Prevalence of Vitamin D Deficiency and Its Related Risk Factor in a Spanish Cohort of Adult HIV-Infected Patients: Effects of Antiretroviral Therapy

Abstract

We analyzed serum 25(OH) cholecalciferol [25(OH)D] levels and factors related to deficiency (<20 ng/ml) or insufficiency (<30 ng/ml) in a cohort of Spanish HIV-infected patients and compared them with an age- and latitude-matched population from another study. We prospectively assessed 25(OH)D deficiency/insufficiency in a cohort of 352 HIV patients during 2009–2010. Predisposing factors were recorded and their relationship to low levels was assessed by logistic regression; a nutritional survey examined intake, nutritional status, and sunlight exposure in a subgroup of 92 patients. We studied the correlation of 25(OH)D with parathyroid hormone (PTH) and alkaline phosphatase. Age-, sex-, and race/ethnicity-adjusted vitamin D deficiency (<20 ng/ml) was 44.0% (95% CI, 38.8–49.4%) and insufficiency (<30 ng/ml) was 71.6% (95% CI, 66.9–76.3). Deficiency was 16.4% more prevalent in our sample than in non-HIV-infected Spaniards. Lower sunlight exposure was the only factor related to lower levels in the lifestyle and nutritional survey (p=0.045). In multiple logistic regression, higher body mass index (BMI), black race/ethnicity, lower seasonal sunlight exposure, men who have sex with men (MSM), and heterosexual transmission categories, efavirenz exposure and lack of HIV viral suppression were independently associated with deficiency/insufficiency. These variables predicted 79% of cases [AUC=0.872 (95% CI, 0.83–0.91)]. Patients receiving protease inhibitors (PIs) [OR 4.0 (95% CI, 1.3–12.3); p=0.014] or NNRTI [OR 3.6 (95% CI, 1.7–11.2); p=0.025] had higher odds of increased PTH levels; this was significant only in 25(OH)D-deficient patients (p=0.004). As in less insolated areas, the prevalence of vitamin D deficiency/insufficiency was high in HIV-infected patients in Spain; among treated patients, levels were higher with PIs than with efavirenz.

Introduction

Vitamin D is considered a hormone with widespread effects; its deficiency has a wide impact, exceeding rickets and osteomalacia. There is evidence concerning its relationship to metabolic syndrome, diabetes, autoimmune diseases, arterial hypertension, and cancer.^1,2 It may even be considered a marker of cardiovascular disease.³ The precise mechanism of the multiple effects of vitamin D in various tissues is unknown; the only common factor is the presence of vitamin D receptor (VDR) in many tissues.

To assess vitamin D status, 25-hydroxycholecalciferol [25(OH)D] is measured. Its serum concentration reflects dietary input, synthesis through sunlight skin exposure, and vitamin D conversion from liver fatty stores.^4,5

Recently reference values have been changed, and a serum level between 30 and 76 ng/ml is considered normal. Using this cutoff point, the prevalence of vitamin D deficiency is as high as 50–80% in the general population.^6,7

The interest in vitamin D in HIV infection relates to its immunomodulatory properties. Nuclear VDRs have been found in immune system cells, such as T, B, and dendritic cells. In immune system cells, 25(OH)D is converted to 1,25(OH)₂D.⁸ Recently, low vitamin D levels have been related to disease progression of HIV-AIDS and its complications.^9,10 Data from the EuroSIDA cohort, which follows 20,000 subjects, highlight the impact of vitamin D deficiency on global and AIDS and non-AIDS-related mortality.¹¹

In recent years data on severe vitamin D deficiency in HIV-infected populations in several countries have been published. Studies differ in their populations, measurement methods, and cutoff points, which hinders comparison and obtaining conclusions. This year, results of the SUN study have been presented.¹² At variance with most previous studies, the number of participants was high (n=672). The study compares vitamin D deficiency and insufficiency according to current criteria and a thorough analysis of potential risk factors for vitamin D deficiency and insufficiency has been performed. Vitamin D deficiency (less then 20 ng/ml) and insufficiency (20–30 ng/ml) rates were high, 29.7% and 70.3%, respectively. In HIV-infected patients, HIV virus-related factors and certain antiretroviral drugs contribute to vitamin deficiency, along with malnutrition, exercise, ethnic background, advanced age, and inadequate sun exposure.

HIV may diminish 25(OH)D through several mechanisms. Proinflammatory cytokines such as tumor necrosis factor (TNF)-α¹³ drop levels, and macrophages and lymphocytes take up vitamin D as disease progresses. It is used in maturation and proliferation of T lymphocytes.¹⁴ Both protease inhibitors (PIs) and nonnucleoside reverse transcriptase inhibitors (NNRTIs) interfere with vitamin D metabolism.^15,16 PIs block hydroxylation of 25(OH)D and bioactivation of 1,25(OH)₂D,^2,14,16 whereas NNRTIs increase 25(OH)D and 1,25(OH)₂D metabolism.¹⁷

Five cross-sectional studies show that efavirenz (EFV)-based highly active antiretroviral therapy (HAART) is related to low vitamin D levels.^{9,12,17
–19} Additionally, more robust evidence has been found in two retrospective and one prospective cohort studies of naive patients.^20
–22 Until the 2011 MONET²³ and ECHO III²⁴ clinical trials, there was no controlled trial-based evidence on a causal relationship of EFV in diminishing 25(OH)D levels. Only a single case series of Spanish HIV patients has been published.²⁵

We undertook this study to assess prevalence of vitamin D deficiency and insufficiency in HIV-infected adults in Spain (40° N) in HAART and to compare it with data on a Spanish noninfected, age-matched population from the same geographic area.²⁶ We have analyzed in our patients risk factors generally associated with vitamin D deficiency. Specifically, we assessed the influence of ethnic background, the use of HAART, and the type of antiretroviral drugs used on vitamin D metabolism.

Materials and Methods

An observational cross-sectional study of a clinical population-based cohort of 450 HIV-infected patients was conducted. In 2009 and 2010, 352 patients diagnosed with HIV infection who were followed-up ay the infectious disease clinic of the Hospital Severo Ochoa, a public urban hospital close to Madrid, were recruited. Patients receiving biphosphonates, vitamin D supplements, or other therapies for bone metabolism were excluded. We analyzed data of blood chemistries, physical examination, and a nutritional and lifestyle survey. The database had been approved by the institutional ethical review board and patients provided verbal informed consent for data collection and use.

For this study we assessed comorbidities related to vitamin D deficiency or insufficiency, as well as HIV infection-related variables.

Laboratory methods

Serum 25(OH)D levels were assessed with a Cobas e 411 multichannel analyzer employing electrochemiluminescence (ECL) technology (Roche), intact PTH with IMMULITE 2000, a solid-phase two-site chemiluminescent immunoassay (Siemens), CD4 count by means of flow cytometry, and HIV viral load using with real time polymerase chain reaction (PCR) (Taqman); less than 50 RNA copies/ml was considered undetectable. Other chemistries were assayed with standard automated techniques.

Based upon previous studies, we chose <30 ng/ml to define vitamin D insufficiency and <20 ng/ml for deficiency. Intact parathyroid hormone (PTH) was considered increased when its plasma level was above 66 pg/ml.

To compare the prevalence of vitamin D deficiency and insufficiency with that of an age-matched Spanish population of the same geographic area we used data published by M. Calatayud.²⁶

In our study a subgroup of cohort subjects (n=92) agreed to participate in a nutritional and lifestyle substudy. A nonvalidated survey about nutrition and lifestyle was administered by a certified nutrition specialist, gathering information on sunlight exposure, use of UVA cabins, dietary vitamin D intake, vitamin supplements, sunscreens, and exercise.

Statistical analysis

Vitamin D deficiency (<20 mg/ml) and insufficiency (<30 ng/ml) estimates with 95% CI were performed, both global and adjusted for age, ethnic background, and gender.

We divided patients in tertiles according to vitamin D levels. Qualitative variables were compared by χ² or Fisher's exact tests; for continuous variables nonparametric Kruskal–Wallis was used.

Correlation between 25(OH)D, PTH, alkaline phosphatase (excluding subjects with alanine aminotransferase ≥40 IU), and CD4⁺ cell count was analyzed with Spearman's correlation test.

The effect of putative vitamin D deficiency predictors was studied by logistic regression. Variables with a p value of <0.20 in univariate analysis were included in the multiple logistic regression model. Thereafter, a reduced model was selected using backward elimination. To include all cases in multivariable analysis, unknown values were replaced by assigned values using the expectation maximization algorithm. Simple linear regression was used to assess the difference of change in the level of vitamin D related to the variables found in the simplified logistic regression. Factors associated with the upper quartile of alkaline phosphatase and iPTH were studied by means of logistic regression.

Associations were considered significant if p<0.05 in a two-tailed test. Statistical analysis was performed with SPSS version 15.0 (SPSS Statistics).

Results

Table 1 shows characteristics of study participants. The median 25(OH)D level was 22 ng/ml (IQR 14.22–31.7 mg/ml). Samples were classified in tertiles according to vitamin D level. The upper tertile included a significant excess of white race subjects (p=0.001) and former intravenous drug users (IDUs) (p=0.0001), and an increased proportion of blood samples drawn in summer (p=0.0001) and patients on antiretroviral therapy (0.006).

Table 1.

Baseline Characteristics of the Study Sample

Characteristic	Number	Percentage
Sex
Male	236	(67)
Female	116	(33)
Age (years)
Median (IQR)	44	(39–49)
≤29	18	(5.1)
30–39	79	(22.4)
40–49	171	(48.6)
≥50	84	(23.9)
Race/ethnicity
White	299	(84.9)
Black	30	(8.5)
Hispanic	23	(6.5)
Transmission category
IDU	155	(44)
Heterosexual	137	(38.9)
MSM	54	(15.3)
Other	6	(1.7)
BMI
Median (IQR), kg/m²	23.81	(21.95–25.80)
≤25	233	(66.2)
25.01–29.99	96	(27.3)
≥30	23	(6.5)
Baseline CD4⁺ count
Median (IQR) cells/μl	501	(334–685)
≤199	36	(10.2)
200–350	62	(17.6)
≥350	254	(72.2)
CD4⁺ count nadir
Median (IQR) cells/μl	188	(80–285)
≤199	188	(53.4)
200–350	116	(34)
≥350	48	(13.6)
Viral load
Median (IQR), copies/ml	49.5	(49.5–49.5)
<50	277	(78.7)
≥50	75	(21.3)
ART exposition
Naive	37	(10.5)
Present exposition	315	(89.5)

IQR, interquartile range; IDU, intravenous drug user; MSM, men who have sex with men; BMI, body mass index; ART, antiretroviral treatment.

Among study participants, the age-, sex-, and race-adjusted prevalence of vitamin D deficiency (25[OH]D <20 ng/ml) was 44.0% (95% CI, 38.8–49.4), and the prevalence of vitamin D insufficiency (<30 ng/ml) was 71.6% (95% CI, 66.9–76.3). Only in 28.4% (95% CI, 23.7–33.1) could vitamin levels be considered optimal (≥30 ng/ml).

Compared with the general Spanish adult population,²⁶ the percentage of subjects with optimal levels of vitamin D was higher in our sample, although the difference was not statistically significant. However, vitamin D deficiency was more prevalent in the study subjects (16.4%; 95% CI, 6.8–26.09; p=0.001).

Table 2 shows results of the dietary intake and lifestyle questionnaires filled out by 92 subjects. Among many factors studied, only increased sunlight exposure showed statistically significant differences toward optimal vitamin D levels (p=0.045).

Table 2.

Results of Nutritional and Lifestyle Survey

Variable	<30 ng/ml (n=75)	>30 ng/ml (n=17)	p
Daily vitamin D intake μg/day, median (IQR)	9.34 (4.50–15.68)	10.94 (5.80–17.46)	0.611
Intake < rda	34.7%	17.6%	0.173
Intake > rda	65.3%	82.4%
Daily sunlight exposure (minutes), median (IQR)	17.14 (4.28–60)	67.5 (9.64–173.57)	0.045
Vitamin supplements			0.249
Yes	21.3%	6.3%
No	78.8%	93.8%
Exercise			0.799
No	56%	64.7%
Sporadic	13.3%	11.8%
Daily	30.7%	23.5%
UV cabins			0.759
Yes	4.2%	5.9%
No	95.8%	94.1%
Tanned last year			0.640
Yes	46.7%	52.9%
No	53.3%	47.1%
Sunscreen use			0.848
Yes	41.2%	38.7%
No	58.8%	61.3%

Rda, recommended dietary allowance.

Of the 352 subject studies, 139 (39.5%) were taking PIs, 129 (36.6%) NNRTIs, and 21 (6%) received coformulated zidovudine/lamivudine/abacavir. There were statistically significant differences in vitamin D levels, which were highest in patients on PIs monotherapy and lowest in those receiving EFV or zidovudine/lamivudine/abacavir. Figure 1 shows the percentage of patients with vitamin D deficiency or insufficiency.

FIG. 1.

Vitamin D levels related to antiretroviral (ART) use.

Univariate and multivariate analysis of vitamin D deficiency- and insufficiency-associated factors in HIV-infected patients (Table 3)

Table 3.

Baseline Demographic and Clinical Factors Associated with Vitamin D Insufficiency or Deficiency

	Vitamin D <30 ng/ml (N=252)	Vitamin D ≥30 ng/ml (N=100)	Nonadjusted OR (95% CI)	Adjusted OR (95% CI)
Sex
Male	82 (72.7%)	34 (29.3%)	0.79 (0.57–1.5)	—
Female	170 (72%)	66 (28%)	Referent
Age, median, IQR	43.5 (37.5–49)	45 (40–49)	0.98 (0.95–1.01)	—
Race/ethnicity
White	208 (69.6%	91 (30.4%)	Referent	Referent
Black	29 (96.7%)	1 (3.3%)	12.6 (1.7–94.5)^*	9.4 (1.11–78.5)^*
Hispanic	15 (65.2%)	8 (34.8%)	0.82 (0.34–2)^*	0.2 (0.05–0.77)^*
Season
Summer	45 (54.9%)	37 (45.1%)	Referent	Referent
Spring	67 (99.5%)	1 (1.5%)	55.1 (7.3–416)^*	65.6 (8.4–511)^*
Fall	73 (55.7%)	58 (44.3%)	1.04 (0.6–1.8)	0.96 (0.5–1.83)
Winter	67 (94.4%)	4 (5.6%)	13.7 (4.6–41.3)^*	16.3 (5.1–52.3)^*
BMI, median, IQR	23.9 (21.9–26.5)	23.5 (21.9–25.1)	1.07 (1.01–1.1)^*	1.08 (1.005–1.16)^*
Years since diagnosis, IQR	11.8 (5.51–17.9)	14.9 (9–19.4)	0.96 (0.93–0.9)^*	1.01 (0.96–1.1)
Years since ART start, IQR	6.4 (2.9–11.7)	7.15 (3.6–11.62)	0.99 (0.94–1.03)	—
AIDS diagnosis
Yes	138 (67.6%)	66 (32.4%)	1.6 (0.9–2.6)	0.93 (0.5–1.81)
No	114 (77%)	34 (23%)	Referent	Referent
CD4⁺ <200	31 (86.1%)	5 (13.9%)	2.69 (1.01–7.2)^*	1.37 (0.4–5)
200–350	44 (71%)	18 (29%)	1.06 (0.58–1.9)	1.39 (0.6–3.1)
≥350	177 (69.7%)	77 (30.3%)	Referent	Referent
CD4⁺ nadir
<200	132 (70.2%)	56 (29.8%)	0.54 (0.25–1.2)	—
200–350	81 (69.8%)	35 (30.2%)	0.53 (0.23–1.22)
≥350	39 (81.2%)	9 (18.8%)	Referent
Antiepileptic drugs
Yes				—
No	6 (66.6%)	3 (33.3%)	0.79 (0.2–3.2)
	246 (71.7%)	97 (28.3%)	Referent
Viral C hepatitis
Yes	102 (60.4%)	67 (39.6%)	0.34 (0.2–0.5)^*	0.78 (0.24–2.5)
No	150 (82%)	33 (18%)	Referent	Referent
Viral B hepatitis
Yes	9 (69.2%)	4 (30.8%)	0.89 (0.27–2.9)	—
No	243 (71.7%)	96 (28.3%)	Referent
ALT
≥40 IU	68 (61.3%)	43 (38.7%)	0.49 (0.3–0.8)^*	0.99 (0.5–1.9)
<40 IU	184 (76.3%)	57 (23.7%)	Referent	Referent
Glomerular filtration rate (MDRD)
<60 ml/min	7 (43.8%)	9 (56.3%)	0.29 (0.1–0.8)	0.52 (1.14–1.96)
≥60 ml/min	245 (72.9	91 (27.3%)	Referent	Referent
HDL
Abnormal	98 (74.8%)	33 (25.2%)	1.29 (0.8–2.1)	—
Normal	154 (69.7%)	67 (30.3%)	Referent
Total cholesterol
≥240 mg/dl	21 (77.8%)	6 (22.2%)	1.42 (0.5–3.6)	—
<240 mg/dl	231 (71.1%)	94 (28.9%)	Referent
Triglycerides
≥150 mg/dl	94 (70.1%)	40 (29.9%)	0.89 (0.5–1.43)	—
<150 mg/dl	158 (72.5%)	60 (27.5%)	Referent
Glucose
≥126 mg/dl	16 (88.9%)	2 (11.2%)	3.3 (0.7–14.1)	4.5 (0.8–24.9)
<126 mg/dl	236 (70.7%)	98 (29.3%)	Referent	Referent
High blood pressure
Yes ≥140/70	78 (76.5%)	24 (23.5%)	1.42 (0.8–2.41)	1.42 (0.7–2.88)
No <140/70	174 (69.6%)	76 (30.4%)	Referent	—
Insulin resistance
HOMA ≥3.8	40 (70.7%)	12 (23.1%)	1.38 (0.7–2.76)	—
HOMA <3.8	212 (70.7%)	88 (29.3%)	Referent
Tobacco
Yes	155 (68.6%)	71 (31.4%)	0.65 (0.4–1.1)	0.89 (0.45–1.2)
No	97 (77%	29 (23%)	Referent	Referent
Treatment
Naive	40 (95.2%)	2 (4.8%)	9.24 (2.2–39)^*	4.3 (0.7–27.7)
Experience	212 (68.4%)	98 (31.6%)	Referent	Referent
Efavirenz exposure
Yes	67 (80.7%)	16 (19.3%)	1.9 (1.04–3.5)^*	2.87 (1.4–5.9)^*
No	185 (68.8%)	84 (31.2%)	Referent
Nevirapine exposure
Yes	27 (65.9%)	14 (34.1%)	0.73 (0.4–1.47)	—
No	225 (72.3%)	86 (27.1%)	Referent
PI exposure
Yes	106 (62.7%)	63 (37.3%)	0.43 (0.3–0.7)^*	1.1 (0.5–2.43)
No	146 (79.8%)	37 (20.2%)	Referent	Referent
Tenofovir exposure
Yes	127 (70.6%)	53 (29.4%)	0.9 (0.6–1.43)	—
No	125 (72.7%)	47 (27.3%)	Referent
NRTI exposure
Yes	15 (71.4%)	6 (28.6%)	0.99 (0.4–2.6)	—
No	237 (71.6%)	94 (28.4%)	Referent
VIH copies
>50	66 (88%)	9 (12%)	3.58 (1.7–7.5)^*	3.5 (1.27–9.9)^*
≤50	186 (67.1%)	91 (32.9%)	Referent	Referent
Transmission category
Heterosexual
MSM	110 (80.3%)	27 (19.79%)	2.79 (1.6–4.7)^*	2.5 (1.2–5.1)^*
Other	45 (83.3%)	9 (16.7%)	3.42 (1.6–7.5)^*	3.4 (1.3–8.6)^*
IDU	5 (83.3%)	1 (16.7%)	3.42 (0.4–30)	8.3 (0.6–124.7)
	92 (59.4%)	63 (40.6%)	Referent	Referent

IQR, interquartile range; BMI, body mass index; ART, antiretroviral treatment; ALT, alanine aminotransferase; HDL, high-density lipoprotein; HOMA, homeostatic model assessment (for insulin resistance); PI, protease inhibitor; NRTI, nucleoside reverse transcriptase inhibitor; MSM, men who have sex with men; IDU, intravenous drug user. ^* p<0.05.

In a univariate analysis, the absence of antiretroviral therapy (ART), nonsuppressed viral load, shorter duration of HIV infection, and lower CD4⁺ cell counts were associated with increased odds of vitamin D deficiency or insufficiency.

Regarding ART, EFV exposure was associated with significantly higher odds of vitamin D insufficiency or deficiency, whereas IPs exposure seemed protective (p=0.014). We did not observe significant associations between vitamin D insufficiency or deficiency and NRTI, tenofovir, or nevirapine exposure.

In univariate analysis of non-HIV-related factors, black race/ethnicity, lesser UV exposure (lower levels in spring/fall than in summer/winter), increased body mass index (BMI), GPT <40 IU, the absence of hepatitis C virus (HCV) coinfection, and estimated GFR ≥60 ml/min/1.73 m² (MDRD) were associated with increased odds of vitamin D insufficiency or deficiency (p<0.05).

In logistic multiple regression, factors independently associated with increased odds of vitamin D insufficiency or deficiency were BMI, black race/ethnicity, less UV exposure in winter and spring, men who have sex with men (MSM) and heterosexual HIV transmission categories, EFV exposure, and unsuppressed HIV viral load (Table 4).

Table 4.

Multiple Linear Regression Analysis of Correlates of 25-Hyroxyvitamin D Concentrations

Correlate	Coefficient	Standard error	p
Male vs. female	−3.88	2.042	0.058
Age	0.081	0.116	0.486
Transmission category
Heterosexual	5.339	3.726	0.153
MSM	5.746	4.345	0.187
Other	−0.216	7.633	0.977
IDU	Referent
Race/ethnicity
Black	−8.39	3.644	0.022^*
Hispanic	14.395	3.790	0.0001^*
White	Referent
BMI	−0.3111	0.145	0.232
Years since HIV diagnosis	−0.339	0.134	0.012^*
Season
Spring	−14.44	2.376	0.0001^*
Fall	−2.004	2.478	0.419
Winter	−10.95	2.360	0.0001^*
Summer	Referent
CD4⁺ count (cells/μl)
≥350	4.687	3.319	0.159
200–350	1.477	3.728	0.692
<200	Referent
HCV yes/no	7.759	2.157	0.0001
ALT (IU) >40 UI vs. <40 UI	4.558	2.095	0.030^*
Glomerular filtration rate MDRD <60 vs. >60	4.526	4.619	0.328
Diabetes yes/no	0.528	4.448	0.906
High blood pressure yes/no	−1.417	2.095	0.499
Tobacco yes/no	−0.981	2.161	0.650
ART no/yes	−11.619	3.794	0.002^*
EFV containing regime	−5.219	2.200	0.018^*
PI containing regime	2.853	2.501	0.257
HIV viral load >50 vs. <50 copies	−6.776	2.885	0.019^*

MSM, men who have sex with men; IDU, intravenous drug user; BMI, body mass index; HCV, hepatitis C virus; ALT, alanine aminotransferase; ART, antiretroviral treatment; EFV, efavirenz; PI, protease inhibitor. ^* p<0.05.

Using this predictive model, which included the aforementioned variables, the sensitivity of predicting vitamin D insufficiency or deficiency (25[OH]D <30 ng/ml) was 87.3%, the specificity was 58%, and the discriminatory power was 79%. The final model has an acceptable power for classification, since it is higher than 75%. Its receiver-operating characteristics (AUC) is 0.872 (95% CI, 0.83–0.91).

In a multiple linear regression model including the same independent variables as the abovementioned logistic regression model, the significance of the association of heterosexual and MSM transmission categories with 25(OH)D deficiency or insufficiency is lost (p=0.153 and 0.187). Years since HIV infection diagnosis and absence of ART are significantly associated with low 25(OH)D, whereas higher levels are found in HCV-infected subjects and in those with ALT ≥40 IU. The remaining significant variables in the logistic regression model retained their independent association with serum 25(OH)D concentration.

Eighty-eight patients (25%) had high PTH levels (upper quartile, ≥66 pg/ml). Among subjects with vitamin D insufficiency, 66 (26.2%) had elevated PTH levels. The odds of high levels of PTH were higher in patients receiving PIs (OR 4.05; 95% CI, 1.33–12.3; p=0.014) or with NNRTI (OR 3.6; 95% CI, 1.7–11.17; p=0.025) than in untreated patients. PTH levels in PI-treated patients were not significantly higher than in NNRTI-treated patients (p=0.72).

Tenofovir-exposed patients had higher PTH levels, but this difference was significant only in patients with vitamin D insufficiency (p=0.004). The significant increase in alkaline phosphatase in patients receiving tenofovir was independent of vitamin D status. There were no differences in phosphate and calcium levels. We observed a negative correlation between serum PTH and 25(OH)D levels (r=0.098; p=0.048) and a positive one with alkaline phosphatase (r=0.0105; p=0.024).

Discussion

Our cohort's vitamin D deficiency or insufficiency rates, 71.59%, are very similar to prevalence rates of other series of HIV-positive patients studied in areas of lower insolation.^12,17,18,21 Another recent paper observed a 74% rate of vitamin D insufficiency in postmenopausal HIV-infected women, similar to the control group of African-American women, the category of highest vitamin D insufficiency prevalence.²⁷

A 71.59% rate of vitamin D deficiency or insufficiency in our cohort of HIV-infected adults of the Hospital Severo Ochoa is high, and close to the prevalence observed in the general population of the Comunidad de Madrid (Spain), as shown by Calatayud.²⁶ This latter study was performed in a sample of similar age recruited in the same geographic area, in May and June. Although the prevalence of vitamin deficiency or insufficiency was similar in both studies, we would like to emphasize the fact that deficiency—probably more clinically important—was significantly more prevalent in HIV patients (16.4%; 95% CI 6.8–26.1, p=0.001). Our findings are similar to rates of the general population elsewhere.^1,7,28

A lower limit of the reference range has recently been set at 30 ng/ml. There are several reasons for this value. In studies performed several years ago, PTH levels were found to be increased in the population whenever 25(OH)D levels dropped below this level.¹ More recent data show optimal calcium absorption when the 25(OH)D level is 32 ng/ml or higher.⁴ Another measure to define an optimal 25(OH)D status is the value at which no further increase of 1,25(OH)₂D is noted after administration of vitamin D. When 25(OH)D levels were below 25 and 30 ng/ml, increases in 1,25(OH)₂D were found.²⁹

Findings from a nutritional and lifestyle survey highlight the relevance of sunlight exposure. Only high exposure, which was linked to outdoor jobs, was related to optimal vitamin D levels.

Considering the prevalence of vitamin D insufficiency, the analysis of the relationship between certain antiretroviral drugs and vitamin D metabolism seems very interesting. Patients on darunavir/ritonavir or lopinavir/ritonavir monotherapy had greater immune recovery, longer HIV infection duration, and closer to optimal vitamin D levels. Our results were similar to those of the MONET study.²³ This exploratory findings should be confirmed in future larger studies on bone health in effective boosted PI monotherapy.

Data on EFV and vitamin D deficiency have accumulated in recent years. Our data show that the odds of vitamin D insufficiency or deficiency increased 2.87 times with the use of this drug, and levels were 5.2 ng/ml lower than with other therapeutic regimes. This is not an NNRTI class effect on vitamin D metabolism. At variance with others,¹² we did not observe a relationship between tenofovir exposure and higher vitamin D levels.

We were unable to show a relationship between HIV infection, degree of immune suppression or AIDS diagnosis, and vitamin D levels, but we did observe lower levels of 25(OH)D in patients with unsuppressed viral load. Other factors significantly associated with 25(OH)D deficiency or insufficiency in multivariate analysis, such as being overweight, black race/ethnicity, and season, are in accordance with previous literature. A surprising finding of our study is higher vitamin D levels in former IDUs. We have no explanation for this; more frequent outdoor jobs and less use of EFV in this transmission category could have a role.

The final model predicts vitamin D deficiency or insufficiency in HIV-infected patients with high discrimination power (79%).

Among multiple comorbidities and drugs, only diabetes mellitus showed an association with lower levels of vitamin D (p<0.10). Although not statistically significant (p=0.089), diabetes mellitus was associated with a clinically significant increase in the odds for vitamin D insufficiency (OR 4.5; 95% CI 0.8–24.9). Osteocalcin production requires adequate vitamin D levels, and recombinant osteocalcin administration may revert to hyperglycemia and hyperinsulinemia³⁰; this points to a link between vitamin D, bone, fat, and glucose metabolism.

Among NRTIs, tenofovir produces major concerns on bone health and vitamin D metabolism. Tenofovir-based regimens in naive^31,32 and nonnaive³³ patients are associated with decreases in bone mineral density (BMD). The precise mechanisms of this loss are not known, but the Swiss Cohort has described an increased fractional excretion of phosphate in the urine in 30–40% of patients, and an increase in alkaline phosphatase, a marker of bone remodeling.³⁴ Two recent cross-sectional studies described a significant increase in median PTH levelsin healthy individuals with vitamin D deficiency.^35,36 We had similar findings: PTH was significantly higher in tenofovir-exposed patients, but only in those with vitamin D insufficiency. However, the significant increase in alkaline phosphatase was independent of vitamin D level. Assessing the contribution of vitamin D, PTH, calcium, phosphate, and certain antiretrovirals such tenofovir to BMD seems advisable.

Although vitamin D insufficiency is widespread, identifying its predictors in HIV-infected patients, such as efavirenz exposure and nonsuppressed viral load, will make it possible to identify those patients at highest risk. The latest European Guidelines for treatment of HIV-infected adults in Europe (October 2011) advise screening patients at risk of vitamin D deficiency.³⁷

Our study has several limitations. The absence of an HIV-negative comparison group limits reaching conclusions on prevalence rates in HIV-positive outpatients. Recruitment of patients in a single hospital with specific geographic and social characteristics in his catchment area may limit the external validity for the whole population living in Spain. The cross-sectional design prevents the establishment of causal relationships. Most patients had fairly good CD4⁺ cell counts, which may have an impact on the results.

Footnotes

Acknowledgments

We have obtained substantial assistance from Susana Pastor in data retrieval and from Rebeca Sanz in performing the nutritional survey.

Author Disclosure Statement

No competing financial interests exist.

References

Holick

. Vitamin D deficiency. N Engl J Med, 2007; 357:266–281.

Wang

, Pencina

, Booth

et al. Vitamin D deficiency and risk of cardiovascular disease. Circulation, 2008; 117:503–511.

Zittermann

, Frisch

, Berthold

et al. Vitamin D supplementation enhances the beneficial effects of weight loss on cardiovascular disease risk markers. Am J Clin Nutr, 2009; 89:1321–1327.

Heaney

. The vitamin D requirement in health and disease. J Steroid Biochem Mol Biol, 2005; 97:13–19.

Ross

, Manson

, Abrams

et al. The 2011 report on dietary reference intakes for calcium and vitamin D from the Institute of Medicine: What clinicians need to know. J Clin Endocrinol Metab, 2011; 96:53–58.

Holick

, Siris

, Binkley

et al. Prevalence of vitamin D inadequacy among postmenopausal North American women receiving osteoporosis therapy. J Clin Endocrinol Metab, 2005; 90:3215–3224.

Ginde

, Liu

, Camargo

Jr . Demographic differences and trends of vitamin D insufficiency in the US population, 1988–2004. Arch Intern Med, 2009; 169:626–632.

Baeke

, Takiishi

, Korf

, Gysemans

, Mathieu

. Vitamin D: Modulator of the immune system. Curr Opin Pharmacol, 2010; 10:482–496.

Welz

, Childs

, Ibrahim

et al. Efavirenz is associated with severe vitamin D deficiency and increased alkaline phosphatase. AIDS, 2010; 24:1923–1928.

10.

Mehta

, Giovannucci

, Mugusi

et al. Vitamin D status of HIV-infected women and its association with HIV disease progression, anemia, and mortality. PLoS One, 2010; 5:e8770.

11.

Viard

, Souberbielle

, Kirk

et al. Vitamin D and clinical disease progression in HIV infection: Results from the EuroSIDA study. AIDS, 2011; 25,10:1305–1315.

12.

Dao

, Patel

, Overton

et al. Low vitamin D among HIV-infected adults: Prevalence of and risk factors for low vitamin D Levels in a cohort of HIV-infected adults and comparison to prevalence among adults in the US general population. Clin Infect Dis, 2011; 52:396–405.

13.

Haug

, Aukrust

, Haug

et al. Severe deficiency of 1,25-dihydroxyvitamin D3 in human immunodeficiency virus infection: Association with immunological hyperactivity and only minor changes in calcium homeostasis. J Clin Endocrinol Metab, 1998; 83:3832–3838.

14.

Villamor

. A potential role for vitamin D on HIV infection? Nutr Rev, 2006; 64:226–233.

15.

Fabbriciani

, De Socio

. Efavirenz and bone health. AIDS, 2009; 23:1181

16.

Cozzolino

, Vidal

, Arcidiacono

et al. HIV-protease inhibitors impair vitamin D bioactivation to 1,25-dihydroxyvitamin D. AIDS, 2003; 17:513–520.

17.

Van Den Bout-Van Den Beukel

, Fievez

, Michels

et al. Vitamin D deficiency among HIV type 1-infected individuals in the Netherlands: Effects of antiretroviral therapy. AIDS Res Hum Retroviruses, 2008; 24:1375–1382.

18.

Rodriguez

, Daniels

, Gunawardene

, Robbins

. High frequency of vitamin D deficiency in ambulatory HIV-positive patients. AIDS Res Hum Retroviruses, 2009; 25:9–14.

19.

Wasserman

, Rubin

. Highly prevalent vitamin D deficiency and insufficiency in an urban cohort of HIV-infected men under care. AIDS Patient Care STDS, 2010; 24,4:223–227.

20.

Brown

, McComsey

. Association between initiation of antiretroviral therapy with efavirenz and decreases in 25-hydroxyvitamin D. Antivir Ther, 2010; 15,3:425–429.

21.

Mueller

, Fux

, Ledergerber

et al. High prevalence of severe vitamin D deficiency in combined antiretroviral therapy-naive and successfully treated Swiss HIV patients. AIDS, 2010; 24:1127–1134.

22.

Conesa-Botella

, Florence

, Lynen

et al. Decrease of vitamin D concentration in patients with HIV infection on a non nucleoside reverse transcriptase inhibitor-containing regimen. AIDS Res Ther, 2010; 7:40.

23.

Fox

, Peters

, Prakash

et al. Improvement in vitamin D deficiency following antiretroviral regime change: Results from the MONET trial. AIDS Res Hum Retroviruses, 2011; 27,1:29–34.

24.

Wohl

, Doroara

, Orkin

et al. Change in vitamin D levels smaller and risk of development of severe vitamin D deficiency lower among HIV-1-infected, treatment-naïve adults receiving TMC278 compared with EFV: 48-week results from the Phase III ECHO Trial. In 18th Conference on Retroviruses and Opportunistic Infections, Boston, MA, 2011.

25.

Garcia Aparicio

, Munoz Fernandez

, Gonzalez

et al. Abnormalities in the bone mineral metabolism in HIV-infected patients. Clin Rheumatol, 2006; 25:537–539.

26.

Calatayud

, Jodar

, Sanchez

, Guadalix

, Hawkins

. [Prevalence of deficient and insufficient vitamin D levels in a young healthy population] Endocrinol Nutr, 2009; 56:164–169.

27.

Stein

, Yin

, McMahon

et al. Vitamin D deficiency in HIV-infected postmenopausal Hispanic and African-American women. Osteoporos Int, 2011; 22,2:477–487.

28.

National Center for Health Statistics CfDCaP: NHANES analytic guidelines. 2004.

29.

Thacher

, Obadofin

, O'Brien

, Abrams

. The effect of vitamin D₂ and vitamin D₃ on intestinal calcium absorption in Nigerian children with rickets. J Clin Endocrinol Metab, 2009; 94:3314–3321.

30.

Overton

, Yin

. The rapidly evolving research on Vitamin D among HIV-infected populations. Curr Infect Dis Rep, 2011; 13:89–93.

31.

Gallant

, Staszewski

, Pozniak

et al. Efficacy and safety of tenofovir DF vs stavudine in combination therapy in antiretroviral-naive patients: A 3-year randomized trial. JAMA, 2004; 292:191–201.

32.

McComsey

, Kitch

, Daar

et al. Bone, limb fat outcomes of ACTG A5224s, a substudy of ACTG A5202: A prospective, randomized, partially blinded phase III trial of ABC/3TC or TDF/FTC with EFV or ATV/r for initial treatment of HIV-1 infection. In 17th Conference on Retroviruses and Opportunistic Infections HIV-1 infection, February 16–19, 2010, San Francisco, CA

33.

Cooper

, Bloch

, Humphries

. Simplification with fixed dose tenofovir-emtricitabine or abacavir-lamivudine adults with suppressed HIV replication (The Steal Study, randomized, open-label, 96-week, non-inferiority trial) In 16th Conference on Retroviruses and Opportunistic Infections, Montreal, Canada, 2009.

34.

Fux

, Rauch

, Simcock

et al. Tenofovir use is associated with an increase in serum alkaline phosphatase in the Swiss HIV Cohort Study. Antivir Ther, 2008; 13:1077–1082.

35.

Childs

, Fishman

, Constable

et al. Short communication: Inadequate vitamin D exacerbates parathyroid hormone elevations in tenofovir users. AIDS Res Hum Retroviruses, 2010; 26:855–859.

36.

Rosenvinge

, Gedela

, Copas

et al. Tenofovir-linked hyperparathyroidism is independently associated with the presence of vitamin D deficiency. J Acquir Immune Defic Syndr, 2010; 54:496–499.

37.

European AIDS Clinical Society (EACS). Guidelines Version 6.0 October 2011. www.europeanaidsclinicalsociety.org/images/stories/EACS-Pdf/eacsguidelines-6.pdf. 2011 November 15.