ORIGINAL ARTICLE JJBMR
Vitamin D Receptor (VDR) Gene Polymorphism IsAssociated With Left Ventricular (LV) Mass andPredicts Left Ventricular Hypertrophy (LVH)Progression in End-Stage Renal Disease (ESRD) PatientsAlessandra Testa,1 Francesca Mallamaci,1 Francesco A Benedetto,2 Anna Pisano,1 Giovanni Tripepi,1
Lorenzo Malatino ,4 Ravi Thadhani ,3 and Carmine Zoccali1
1CNR-IBIM, Research Unit of Clinical Epidemiology and Physiopathology of Renal Disease and Hypertension, Reggio Calabria, Italy2Cardiology Unit, Morelli Hospital, Reggio Calabria, Italy3Division of Internal Medicine, University of Catania, c/o Ospedale Cannizzaro, Catanua, Italy4Renal Unit and the Center for D-Receptor Activation Research, Massachusetts General Hospital, Harvard Medical School, Boston,MA, USA
ABSTRACTLeft ventricular hypertrophy (LVH) is a strong cardiovascular risk marker in end-stage renal disease (ESRD) patients. Vitamin D deficiency
and/or disturbed vitamin D signaling has been implicated in LVH in experimental models. Because the BsmI vitamin D receptor VDR gene
polymorphism may alter VDR function, we performed a cross-sectional and longitudinal study in a cohort of 182 dialysis patients to
investigate (1) the relationship between BsmI VDR gene polymorphism and left ventricular mass index (LVMI) measured by
echocardiography and (2) the predictive power of this polymorphism for progression in LVH over a 18� 2 months of follow-up. As
a reference group, we used 175 healthy subjects matched to the study population as for age and sex. The distribution of BsmI genotypes
did not significantly deviate from Hardy-Weinberg equilibrium either in patients or in the control group of healthy subjects. The
frequency of the B allele of BsmI polymorphism (40.4%) in dialysis patients was similar to that of healthy control subjects (38.6%), and the
number of B alleles was directly related to LVMI (r¼ 0.20, P¼ .007). This relationship remained robust (b¼ 0.19, P¼ .006) in multivariate
analysis adjusting for traditional and nontraditional risk factors and antihypertensive and calcitriol treatment. In the longitudinal study,
LVMI rose from 60.1� 17.9 to 64.2� 19.3 g/m2.7 (P< .001), and again, the number of B alleles was associated with LVMI changes both in
crude and in fully adjusted analyses. These cross-sectional and longitudinal observations coherently support the hypothesis that altered
vitamin D signaling is implicated in LVH in ESRD patients. � 2010 American Society for Bone and Mineral Research.
KEY WORDS: VITAMIN D; POLYMORPHISM; ASSOCIATION; LEFT VENTRICULAR HYPERTROPHY
Introduction
Left ventricular hypertrophy (LVH) is recognized as one of the
strongest risk factors for all-cause and cardiovascular
mortality in patients with end-stage renal disease (ESRD).(1,2)
The pathogenesis of LVH in ESRD is multifactorial, and arterial
hypertension, hyperparathyroidism, severe anemia, hypoalbu-
minemia, chronic volume overload,(3) sympathetic overactivity,(4)
and accumulation of the endogenous inhibitor of nitric oxide
(NO) synthase asymmetric dimethylarginine (ADMA)(5) have
been implicated as causative mechanisms responsible for LVH in
these patients. However, collectively, these risk factors explain
only in part the variability in left ventricular (LV) mass in the
dialysis population.
Received in original form November 27, 2008; revised form February 4, 2009; acce
Address correspondence to: Professor Carmine Zoccali, CNR-IBIM, CNR e Nefrologia,
89124 Reggio Calabria, Italy. E-mail: [email protected]
Journal of Bone and Mineral Research, Vol. 25, No. 2, February 2010, pp 313–319
DOI: 10.1359/jbmr.090717
� 2010 American Society for Bone and Mineral Research
Vitamin D insufficiency or deficiency is pervasive in ESRD
patients and may have substantial health implications in this
population.(6) The myocardium is an important target of vitamin
D. Vitamin D receptor (VDR) knockout mice show myocardial
renin overexpression and marked cardiomyocyte hypertrophy.(7)
In vitro 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] attenuates
cardiomyocyte proliferation(8) and hypertrophy,(9) and treatment
with paricalcitol attenuates the development of LVH and LV
dysfunction in Dahl rats,(10) a reliable animal model of LVH and
vitamin D deficiency. Retrospective analyses in hemodialysis
patients and small clinical trials show that treatment with a
vitamin D analogue may regress LVH in these patients,(11–13)
providing further rationale for a full-fledged randomized clinical
trial in ESRD. Until now, studies in the VDR knockout mice
pted July 6, 2009. Published online July 13, 2009.
Ospedali Riuniti, c/o EUROLINE di Ascrizzi Vincenzo, Via Vallone Petrara 55-57,
313
represent the most convincing evidence of the fundamental role
of vitamin D in the regulation of LV mass.
While the clinical trial remains the ultimate test for establishing
whether or not vitamin D is implicated in LVH in ESRD, examining
the association between VDR gene polymorphisms and LV mass
represents an additional relevant step for unraveling the role of
vitamin D in cardiac disease in these patients. An important
advantage of genetic association studies is that the transmission
of genes occurs at random (Mendelian randomization), thus
mimicking the design of classical intervention trials.(14) Three
common polymorphisms (BsmI, ApaI, and TaqI) at the 3’ end of
the VDR have been intensively investigated, and one of these
(BsmI) already has been associated with survival in the dialysis
population,(15) suggesting that this polymorphism may be
implicated in the high risk of this population. However, the
association between the BsmI polymorphism and LVH has never
been tested. Therefore, we performed an association study
investigating the relationship between this polymorphism and
LV mass in a cohort of ESRD patients. To minimize the risk that
the observed association are merely due to chance, we
performed a study including both a cross-sectional analysis of
the association between BsmI VDR gene polymorphism and LV
mass index (LVMI) and a longitudinal observation aimed at
establishing whether or not this polymorphism predicts LVH
progression in ESRD. For the first time, we demonstrate that the
BsmI VDR gene polymorphism is independently related to LVH
and to LVH progression in ESRD patients.
Methods
The study protocol was in conformity with the local ethical
guidelines of our institution and informed consent was obtained
from each participant.
Patients
One-hundred and eighty-two hemodialysis patients (104 males
and 78 females, all Caucasians) who had been on regular dialysis
treatment (RDT) for at least 6 months and who were free of overt
infections (i.e., fever, infected vascular access, or peritonitis or exit-
site infection) were recruited for the study. Patients were being
treated thrice weekly with standard bicarbonate dialysis (Na
138mmol/L, HCO3 5mmol/L, K 1.5mmol/L, Ca 1.25mmol/L, Mg
0.75mmol/L) with either Cuprophan or semisynthetic membranes.
Control subjects
As a control group, we enrolled a series of 175 age- and sex-
matched healthy blood donors from the same geographic area
of patients.
Genotyping of the BsmI VDR gene polymorphism
The participants were genotyped for the G ! A change
identified as BsmI polymorphism (bB alleles) in intron 8 of the
VDR gene on chromosome 12. This polymorphism, described
under identification number rs1544410, was studied by high-
throughput TaqMan allelic discrimination assay. Genomic DNA
was extracted from peripheral blood leukocytes by the salting-
314 Journal of Bone and Mineral Research
out technique.(16) The assay mix (including unlabeled PCR
primers FAM and VIC dye-labeled TaqMan MGB probes) of the
Assays-on-Demand was designed and provided by Applied
Biosystems (Applied Biosystems, Inc., Foster City, CA, USA). The
reaction system contained 1 to 5 ng of genomic DNA, 12.5mL of
TaqMan Universal PCR Master Mix, 2�No AmpErase UNG, and
1.25mL 40�Assay Mix and was adjusted with H2O for a total
volume of 25mL. The genotyping was performed on an ABI Prism
7300, and a random 5% of samples were repeated indepen-
dently to confirm genotyping results. The genotype results for
these samples were completely consistent.
Laboratory measurements
A fasting blood sampling was performed during the midweek
nondialysis day. Serum cholesterol, albumin, phosphate, para-
thormone (PTH), and hemoglobin measurements were made
using standard methods in a routine clinical laboratory. Serum C-
reactive protein (CRP), plasma total homocysteine, and asym-
metric dimethylarginine (ADMA) were measured as previously
reported.(17) The plasma concentration of norepinephrine was
measured by a commercially available radioimmunoassay (RIA)
kit (Amicyl-test TM, Immunological Laboratories, Hamburg,
Germany). The intraassay coefficient of variation was 7% to 15%.
Echocardiography and blood pressure
These studies were performed on a nondialysis day within 2
hours of blood sampling. All echocardiographic measurements
were carried out according to the recommendations of the
American Society of Echocardiography by an observer unaware
of biochemical results. Left ventricular mass (LVM) was calculated
according to the Devereux formula and indexed to height2.7
(LVMI).(18) LVH was defined by an LVMI of over 47 g/m2.7 in
women or over 50 g/m2.7 in men. The height-based indexing of
LVM was specifically chosen to minimize any potential distortion
attributable to extracellular volume expansion (surface area
indexing being weight sensitive).(2)
Blood pressure (BP) was measured before dialysis and during
all dialysis sessions of the month (i.e., 12 measurements)
preceding the baseline echocardiographic study. Average values
were considered for statistical analysis. Average monthly BP is
associated with LVmass equally well as 24 hour ambulatory BP in
dialysis patients.(19)
Patients who repeated the echocardiographic study
Thirty-four of the 182 patients who entered into this study did
not repeat the second echocardiography because of death.
Therefore, 148 patients were left for the longitudinal part of this
study aimed at examining the relationship between the BsmI
polymorphism and changes in LVMI. These patients did not
materially differ from the original study population (data not
shown).
Statistical analysis
Power analysis
We assumed additivity of the effects of the B allele on LV mass
index. Such an assumption is preferred when allele dominance
TESTA ET AL.
cannot be assumed.(20) Nonrandomized parallel studies indicate
that the lack of 1,25(OH)2D3 treatment may be responsible for a
15% increase in LV mass in ESRD patients.(13) We hypothesised
that homozygotes for the protective allele of the BmsI
polymorphism gene (bb) have a LV mass that is 15% lower
than homozygotes for the risk allele (BB) and that LV mass in
heterozygotes (Bb) takes an intermediate value (7.5% higher
than in bb homozygotes). We estimated that LVMI in Bb patients
would be 61 g/m2.7 (� SD 15), that is the average value of LVMI
in hemodialysis patients in our center. On the basis of these data,
we calculated that the average value of LVMI would be 56 g/m2.7
in bb (7.5% lower than in Bb) and 66 g/m2.7 in BB (7.5% higher
than in Bb) genotypes. Power analysis based on these data
indicated that a sample size of 180 dialysis patients had an 80%
probability to detect as statistically significant (a error¼ 0.05) a
5 g/m2.7 average increase in LVMI for each unitary increase in the
number of B alleles.
Statistical tests
The relationship between the number of B alleles of BsmI VDR
polymorphism and LV mass at baseline and with LV mass
changes during the follow-up was investigated by univariate and
multiple linear regression analyses (Models 1 to 4 in Tables 2
and 3). As potential confounders we considered Framingham risk
factors (e.g., age, sex, smoking, diabetes, systolic pressure and
antihypertensive treatment, cholesterol, and previous cardio-
Table 1. Main Demographic, Clinical, Biochemical, and Echocardiogr
Vita
bb (n¼ 61
Age (years) 60� 13
Duration of RDT (months) 47 (21–12
Male sex, n (%) 34 (56%)
Smoking, n (%) 22 (36%)
Diabetes, n (%) 10 (16%)
Patients with previous cardiovascular events, n (%) 33 (54%)
On antihypertensive treatment, n (%) 23 (38%)
On EPO treatment, n (%) 29 (47%)
On calcitriol treatment, n (%) 28 (46%)
Systolic pressure (mmHg) 137� 25
Diastolic pressure (mmHg) 74� 14
Heart rate (b/min) 80� 10
Cholesterol (mmol/L) 5.26� 1.6
Haemoglobin (g/L) 107� 22
Albumin (g/L) 41� 6
Calcium phosphate (mmol2/L2) 4.5� 1.3
Phosphate (mmol/L) 1.96� 0.5
PTH (pg/mL) 161 (64–34
Norepinephrine (nmol/L) 3.3 (2.0–6.
CRP (mg/L) 9.5 (3.4–20
ADMA (mmol/L) 23.2 (17.4–3
Homocysteine (mmol/L) 2.42 (1.55–3
Kt/V 1.23� 0.2
LVMI (g/m2.7) 56.4� 17.
Data are expressed as mean� SD, median and interquartile range, or as pe
VITAMIN D POLYMORPHISM AND LVH IN ESRD
vascular events), risk factors peculiar to ESRD (e.g., duration of
dialysis treatment, haemoglobin, albumin, phosphate, PTH, and
fractional urea clearance [Kt/V]), and emerging risk factors (e.g.,
CRP, homocysteine, ADMA, and norepinephrine). To control for
the potential confounding effect of current calcitriol therapy, we
also included this factor in the multiple regression analysis. Data
are expressed as standardized regression coefficient b and P
value. The deviation of Hardy-Weinberg equilibrium was
investigated by calculating the chi-square test between
observed and expected frequencies. Since we did not test
other VDR single-nucleotide polymorphisms (SNPs), no adjust-
ment for multiple testing was done.
Data are expressed as mean� SD (normally distributed data),
median and interquartile range (nonnormally distributed data),
or percent frequency, as appropriate, and by trend analysis.
Positively skewed distributed data (e.g., CRP, homocysteine, and
ADMA) were log transformed (lg10) before data analysis.
All calculations were done using two standard statistical
packages (SPSS for Windows, Version 9.0.1,� SPSS Inc., Chicago,
IL, USA, and NCSS PASS (2004), Number Cruncher Statistical
Systems, Kaisville, UT, USA).
Results
In the entire study population (see Table 1), the median duration
of RDT was 45 months (interquartile range 20 to 110 months).
The average urea Kt/V in these patients was 1.22� 0.28.
aphic Data in Hemodialysis Patients
min D receptor (BsmI polymorphism)
) Bb (n¼ 95) BB (n¼ 26) P for trend
57� 16 60� 14 .63
1) 43 (20–92) 46 (14–121) .72
54 (57%) 16 (62%) .65
38 (40%) 8 (31%) .83
13 (14%) 3 (11%) .52
44 (46%) 14 (54%) .74
38 (40%) 8 (31%) .69
59 (62%) 12 (46%) .62
55 (58%) 12 (46%) .61
143� 24 147� 22 .05
78� 13 78� 12 .12
80� 10 76� 9 .27
5 5.38� 1.37 5.75� 1.47 .19
106� 17 107� 20 .96
42� 4 41� 6 .47
4.6� 1.2 4.4� 1.8 .95
0 2.02� 0.44 1.99� 0.49 .73
5) 171 (70–405) 145 (53–436) .51
4) 3.1 (1.6–5.3) 3.7 (1.9–5.6) .60
.1) 8.1 (3.4–15.0) 8.0 (3.4–21.1) .66
5.2) 28.4 (21.0–44.5) 25.2 (19.3–36.7) .34
.88) 2.88 (1.68–4.08) 3.33 (1.38–4.15) .23
9 1.20� 0.25 1.27� 0.32 .74
7 62.7� 19.1 66.9� 15.4 .007
rcent frequency, as appropriate.
Journal of Bone and Mineral Research 315
Fig. 1. Distribution of left ventricular mass index (LVMI) in the study
cohort.
Twenty-six patients were diabetics, and 68 were habitual
smokers (22� 16 cigarettes/day). Sixty-nine patients were on
antihypertensive drugs [48 on monotherapy with angiotensin-
converting enzyme (ACE) inhibitors, AT-1 antagonists, calcium
channel blockers, and alpha and beta blockers and the remaining
21 on double or triple therapy with various combinations of
these drugs]. One hundred patients were on treatment with
erythropoietin and 95 with calcitriol. At baseline, LVMI was on
average 61� 18 g/m2.7. The distribution of LVMI in the study
population is presented graphically in Fig. 1.
The distribution of BsmI genotypes did not significantly
deviate from Hardy-Weinberg equilibrium either in patients (bb
genotype 33.5%, Bb genotype 52.2%, and BB genotype 14.3%;
x2¼ 1.29, P¼NS) or in controls (bb genotype 37.8%, Bb
genotype 47.4%, and BB genotype 14.9%; x2¼ 0.01, P¼NS).
The frequency of the B allele (38.6%) in healthy subjects was
almost identical to that of dialysis patients (40.4%). Data analysis
according to BsmI genotypes (see Table 1) revealed that there
was a dose-dependent relationship between the number of BsmI
B alleles and systolic arterial pressure (bb genotype
137� 25mmHg, Bb genotype 143� 24mmHg, BB genotype
147� 22mmHg; P for trend¼ 0.048) (see Table 1). No significant
difference among the three groups was observed for the
remaining variables listed in Table 1.
Fig. 2. Relationship between BsmI polymorphism and LVMI and with LVMI ch
316 Journal of Bone and Mineral Research
BsmI genotypes and baseline LVMI: linear regressionanalyses
On univariate analysis, the number of B alleles was directly and
significantly related to LVMI (r¼ 0.20, P¼ .007) (Fig. 2, left panel).
The independent association between the BsmI polymorphism
and LVMI was tested in multiple linear regression models of
increasing complexity (Table 2). Data adjustment for age, sex,
and duration of RDT did not modify the strength of the B allele–
LVMI link (b¼ 0.21, P¼ .004) (Model 1), and this also was true
when we added into the model other Framingham risk factors
(e.g., smoking, systolic pressure, diabetes, cholesterol, and
previous cardiovascular events), as well as antihypertensive
and calcitriol treatment (b¼ 0.20, P¼ .004) (Model 2). Further
data adjustment for factors peculiar to ESRD (e.g., hemoglobin,
albumin, phosphate, PTH, and Kt/V) (Model 3) and nontraditional
risk factors (e.g., CRP, homocysteine, norepinephrine, and ADMA)
(Model 4) did not change the strength of the association. In the
fully adjusted multiple regression model (Model 4), besides the
BsmI polymorphism, serum albumin (b¼�0.24, P¼ .003),
systolic arterial pressure (b¼ 0.24, P¼ .002), and plasma ADMA
(b¼ 0.17, P¼ .02) also were significantly and independently
related to LVMI.
BsmI genotypes and changes in LVMI
Since baseline LVMI was inversely associated with LVMI changes
(r¼�0.18, P¼ .03), indicating regression to the mean, LVMI
changes therefore were adjusted for the corresponding baseline
values before analyses. As shown in Table 3, the number of B
alleles was associated with LVMI changes both in crude (see
Fig. 2, right panel) and in adjusted analyses, the correlation
coefficient of this relationship remaining virtually unchanged
after full data adjustment across models of increasing complexity
(b¼ 0.19, P¼ .03).
Discussion
This study shows that in chronic dialysis patients the frequency
of the B allele of the BsmI VDR gene polymorphism is
independently related to LVH and associated with higher
progression rate of LVH. Overall, these genetic associations are
compatible with the hypothesis that altered vitamin D
anges. Data are mean� SD.
TESTA ET AL.
Table 2. Multiple Linear Regression Models for Baseline LVMI
Dependent variable: LVMI
Unadjusted r (P) Model 1 b (P) Model 2 b (P) Model 3 b (P) Model 4 b (P)
BsmI polymorphism 0.20 (.007) 0.21 (.004) 0.20 (.004) 0.19 (.004) 0.18 (.006)
Age 0.27 (<.001) 0.21 (.06) 0.13 (.10) 0.12 (.12)
Sex 0.07 (.37) 0.07 (.40) 0.13 (.10) 0.09 (.28)
Duration of RDT 0.04 (.58) 0.08 (.28) 0.07 (.35) 0.03 (.67)
Smoking 0.01 (.85) 0.04 (.60) 0.07 (.35)
Diabetes 0.05 (.48) 0.10 (.16) 0.09 (.21)
Cholesterol �0.12 (.09) 0.02 (.80) �0.04 (.58)
Systolic pressure 0.23 (.003) 0.22 (.003) 0.24 (.002)
Previous cardiovascular events 0.18 (.02) 0.16 (.04) 0.14 (.07)
Antihypertensive treatment 0.12 (.12) 0.08 (.31) 0.05 (.55)
On treatment with calcitriol �0.03 (.62) �0.01 (.83) �0.05 (.47)
Hemoglobin �0.12 (.09) �0.11 (.12)
Albumin �0.28 (.001) �0.24 (.003)
Phosphate 0.10 (.15) 0.08 (.22)
PTH 0.12 (.08) 0.07 (.31)
Kt/V �0.001 (.98) �0.02 (.75)
CRP �0.008 (.91)
Homocysteine �0.008 (.91)
ADMA 0.17 (.02)
Norepinephrine 0.06 (.43)
Data are expressed as beta regression coefficients (b) and P values.
metabolism exerts an important influence on the control of LV
mass in this population.
LVH is considered to be a clinical indicator integrating the
long-term exposure not only to pressure overload but also to
Table 3. Multiple Linear Regression Models for Changes in LVMI
Dependent variable: chang
Unadjusted b (P) Model 1 b
BsmI polymorphism 0.19 (.02) 0.19 (.0
Age 0.01 (.8
Sex 0.003 (.9
Duration of RDT �0.03 (.6
Smoking
Diabetes
Cholesterol
Systolic pressure
Previous cardiovascular events
Antihypertensive treatment
On treatment with calcitriol
Hemoglobin
Albumin
Phosphate
PTH
Kt/V
CRP
Homocysteine
ADMA
Norepinephrine
Data are expressed as beta regression coefficients (b) and P values.
VITAMIN D POLYMORPHISM AND LVH IN ESRD
several hemodynamic and nonhemodynamic factors. Regression
of LVH is associated with a 59% lower risk of subsequent adverse
events compared with the persistence or new development of
LVH.(21) LVH is exceedingly frequent in dialysis patients, with a
es in LVMI (g/m2.7/month) adjusted for LVMI at baseline
(P) Model 2 b (P) Model 3 b (P) Model 4 b (P)
2) 0.18 (.04) 0.19 (.03) 0.19 (.03)
9) 0.04 (.68) 0.05 (.62) 0.07 (.51)
6) �0.05 (.62) �0.10 (.32) �0.12 (.30)
9) �0.05 (.58) �0.04 (.69) �0.06 (.57)
0.06 (.53) 0.03 (.76) 0.04 (.70)
�0.11 (.19) �0.12 (.17) �0.10 (.31)
0.08 (.33) �0.12 (.21) �0.15 (.15)
0.03 (.74) 0.04 (.70) 0.03 (.74)
�0.02 (.82) 0.004 (.97) �0.02 (.80)
0.007 (.94) 0.04 (.68) 0.03 (.74)
0.15 (.07) 0.16 (.07) 0.16 (.07)
0.12 (.20) 0.11 (.25)
0.10 (.34) 0.08 (.47)
�0.08 (.35) �0.07 (.48)
0.17 (.05) 0.12 (.19)
�0.06 (.57) �0.06 (.56)
�0.003 (.97)
0.13 (.19)
0.02 (.85)
0.07 (.49)
Journal of Bone and Mineral Research 317
prevalence ranging from 60% to 78%. In line with studies in other
conditions, cohort studies in dialysis patients solidly link LVH
progression and mortality(22) and improved survival when LVH
regresses.(23) Even though several causative factors are impli-
cated in high LV mass in the dialysis population, regression of
LVH in this population is difficult to achieve, and even a most
intensive treatment strategy including long nocturnal dialysis
failed to normalize LV mass in several patients.(24)
Even though experimental evidence linking vitamin D
deficiency and alterations in LV mass and function appears
coherent and convincing,(7–9) the issue received only scant
attention in dialysis patients. In an uncontrolled series of 12
dialysis patients,(11) treatment with 1a-hydroxycholecalciferol for
6 weeks elicited an increase in LV contractility, as measured by
fractional fiber shortening and mean velocity of fiber shortening.
In a nonrandomized parallel study, intravenous calcitriol caused
a marked reduction in LVH, whereas LVH did not regress in
untreated patients. Apart from these small nonrandomized
studies, until now, there has been no study exploring the link
between the vitamin D system and LV mass in this population.
The response to vitamin D compounds critically depends on
the VDR functioning. In humans, three common polymorphisms
(BsmI, ApaI, and TaqI) at the 3’ end of the VDR have been
identified, and these polymorphisms have been linked to a
variety of diseases, from low bone mass density (BMD)(25) to type
1 diabetes and cancer.(26) In ESRD, the B allele of the BsmI
polymorphism was associated with a low BMD in patients
younger than 65 year of age(26) and with inflammation, anemia,
and resistance to erythropietin.(27) Of note, patients homozygous
for the B allele display a substantially shorter survival than
heterozygotes or homozygotes for the b allele.(15) Given the
exceedingly high rate of cardiovascular complications in ESRD, it
is plausible that such an association underlies disturbed control
of a vitamin D–dependent mechanism affecting the cardiovas-
cular system. To date, there have been no studies investigating
the BsmI polymorphism as related to cardiovascular disease
neither in the general population nor in the dialysis population.
The consequences of vitamin D deficiency and vitamin D
resistance in ESRD depend on a large number of factors.
Therefore, confounding is a real possibility in observational
studies based on the measurement of vitamin D plasma levels or
on vitamin D treatment status (treated versus untreated).
Mendelian randomization(14) is an interesting option to explore
the nature of the LV mass– vitamin D association. Unlike classic
epidemiology studies, SNP association studies are uncon-
founded by behavioral and environmental factors because
these factors usually do not alter genotype,(14) thus representing
a useful approach in studies focusing on the ESRD population.(15)
As predicated by Mendialian randomization theory, with the
exception of blood pressure (see below), risk factors were evenly
distributed among patients categorized on the basis of the B
allele. We hypothesised that the B allele of the BmsI
polymorphism may serve as a marker of altered vitamin D
signaling in ESRD patients and assumed that this alteration in BB
homozygotes produces an increase in LV mass quantitatively
similar to the difference in LV mass between vitamin D–treated
and untreated patients.(13) At baseline, in our cohort we found an
association between the B allele and LVMI. This association was
318 Journal of Bone and Mineral Research
almost unmodified after extensive statistical adjustment for
other risk factors, including calcitriol treatment, serum phos-
phate, and PTH. The independent association of the B allele with
systolic blood pressure is of interest in that hypertension is a
phenotypic characteristic of the VDR knockout model and
because a link between this polymorphism and BP has been
noted very recently in another study in ESRD.(28) This
phenomenon suggests that altered signaling may also con-
tribute to raise BP in ESRD patients. Both 25-hydroxycholecalci-
ferol and 1,25(OH)2D3 levels are strongly associated with arterial
distensibility, pulse wave velocity, and endothelial function, as
measured by the forearm blood flow response to hand warming
in ESRD patients,(29) indicating a relevant impact of vitamin D
status on the control of large and small arterial vessels. Beyond
baseline associations, we also found that the B allele predicts
LVH progression. This relationship is remarkable in that it also
persisted virtually unchanged in a multivariate model consider-
ing the regression-to-the-mean phenomenon, which is an
important confounder in longitudinal studies of LV mass in
ESRD patients.(22) Over half the patients in this study were on
long-term treatment with calcitriol, but apparently this treatment
had no effect on LV mass index nor modified the association
between the B allele and this index. Because confounding by
indication cannot be eliminated safely by statistical adjustment,
we believe that no conclusion can be drawn on the basis of our
data on the effect of treatment with activated vitamin D
compounds on LVH in ESRD. An ongoing double-blind, placebo-
controlled, randomized clinical trial testing the effect of an active
form of vitamin D (paracalcitol) on LVMI in ESRD patients
(http://clinicaltrials.gov/ct2/show/NCT00616902?term¼PRIMO&
rank ¼2) will provide a definitive answer to the question of
whether vitamin D administration can reverse LVH in ESRD
patients.
In conclusion, in dialysis patients, the B allele of the BsmI VDR
gene polymorphism is strongly and independently related to
LVH and is associated with a higher progression rate of LVH.
Since genes are randomly transmitted (Mendelian randomiza-
tion), our cross-sectional and longitudinal observations consis-
tently support the hypothesis that altered vitamin D signaling is
implicated in LVH in ESRD patients.
Disclosures
The authors state that they have no conflicts of interest.
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