with any positive serological tests, 4 to 6 duodenal biopsies were obtained. An experienced pathologist who was blinded to the patient’s history and antibody assay results assessed the mucosal biopsy sections for pathologic features of CD. Diagnosis of CD was made when there were an increased number of intraepithelial lymphocytes with associated subtotal or total villous atrophy (32). Histological grading was done according the conventional system which grades the mucosal findings as normal, slight partial villous atrophy, marked partial villous atrophy, subtotal and total villous atrophy and Marsh system (34) which classified the histological changes of CD as Type 0 or preinfiltrative stage (normal), Type 1 or infiltrative lesion (increased intraepithelial lymphocytes), Type 2 or hyperplastic lesion (Type 1+ hyperplastic crypts), Type 3 or destructive lesion (Type 2 + variable degree of villous atrophy) and Type 4 or hypoplastic lesion (total villous atrophy with crypt hypoplasia). Type 3 has been modified to include Type 3a (partial villous atrophy), Type 3b (subtotal villous atrophy) and Type 3c (total villous atrophy) (35).

Antigliadin antibody assay (36)

IgA and IgG AGA titres were determined by means of enzyme-linked immunosorbent assay (ELISA). Gliadin was prepared from wheat gluten (product no. G-3375; Sigma Chemical Company, St. Louis, Mo.) in 70% ethanol (1 mg/mL). A 200-µL aliquot of gliadin (5 µg/mL in carbonate buffer [0.015M sodium carbonate + 0.03M sodium bicarbonate adjusted to pH 9.6 in water]) was added to each well of a microtitre ELISA plate (Falcon 3915; Becton Dickinson Labware, Lincoln Park, NJ) and kept overnight at 4°C. Subsequently, 200 µL of PBS–BSA 1% Tween (phosphate-buffered saline [PBS; Gibco, Grand Island, NY], bovine serum albumin [BSA; Sigma] and Tween 20 [Sigma]) were added to each well, and the plate was left at room temperature for 11.2 hours. Each serum sample (100 µL) to be tested was added to the wells in quadruplicate at a 1:100 dilution in PBS–BSA 1% Tween. Subsequently, 100 µL of either peroxidase conjugated goat antihuman IgA or IgG (Sigma) was added at a concentration of 1:500 or 1:20 000, respectively, in PBS–BSA 1% Tween, each in duplicate wells. After a 1-hour incubation at 37°C, 100 µL of OPD (ophenylmediamine dihydrochloride) substrate solution (10 mL of buffer [0.2M sodium hydrogen phosphate + 0.1M citric acid] adjusted to pH 5.0), 4 µL of hydrogen peroxide and 4 mg of OPD (Sigma) were added to each well. The plates were then incubated at room temperature, without exposure to light, for 30 minutes. Washing steps (× 3) with PBS-Tween were incorporated after each of the above interaction stages to remove any nonimmobilized species. To stop the reaction, 25 µL of 4N sulfuric acid was added to each well. Optical density was read at 492 nm using an automated ELISA detector. The optimal discriminative ability of IgA AGA and IgG AGA using this method was at an optical density of 0.25 and 0.30, respectively. The celiac patient’s serum was used as a positive control with each batch, to confirm the reproducibility of the AGA assays on different days. Furthermore, appropriate negative controls were routinely carried out, using the anti-human immunoglobulin antibodies without serum. The background optical densities from these negative control wells were subtracted from results with each patient’s serum.Endomysial IgA antibody test: (ImmuGlo™ Anti-Endomysial Antibody (EMA) Test System: indirect immunofluorescence): IMMCO Diagnostics, Inc. 60 Pineview Drive Buffalo, NY 14228-2120 USA. Patient serum (diluted 1 in 10 in 0.5 mol/L phosphate-buffered saline (PBS; pH 7.2) containing 0.2% bovine serum albumin (BSA)) is incubated at room temperature on tissue sections(5 mum cryostat primate smooth muscle sections attached to glass microscope slides coated with poly-L-lysine (Sigma)) for 20 minutes to allow binding of antibodies to the substrate. Any antibodies not bound are removed by rinsing. Bound antibodies of the IgA and IgG class are detected by incubation of the substrate with fluorescein-labeled, anti-human immunoglobulin (Ig A) conjugate (Fluorescein isothiocynate (FITC)-conjugated rabbit anti-human IgA (Dako, Denmark), diluted 1 in 50 with PBS). Reactions are observed under a fluorescence microscope equipped with appropriate filters. The presence of EMA is demonstrated by an apple green fluorescence of the endomysial lining of smooth muscle bundles. The titer (the reciprocal of the highest dilution giving a positive reaction) of the antibody is then determined by testing serial dilutions (26, 27).IgA Anti-tissue transglutaminase:

The anti-tTG IgA antibodies were determined in duplicate using an ELISA-based commercially available kit (Eu-tTG Eurospital, Trieste, Italy) that uses recombinant human/E coli tTG antigen preparation as the coating antigen, and anti-human IgA peroxidase Goat HRP conjugate as the secondary antibody; as indicated by the manufacturer. TMP was used as substrate. Sera with a concentration > 5 arbitrary units (AU)/ml were considered positive.

IgG anti-tissue transglutaminase antibodies (IgG-ANTI-tTG) detection and estimation:IgG-anti-tTG were detected using tTG coated 96 well plates (100 ng/well) from a commercially available IgA-anti-tTG detection kit (Immunopharmacology Research, Catania, Italy) activated with CaCl2 (5 mM), as suggested by Sulkanen and colleagues and Dieterich and colleagues. After four washes with phosphate buffered saline (PBS) 0.15 M, 0.1% human serum albumin (HSA), and 0.05 % Tween 20, the wells were blocked by incubation with HSA (2% in PBS) for two hours at room temperature. After three washes, sera diluted 1:250 in PBS were incubated at room temperature for two hours. The presence of IgG anti-tTG autoantibodies were evaluated after incubation with horseradish-peroxidase conjugated antihuman IgG (1:6000 in PBS, 0.05% HSA, one hour at room temperature) and substrate (1 mg/ml ortho-phenylendiamine (Sigma) in sodium citrate 1 M, citric acid 1 M, and 0.06% H202, 30 minutes at room temperature) as absorbance values of blocked reactions (0.3 M sulphuric acid) at 492 nm were measured on an ELISA reader. Sera were considered positive for IgG -anti-tTG when absorbance (ABS) of a sample was twofold greater than that of the calculated cut off value ((positive control ABS+ negative control ABS)/2). Positive and negative control values were, respectively, the absorbance of pool EMA-IgG positive and negative sera after background subtraction. The antibody levels in patient diluted serum samples were estimated by comparison with the levels on a standard curve (antibody concentration range, 0 to 100 U/ml), and samples yielding a result greater than 100 U/ml were reinvestigated by the use of higher dilutions.

Patients can be considered as one of two groups - 'silent' celiac disease when the patients are symptom free and serologically negative but bear the hallmarks of the disease on histological grounds, and 'latent' celiac disease when serological tests are positive but there are no or minimal histological changes, such as increased density of intraepithelial lymphocytes (IELs) (25).Statistical analysisResults were analysed using SPSS 12 for windows software.

Results:This study included 320 patients; 184 males and 136 females. Age ranged from 18-56 years. The most common clinical presentations in all patients, according sex and seropositivity were shown in table 1 and 2. The result of serology for coeliac disease is shown in table 3. Table 1: clinical presentation in all patients.

Complaints MalesN=184

FemalesN=136

TotalN=320

Complaints MalesN=184

FemalesN=136

TotalN=320

Abdominal painFrequent or loose stoolHard or infrequent stoolAltered stool habitsOther stool abnormalities*

7835212632

9816462027

17651674695

Dyspepsia Postprandial distress. Epigastric painAbdominal distension Eructation. Chronic Anorexia

52386441184

221847452111

7456

101863915

Other stool abnormalities*: straining during a bowel movement ,urgency (having to rush to have a bowel movement) , feeling of incomplete bowel movement or anorectal obstruction, passing mucus (white material) during bowel movement , abdominal fullness, bloating or swelling.

Table 2: clinical presentation in all patients according sero-positivity.Complaints Serologically

negative (n=305)

Serologically positive (n=15)

Complaints Serologically negative (n=305)

Serologically positive(n=15)

Abdominal painFrequent or loose stoolHard or infrequent stoolAltered stool habitsOther stool abnormalities

164 (53.8%)43(14.1%)66(21.6%)45(14.8%)

93(30.5%)+

12 (80.0%)*8 (53.3%)*1 (6.7%)1 (6.7%)2 (13.3%)

Dyspepsia Postprandial distress. Epigastric painAbdominal distensionEructation. Chronic Anorexia

64(21.0%)55(18.0%)91(29.8%)87(28.5%)34(14.1%)11(3.6%)

10 (66.7%)*11 (73.3%)*10 (66.7%)*9 (60.0%)*5 (33.3%)*4 (26.7%)*

Significantly higher in comparison to serologically negative (P < 0.05).

Table 3: clinical presentation in sero-positive patients before and after Gluten Free Diet (GFD):

ComplaintsSerologically positive (n=15)

ComplaintsSerologically positive (n=15)

Before GFD After GFD Before GFD After GFDAbdominal painFrequent or loose stool

12 (80.0%)8 (53.3%)

3(20.0%) *2(13.3%) *

Dyspepsia Postprandial distress.

10 (66.7%)11 (73.3%)

4 (26.7%) * 2 (13.3%) *

Hard or infrequent stoolAltered stool habitsOther stool abnormalities

1 (6.7%)1 (6.7%)2 (13.3%)

2 (13.3%)3 (20.0%)4 (26.7%)

Epigastric painAbdominal distensionEructation. Chronic Anorexia

10 (66.7%)9 (60.0%)5 (33.3%)4 (26.7%)

3 (30.0%) *2 (13.3%) *

1 (6.7%)1 (6.7%)

* Significant decrease in the number of patients after GFD in comparison to before GFD (P < 0.05). Table 4: Haemoglobin, serum iron, and liver enzymes in serologically negative and serologically positive patients before and after gluten free diet.Factor Serologically negative patients Serologically Positive patients

At diagnosis At diagnosis After GFDHb (g/dl) 14.53 ± 03.45 13.21 ± 3.72* 14.25 ± 3.25+S. iron (μg/dl) 72.63 ± 29.32 55.65 ±23.72* 65.34 ±20.76+ALT (units/dl) 25.61 ± 11.34 27.23 ± 13.52 26.53 ± 13.84AST (units/dl) 22.16 ± 10.35 26.35 ± 11.72 24.73 ± 11.51Alk. Phosphatase (U/dl)

* Significant differences in comparison with serologically negative patients at diagnosis (P< 0.05).+ Significant differences in comparison with serologically positive patients at diagnosis (P< 0.05).

Table 5: Blood levels of albumin, calcium, cholesterol and triglycerides in serologically negative and serologically positive patients before and after gluten free diet.Factor Serologically negative patients Serologically Positive patients

At diagnosis At diagnosis After GFDS. albumin (g/dl) 4.30 ± 00.65 3.51 ± 00.71* 4.22 ± 0.51+S calcium (mg/dl) 10.13 ± 1.20 8.35 ± 1.41* 9.02 ± 4.52+Blood Cholesterol (mg/dl) 169.50 ±21.37 151.28±23.56 158.40±22.64Triglycerides (mg/dl) 153.28 ± 32.62 146.53±28.35 150.24 ±23.52

* Significant differences in comparison with serologically negative patients at diagnosis (P< 0.05).+ Significant differences in comparison with serologically positive patients at diagnosis (P< 0.05).

Table 6: the results of serology in all patientsSerological tests total Positivity Positivity (males) Positivity (female)Anti-gliadin antibodies (IgA/G) 15/320 (4.69% 7 (2.19%) 8 (2.50%)EMA IgA 13/320 (4.06%) 6 (1.88%) 7 (2.19%)Anti-TG2 IgA 12/320 (3.76%) 5 (1.57%) 7(2.19%)Anti- TG2 IgG 13/320 (4.06%) 6 (1.88%) 7 (2.19%)

Table 7: IgA and IgG anti-gliadin antibody titres in those with positive tests (15 patients):IgA anti-gliadin titre IgG anti-gliadin titre

Optical density

Number of patients Optical density

Number of patientsAt the diagnosis After GFD At the diagnosis After GFD

>0.8750.75-0.8750.625–0.750.5–0.6250.375–0.50.25–0.3750.125–0.250.0–0.125 (-)*

52133001

01122125

>1.00.9-1.00.8–0.90.7–0.80.6–0.70.5–0.60.4–0.50.3–0.40.1-0.30.0-0.1 (-)*

3221121102

0011013117

Titre is expressed as optical density. GFD: gluten free diet. 14 patients were positive for IgA, and 13 were positive for IgG. Those who were negative for any of them are positive for the other. * = negative values

Table 8: Serology and histopathology in all positive cases (Before/After GFD):

AGA EMA IgA

Anti-TG2Histopathology (Marsh

classification)**IgA IgG IgA IgG At diagnosis After GFD*

12

3 +45678910

11+12131415

Totalpositivity

+/++/-+/++/-+/-+/++/++/-+/--/-+/++/++/++/++/-

14/8

+/++/++/++/+-/-+/++/++/-+/-+/-+/++/+-/-+/++/-

13/9

+/++/++/++/-+/-+/++/++/-+/+-/-+/++/+-/-+/++/+

13/10

+/-+/++/-+/+-/-+/-+/++/++/+-/++/-+/--/-+/++/-

12/7

+/-+/++/++/++/-+/++/++/-+/--/++/-+/+-/-+/++/-

13/8

Type 1Type2Type3aType 1NormalType3bType2Type2 Type 1NormalType1Type2

NormalType2

Normal11

NormalType 2NormalNormalNormalType 2Type 2Type 1Type2

NormalNormalType 1NormalType1

Normal7

After gluten free died for about 6 months, seroconversion to negative tests occurred in 6 patients for AGA-IgA, 4 for AGA- IgG, 3 for EMA IgA, 5 for Anti-TG2 IgA and 5 for Anti-TG2 IgG. * GFD: gluten-free diet. + these patients have family history of coeliac disease in first degree relatives.

It is clear from these results that patients number 10, and 13 were only positive for one test (IgG-AGA and IgA-AGA respectively) and patient number 5 is positive for three of the five tests; IgA AGA, EMA and IgG anti-TG2. The three patients showed normal histopathology in addition to patient number15. Thus, only 11 patients (74.44% of those with positive serology and 3.49% of total patients) were diagnosed by biopsies as compatible with coeliac disease. Two patients have family history of coeliac disease in first degree relatives. One of them; patient number 3 was found to have positive serology for all tests and type 3a histopathology. The other patient had similar serology but type 1 histopathology. All patients with positive serology were subjected to GFD for at least 6 months. Then all serological tests and tissue biopsies were repeated and showed some improvement.

Table 9: Summery of histopathological findings in the 15 AGA positive patients:Normal Positive histopathology

Type 1 Type 2 Type 3 Type 4 TotalAt diagnosis 4 4 5 2 0 11 (74.44%)After GFD 8 3 4 0 0 7 (46.67%) The grade of histopathology showed complete healing in 4 patients and improvement to lower grades

in 4 patients (one type 3 changed to type 1 and 3 cases of type 2 changed to type 1). Worsening occurred in one case from type 1 to type 2. 7 cases still showed type 1 or type 2 diseases.

Discussion:Coeliac disease is a T-lymphocyte-mediated autoimmune gastrointestinal disorder induced

by ingestion of gluten found in wheat, rye and barley.[44,45] It meets the World Health Organization (WHO) criteria for mass screening because the disease is common, difficult to detect based on clinical symptoms, has sensitive and fairly specific screening tests for

diagnosis, it can be treated effectively, and, if left untreated, substantial morbidity and even severe complications may ensue. However, uncertainties about the natural course and management of subclinical CD have turned mass screening into a controversial issue (51).

The active disease is characterized by gluten-dependent autoantibodies against endomysium (EMA), a complex connective tissue structure surrounding smooth muscle cells, and more precisely, against the protein type 2 ('tissue') transglutaminase (TG2), the coeliac autoantigen anchored to endomysial collagen by fibronectin.[46,47] Detection of these autoantibodies in the serum is a useful means of identifying new coeliac patients presenting with only mild gastrointestinal symptoms, non-specific general complaints or extraintestinal manifestations, or in populations in general.[47-50] However, the benefits of serologic screening for coeliac disease in asymptomatic individuals are debatable. Symptoms may be similar to that of IBS.[6,7] and both diseases might have similar natural history. Many studies suggest that the prevalence of coeliac disease is 3-11% [6,7,8] among patients diagnosed with IBS, compared with 0.02-1.0%[6,7,8,9,40,41] in the general population. Screening studies have further shown that many patients suffer only minimal if any symptoms and the prevalence of this disease thus remains underestimated, when only patients with classical coeliac disease are recognized (42,43). The purpose of identifying coeliac disease in such patients with suspected IBS is to improve quality of life and decrease the cost of un-needed tests, non-specific medicines, and frequent consultation.

In our study, patients who attended GIT clinic with manifestations indicating primary diagnosis of IBS were selected and investigated for coeliac disease. Those who showed positive serology were subjected to upper endoscopy for histopathological confirmation of the disease to be given a course of gluten free diet for at least 6 months. The impact of such therapeutic modality on patient symptomatology and blood parameters was studied.

A total number of 320 patients with highly suggestive, symptoms-based diagnosis of IBS were enrolled. Positive serology was found in 15 cases (4.69%). The anti-gliadin antibodies showed the highest positivity and sensitivity when both IgG and IgA were performed together. Other tests showed slightly lower positivity. However, tissue examination showed positive diagnosis in only 11 cases (74.44% of those with positive serology and 3.49% of total patients). Anti-TG2 was the most specific one particularly IgA antibody which showed matching with tissue diagnosis in 14/15 cases. The differences between these tests were not statistically significant because the number of positive cases was small. Therefore, from this study we can not determine the tests with higher sensitivity or specificity in the diagnosis of coeliac disease. These findings were similar to the previous studies showing prevalence of 3-11% in patients with IBS (6,7,8)

.Symptoms, with some overlap related to IBS or coeliac disease, were significantly higher

in patients with positive serology than those with negative serology for coeliac disease. These symptoms included abdominal pain, post-prandial stress, dyspepsia, epigastric pain, abdominal distension, diarrhea, eructation and anorexia in this order. Thus, the high prevalence of these symptoms and the presence of many of them could be considered as premonitory or green light for testing for coeliac disease.

Haemoglobin, serum iron, calcium, albumin levels were found to be significantly lower in

those with positive serology and histopathology than negative patients. Also the levels of these parameters were significantly elevated after the induction of gluten free diet. However, this increase did not reach the level in those with negative serology. These parameters are reflective of the nutritional deficiencies found in these patients. But because being mild reductions, such

deficiencies were not clinically manifest by these patients. After putting the patients on GFD, their levels started to increase significantly. Cholesterol and triglycerides were found to be lower in serologically positive patients with some increase after GFD but all these changes were statistically insignificant.

Also, liver transaminases were found to be lower in serologically positive patients but statistically insignificant. Liver enzymes elevations were found in coeliac disease in many studies (37,38,39). But because of the subclinical nature of patients of this study, the elevation was not above the normal range. The liver might be involved in this disease as a cryptogenic nature, primary biliary cirrhosis, and occasionally severe liver disease and cirrhosis due to autoimmune hepatitis.

In this study also, the disease activity in those with subtle symptoms was mild and found to be of type1 and 2 in 9/11 cases with only 2 cases of type 3. This finding may explain the minimal symptomatology in these patients. After a period of gluten free diet for these patients with positive serology of about 6-12 months, their tissue pathology has improved in most cases. The grade of histopathology has shifted to the left with complete healing in 4 patients and improvement to lower grades in 4 patients (one type 3 changed to type 1 and 3 cases of type 2 changed to type 1). Worsening occurred only in one case from type 1 to type 2. This may be due to non-compliance or non-adherence to gluten free diet. Still 7 cases had type 1 or type 2 diseases. The improvement; however, was not satisfactory in these patients. This might be due to the mild nature of the disease manifestations and previous diagnosis of IBS and therefore; the patients did not stick to gluten free diet.

It is concluded from this study that minimally symptomatic coeliac disease can easily be mistaken for IBS. The presence of many persistent gastrointestinal symptoms in addition to signs and lower serum levels of some nutritional parameters must alert the physicians to screen for coeliac disease. Any serological test can be used for the screening but this must be confirmed by tissue diagnosis which is the gold standard for diagnosis of this disease. Finally, screening for this disease among patients with IBS must be considered to offer better prognosis to these patients simply by gluten free diet.

معدل وجود مرض داء الزلىقى فى مرضى القولون العصبى السعوديين البالغين

شندى محمد شندى شريف*، نهال العسلى**و نعيمة العشرى** ىقسم المراض المتوطنة*و ىقسم الكيمييياء اللكلينيكييية**,معهد تيودور حبلهارس للحبحاث.

لقد أوجدت القليل من الدراسات الحديثة معدل أعلى لميرض داء الزلىقيى فيى مرضيى القوليون العصيبى حبالمقارنية حبعامية النياس وذليك %. لكما أوجدت دراسات مماثليية معييدل مميياثل للمييرض فييى دول۲,٠-٦,٠ % حبالمقارنة حبالمعدل العام ماحبين 11-3حبمعدل يتراوح حبين

الشرق الوسط والدول الورحبية وذلك فى العامة واللذين هم معرضين ألكثر للمرض. ولكان الهدف من هذا البحث هو تحديد معدل مرض داء الزلىقى والتواحبع اللكلينيكة الممكنة للتختبارات المصلية له فى مرضى القولييون

مريضا من البالغين تم أتخذ التاريخ المرضى لهم وفحصهم فحصا سريريا وعمييل320العصبى السعوديين البالغين. وىقد شملت الدراسة التحاليل الضرورية ثم التحاليل الخاصة حبداء الزلىقى. وتم عمل منظار الجهاز الهضمى العاوي للحالت اليجاحبية لتخذ تخييذع ميين الثنييى عشر لفحصها مجهريا مع منع تناول الذغذية التى تحتوى على الجلوتين ثم إعادة هذه الفحوص حبعد ستة أشهر إليى أثنييى عشير شيهرا

حاليية (15السيييرولوجى) ولكيانت لكالتييالى: 0 حاليية ايجاحبيية حبالتحلييل المصييلي 320/15من هذه التغذية. وىقد أظهيرت الدراسية وجيود حالة فقيط حبيالفحص المجهيرى.11 حالة حبالجسام المضادة التخرى.و 13-12%) حباستخدام الجسام المضادة ضد الجليادين و 4.69

ولكانت أعراض المرض مثل ألم البطن و والسهال وعسر الهضم وذغيرها ألكثر شيييوعا فييى هييؤلء المرضييى عنهييا فييى المرضييى ذات التحاليل السلبية. لكما وجد ذلك أيضا حبالنسبة لنخفاض معدل الهيموجلوحبين والحديد و الزلل و الكالسيوم فى الدم. وىقييد تحسيينت جميييع العوامل تحسنا ذات دللة إحصائية حبعد منع تناول الذغذية التى تحتوى على الجليوتين سيتة أشيهر إليى أثنييى عشير شيهر. لكميا تحيولت

حيالت أتخيري اليى درجيات أىقيل فيى4 حالت شفاءا تاما وتحسينت 4 حالت حسب نوع التحليل. وتم شفاء 6-3النحاليل الى سلبية ىقى حين ساءت حالة واحدة حسب الفحص المجهري لخذع الثنى عشر. يستخلص ميين هييذا البحييث أن مييرض داء الزلىقييى ذات العييراض الضئيلة ذغالبا ما يشخص تخطئا لكقولون عصبى (متلزمة المعاء المتهيجة). ولكن الوجود الييدائم لعييدد ميين أعييراض الجهياز الهضيمى حبالضافة لنقص معدل الهيموجلوحبين والحديد و الزلل و الكالسيوم فى الدم لكدللت سوء التغذية يجب أن تكون مؤشييرا لتنييبيه الطبيياء

المعالجين للبحث عن المرض وذلك بأي من التحاليل المصلية لهذا الداء وتأكيد ذلك بالفحص المجهري لخذع المعاء الدقيقة وذلك مممنأجل التحسن الضفضل علي المدي الطويل لهؤلء المرضى وذلك بمنع تناول الذغذية التى تحتوى على الجلوتين.

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