association of 25-hydroxy vitamin d deficiency …...the active form of vitamin d, its serum levels...

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ASSOCIATION OF 25-HYDROXY VITAMIN D DEFICIENCY WITH NT-PRO BNP LEVELS IN ACUTE MYOCARDIAL INFARCTION PATIENTS

By

Rajyalakshmi Gadi

A Thesis Submitted to the Graduate Faculty of

WAKE FOREST UNIVERSITY GRADUATE SCHOOL OF ARTS AND SCIENCES

in Partial Fulfillment of the Requirements

for the Degree of

MASTER OF SCIENCE

in the Health Sciences Research Program

August, 2010

Winston-Salem, North Carolina

Approved by:

Michelle J Naughton, Ph.D., Advisor

Examining Committee:

David C Goff, MD., Ph.D., FACP, FAHA. Chair

John Spertus, MD., MS., FACC.

Doug Case, Ph.D.

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ACKNOWLEDGMENTS

I would like to thank the following people for their help and support with this

project. Dr. John Spertus has been my mentor and advisor since I came to Kansas for

my fellowship. His feedback has been invaluable in every phase of this

project. His mentorship has helped me enormously in improving both my research and

writing skills needed to finish this thesis project successfully.

I would also like to thank Dr. James Wetmore and Fengming Tang who assisted in

editing the manuscript in preparation for publication. I would like to extend my

appreciation to Drs. David Goff and Doug Case for taking the time from their busy

schedules to serve on my thesis committee.

Finally I would like to thank Dr. Michelle Naughton for her continued support and

encouragement as the thesis advisor.

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TABLE OF CONTENTS

LIST OF ILLUSTRATIONS V

LIST OF ABBREVIATIONS VI

ABSTRACT VII

CHAPTER 1: BACKGROUND

Introduction 1

Studies of Vitamin D Deficiency in AMI Patients

Study 1 3

Study 2 4

Studies of Vitamin D Deficiency and Heart failure

Study 1 6

Study 2 7

Study 3 7-8

Study 4 8-9

Summary and Specific Aims 10

Significance of the Proposed Research 11

Chapter 1 references 12-15

CHAPTER 2: ASSOCIATION OF 25-HYDROXY VITMAIN D DEFICIENCY

WITH NT-PRO BNP LEVELS IN AMI PATIENTS

Abstract 17

Introduction 18

Materials and Methods 19

iv

Results 23

Discussion 24


CHAPTER 3: DISCUSSION

Project summary 37

Additional analysis 38

Implications of the research project 40

Future directions 41


CURRIUCULUM VITAE 46

v

LIST OF ILLUSTRATIONS

CHAPTER 1:

TABLES

Table 1: Relative risk (95% CI) of myocardial infarction associated with

quartile plasma levels of 25-hydroxyvitamin D3 4

CHAPTER 2:

TABLES

Table 1: Baseline characteristics of study participants, by 25(OH)D groups 30

Table 2: Association of 25(OH)D with NT-proBNP levels 31

Table 3: Results of multivariable regression analysis 32

FIGURES

Figure 1: Spearman correlation between 25(OH)D (ng/ml) 29

and NT-proBNP levels

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LIST OF ABBREVIATIONS

25(OH)D – 25 Hydroxy vitamin D

1,25(OH)2D – 1,25-dihydroxy vitamin D

VDR – vitamin D receptors

CVS – cardiovascular system

HTN – hypertension

DM-2 – type2 diabetes

AMI – acute myocardial infarction

LVDF – left ventricular dysfunction

ANP – atrial natriureitc peptide

BNP – Brain natriuretic peptide

PTH – parathyroid hormone

SHPT– secondary hyperparathyroidism

NT-proBNP – N-terminal pro-brain natriuretic peptide

CHF – Congestive heart failure

ESRD – end stage renal disease

CKD – chronic kidney disease

GFR – glomerular filtration rate

NSTEMI – non ST segment myocardial infarction

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ABSTRACT

Background and objectives: Nutritional vitamin D deficiency is an emerging risk factor

for acute myocardial infarction (AMI) and heart failure. The association between 25-

hydroxyvitamin D levels and N-terminal pro B-type natriuretic peptide (NT-proBNP), a

robust prognostic marker for post-MI mortality and heart failure is unknown and could

illuminate a potential pathway for adverse outcomes among post-MI patients with 25-

hydroxyvitamin D deficiency.

Design, setting, participants and measurements: In a cross sectional analysis, we

studied 238 AMI patients from 21 US centers to test the association of nutritional

vitamin D (25-hydroxyvitamin D [25(OH)D]) deficiency with NT-proBNP levels. Patients’

25(OH)D levels were categorized as normal (≥30 ng/ml), insufficient (>20 ─ <30 ng/ml),

deficient (>10 ─ ≤ 20 ng/ml), and severely deficient (≤10 ng/ml) groups.

Results: 96% of AMI patients had low 25(OH)D levels, with 75% having 25(OH)D

deficiency and 21% having insufficiency. No significant trends for higher mean log NT-

proBNP levels in severely deficient (6.9 ± 1.3 pg/ml), deficient (6.9 ± 1.2 pg/ml) and

insufficient (6.9 ± 0.9 pg/ml) groups were observed as compared with patients having

normal (6.1 ± 1.7 pg/ml) levels, P = 0.165. In multivariate regression model after

adjusting for several covariates, 25(OH)D was not associated with NT-proBNP levels.

Conclusion: Potential associations between nutritional vitamin D deficiency and

prognosis in the setting of AMI are unlikely to be mediated through NT-proBNP

pathways. Future studies should examine other mechanisms such as inflammation and

vascular calcification by which 25(OH)D deficiency could mediate adverse outcomes

such as heart failure and mortality post AMI.

1

CHAPTER 1: BACKGROUND

Introduction

Vitamin D deficiency is highly prevalent in the United States and worldwide.1

NHANES III reported that the prevalence of 25 hydroxy vitamin D [25(OH)D] deficiency

in the US population is between 25%-57%.2 25(OH)D is the principal circulating storage

form of vitamin D in the human body. Vitamin D in its active form, 1,25-dihydroxy

vitamin D [1,25(OH)2D] is a hormone as it is primarily produced in the kidneys and

circulates in blood exerting its effects on various tissues throughout the body via the

vitamin D receptors (VDR). VDR’s have a broad tissue distribution that includes

vascular smooth muscle, endothelium and cardiomyocytes.1;3 Although 1,25(OH)2D is

the active form of vitamin D, its serum levels do not correlate with the overall vitamin D

status and are generally not clinically useful.4 On the other hand serum 25(OH)D

concentrations reflect both vitamin D intake and endogenous production and are more

reflective of an individual’s overall vitamin D status. Hence serum 25(OH)D levels are

frequently used in clinical settings to assess vitamin D status Although a consensus

regarding the optimal level of serum 25(OH)D has not been established most experts

define 25(OH)D deficiency as a level <20 ng/ml and vitamin D insufficiency as 21-29

ng/ml.5;6 For studied end points such as incident MI or all cause mortality a level of ≥

30ng/ml is considered optimal.

It is now increasingly recognized that adequate vitamin D status is not only

important for bone health and the prevention of osteoporosis but also for optimal

function of many other organs and tissues throughout the body, including the

cardiovascular (CV) system.3 Cardiac myocytes have cytosolic vitamin D receptors

2

(VDR)4 that bind active vitamin D (1,25 dihydroxy vitamin D), but unlike vascular smooth

muscle cells, cardiac myocytes lack 1α-hydroxylase activity, 7 an enzyme that converts

inactive vitamin D(25 hydroxy vitamin D) to active vitamin D. Hence cardiac muscle is

strongly dependent upon circulating active vitamin D or calcitriol levels. In the past,

several in vitro studies have shown that calcitriol regulates intracellular calcium

metabolism and thus myocardial contractility. 8-10 Consequently, 25(OH)D deficiency

has been associated with aberrant cardiac contractility, cardiomegaly, and increased

ventricular mass due to myocardial collagen deposition,9;10 independent of its known

effects on blood pressure.11

Apart from its effects on the myocardium, 25 (OH)D deficiency also leads to

enhanced atherosclerosis secondary to vascular smooth muscle cell (VSMC)

proliferation12;13 and increased production of pro inflammatory cytokines (IL-6 and TNF-

alfa).14

There is growing body of evidence from clinical studies that 25(OH)D deficiency

also plays an important role in the genesis of coronary risk factors, including

hypertension (HTN), type-2 diabetes (DM-2) and the metabolic syndrome.4;15-18

Furthermore, large epidemiological studies, including the Health Professionals Study 19

and the Framingham Offspring Study, 20 have shown that low 25(OH)D levels

(<15ng/ml) as compared with levels ≥ 30ng/ml, were independently associated with

twice the risk of incident myocardial infarction (MI) and a higher risk of incident

cardiovascular events including fatal and nonfatal stroke. Moreover, the risk of all-

cause mortality was higher among subjects with vitamin D levels <17.8ng/ml in

NHANES III as compared with subjects having levels >32.21 Consequently, circulating

3

25(OH) D levels do not just represent the vitamin D status of an individual but can

potentially serve as a biomarker for cardiovascular risk. Despite these studies

emphasizing the importance of 25(OH)D deficiency and CV disease, there is sparse

data in the literature about the prevalence of 25(OH)D deficiency in acute myocardial

infarction (AMI) patients, who comprise a high risk population. A synopsis of the studies

on Vitamin D deficiency in AMI patients is presented below.

Studies of Vitamin D Deficiency in AMI Patients

Two small case control studies have examined vitamin D levels in AMI patients.

Study 1: Scragg R, Jackson R, Holdaway IM, Lim T, Beaglehole R: Myocardial

infarction is inversely associated with plasma 25-hydroxyvitamin D3 levels: a

community-based study. Int J Epidemiol 19:559-563, 1990

A study by Scragg and colleagues 22 was conducted in the Central Auckland area

of New Zealand between March 1986 and 1988. Cases were patients between 35-64

years of age, who were diagnosed with AMI per the WHO-MONICA project criteria. 23

Age and sex matched controls were obtained by random sampling of the general

population. About 179 case-control pairs were analyzed. The results indicated that the

mean plasma 25(OH)D level in the cases was lower compared to the controls (32

nmol/lit vs. 35 nmol/lit, p value=0.017).The odds ratio of AMI decreased with increasing

quartiles of vitamin D as shown in the table below.

4

Table1. Relative risk (95% CI) of myocardial infarction associated with quartile

plasma levels of 25-hydroxyvitamin D3

Level of

25-hydroxy vitamin D3

(nmol/L) Relative risk (95% CI)

<25 1.00

25−32 0.56 (0.30, 1.03)

33−42 0.33 (0.17, 0.64)

≥43 0.30 (0.15, 0.61)

Study 2: Lund B, Badskjaer J, Lund B, Soerensen OH: Vitamin D and ischaemic heart

disease. Horm Metab Res 10:553-556, 1978

In this study by Lund et al,24 serum 25 (OH)D levels were measured in 128 patients with

ischemic heart disease admitted to the Frederiksberg Hospital in Denmark. 53 of these

patients had a diagnosis of AMI and 75 had angina pectoris. Seasonal variations in the

vitamin D levels were compared to 409 controls. The mean 25(OH)D levels in the AMI

and angina patients were 24 ± 10ng/ml and 23.5 ± 9.6 ng/ml respectively, which were

not statistically different from the level of 28.8 ± 12.3 recorded in controls. However, the

mean vitamin D levels in the cases were lower compared to the controls in the months

of May-June (P <0.01) and July-August (P <0.05), indicating that serum vitamin D levels

change with sun exposure. In summary, the studies of vitamin D deficiency in AMI

patients were not recent and were limited by their small sample sizes and were not

based in the U.S., where better nutrition and fortification of milk is common.

Studies of Vitamin D Deficiency and Heart failure

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Apart from the paucity of studies on vitamin D deficiency in AMI patients, gaps

exist in the literature regarding the role of vitamin D deficiency in the development of left

ventricular dysfunction (LVDF) or heart failure, an important complication post MI.

As enumerated in several in vitro studies, 25(OH)D deficiency is associated with

aberrant cardiac contractility, cardiomegaly, and increased ventricular mass due to

myocardial collagen deposition,9;10 all of which ultimately lead to heart failure. Further, it

has been shown that activation of nuclear vitamin D receptors by 1,25(OH)2D3

suppresses the expression and secretion of atrial natriuretic peptide (ANP) and Brain

natriuretic peptide (BNP) in cardiac myocytes,25-27 both of which are biomarkers of heart

failure. In addition to its direct effects on myocardium, vitamin D deficiency may exert

indirect effects on the myocardium by elevated serum parathyroid hormone (PTH)

levels4 leading to secondary hyperparathyroidism (SHPT). PTH is released from the

parathyroid gland in response to low calcium levels from vitamin D deficiency. PTH

binds to its receptors on the myocardium and impairs the energy metabolism of

myocardial cells 28 and induces cardiomyocyte hypertrophy, 29 leading to heart failure or

left ventricular dysfunction.30

In recent years, it has been established that N-terminal pro-brain natriuretic

peptide (NT-proBNP), a pro-hormone of BNP released from cardiac ventricles, is

associated with the severity of left ventricular dilatation and dysfunction after MI.31;32 In

addition, NT-pro BNP is a sensitive and robust prognostic biomarker of mortality in

acute MI,33-35 heart failure(HF)36;37 and chronic hemodialysis38;39 patients. Moreover, NT-

pro BNP levels have direct clinical implications, as they are used to guide therapeutic

interventions in HF patients, leading to improved outcomes as compared to routine

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clinical treatment.40 A synopsis of the studies examining the association of vitamin D

and NT-proBNP in other clinical settings is given below.

Study 1: Zittermann A, Schleithoff SS, Tenderich G, Berthold HK, Korfer R, Stehle P:

Low vitamin D status: a contributing factor in the pathogenesis of congestive heart

failure? J Am Coll Cardiol 41:105-112, 200

This was a case-control study that examined the association between plasma

NT-proANP, a marker of heart failure severity, and vitamin D metabolites. The study

was comprised of 54 cases and 34 controls recruited at the Heart and Diabetes Center

in Bonn, Germany from Nov 2000 to March 2001. Among the cases, 20 patients were <

50 years of age and 34 were ≥ 50 years, and all the controls were above 50 years old.

All cases had ≥ class II New York Heart Association (NYHA) congestive heart failure.

25 hydroxy vitamin D levels were lower in both patient groups compared to the controls

(9 ng/ml and 11ng/ml in cases vs. 18 ng/ml in controls,(p< 0.001). Both groups of CHF

patients had markedly elevated NT-proANP levels and parathyroid hormone levels (p

<0.001) compared to controls. In a nonlinear regression analysis, 25(OH)D inversely

correlated with NT-proANP (r2= 0.16, p<0.001).41 This study concluded that low vitamin

D levels could contribute to myocardial dysfunction in patients with congestive heart

failure.

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Study 2: Matias PJ, Ferreira C, Jorge C, Borges M, Aires I, Amaral T, Gil C, Cortez

J,Ferreira A: 25-Hydroxyvitamin D3, arterial calcifications and cardiovascular risk

markers in hemodialysis patients. Nephrol Dial Transplant 24:611-618, 2009.

This was a cross-sectional study to determine the association of Vitamin D

deficiency on BNP and other vascular calcification parameters in 223 hemodialysis

patients in a single hemodialysis unit in Portugal. The mean serum 25(OH)D levels in

these patients was low at 21.6 ± 12.2ng/ml. In an unadjusted analysis, serum 25(OH)D

levels negatively correlated with BNP(r= -0.22, p= .002) and vascular

calcification(r=0.26, p<0.001).On multivariate analysis, lower levels of 25(OH)D were

independently associated with higher BNP levels (p=0.005) and higher vascular

calcification scores (≥3), (p =0.002).42 This study thus concluded that low 25(OH)D

levels are a cardiovascular risk marker in hemodialysis patients and that the effects of

its repletion on cardiovascular morbidity and mortality needs to be clarified in large

randomized controlled trials.

Study 3: Singh NP, Sahni V, Garg D, Nair M: Effect of pharmacological suppression of

secondary hyperparathyroidism on cardiovascular hemodynamics in predialysis CKD

patients: A preliminary observation. Hemodial Int 11:417-423, 2007

In humans it is known that the administration of active vitamin D [125(OH)D3] to

end stage renal disease (ESRD) patients improves left ventricular function.43-45

In this study, 20 pre-dialysis CKD patients (e GFR <30 ml/min/1.73m2) with

secondary hyperparathyroidism (SHPT) (i.e.; serum intact PTH levels >180 pg/ml), were

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treated with active vitamin D (calcitriol) for 12 weeks and 10 of similar patients were

treated with placebo. Echocardiography assessment of cardiac function was performed

at baseline and after 12 weeks of treatment. No significant change in LV dimensions or

ejection fraction was observed after 12 weeks of treatment, but there was significant

improvement in LV diastolic parameters, namely the A wave velocity (0.69 ± 0.089 to

0.68 ± 0.084, p = 0.001)) and E/A ratio ( 1.193 ± 0.21 to 1.238 ± 0.18,P = 0.001). No

change was observed in the placebo group.46 The study concluded that diastolic

dysfunction seen in pre-dialysis CKD patients could possibly be improved by active

vitamin D therapy, although larger and longer duration studies are warranted to

substantiate these findings.

Study 4: Pilz S, Marz W, Wellnitz B, Seelhorst U, Fahrleitner-Pammer A, Dimai HP,

Boehm BO, Dobnig H: Association of vitamin D deficiency with heart failure and sudden

cardiac death in a large cross-sectional study of patients referred for coronary

angiography. J Clin Endocrinol Metab 93:3927-3935, 2008

This study included 3,299 patients from the LUdgwigshafen RIsk and

Cardiovascular Health Study (LURIC). This prospective cohort study included patients

routinely referred to coronary angiography from a single tertiary care center in

Southwest Germany, and who had baseline 25(OH)D levels measured, between July

1997 and January 2000. The cross sectional associations between baseline 25(OH)D

levels and the heart failure marker, NT-proBNP, was studied. In addition the hazard

ratio for heart failure deaths according to vitamin D status was studied prospectively.

25(OH)D levels correlated negatively with NT-proBNP levels( r = - 0.190, P <0.001).

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Patients in higher NYHA classes had significantly lower 25(OH)D levels (P <0.001). In

multiple linear regression analysis after adjusting for age, race, gender, BMI, smoking

status, co-morbidities, renal function(GFR) and C-reactive protein, 25(OH)D remained

independently associated with NT–proBNP (β coefficient =-0.082, p <0.001).

After a mean follow-up of 7.7 years, 116 patients died due to heart failure and

188 died due to sudden cardiac death (SCD). After adjustment for cardiovascular risk

factors, the hazard ratios for death due to heart failure and SCD were 2.84 (95% CI: 1.2

- 6.74) and 5.05 (2.13-11.97), respectively, when comparing patients with severe

vitamin D deficiency (<25nmol/l) with persons in normal range (≥ 75nmol/l). The study

concluded that vitamin D deficiency is associated with prevalent myocardial dysfunction

and deaths due to heart failure and sudden cardiac death.

Summary and Specific Aims

There are no studies in the U.S. that have examined the vitamin D status of

patients admitted with acute myocardial infarction. Studies completed outside of the

Untied States indicate that both 25(OH)D and NT-proBNP are associated with LV

dysfunction, and that low circulating levels of 25(OH)D could potentially contribute to, or

potentiate, the development of left ventricular dysfunction (LVDF) and heart failure.

However, all of the above studies were not based on patients living in the U.S., where

better nutrition and fortification of milk is common.47

Given the adverse health implications of 25(OH)D deficiency and the lack of

studies in patients who have had a MI (a particularly high-risk group), we will examine

the prevalence of vitamin D deficiency and the association of 25(OH)D deficiency with

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NT-pro BNP levels in a multicenter cohort of AMI patients. To study these important

gaps in knowledge, we will use a unique multicenter prospective registry called

TRIUMPH (Translational Research Investigating Underlying disparities in recovery from

acute Myocardial infarction: Patients' Health status). This data base has a nationally

representative sample of AMI patients with detailed socio-demographic and clinical data

in addition to laboratory values (including 25(OH)D levels) and clinical health status.

These data were collected between June and December 2008. The specific aims for

the thesis research project are:

Specific aim 1: To describe the prevalence and patient characteristics associated with

25(OH)D deficiency in AMI patients. Using the TRIUMPH database we will describe the

distribution of 25(OH)D levels both as a continuous measure and categorized as those

with levels below 10 ng/ml, 10 to ≤ 20 ng/ml, >20 and <30ng/ml and ≥ 30 ng/ml.

Specific aim 2: To determine the association of 25(OH)D deficiency with NT-pro BNP

levels in AMI patients. We hypothesize that patients with lower vitamin D levels will

have higher NT–pro BNP levels and that vitamin D deficiency is independently

associated with NT-proBNP levels.

Significance of the Proposed Research

The current analyses will add to our understanding of the prevalence of 25(OH)D

deficiency in AMI patients, a high risk population and also provide novel insights into

those clinical characteristics which are most strongly associated with 25(OH)D

deficiency in AMI patients. For example, if 25(OH)D deficiency in AMI patients is found

11

to be significantly associated more with non ST segment elevation myocardial infarction

(NSTEMI) than ST segment elevation myocardial infarction (STEMI) in our cross –

sectional analysis, it might form the basis for future studies to determine the protective

role of 25(OH)D in preventing plaque rupture leading to partial occlusion of a coronary

artery which is the physiology behind a NSTEMI. Apart from the potential mechanistic

insights that might be inspired by our findings, our data will define the prevalence of a

novel adverse risk factor among AMI patients and could lead to studies that seek to

develop innovative strategies for the recognition and treatment of 25(OH)D deficiency

at the time of AMI and prior to hospital discharge.

In addition, the discovery of an association between circulating levels of 25(OH)D

and NT-proBNP would not only suggest a potential pathway for adverse outcomes

among post-MI patients with 25(OH)D deficiency, but could identify a potentially novel

therapeutic target (i.e., nutritional vitamin D supplementation) to reduce NT-proBNP

levels in the hopes of improving prognosis after MI.

Finally, with longer follow-up of this patient population (beyond the scope of this

proposed project) we will be able to determine whether 25(OH)D deficiency further risk

stratifies long-term clinical outcomes post MI after adjusting for currently used

prognostic schemes.

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REFERENCES

1. Holick MF: High prevalence of vitamin D inadequacy and implications for health. Mayo Clin Proc 81:353-373, 2006

2. Martins D, Wolf M, Pan D, Zadshir A, Tareen N, Thadhani R, Felsenfeld A, Levine B, Mehrotra R, Norris K: Prevalence of cardiovascular risk factors and the serum levels of 25-hydroxyvitamin D in the United States: data from the Third National Health and Nutrition Examination Survey. Arch Intern Med 167:1159-1165, 2007

3. Zittermann A, Schleithoff SS, Koerfer R: Putting cardiovascular disease and vitamin D insufficiency into perspective. Br J Nutr 94:483-492, 2005

4. Holick MF: Vitamin D deficiency. N Engl J Med 357:266-281, 2007

5. Bischoff-Ferrari HA, Giovannucci E, Willett WC, Dietrich T, wson-Hughes B: Estimation of optimal serum concentrations of 25-hydroxyvitamin D for multiple health outcomes. Am J Clin Nutr 84:18-28, 2006

6. Holick MF: Vitamin D Status: Measurement, Interpretation, and Clinical Application. Ann Epidemiol 2008

7. Hewison M, Zehnder D, Chakraverty R, Adams JS: Vitamin D and barrier function: a novel role for extra-renal 1 alpha-hydroxylase. Mol Cell Endocrinol 215:31-38, 2004

8. Weishaar RE, Simpson RU: Vitamin D3 and cardiovascular function in rats. J Clin Invest 79:1706-1712, 1987

9. Weishaar RE, Simpson RU: Involvement of vitamin D3 with cardiovascular function. II. Direct and indirect effects. Am J Physiol 253:E675-E683, 1987

10. Weishaar RE, Kim SN, Saunders DE, Simpson RU: Involvement of vitamin D3 with cardiovascular function. III. Effects on physical and morphological properties. Am J Physiol 258:E134-E142, 1990

11. Weishaar RE, Simpson RU: The involvement of the endocrine system in regulating cardiovascular function: emphasis on vitamin D3. Endocr Rev 10:351-365, 1989

12. Mitsuhashi T, Morris RC, Jr., Ives HE: 1,25-dihydroxyvitamin D3 modulates growth of vascular smooth muscle cells. J Clin Invest 87:1889-1895, 1991

13. Mohtai M, Yamamoto T: Smooth muscle cell proliferation in the rat coronary artery induced by vitamin D. Atherosclerosis 63:193-202, 1987

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14. Muller K, Haahr PM, Diamant M, Rieneck K, Kharazmi A, Bendtzen K: 1,25-Dihydroxyvitamin D3 inhibits cytokine production by human blood monocytes at the post-transcriptional level. Cytokine 4:506-512, 1992

15. Forman JP, Bischoff-Ferrari HA, Willett WC, Stampfer MJ, Curhan GC: Vitamin D intake and risk of incident hypertension: results from three large prospective cohort studies. Hypertension 46:676-682, 2005

16. Forman JP, Curhan GC, Taylor EN: Plasma 25-hydroxyvitamin D levels and risk of incident hypertension among young women. Hypertension 52:828-832, 2008

17. Lind L, Hanni A, Lithell H, Hvarfner A, Sorensen OH, Ljunghall S: Vitamin D is related to blood pressure and other cardiovascular risk factors in middle-aged men. Am J Hypertens 8:894-901, 1995

18. Scragg R, Sowers M, Bell C: Serum 25-hydroxyvitamin D, diabetes, and ethnicity in the Third National Health and Nutrition Examination Survey. Diabetes Care 27:2813-2818, 2004

19. Giovannucci E, Liu Y, Hollis BW, Rimm EB: 25-hydroxyvitamin D and risk of myocardial infarction in men: a prospective study. Arch Intern Med 168:1174-1180, 2008

20. Wang TJ, Pencina MJ, Booth SL, Jacques PF, Ingelsson E, Lanier K, Benjamin EJ, D'Agostino RB, Wolf M, Vasan RS: Vitamin D deficiency and risk of cardiovascular disease. Circulation 117:503-511, 2008

21. Melamed ML, Michos ED, Post W, Astor B: 25-hydroxyvitamin D levels and the risk of mortality in the general population. Arch Intern Med 168:1629-1637, 2008

22. Scragg R, Jackson R, Holdaway IM, Lim T, Beaglehole R: Myocardial infarction is inversely associated with plasma 25-hydroxyvitamin D3 levels: a community-based study. Int J Epidemiol 19:559-563, 1990

23. The World Health Organization MONICA Project (monitoring trends and determinants in cardiovascular disease): a major international collaboration. WHO MONICA Project Principal Investigators. J Clin Epidemiol 41:105-114, 1988

24. Lund B, Badskjaer J, Lund B, Soerensen OH: Vitamin D and ischaemic heart disease. Horm Metab Res 10:553-556, 1978

25. Wu J, Garami M, Cao L, Li Q, Gardner DG: 1,25(OH)2D3 suppresses expression and secretion of atrial natriuretic peptide from cardiac myocytes. Am J Physiol 268:E1108-E1113, 1995

26. Chen S, Nakamura K, Gardner DG: 1,25-dihydroxyvitamin D inhibits human ANP gene promoter activity. Regul Pept 128:197-202, 2005

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27. Bidmon HJ, Gutkowska J, Murakami R, Stumpf WE: Vitamin D receptors in heart: effects on atrial natriuretic factor. Experientia 47:958-962, 1991

28. Baczynski R, Massry SG, Kohan R, Magott M, Saglikes Y, Brautbar N: Effect of parathyroid hormone on myocardial energy metabolism in the rat. Kidney Int 27:718-725, 1985

29. Schluter KD, Piper HM: Cardiovascular actions of parathyroid hormone and parathyroid hormone-related peptide. Cardiovasc Res 37:34-41, 1998

30. Horl WH: The clinical consequences of secondary hyperparathyroidism: focus on clinical outcomes. Nephrol Dial Transplant 19 Suppl 5:V2-V8, 2004

31. Crilley JG, Farrer M: Left ventricular remodelling and brain natriuretic peptide after first myocardial infarction. Heart 86:638-642, 2001

32. Bettencourt P, Ferreira A, Pardal-Oliveira N, Pereira M, Queiros C, Araujo V, Cerqueira-Gomes M, Maciel MJ: Clinical significance of brain natriuretic peptide in patients with postmyocardial infarction. Clin Cardiol 23:921-927, 2000

33. Morrow DA, Braunwald E: Future of biomarkers in acute coronary syndromes: moving toward a multimarker strategy. Circulation 108:250-252, 2003

34. Omland T, Aakvaag A, Bonarjee VV, Caidahl K, Lie RT, Nilsen DW, Sundsfjord JA, Dickstein K: Plasma brain natriuretic peptide as an indicator of left ventricular systolic function and long-term survival after acute myocardial infarction. Comparison with plasma atrial natriuretic peptide and N-terminal proatrial natriuretic peptide. Circulation 93:1963-1969, 1996

35. Omland T, de Lemos JA, Morrow DA, Antman EM, Cannon CP, Hall C, Braunwald E: Prognostic value of N-terminal pro-atrial and pro-brain natriuretic peptide in patients with acute coronary syndromes. Am J Cardiol 89:463-465, 2002

36. Bayes-Genis A, Lopez L, Zapico E, Cotes C, Santalo M, Ordonez-Llanos J, Cinca J: NT-ProBNP reduction percentage during admission for acutely decompensated heart failure predicts long-term cardiovascular mortality. J Card Fail 11:S3-S8, 2005

37. Januzzi JL, van KR, Lainchbury J, Bayes-Genis A, Ordonez-Llanos J, Santalo-Bel M, Pinto YM, Richards M: NT-proBNP testing for diagnosis and short-term prognosis in acute destabilized heart failure: an international pooled analysis of 1256 patients: the International Collaborative of NT-proBNP Study. Eur Heart J 27:330-337, 2006

38. Madsen LH, Ladefoged S, Corell P, Schou M, Hildebrandt PR, Atar D: N-terminal pro brain natriuretic peptide predicts mortality in patients with end-stage renal disease in hemodialysis. Kidney Int 71:548-554, 2007

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39. Roberts MA, Srivastava PM, Macmillan N, Hare DL, Ratnaike S, Sikaris K, Ierino FL: B-type natriuretic peptides strongly predict mortality in patients who are treated with long-term dialysis. Clin J Am Soc Nephrol 3:1057-1065, 2008

40. Troughton RW, Frampton CM, Yandle TG, Espiner EA, Nicholls MG, Richards AM: Treatment of heart failure guided by plasma aminoterminal brain natriuretic peptide (N-BNP) concentrations. Lancet 355:1126-1130, 2000

41. Zittermann A, Schleithoff SS, Tenderich G, Berthold HK, Korfer R, Stehle P: Low vitamin D status: a contributing factor in the pathogenesis of congestive heart failure? J Am Coll Cardiol 41:105-112, 2003

42. Matias PJ, Ferreira C, Jorge C, Borges M, Aires I, Amaral T, Gil C, Cortez J, Ferreira A: 25-Hydroxyvitamin D3, arterial calcifications and cardiovascular risk markers in haemodialysis patients. Nephrol Dial Transplant 24:611-618, 2009

43. Coratelli P, Petrarulo F, Buongiorno E, Giannattasio M, Antonelli G, Amerio A: Improvement in left ventricular function during treatment of hemodialysis patients with 25-OHD3. Contrib Nephrol 41:433-437, 1984

44. Kim HW, Park CW, Shin YS, Kim YS, Shin SJ, Kim YS, Choi EJ, Chang YS, Bang BK: Calcitriol regresses cardiac hypertrophy and QT dispersion in secondary hyperparathyroidism on hemodialysis. Nephron Clin Pract 102:c21-c29, 2006

45. Kovesdy CP, Ahmadzadeh S, Anderson JE, Kalantar-Zadeh K: Association of activated vitamin D treatment and mortality in chronic kidney disease. Arch Intern Med 168:397-403, 2008

46. Singh NP, Sahni V, Garg D, Nair M: Effect of pharmacological suppression of secondary hyperparathyroidism on cardiovascular hemodynamics in predialysis CKD patients: A preliminary observation. Hemodial Int 11:417-423, 2007

47. Moore C, Murphy MM, Keast DR, Holick MF: Vitamin D intake in the United States. J Am Diet Assoc 104:980-983, 2004

16

CHAPTER 2

Association of 25-hydroxyvitamin D deficiency with NT-proBNP levels in AMI

patients

Authors : Rajyalakshmi Gadi, 1, 3 James B Wetmore, 1 John H Lee, 2 James H O’Keefe, 2

Paul S Chan, 2 Fengming Tang, 2 and John A Spertus 2

From: 1 Division of Nephrology and Hypertension, University of Kansas Medical Center,

Kansas City, KS, USA, and 2 Department of Cardiovascular Research, Saint Luke’s

Hospital Mid-America Heart Institute, Kansas City, MO, USA

3To whom correspondence should be addressed: Rajyalakshmi Gadi, MD,

University of Kansas Medical Center, MS 3002, 3901 Rainbow Blvd., Kansas City, KS

66160; Tel: 913-588-6074; Fax: 913-588-3867; email: [email protected].

Running Title : nutritional vitamin D deficiency and biomarkers in AMI

Word count , abstract: 247

Word count , text: 2992

17

ABSTRACT

Background and objectives: Nutritional vitamin D deficiency is an emerging risk factor

for acute myocardial infarction (AMI) and heart failure. The association between 25-

hydroxyvitamin D levels and N-terminal pro B-type natriuretic peptide (NT-proBNP), a

robust prognostic marker for post-MI mortality and heart failure is unknown and could

illuminate a potential pathway for adverse outcomes among post-MI patients with 25-

hydroxyvitamin D deficiency.

Design, setting, participants and measurements: In a cross sectional analysis, we

studied 238 AMI patients from 21 US centers to test the association of nutritional

vitamin D (25-hydroxyvitamin D [25(OH)D]) deficiency with NT-proBNP levels. Patients’

25(OH)D levels were categorized as normal (≥30 ng/ml), insufficient (>20 ─ <30 ng/ml),

deficient (>10 ─ ≤ 20 ng/ml), and severely deficient (≤10 ng/ml) groups.

Results: 96% of AMI patients had low 25(OH)D levels, with 75% having 25(OH)D

deficiency and 21% having insufficiency. No significant trends for higher mean log NT-

proBNP levels in severely deficient (6.9 ± 1.3 pg/ml), deficient (6.9 ± 1.2 pg/ml) and

insufficient (6.9 ± 0.9 pg/ml) groups were observed as compared with patients having

normal (6.1 ± 1.7 pg/ml) levels, P = 0.165. In multivariate regression model after

adjusting for several covariates, 25(OH)D was not associated with NT-proBNP levels.

Conclusion: Potential associations between nutritional vitamin D deficiency and

prognosis in the setting of AMI are unlikely to be mediated through NT-proBNP

pathways. Future studies should examine other mechanisms such as inflammation and

vascular calcification by which 25(OH)D deficiency could mediate adverse outcomes

such as heart failure and mortality post AMI.

18

Keywords : vitamin D, N-terminal proBNP, acute myocardial infarction

Introduction

Nutritional vitamin D deficiency is highly prevalent, occurring in approximately

30%-50% of the general population (1,2). In several studies, 25-hydroxyvitamin D

[25(OH)D] deficiency has been independently associated with both incident acute

myocardial infarction (AMI) (3) and heart failure (HF) (4,5), suggesting that 25(OH)D

plays an important role in cardiac function. Supporting this hypothesis, several in vitro

studies have shown that calcitriol (1,25(OH)2D3), an active form of vitamin D, regulates

intracellular calcium metabolism and myocardial contractility through specific vitamin D

receptors on cardiac myocytes (6-8). Consequently, 25(OH)D deficiency has been

associated with aberrant cardiac contractility, cardiomegaly, and increased ventricular

mass due to myocardial collagen deposition (7,8), independent of its known effects on

blood pressure (9). In studies of experimental animals, activation of nuclear vitamin D

receptors by 1,25(OH)2D3 suppresses the expression and secretion of atrial natriureitc

peptide (ANP) and brain natriuretic peptide (BNP) in cardiac myocytes (10-12), while in

clinical studies of predialysis chronic kidney disease patients, individuals treated with

1,25(OH)2D3 for 12 weeks were observed to have improved left ventricular diastolic

function as compared with placebo (13). Collectively, these findings suggest that low

circulating levels of 25(OH)D could potentially contribute to, or potentiate, the

development of left ventricular dysfunction (LVDF) and heart failure after AMI.

In recent years, N-terminal pro-brain natriuretic peptide (NT-proBNP), a

prohormone of BNP released from cardiac ventricles, has been associated with the

19

severity of left ventricular dilatation and dysfunction after AMI (14,15). In addition, NT-

proBNP is a sensitive and robust prognostic biomarker of mortality in AMI (16-18), HF

(19,20) and chronic hemodialysis (21,22) patients. Moreover, NT-proBNP levels have

direct clinical implications, as they are used to guide therapeutic interventions in HF

patients, leading to improved outcomes as compared with routine clinical treatment (23).

Thus, while both 25(OH)D and NT-proBNP are both known to be associated with

LV dysfunction after AMI, it is not known whether there is a correlation between levels of

25(OH)D and NT-proBNP. In clinical studies, inverse associations between 25(OH)D

levels and NT-pro ANP in HF (4) and between 25(OH)D and BNP in dialysis patients

have been suggested (24,25). However, these studies had small sample sizes and

were not based on patients living in the U.S., where better nutrition and fortification of

milk is common (26). Given the adverse health implications of 25(OH)D deficiency and

the lack of studies in AMI patients (a particularly high-risk group), we examined the

association of 25(OH)D deficiency with NT-proBNP levels in a multicenter cohort of AMI

patients. Discovery of an association between circulating levels of 25(OH)D and NT-

proBNP would not only suggest a potential pathway for adverse outcomes among post-

AMI patients with 25(OH)D deficiency, but could also identify a potentially novel

therapeutic target (i.e., nutritional vitamin D supplementation) to reduce NT-proBNP

levels in the hopes of improving prognosis after MI.

Materials and Methods

Study Population

20

This study is a cross sectional analysis of a cohort study. Participants were

drawn from the TRIUMPH (Translational Research Investigating Underlying disparities

in recovery from acute Myocardial infarction: Patients' Health status) study, a

prospective multicenter cohort study of AMI patients across 21 U.S. centers. Patients

were eligible for TRIUMPH if they were ≥ 18years of age and had a diagnosis of AMI.

AMI was diagnosed by the presence of either a CK–MB elevation greater than twice

normal or Troponin-I elevation of >0.1mg/ml within 24 hours of arrival to the hospital

with a clinical presentation suggestive of an AMI (e.g., prolonged ischemic signs or

chest pain symptoms, at least one EKG with ST- wave elevation or ST-wave depression

in 2 or more consecutive leads, and no alternative explanation for the presence of

elevated serum cardiac markers). Patients were excluded if they transferred to the

participating hospital from another facility greater than 24 hours after their original AMI

presentation, if they refused or could not provide informed consent, or if they were

receiving hospice care. For this study, we included the last 250 TRIUMPH patients, in

each of whom vitamin D levels were assessed in addition to the standard data collected.

TRIUMPH complied with the Declaration of Helsinki and was approved by the

institutional review boards of each participating institution. Written informed consent was

obtained from all participants.

Data Collection

During index hospitalization, trained data collectors performed a patient interview

and detailed chart abstraction within 24-72 hours of admission. Patient data at AMI

presentation, including demographic features, socioeconomic status, co-morbidities,

21

severity of AMI (ST elevation vs. non-ST elevation AMI), Killip class, Rose dyspnea

index, vital signs, and laboratory values were abstracted. The Rose dyspnea score is a

validated self-reported question in AMI patients that is scored from 0-4, with higher

scores indicating worse dyspnea (27). Regional and seasonal data were collected,

given their association with vitamin D levels (3,28,29). Patients were classified by their

residing states in to 5 geographic regions, namely northeast (NE), southeast (SE),

southwest (SW), mid-west (MW) and west (W). Months of April -June, July-August and

September-December were classified as summer, fall, and early winter seasons,

respectively.

Left ventricular systolic function was classified as normal, mild, moderate or

severe as assessed by echocardiography, angiography or nuclear imaging and as

documented in the hospital record. Finally, reperfusion therapy (coronary angiography,

percutaneous intervention [PCI], and/or coronary artery bypass surgery [CABG]) and

other acute therapies as well as medications prescribed at discharge were recorded.

Blood samples from all consenting patients were sent to a core laboratory (Clinical

Reference Laboratories, Lenexa, KS) for NT-proBNP measurement (Roche

Diagnostics, Indianapolis, IN) and 25(OH)D assays, as described below.

Laboratory Measurements

An in vitro radioimmunoassay (RIA) assay (DiaSorin, Stillwater, MN) was used

for quantitative determination of 25(OH)D and other hydroxylated vitamin D metabolites

in human serum. The DiaSorin 25(OH)D assay comprises a two-step procedure

involving both a rapid extraction of 25(OH)D and other hydroxylated metabolites from

22

serum or plasma with acetonitrile and a 25(OH)D-specific antibody and tracer while

incubating for 90 minutes at 20-25°C, and phase separat ion with a second antibody-

precipitating complex. The total (intra- and inter-assay) precision of the assay has a

coefficient of variation (CV) of 9.4% and 11% for control values of 8.6ng/ml and

49.0ng/ml, respectively.

For measurement of circulating NT-proBNP levels, the Roche electro-

chemiluminescence immunoassay for the in vitro quantization was used (ECLIA Roche

diagnostics GmbH, Mannheim, Germany). The precision of this assay was represented

by a CV of 3.2% and 2.3% for control values of 175pg/ml and 4962 pg/ml, respectively.

Blood samples for NT-proBNP were drawn prior to hospital discharge. The mean time

for NT-proBNP blood draw was 3.5 ± 4 days in this cohort. Glomerular filtration rate

(eGFR) was estimated using the four variable Modified Diet in Renal Disease (MDRD)

study equation (30).

Statistical analysis

Participants were classified into clinically-relevant categories on the basis of

25(OH)D levels. AMI patients with levels ≥30 ng/ml were classified as normal, while

levels >20ng/ml and <30ng/ml were considered insufficient, levels of >10 to ≤ 20ng/ml

were deficient, and levels ≤10ng/ml were severely deficient. Median NT-proBNP levels

were compared using the non-parametric Kruskal-Wallis test, due to the positively

skewed distribution of NT-proBNP. Log NT-proBNP and quartiles of NT-proBNP were

compared across the four 25(OH)D strata using ANOVA and chi-square tests,

respectively. Spearman rank correlation was obtained between NT-proBNP and

23

25(OH)D levels. Multivariable linear regression analysis to determine the association of

25(OH)D levels with log NT-proBNP was performed, adjusting for relevant covariates

and confounders. A p value of <0.05 was considered statistically significant. All

analyses were conducted using SAS v9.1 software (Cary, NC, U.S.).

Results

Characteristics of participants across 25(OH)D groups are shown in Table 1. Of

the 238 enrolled patients, the median 25(OH)D concentration was 16 ng/ml

(interquartile range [IQR] 12-21). Classifying 25(OH)D levels into clinically-interpretable

ranges, 40 (16.8%) were found to be severely deficient, 138(57.9%) deficient and 50

(21.0%) insufficient. Only 4.2% of participants in the study had normal 25(OH)D levels,

with none of the African-American participants having normal 25(OH)D levels.

No statistically significant differences by age or gender were noted across

25(OH)D groups. Deficiency in 25(OH)D was associated with a history of recent

smoking, low physical activity, lack of medical insurance, and poor social support. No

difference in those with or without 25(OH)D deficiency were observed for hypertension,

diabetes, history of MI, HF, or the type of AMI (STEMI vs. NSTEMI). Similarly, Killip

class at arrival and Rose dyspnea scores did not differ by 25(OH)D levels. During the

study period, no statistically significant regional variations were observed in patients’

enrollment across 25(OH)D groups (p = 0.79), nor did estimated glomerular filtration

rates differ (p = 0.24). There was a trend towards seasonal variation in 25(OH)D levels

(p = 0.064). As expected, lower serum calcium and higher parathyroid hormone levels

(PTH) were significantly associated with 25(OH)D deficiency (p values of 0.03 and

24

0.004, respectively). Likewise the use of Omega 3 supplements was higher in patients

with normal 25(OH)D levels (50%) compared to those who had insufficient (36%),

deficient (22.5%) and severely deficient (7.5%) levels (p value = 0.002).

No statistically significant correlation between 25(OH)D and log NT-proBNP

levels was observed (rho = - 0.0025, p = 0.97; Figure 1). No significant trends for higher

mean log NT-proBNP levels in severely deficient (6.9 ± 1.3 pg/ml), deficient (6.9 ± 1.2

pg/ml) and insufficient (6.9 ± 0.9 pg/ml) groups were observed as compared with

patients having normal (6.1 ± 1.7 pg/ml) levels, p = 0.165.(Table 2).

In the multivariable linear regression model, after adjusting for age, race, gender,

BMI, social support, medical insurance, smoking status, seasons, co morbidities such

as diabetes and history of prior MI, in-hospital percutaneous coronary intervention (PCI)

and coronary artery bypass graft (CABG), high sensitivity C reactive protein, e GFR and

omega 3 supplements use, 25(OH)D levels were not significantly associated with log

NT-proBNP levels. (Table 3).However, NT-proBNP levels remained independently

associated with hs CRP, a marker of inflammation, even after adjusting for 25(OH)D

levels.

Discussion

In this cross sectional observational study, we found no evidence of an

association between 25(OH)D levels and NT-proBNP levels, nor other clinical markers

of acute LV dysfunction, including Killip class or Rose dyspnea score. To our

knowledge, this is the first study to compare levels 25(OH)D and NT-proBNP in an AMI

population. Thus, while both 25(OH)D levels and NT-proBNP are associated with

25

cardiovascular disease and heart failure, they appear to impact prognosis through

different mechanisms in the setting of AMI.

Importantly, however, we found exceedingly low levels of nutritional vitamin D in

this cohort of AMI patients, such that few patients had normal 25(OH)D levels. In fact,

74.7% of these AMI patients had significant 25(OH)D deficiency with another 21%

having insufficiency, a prevalence much higher than the national estimates of 25%-57%

in the general population (31-33). Consistent with previous reports, we also found that

25(OH)D deficiency was associated with older age, higher BMI, smoking, low physical

activity and lesser use of omega 3 supplements (1,28,34). Moreover, we were able to

extend these previously known clinical correlations by finding that 25(OH)D deficiency is

associated with low social support and lack of medical insurance, both of which are

associated with higher morbidity and mortality after MI (34-36). Thus although our data

do not support an association between 25(OH)D deficiency and a biomarker of LV

dysfunction in the setting of AMI, the high prevalence of 25(OH)D deficiency in this

cohort of AMI patients is noteworthy. Given that 25(OH)D deficiency has been

associated with incident MI (3) and CV events (47) in prior observational studies, it

would be important for future investigators to examine whether rectifying vitamin D

levels in post-AMI patients is associated with an improvement in subsequent outcomes.

Our findings extend and confirm the preliminary insights provided by Pilz et al,

who studied patients with established CAD referred to coronary angiography. While they

found that 25(OH)D deficiency was significantly associated with higher NYHA functional

HF classes in unadjusted analysis, this association was no longer significant after

adjustment for other clinical variables (37). In contrast, our findings are dissimilar from

26

prior observations that in hemodialysis patients,where lower 25(OH)D levels were

found to be associated with significantly higher log BNP levels (24). NT-proBNP is a

more stable form of BNP and correlates well with BNP levels in heart failure patients

(38). Similar reports in chronic HF patients have also suggested an inverse, nonlinear

association between 25(OH)D and NT-proANP levels (r2 = 0.16, p<0.001) (4), although

a clinical trial of nutritional vitamin D supplementation failed to alter patients’ NT-proBNP

levels (39). More recently, a large cohort of patients referred to coronary angiography in

the LU dgwigshafen RIsk and Cardiovascular Health (LURIC) study suggested that

25(OH)D levels remained independently associated with NT-proBNP levels in a

multivariable model (β = -0.180, p <0.001) (37). It is unknown whether the discrepancy

in our findings and these previous studies is due to unmeasured confounding in prior

reports, or whether the lack of an association is due to the impact of acute myocardial

injury on NT-proBNP levels (40,41).

In recent years there has been growing interest in the role of 25(OH)D and

optimal organ function, including that of the cardiovascular (CV) system. Cardiac

myocytes have cytosolic vitamin D receptors (VDR) (42) that bind active vitamin D, but

lack 1α-hydroxylase activity (43). Hence cardiac muscle is strongly dependent upon

circulating calcitriol level, underscoring the importance of studying 25(OH)D levels in

patients with cardiovascular disease. For example, several studies have shown in-vitro

25(OH)D deficiency to be associated with impaired cardiac inotropy, increased left

ventricular mass due to myocardial collagen deposition (7,8), enhanced atherosclerosis

secondary to vascular smooth muscle cell (VSMC) proliferation (44,45) and pro-

inflammatory cytokines (IL-6 and TNF-alfa) (46). Furthermore, large epidemiological

27

studies, including the Health Professionals study (3) and Framingham offspring study

(47) have shown low 25(OH)D levels (<15ng/ml vs. ≥30ng/ml) to be independently

associated with a doubling of risk for incident MI, both fatal and nonfatal events.

Moreover, the risk of all-cause mortality was higher among subjects with 25(OH)D

levels <17.8ng/ml in NHANES III as compared with subjects having levels >32ng/ml.

Finding no association between vitamin D levels and NT-proBNP in our study suggests

that the association of 25(OH)D with AMI mortality is unlikely to be mediated through

NT-proBNP, despite the prognostic association between both biomarkers and survival.

The results of our study should be interpreted in the context of some potential

limitations. Our sample size was small, and therefore was underpowered for some

analyses. For example, we did not find an association with hypertension (HTN) or

diabetes (28,47), chronic kidney disease (42,48) or heart failure history (4,5) as reported

in prior studies. However, in this study of 238 patients we had 80% power to observe an

unadjusted correlation of 0.18 between 25(OH)D and NT-proBNP. Importantly, there

were very few patients (n =10) who had normal 25(OH)D levels, and if there is a non-

linear association between lower and normal 25(OH)D levels, we may have been

underpowered to detect this. It is also possible that the association between the

biomarkers we tested might have been stronger at a time of clinical stability or at the

time of acute presentation, since our samples for both 25(OH)D and NT-proBNP were

taken prior to discharge; however the prognostic importance of NT-proBNP at the time

of MI has consistently been shown to be strong.(17,18)

In conclusion, the mechanism by which nutritional vitamin D deficiency mediates

outcomes in AMI patients does not appear to be through its effects on, or a relationship

28

with, NT-pro BNP. Future studies should better clarify the clinical mechanism by which

25(OH)D deficiency is associated with outcomes in AMI patients. Potential candidates

might include more long-term processes such as inflammation and vascular

calcification. While we did not observe an association between levels of 25(OH)D and

NT-proBNP, we did find a remarkably high prevalence of 25(OH)D deficiency among

AMI patients. Hospitalization for an AMI offers clinicians an opportunity to not only

identify modifiable risk factors and optimize medications for secondary prevention but

also to address other co morbidities, such as 25(OH)D deficiency. Given that nutritional

vitamin D is readily available, inexpensive, and has a good safety profile, future studies

should investigate whether addressing 25(OH)D deficiency might improve outcomes in

AMI patients, regardless of the mechanism of its known association with post-MI risk.

Acknowledgments

TRIUMPH was supported by the NHLBI Specialized Center of Clinically Oriented

Research in Cardiac Dysfunction and Disease (grant no. P50 HL077113) and

Cardiovascular Outcomes, Inc, Kansas City, Missouri. Dr.Gadi was supported by

Genzyme research fellowship award.

Disclosures

None

29

Figure1. Spearman correlation between 25(OH)D (ng/ml) and NT-proBNP levels(pg/ml)

NT

-pro

BN

P (

pg/m

l)

0

10000

20000

30000

40000

25-hydroxyvitamin D (ng/ml)

0 10 20 30 40

Spearman's rho = - 0.0025, P =0.97

30

Table1. Baseline characteristics of study participants, by 25(OH)D groups

25(OH)Dgroup, (ng/ml)

Characteristic 0 −10 >10 − ≤20 >20 − <30 ≥ 30 P-Value

(n= 40) (n = 138) (n = 50) (n = 10) Demographics Age(yr)a 55.0±11.2 57.8±11.8 58.8±10.2 62.4±11.3 0.209 Male, n (%) 26(56) 104(75.4) 38 (76) 8 (80) 0.569 Caucasian, n (%) 23 (57.5) 103 (74.6) 41 (82) 10 (100) 0.012 BMIb (kg/m2) 32.1 ± 5.6 30.9 ± 6.9 29.5 ± 5.6 26.5 ± 4.6 0.066 Socio-economic status Low social support, n (%) 11 (27.5) 20 (14.9) 3 (6) 0 (0) 0.023 No health insurance, n (%) 14 (35) 28 (20.9) 5 (10.4) 1 (10) 0.036 Lifestyle Smoked within 1 month, n (%) 24 (60) 49 (35.5) 15 (30) 0(0) <0.001 Leisure time activity, n (%) 0.047 Mainly sedentary 20 (50) 56 (40.9) 19 (38) 1 (10) Mild exercise 13 (32.5) 49 (35.8) 11 (22) 3 (30) Moderate exercise 6 (15) 27 (19.7) 17 (34) 4 (40) Strenuous exercise 1 (2.5) 5 (3.6) 3 (6) 2 (20) Geographical region, n (%) 0.79 Midwest 18 (56.3) 53 (45.3) 15 (34.1) 5 (62.5) Northeast 3 (9.4) 15 (12.8) 5 (11.4) 0 (0) Southeast 5 (15.6) 23 (19.7) 9 (20.5 ) 1 (12.5) Southwest 4 (12.5) 22 (18.8) 12 (27.3) 2 (25) West 2 (6.3) 4 (3.4) 3 (6.8 ) 0 (0) Season, n (%) 0.064 Apr-Jun 1 (2.5) 1 (0.7) 0 (0) 0 (0) Jul-Sep 10 (25) 56 (40.6) 24 (48) 7 (70) Oct-Dec 29 (72.5) 81 (58.7) 26 (52) 3 (30) Co-morbidities, n (%) Diabetes 13 (32.5) 41 (29.7) 8 (16) 2 (20) 0.209 Hypertension 26 (65) 89 (64.5) 33 (66) 6 (60) 0.99 Prior MI 2 (5) 36 (26.1) 9 (18) 2 (20) 0.019 CKDb 5 (12.5) 4 (2.9) 2 (4) 1 (10) 0.06 Chronic heart failure 3 (7.5) 6 (4.3) 1 (2) 1 (10) 0.373 LVH on admission EKG, n (%) 7 (17.9) 9 (6.7) 4 (8.2) 0 (0) 0.171 Killip class on arrival, n (%) 0.347 I 36 (94.7) 128 (94.8) 43 (86) 9 (90) II 2 (5.3) 4 (3) 5 (10) 1 (10) III 0 (0) 2 (1.5) 2 (4) 0 (0) IV 0 (0) 1 (0.7) 0 (0) 0 (0) Rose dyspnea score 1.0 ± 0.9 0.9 ± 0.9 0.8 ± 0.8 0.6 ± 1.0 0.43 Final MI diagnosis, n (%) 0.352 STEMIb 15 (37.5) 64 (46.4) 23 (46) 2 (20) NSTEMI 25 (62.5) 74 (53.6) 27 (54) 8 (80) LV systolic function, n (%) 0.533 Normal 24 (61.5) 88 (63.8) 36 (72) 6 (60) Mild 8 ( 20.5) 24 (17.4) 10 (20) 1 (10) Moderate 4 (10.3) 16(11.6) 4 (8) 1 (10) Severe 3 (7.7) 10 (7.2) 0 (0) 2 (20) Laboratory values Troponin I (ng/ml)a 1.4 ±1.9 1.7 ± 1.8 1.6 ± 1.7 0.7 ± 1.2 0.3 Total Cholesterol (mg/dl)a 150.6 ± 47 157.3 ± 29 150.5 ± 29 189.7 ± 47 0.011

31

HDL(mg/dl)a 37.9 ± 10.6 38.4 ± 8.7 40.6 ± 10.7 43.1 ± 25.2 0. 353 LDL (mg/dl)a 92.2 ± 36.7 97.4 ± 25 90.4 ± 25.7 119.2 ± 42.8 0.0 32 Triglycerides(mg/dl)a 153.2 ± 74 185.6 ± 145 143.6 ± 79 195.4 ± 169 0.1 61 Calcium (mg/dl) 8.9 ± 0.6 8.8 ± 0.6 9.0 ± 0.4 9.3 ± 0.6 0.038 Intact PTHb (pg/ml) 59.8 ± 67.7 40.1 ± 28.1 32.8 ± 15.3 32.0 ± 26.5 0.004 Phosphate (mg/dl) 3.6 ± 0.7 3.5 ± 0.8 3.5 ± 0.7 3.7 ± 0.9 0.683 hs CRPb (mg/l) 5.2 ± 5.6 3.8 ± 5.3 3.8 ± 4.5 0.9 ± 0.8 0.1 12 e GFRb 84.7 ± 28.1 88.3 ± 27.3 80.3 ± 18.9 80.1 ± 18.8 0. 241 Medications ACEI/ARBb, n (%) 11 (27.5) 39 (28.3) 18 (36) 1 (10) 0.424 Diuretic, n (%) 9 (22.5) 31 (22.5) 15 (30) 10(10) 0.571 Omega-3 supplements, n (%) 3 (7.5) 31 (22.5) 18 (36) 5 (50) 0.002

aData shown as mean ± standard deviation. bBMI, body mass index; CKD, chronic kidney disease; STEMI, ST-segment elevation myocardial infarction; eGFR, estimated glomerular filtration rate; PTH, parathyroid hormone; hs CRP, high sensitivity C-reactive protein; ACEI, angiotensin converting enzyme inhibitors; ARB, angiotensin receptor blockers

Table2. Association of 25(OH)D with NT-proBNP levels

25(OH)D group, ng/ml

0 −10 >10 − ≤20 >20 − <30 ≥ 30 Characteristic (n = 40) (n = 138) (n = 50) (n= 10) P -Value

Median NT-proBNP (pg/ml) 1094.5 897.5 1065.5 757.5 0.457

Log NT-pro BNPa 6.9 ± 1.3 6.9 ± 1.2 6.9 ± 0.9 6.1 ± 1.7 0.165

NT-pro BNPb, n (%) 0.579

Quartile 1 (5 to <488) 11 (27.5) 34 (24.6) 10 (20) 3 (30)

Quartile 2 (488 to <924.5) 8 (20) 38 (27.5) 13 (26) 2 (20)

Quartile 3 (924.5 to <1882) 9 (22.5) 31 (22.5) 14 (28) 5 (50)

Quartile 4 (1882 to 31230) 12 (30) 35 (25.4) 13 (26) 0(0) aValues shown are means ± standard deviation. bNT-proBNP, N terminal pro brain natriuretic peptide.

32

Table3. Results of Multivariable regression analysis

aBMI, body mass index; MI, myocardial infarction; PCI, percutaneous coronary intervention; CABG, coronary artery bypass graft; hs CRP, high sensitivity c reactive protein; e GFR, estimated glomerular filtration rate;25 (OH)D,25 hydroxy vitamin D. b log BNP change for 1ngm/l change in 25(OH)D

Covariate β Coefficient P -Value

Age per 10yr increment 0.256 0.001

Caucasian race 0.374 0.03

Male sex -0.179 0.27

BMIa per 5 units increment -0.115 0.06

Lack of social support -0.205 0.33

No health insurance 0.275 0.17

Smoked within 1month 0.027 0.87

Seasons

Nov-Feb vs. Mar-Oct 0.170 0.25

History of Diabetes 0.006 0.97

History of prior MIa 0.244 0.16

In hospital PCIa 0.164 0.35

In hospital CABGa -0.152 0.55

hs CRPa 0.439 <0.0001

e GFRa -0.040 0.008

Omega 3 supplements -0.217 0.2

25(OH)Da ng/ml

<10 vs. ≥30 0.907b 0.06

>10- ≤ 20 vs. ≥30 0.848 0.05

>20-<30 vs. ≥30 0.764 0.08

33

References

1. Lee JH, O'Keefe JH, Bell D, Hensrud DD, Holick MF: Vitamin D deficiency an important, common, and easily treatable cardiovascular risk factor? J Am Coll Cardiol 52:1949-1956, 2008




5. Zittermann A, Schleithoff SS, Koerfer R: Vitamin D insufficiency in congestive heart failure: why and what to do about it? Heart Fail Rev 11:25-33, 2006

6. Weishaar RE, Simpson RU: Vitamin D3 and cardiovascular function in rats. J Clin Invest 79:1706-1712, 1987



9. Weishaar RE, Simpson RU: The involvement of the endocrine system in regulating cardiovascular function: emphasis on vitamin D3. Endocr Rev 10:351-365, 1989

10. Wu J, Garami M, Cao L, Li Q, Gardner DG: 1,25(OH)2D3 suppresses expression and secretion of atrial natriuretic peptide from cardiac myocytes. Am J Physiol 268:E1108-E1113, 1995

11. Chen S, Nakamura K, Gardner DG: 1,25-dihydroxyvitamin D inhibits human ANP gene promoter activity. Regul Pept 128:197-202, 2005

12. Bidmon HJ, Gutkowska J, Murakami R, Stumpf WE: Vitamin D receptors in heart: effects on atrial natriuretic factor. Experientia 47:958-962, 1991

13. Singh NP, Sahni V, Garg D, Nair M: Effect of pharmacological suppression of secondary hyperparathyroidism on cardiovascular hemodynamics in predialysis CKD patients: A preliminary observation. Hemodial Int 11:417-423, 2007

34






19. Bayes-Genis A, Lopez L, Zapico E, Cotes C, Santalo M, Ordonez-Llanos J, Cinca J: NT-ProBNP reduction percentage during admission for acutely decompensated heart failure predicts long-term cardiovascular mortality. J Card Fail 11:S3-S8, 2005

20. Januzzi JL, van KR, Lainchbury J, Bayes-Genis A, Ordonez-Llanos J, Santalo-Bel M, Pinto YM, Richards M: NT-proBNP testing for diagnosis and short-term prognosis in acute destabilized heart failure: an international pooled analysis of 1256 patients: the International Collaborative of NT-proBNP Study. Eur Heart J 27:330-337, 2006

21. Madsen LH, Ladefoged S, Corell P, Schou M, Hildebrandt PR, Atar D: N-terminal pro brain natriuretic peptide predicts mortality in patients with end-stage renal disease in hemodialysis. Kidney Int 71:548-554, 2007

22. Roberts MA, Srivastava PM, Macmillan N, Hare DL, Ratnaike S, Sikaris K, Ierino FL: B-type natriuretic peptides strongly predict mortality in patients who are treated with long-term dialysis. Clin J Am Soc Nephrol 3:1057-1065, 2008

23. Troughton RW, Frampton CM, Yandle TG, Espiner EA, Nicholls MG, Richards AM: Treatment of heart failure guided by plasma aminoterminal brain natriuretic peptide (N-BNP) concentrations. Lancet 355:1126-1130, 2000

24. Matias PJ, Ferreira C, Jorge C, Borges M, Aires I, Amaral T, Gil C, Cortez J, Ferreira A: 25-Hydroxyvitamin D3, arterial calcifications and cardiovascular risk markers in haemodialysis patients. Nephrol Dial Transplant 24:611-618, 2009

25. Obineche EN, Saadi H, Benedict S, Pathan JY, Frampton CM, Nicholls MG: Interrelationships between B-type natriuretic peptides and vitamin D in patients on maintenance peritoneal dialysis. Perit Dial Int 28:617-621, 2008

26. Moore C, Murphy MM, Keast DR, Holick MF: Vitamin D intake in the United States. J Am Diet Assoc 104:980-983, 2004

35

27. Arnold SV, Spertus JA, Jones PG, Xiao L, Cohen DJ: The impact of dyspnea on health-related quality of life in patients with coronary artery disease: results from the PREMIER registry. Am Heart J 157:1042-1049, 2009


29. Zittermann A, Schleithoff SS, Koerfer R: Putting cardiovascular disease and vitamin D insufficiency into perspective. Br J Nutr 94:483-492, 2005

30. Murthy K, Stevens LA, Stark PC, Levey AS: Variation in the serum creatinine assay calibration: a practical application to glomerular filtration rate estimation. Kidney Int 68:1884-1887, 2005

31. Holick MF: High prevalence of vitamin D inadequacy and implications for health. Mayo Clin Proc 81:353-373, 2006

32. Looker AC, wson-Hughes B, Calvo MS, Gunter EW, Sahyoun NR: Serum 25-hydroxyvitamin D status of adolescents and adults in two seasonal subpopulations from NHANES III. Bone 30:771-777, 2002

33. Tangpricha V, Pearce EN, Chen TC, Holick MF: Vitamin D insufficiency among free-living healthy young adults. Am J Med 112:659-662, 2002

34. Mookadam F, Arthur HM: Social support and its relationship to morbidity and mortality after acute myocardial infarction: systematic overview. Arch Intern Med 164:1514-1518, 2004

35. Burg MM, Barefoot J, Berkman L, Catellier DJ, Czajkowski S, Saab P, Huber M, DeLillo V, Mitchell P, Skala J, Taylor CB: Low perceived social support and post-myocardial infarction prognosis in the enhancing recovery in coronary heart disease clinical trial: the effects of treatment. Psychosom Med 67:879-888, 2005

36. Rahimi AR, Spertus JA, Reid KJ, Bernheim SM, Krumholz HM: Financial barriers to health care and outcomes after acute myocardial infarction. JAMA 297:1063-1072, 2007

37. Pilz S, Marz W, Wellnitz B, Seelhorst U, Fahrleitner-Pammer A, Dimai HP, Boehm BO, Dobnig H: Association of vitamin D deficiency with heart failure and sudden cardiac death in a large cross-sectional study of patients referred for coronary angiography. J Clin Endocrinol Metab 93:3927-3935, 2008

38. Mueller T, Gegenhuber A, Poelz W, Haltmayer M: Head-to-head comparison of the diagnostic utility of BNP and NT-proBNP in symptomatic and asymptomatic structural heart disease. Clin Chim Acta 341:41-48, 2004

39. Schleithoff SS, Zittermann A, Tenderich G, Berthold HK, Stehle P, Koerfer R: Vitamin D supplementation improves cytokine profiles in patients with congestive heart failure: a double-blind, randomized, placebo-controlled trial. Am J Clin Nutr 83:754-759, 2006

40. de Lemos JA, Morrow DA: Brain natriuretic peptide measurement in acute coronary syndromes: ready for clinical application? Circulation 106:2868-2870, 2002

36

41. Morrow DA, de Lemos JA, Sabatine MS, Murphy SA, Demopoulos LA, DiBattiste PM, McCabe CH, Gibson CM, Cannon CP, Braunwald E: Evaluation of B-type natriuretic peptide for risk assessment in unstable angina/non-ST-elevation myocardial infarction: B-type natriuretic peptide and prognosis in TACTICS-TIMI 18. J Am Coll Cardiol 41:1264-1272, 2003


43. Hewison M, Zehnder D, Chakraverty R, Adams JS: Vitamin D and barrier function: a novel role for extra-renal 1 alpha-hydroxylase. Mol Cell Endocrinol 215:31-38, 2004

44. Mitsuhashi T, Morris RC, Jr., Ives HE: 1,25-dihydroxyvitamin D3 modulates growth of vascular smooth muscle cells. J Clin Invest 87:1889-1895, 1991

45. Mohtai M, Yamamoto T: Smooth muscle cell proliferation in the rat coronary artery induced by vitamin D. Atherosclerosis 63:193-202, 1987

46. Muller K, Haahr PM, Diamant M, Rieneck K, Kharazmi A, Bendtzen K: 1,25-Dihydroxyvitamin D3 inhibits cytokine production by human blood monocytes at the post-transcriptional level. Cytokine 4:506-512, 1992


48. Gonzalez EA, Sachdeva A, Oliver DA, Martin KJ: Vitamin D insufficiency and deficiency in chronic kidney disease. A single center observational study. Am J Nephrol 24:503-510, 2004

37

CHAPTER 3: DISCUSSION

Project Summary

In this study we evaluated the association of nutritional vitamin D deficiency with

NT-proBNP levels in acute myocardial infarction patients. Nutritional vitamin D [25-

hydroxyvitamin D 25(OH)D] deficiency is highly prevalent, occurring in approximately

30%-50% of the general population.1;2 In several studies, 25-hydroxyvitamin D

[25(OH)D] deficiency has been independently associated with both incident acute

myocardial infarction (AMI) 3 and heart failure (HF) 4;5, suggesting that 25(OH)D plays

an important role in cardiac function. As shown in several in-vitro studies, 25(OH)D

deficiency has been associated with aberrant cardiac contractility, cardiomegaly, and

increased ventricular mass due to myocardial collagen deposition. 6;7 On the other

hand, N-terminal pro-brain natriuretic peptide (NT-proBNP), a prohormone of BNP

released from cardiac ventricles, has been shown to be a sensitive and robust

prognostic biomarker for post-MI mortality and heart failure.8-12 Thus, low circulating

levels of 25(OH)D could potentially contribute to, or potentiate, the development of left

ventricular dysfunction (LVDF) and heart failure after AMI. We therefore hypothesized

that low levels of 25(OH)D would be associated with higher NT-proBNP levels which is

a marker of heart failure.

We prospectively studied 238 acute myocardial infarction (AMI) patients from 21

US centers to test the association of nutritional vitamin D deficiency (25-hydroxyvitamin

D [25(OH)D]) with NT-proBNP levels. Patients’ 25(OH)D levels were categorized in to

clinically interpretable groups such as normal (≥30ng/ml), insufficient (<30ng/ml and

38

>20ng/ml), deficient (≤ 20ng/ml and >10ng/ml), and severely deficient (≤ 10ng/ml)

groups.

Surprisingly 96% of AMI patients were found to have low 25(OH)D levels, of

which 40 (16.8%) were severely deficient, 138(57.9%) deficient and 50 were (21.0%)

insufficient. The median 25(OH)D concentration was 16 ng/ml (interquartile range [IQR]

12-21). Notably, none of the African American patients in the study had normal vitamin

D levels. Patient reported indicators of heart failure such as Killip class at arrival and

Rose dyspnea scores did not differ by 25(OH)D levels.

The main finding of the study was that no statistically significant correlations

between 25(OH)D and log NT-proBNP levels were observed (rho = - 0.0025, p = 0.97).

Similarly, no significant associations between the log-transformed NT-proBNP levels

and 25(OH)D categories (6.9 ± 1.3 pg/ml in severely deficient vs. 6.1 ± 1.7 pg/ml in

replete group, P = 0.165) were found. In multivariable regression model after adjusting

for several covariates, 25(OH)D was not associated with NT-proBNP levels.

Additional analyses

Bivariate associations between NT-proBNP and patient characteristics were

obtained to determine the variables that needed adjustment in the multivariable model

(Table 4). These results are being presented as additional analyses that were

completed for this thesis project, but were not appropriate to include in the manuscript

(Chapter 2). This analysis was a precursor to the multivariable model that is presented

in Table 3 of Chapter 2

39

Table4. Association of NT-proBNP with patient characteristics at baseline

NT-proBNP levels

Characteristic Quartile 1 (5 - <488)

Quartile 2 (488 - <924.5)

Quartile 3 (924.5- <1882)

Quartile 4 (1882- 31230) P- Value

n= 58 n = 61 n= 59 n = 60

Demographics

Age(yr)a 54.2 ± 10.4 56.1 ± 10.8 57.2 ± 10.8 63.3 ± 11.3 < 0.001

Male, n (%) 44 (75.9) 51 (83.6) 42 (71.2) 39 (65) 0.122

Caucasian, n (%) 40 (69) 47 (77) 41 (69.5) 49 (81.7) 0.317

BMIb (kg/m2) 32.4 ± 6.2 29.9 ± 6.1 31.4 ± 7.3 28.8 ± 5.9 0.017

Smoked within 1 month, n (%) 20 (34.5) 27 (44.3) 21 (35.6) 20 (33.3) 0.586

Comorbidities

Atrial Fibrillation, n (%) 4 (6.9) 0(0) 3 (5.1) 9 (15) 0.006

Chronic Heart Failure, n (%) 0(0) 0(0) 6 (10.2) 5 (8.3) 0.003

Hypertension, n (%) 39 (67.2) 31 (50.8) 41 (69.5) 43 (71.7) 0.066

Diabetes, n (%) 19 (32.8) 9 (14.8) 16 (27.1) 20 (33.3) 0.077

Prior MIb, n (%) 11 (19 ) 12 (19.7) 9 (15.3 ) 17 (28.3) 0.34

Prior PCIb, n (%) 12 (20.7) 13 (21.3) 11 (18.6) 12 (20) 0.986

Prior CABGb, n (%) 5 (8.6) 6 (9.8) 7 (11.9) 9 (15) 0.71

Final MI diagnosis, n (%) 0.017

STEMIb 17 (29.3) 32 (52.5) 32 (54.2) 23 (38.3)

NSTEMIb 41 (70.7) 29 (47.5) 27 (45.8) 37 (61.7)

Arrival Killip Class, n (%) 0.928

I 53 (94.6) 55 (93.2) 53 (89.8) 55 (93.2)

II 2 (3.6) 4 (6.8) 3 (5.1) 3 (5.1)

III 1 (1.8) 0 (0) 2 (3.4) 1 (1.7)

IV 0 (0) 0 (0) 1 (1.7) 0 (0)

Rose dyspnea score 0.7 ± 0.8 0.7 ± 0.7 0.9 ± 0.9 1.2 ± 1.0 0.007

LV Systolic Function, n (%) < 0.001

Normal 47 (81) 45 (73.8) 35 (60.3) 27 (45)

Mild 9 (15.5) 9 (14.8) 14 (24.1) 11 (18.3)

Moderate 2 (3.4) 6 (9.8) 5 (8.6) 12 (20)

Severe 0 (0) 1 (1.6) 4 (6.9) 10 (16.7)

In hospital PCIb, n (%) 32 (55.2) 50 (82) 45 (76.3) 40 (66.7) 0.009

In hospital CABGb, n (%) 9 (15.5) 2 (3.3) 4 (6.8) 8 (13.3) 0.085

Laboratory values

Troponin I (ng/ml)a 0.6 ± 0.7 1.7 ± 1.6 2.0 ± 1.8 2.0 ± 2.2 < 0.001

e GFRb 89.4 ± 22.3 91.3 ± 17.7 85.8 ± 27.4 76.2 ± 31.3 0. 006

Calcium(mg/dl) 9.0 ± 0.6 9.0 ± 0.6 8.9 ± 0.5 8.6 ± 0.5 < 0.001

Intact PTHb 32.3 ± 24.8 40.6 ± 27.5 39.9 ± 29.2 54.1 ± 56.4 0. 016

Phosphate(mg/dl) 3.8 ± 0.8 3.4 ± 0.8 3.6 ± 0.7 3.3 ± 0.8 0.007

Total Cholesterol (mg/dl)a 158.7 ± 35.4 165.1 ± 31.0 154.3 ± 36.0 146.0 ± 33. 4 0.024

hsCRPb(mg/l) 1.7 ± 2.1 2.7 ± 2.7 4.3 ± 5.4 7.0 ± 7.0 < 0. 001

40

Medications

ACEI/ARBb, n (%) 18 (31) 14 (23) 14 (23.7) 23 (38.3) 0.209

Diuretics, n (%) 12 (20.7) 6 (9.8) 14 (23.7) 24 (40) 0.001

SF-12 PCSb 43.1 ± 12.8 45.9 ± 11.1 41.1 ± 13.3 39.2 ± 13.7 0. 034 aData shown as mean ± standard deviation bBMI, body mass index; MI, myocardial infarction; PCI, percutaneous coronary intervention; CABG, coronary artery bypass graft; STEMI, ST-segment elevation myocardial infarction; eGFR, estimated glomerular filtration rate; PTH, parathyroid hormone; hs CRP, high sensitivity C-reactive protein; ACEI, angiotensin converting enzyme inhibitors; ARB, angiotensin receptor blockers;SF-12 PCS, short form -12 physical component score

Implications of the research project

The results of this study have several important implications. This is the first

study in the US to report the prevalence of nutritional vitamin D deficiency in a

multicenter cohort of AMI patients. Vitamin D deficiency was highly prevalent in this

cohort of AMI patients with only 4.2% of participants in the study having normal

25(OH)D levels. Given that 25(OH)D deficiency has been associated with incident MI 3

and CV events14 in prior observational studies, it would be important for future

investigators to examine whether rectifying vitamin D levels in post-AMI patients is

associated with an improvement in subsequent outcomes..

Secondly, this study is unique as it identifies a potential mechanism by which

25(OH)D deficiency mediates worse prognosis after AMI, which is currently an area of

intense research. If the findings of this study could be replicated in a larger cohort of

AMI patients or post-AMI heart failure patients, especially during the recovery period, it

would further strengthen our conclusion that nutritional vitamin D deficiency may not be

mediating prognosis after AMI through effects on NT-proBNP pathways. This could lead

to studies that would explore other potential mechanisms such as inflammation and

41

vascular calcification by which 25(OH)D deficiency could mediate adverse outcomes in

AMI patients.

Future Directions

Future studies with longer follow-up of AMI patients could help determine

whether 25(OH)D deficiency further risk stratifies long-term clinical outcomes post-MI

after adjusting for currently used prognostic schemes such as Thrombolysis in

Myocardial Infarction (TIMI) or Global registry of Acute Coronary Events (GRACE) risk

scores.17 Also given that nutritional vitamin D is readily available, inexpensive, and has

a good safety profile, future randomized controlled trials should investigate whether

treating 25(OH)D deficiency might improve hard outcomes, such as mortality and

health status in AMI patients, regardless of the mechanism of its known association with

post-MI risk.

Currently I am a 3rd year nephrology fellow. With the experience that I have

gained through this thesis project, I plan to pursue a career development award on a

related subject. One of the areas of my interest is the role of 25-hydroxy vitamin D

deficiency in the progression of chronic kidney disease and its impact on cardiac

dysfunction in predialysis patients. Chronic kidney disease (CKD) affects nearly 20

million individuals in the US 18;19 and is associated with significant cardiovascular

morbidity and all-cause mortality.20-22 In addition an estimated 300,000 patients have

end stage renal disease (ESRD) and are currently undergoing dialysis in the US.

Although it has been known for several decades that hypertension, diabetes and

albuminuria are the risk factors for progression to ESRD, gaps exist in our knowledge

42

about the role of novel risk factors such as vitamin D, inflammation and uric acid that

could effect progression of CKD to ESRD.23 Vitamin D has anti-proliferative and

immunomodulatory properties, and functions as an endocrine regulator of the renin-

angiotensin system.24;25 Animal and cell culture studies have shown that vitamin D

suppresses the transcription of renin, decreased circulating angiotensin II levels,

preventing podocyte loss26 and glomerulosclerosis,27 and thus decreasing albuminuria

which is an established predictor of CKD progression. As such, low levels of vitamin D

are a potential novel risk factor for progression of CKD.28 In addition, nutritional vitamin

D deficiency has been associated with incident MI 3 and heart failure 4;5 both of which

are highly prevalent in CKD patients. Thus, understanding the renal and cardio-

protective role of vitamin D in cohort studies would not only provide mechanistic insights

but also lay the foundation for future randomized controlled trials with vitamin D, a safe

and inexpensive treatment.

43

References

1. Lee JH, O'Keefe JH, Bell D, Hensrud DD, Holick MF: Vitamin D deficiency an important, common, and easily treatable cardiovascular risk factor? J Am Coll Cardiol 52:1949-1956, 2008




5. Zittermann A, Schleithoff SS, Koerfer R: Vitamin D insufficiency in congestive heart failure: why and what to do about it? Heart Fail Rev 11:25-33, 2006







44




15. Gonzalez EA, Sachdeva A, Oliver DA, Martin KJ: Vitamin D insufficiency and deficiency in chronic kidney disease. A single center observational study. Am J Nephrol 24:503-510, 2004


17. de Araujo GP, Ferreira J, Aguiar C, Seabra-Gomes R: TIMI, PURSUIT, and GRACE risk scores: sustained prognostic value and interaction with revascularization in NSTE-ACS. Eur Heart J 26:865-872, 2005

18. Coresh J, Astor BC, Greene T, Eknoyan G, Levey AS: Prevalence of chronic kidney disease and decreased kidney function in the adult US population: Third National Health and Nutrition Examination Survey. Am J Kidney Dis 41:1-12, 2003

19. Coresh J, Selvin E, Stevens LA, Manzi J, Kusek JW, Eggers P, Van LF, Levey AS: Prevalence of chronic kidney disease in the United States. JAMA 298:2038-2047, 2007

20. Culleton BF, Larson MG, Wilson PW, Evans JC, Parfrey PS, Levy D: Cardiovascular disease and mortality in a community-based cohort with mild renal insufficiency. Kidney Int 56:2214-2219, 1999

21. Go AS, Chertow GM, Fan D, McCulloch CE, Hsu CY: Chronic kidney disease and the risks of death, cardiovascular events, and hospitalization. N Engl J Med 351:1296-1305, 2004

22. Parfrey PS, Foley RN: The clinical epidemiology of cardiac disease in chronic renal failure. J Am Soc Nephrol 10:1606-1615, 1999

23. Hsu CY, Iribarren C, McCulloch CE, Darbinian J, Go AS: Risk factors for end-stage renal disease: 25-year follow-up. Arch Intern Med 169:342-350, 2009

24. Li YC: Vitamin D regulation of the renin-angiotensin system. J Cell Biochem 88:327-331, 2003

45

25. Li YC, Qiao G, Uskokovic M, Xiang W, Zheng W, Kong J: Vitamin D: a negative endocrine regulator of the renin-angiotensin system and blood pressure. J Steroid Biochem Mol Biol 89-90:387-392, 2004

26. Kuhlmann A, Haas CS, Gross ML, Reulbach U, Holzinger M, Schwarz U, Ritz E, Amann K: 1,25-Dihydroxyvitamin D3 decreases podocyte loss and podocyte hypertrophy in the subtotally nephrectomized rat. Am J Physiol Renal Physiol 286:F526-F533, 2004

27. Schwarz U, Amann K, Orth SR, Simonaviciene A, Wessels S, Ritz E: Effect of 1,25 (OH)2 vitamin D3 on glomerulosclerosis in subtotally nephrectomized rats. Kidney Int 53:1696-1705, 1998

28. de B, I, Ioannou GN, Kestenbaum B, Brunzell JD, Weiss NS: 25-Hydroxyvitamin D levels and albuminuria in the Third National Health and Nutrition Examination Survey (NHANES III). Am J Kidney Dis 50:69-77, 2007

46

CURRICULUM VITAE

Rajyalakshmi Gadi

CURRENT APPOINTMENT

09/2008- 08/2011: Nephrology fellow at University of Kansas Medical centre

University of Kansas Medical Center

3901 Rainbow Boulevard, MSN 1039

Kansas City, Kansas 66160

09/2008-08/2009: Outcomes Research fellow at Mid-America Heart Institute,

Saint Luke’s Hospital, MO.

EDUCATION

11/1996 - 06/2002: Gandhi Medical College - (MBBS) Bachelor of

Medicine and Surgery (Honors).

06/2004- 06/2007: Internal Medicine residency

University of Arkansas for Medical sciences

Little Rock, AR.

08/2007-08/2010: Master of Science in Health Sciences Research

Wake Forest University

Winston –Salem, NC, 27157.

47

WORK EXPERIENCE

08/2007 – 06/2008: Teaching attending and preceptor of Internal Medicine in

Ambulatory Medicine Clinic.

Wake Forest University Baptist Medical Center

Winston-Salem, NC.

06/2004 – 06/2007: Internship and Residency at University of Arkansas

for Medical Sciences (UAMS)

Department of Internal Medicine

Little Rock, AR

08/2002 -- 05/2003: House officer at Care cardiac hospital,

Division of Cardiac care and surgical services,

Hyderabad, AP, India.

06/2001-- 06/2002: Internship and House Officer,

Gandhi Hospital,

Hyderabad, AP, India.

BOARD EXAMINATIONS

03/2003: Passed USMLE Step 1 (99 Percentile) 10/2003: Passed USMLE Step 2 (95 Percentile) 11/2003: Passed CSA (clinical skills examination)

48

04/2004: Passed USMLE Step 3 (85 Percentile) 01/2004: ECFMG certified 08/2007: Certified in ABIM examination (American Board of Internal Medicine)

MEDICAL LICENCE

Kansas State Medical Board of Healing Arts.

RESEARCH INTERESTS

Cardiovascular outcomes in chronic kidney disease patients. Role of vitamin D deficiency in acute myocardial infarction patients and outcomes. Hypertension control in chronic kidney disease patients and outcomes.

PUBLICATIONS

(1) Song EY, McClellan WM, McClellan A, Gadi R, Hadley AC, Krisher J, Clay M, Freedman BI. Effect of Community Characteristics on Familial Clustering of End-Stage Renal Disease. Am J Nephrol 2009 September 30; 30(6):499-504.

(2) Rajyalakshmi Gadi, John H. Lee, James H. O'Keefe, Paul Chan, Fengming Tang,

John A. Spertus. Association of Vitamin D Deficiency with NT-proBNP Levels in AMI Patients. J Am Soc Nephrol Abstracts Issue 20:2009. Accessed online at http://www.asn-online.org/education_and_meetings/renal_week/2009/digital-abstract.aspx.

(3) Ahmad B. Malik, Rajyalakhshmi Gadi, Sundraraman Swaminathan. Etiology, Risk

Factors and Outcomes of Acute interstitial Nephritis: Epidemiological Update. J Am Soc Nephrol Abstracts Issue 10:2007. Accesses online at http://www.abstracts2view.com/asn07/sessionindex.php.

SOCIETY MEMBERSHIPS

American College of Physicians

American Society of Nephrology

association of 25-hydroxy vitamin d deficiency …...the active form of vitamin d, its serum levels...

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