Introduction

Hypertension is a modifiable risk factor risk factor that is associated with cardiovascular and cerebrovascular disease and mortality [1, 2]. Approximately 90–95% of cases result from primary hypertension with many etiological factors, such as age, race, family history, obesity, lifestyle, use of tobacco, high salt intake, stress, consumption of alcohol in large quantities, and nonspecific lifestyle and genetic factors [3, 4]. Worldwide, nearly one billion adults (25%) are suffering from primary hypertension, and the prevalence may increase by 29% by 2025 [3, 4]. Recent studies have established an association of serum vitamin D deficiency with primary hypertension [5]. Vitamin D is important for the human body to maintain a balance between calcium and phosphorus. Vitamin D deficiency can cause weakness, increased bone loss, and low levels of bone mineralization [6]. Serum 25-hydroxyvitamin D is a marker of vitamin D status in the human body [7]. The aim of this study was to investigate the association between serum 25-hydroxyvitamin D deficiency and primary hypertension in south Indian subjects. Very limited studies are available from the Indian subcontinent.

Materials and methods

We prospectively enrolled 400 hypertensive subjects and age- and sex-matched normotensive subjects at the Yashoda Hospital Hyderabad. Yashoda Hospital is a tertiary care center that serves the states of Andhra Pradesh and Telangana in south India. The study period was from January 2015 to December 2017, and the study was approved by the Institutional Ethics Committee. We enrolled both groups in three seasons (summer season: March to mid-June, rainy season: mid-June to October and winter season: November to February).

Selection of cases

Hypertensive subjects were identified by systolic blood pressure (SBP) ≥140 mmHg and diastolic blood pressure (DBP) ≥90 mmHg, or subjects with self-reported hypertension. Incident hypertension was considered a recently developed hypertension with SBP ≥140 mmHg and DBP ≥90 mmHg at baseline and three consecutive visits or based on the Joint National Committee VIII criteria [2].

Selection of normotensive (controls)

Normotensive subjects were defined as those with SBP ≤120 mmHg and DBP ≤80 mmHg at baseline and three consecutive visits without the use of antihypertension medications.

Inclusion and exclusion criteria

Hypertensive subjects older than 40 years and age- and sex-matched normotensive subjects were included in the study. Individuals with vitamin D supplementation, tuberculosis, any bone diseases, chronic kidney disease, muscle weakness, secondary hypertension, steroids, cardiovascular and cerebrovascular disease, chronic liver disease, anti-epileptic drug use, cholestyramine drugs, antacid medications, orlistat and osteoporosis drugs or white coat syndrome were excluded from the study for both groups (hypertensive and normotensive).All subjects’ data were collected through face-to-face interviews, and present and past medical histories were collected. A standardized questionnaire was adapted from the behavioral risk factor surveillance system from the Centers for Disease Control and Prevention [7]. Subjects in both groups underwent fasting blood sugar, lipid profile, liver function, serum calcium, alkaline phosphatase, phosphorous, 25-hydroxyvitamin D, and C-reactive protein (CRP) analyses.

Risk factor analysis

Hypertension was defined as ≥140 mmHg SBP and ≥90 mmHg DBP or the use of antihypertension medications, diabetes was diagnosed if subjects were taking antidiabetic medications, dyslipidemia was considered in subjects having ≥200 mg/dL or who were on statins, alcoholism was defined as alcohol consumption >50 g/day, smokers were defined as those reporting a daily smoking habit, ex-smokers and occasional smokers were categorized as nonsmokers [4, 8, 9], CRP positivity was considered when the CRP level was ≥0.6 mg/dL [10], and alanine transaminase (ALT) levels ≥70 U/L and aspartate transaminase (AST) levels ≥47 U/L were treated as abnormal levels [11].

Blood pressure monitoring

Ambulatory blood pressure monitoring was performed in cases and controls. Blood pressure was measured by a trained physician with a mercury sphygmomanometer on the right arm with the participant in a seated position after 5 min of rest.

Assessment of 25-hydroxyvitamin D

In our laboratory, we used a chemiluminescent microparticle immunoassay with an automated instrument for the estimation of 25-hydroxyvitamin D, and the sensitivity and specificity of the instrument were 53% and 90.5%, respectively. Our lab manual and the current literature state that serum 25-hydroxyvitamin D values ≤20 ng/mL indicate deficiency and that values ≥20.1 ng/mL are normal [12, 13].

Statistical analysis

Statistical analysis was performed using SPSS Inc. (Statistical Package for the Social Sciences) 21.0 software. The mean and standard deviation were calculated. The paired t-test was performed to assess differences in continuous variables. Multiple logistic regression analyses were performed before and after adjustment for potential confounders (sex, dyslipidemia, CRP positivity, obesity, diabetes, alcoholism, and smoking). We assessed the receiver operating characteristic (ROC) curve to identify the cut-off value of serum 25-hydroxyvitamin D, which may predict the outcome values. All tests were two-sided, and a p value < 0.05 was considered statistically significant.

Results

A total of 400 hypertensive subjects and 400 normotensive subjects were included in the study. Men accounted for 63.7% of the hypertensive patients and 63.7% of the control subjects. The mean age was similar in both groups (52.4 ± 4.43 years in the hypertensive subjects and 53.1 ± 5.12 years in the normotensive subjects). Hypertensive subjects had a significantly higher proportion of vitamin D deficiency (p < 0.0001), low mean serum calcium (p < 0.0001), elevated mean parathyroid hormone (PTH) (p < 0.0001), low mean serum phosphorous (p < 0.0001), abnormal ALT (p = 0.0015), abnormal AST (p = 0.0056), and high mean alkaline phosphatase (p < 0.0001) (Table 1).

Table 1 Baseline characteristics
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We compared various factors based on vitamin D status in hypertensive subjects. Long mean duration of hypertension (p = 0.02), elevated mean SBP (p < 0.0001) and mean DBP (p < 0.0001), abnormal ALT (p = 0.0064), and abnormal AST (p = 0.0034) were significantly associated with vitamin D deficiency (Table 2).

Table 2 Difference between 25-hydroxyvitamin D deficiency and normal 25-hydroxyvitamin in hypertensive subjects
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After multivariate analysis, we established that the major predictors of hypertension were serum 25-hydroxyvitamin D (odds 1.78; 95% CI: 1.31–2.41), abnormal ALT (odds: 1.2; 95% CI: 0.98–1.94), abnormal AST (odds: 1.1; 95% CI: 0.89–1.72), and CRP positivity (odds 1.48; 95% CI: 1.18–2.12) (Table 3).

Table 3 Multiple regression analysis before and after adjustment
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We performed an ROC curve to identify the cut-off value of serum 25-hydroxyvitamin D, which may be used to predict primary hypertension. The maximum area under the curve was 0.71 with a 25-hydroxyvitamin D value of 18 ng/ml with a sensitivity of 73.4 (61.7–82.1) and specificity 54.6 (51.4–65.3) (p < 0.0001) (Fig. 1).

Fig. 1
figure 1

Receiver operating characteristic curve of serum 25-hydroxyvitamin D levels in hypertensive subjects to predict a positive outcome

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Discussion

In the present study, a low mean serum 25-hydroxyvitamin D level was significantly more common in hypertensive subjects than in controls (20.5 ± 6.3 ng/mL vs 26.6 ± 9.0 ng/mL) (p < 0.0001), and our findings were in agreement with others [5, 14,15,16,17]. Akbari found a mean serum 25-hydroxyvitamin D value of 24.5 ± 19.4 ng/mL for patients and 15.3 ± 12.2 ng/mL for controls [14]. Ahmad et al. showed mean vitamin D levels of 24.1 ± 16.3 ng/mL in cases and 39.8 ± 11.1 ng/mL in controls [15]. Holick found vitamin D levels of 26.4 ± 4.9 ng/mL in cases and 36 ± 5 ng/mL in controls [16]. Priya et al. found vitamin D levels of 15.15 ± 12.51 ng/mL in cases and 33.59 ± 16.69 ng/mL in controls in her study [5]. Gowda and Khan showed a mean value of vitamin D 19.9 of ng/mL in cases and 32.2 ng/mL in controls [17].

A recent study observed a rate of 50% deficiency in cases and 33.3% deficiency in control subjects [18]. In our study, we found 40.2% deficiency in hypertensive subjects and 27.7% deficiency in controls, which is similar to the findings noted by others [5]. African Americans have a higher prevalence of vitamin D deficiency among hypertension subjects due to lower serum vitamin synthesis due to skin pigmentation [19]. In the current study, we established that serum 25-hydroxyvitamin D deficiency was independently associated with hypertension among the subjects (odds: 1.78; 95% CI: 1.31–2.41); our findings were supported by others [20,21,22,23]. Qi et al. noted the same finding in his study (odds: 1.22; 95% CI: 1.01–1.48) [20]. Forman et al. (odds: 2.67; 95% CI: 1.05–6.79) [20] and Jode et al. also noted similar findings (odds: 1.22; 95% CI: 0.87–1.72) [23]. Pittas et al. established that the risk increased by 6.1 times in men and 2.6 times in women [22].

Several studies have shown possible mechanisms that link vitamin D to hypertension; vitamin D negatively regulates the renin–angiotensin system and is associated with endothelial vasodilator dysfunction [20]. Vitamin D mediates the activation of the renin–angiotensin–aldosterone system and causes hypertension [24]. However, few studies have found that serum 25-hydroxyvitamin D is not significantly associated with hypertension [25, 26].

Systolic BP

In the present study, we found that high mean SBP was significantly associated with 25-hydroxyvitamin D deficiency hypertension 151.8 ± 18.2 mmHg and controls 141.2 ± 10.2 mmHg (p < 0.0001), a similar findings noted by others [18, 27, 28].

Kota et al. demonstrated a mean SBP of 173.7 ± 18.8 mmHg among subjects in his study [27]. Qureshi et al. noted that elevated SBP was significantly associated with vitamin D deficiency [18]. Duprez et al. noted that low levels of serum 25-hydroxyvitamin D were associated with SBP [28]. Randomized studies have explored the effects of calcium with vitamin D 400 IU supplementation over a period of time, and SBP was significantly reduced [18, 29].

Diastolic BP

In the current study, we demonstrated that the mean DBP was significantly associated with serum 25-hydroxyvitamin D in hypertensive subjects (97.4 ± 8.4 mmHg vs 89.5 ± 7.5 mmHg, p < 0.0001). Similar results were reported by Kota et al. (mean 108.1 ± 11.8) [27]. Studies have demonstrated that vitamin D supplementation can reduce DBP [29, 30]. However, a recent study noted no significant reduction in diastolic pressure as a result of vitamin D supplementation [3].

Seasonal variation

In our study, we found that serum 25-hydroxyvitamin D was not significantly associated with hypertension, and these findings were supported by previous studies [6, 7, 9, 10, 12]. However, few studies have found a significant association with seasonal variations [18, 31].

Parathyroid hormone

In the present study, we found that elevated PTH was significantly associated with hypertension and that PTH was not significantly associated with serum 25-hydroxyvitamin D deficiency in hypertensive subjects. Our findings were in agreement with others [26]. In contrast, few studies have noted that high levels of PTH and vitamin D deficiency are independently associated with hypertension [32].

Abnormal transaminase enzyme levels

Elevated liver enzymes (ALT and AST) are indicators of liver disease and the risk of high blood pressure [11, 33]. In the present study, we found that abnormal ALT and AST enzymes were significantly associated with serum 25-hydroxyvitamin D in hypertensive subjects (p = 0.0015, and p = 0.0034). Similar findings were found in other studies. Zelber-Sagi et al. noted that the elevated ALT levels were associated with vitamin D deficiency in the general population [34], and Bahrynian et al. noted in his study that elevated liver enzymes (ALT and AST) were associated with vitamin D deficiency in adolescents with hypertension [35].

The current study established abnormal ALT (odds: 1.2; 95% CI: 0.98–1.94) as being independently associated with hypertension, but the same results were not found for AST (odds: 0.98; 95% CI: 0.59–1.32); these findings are in agreement with others [36]. However, a few studies have noted no significant association between vitamin D and elevated ALT and AST enzymes [37, 38].

C-reactive protein

In the current study, we established that CRP was significantly positively associated with hypertension compared with the CRP levels of normotensive subjects, and our findings were in agreement with others [39,40,41]. Cross-sectional studies have demonstrated elevations in inflammatory markers among individuals with high blood pressure [39, 41]. In the present study, we established CRP positivity as an independent risk factor for hypertension (odds: 1.48; 95% CI: 1.18–2.12), which was supported by previous studies [42].

CRP functions as a pro-atherosclerotic factor by upregulating angiotensin type 1 receptor expression [39]. Hypertension may lead to inflammatory stimuli in the vessel wall and support the protection of proinflammatory cytokines (tumor necrosis factor-α, interleukin-6) [43]. CRP decreases the production of nitric oxide and increases plasminogen activator inhibitor-1 and endothelin-1 activity in endothelial cells, and promotes vasoconstriction, platelet activation, and thrombosis formation. In addition, CRP upregulates angiotensin receptor-1 and increases angiotensin-II activity and increases blood pressure [40]. However, a few studies have found no significant relationship between CRP positivity and hypertension [44].

Pitfalls of study

The strengths of this study include that the laboratory tests of both groups of subjects (hypertensive and normotensive) were performed in one lab. We applied multiple logistic regression analysis before and after adjustment for confounders to identify independent associations and performed ROC curves. The limitations of the study include that we were unable to analyze the presence of metabolic syndrome, the prevalence of postmenopausal women, the estimation of sun exposure, physical activity, and the dietary intake of vitamin D in either group. In our study, we measured aminotransferase levels one time in both groups.

Conclusion

In the present study, we established that low serum 25-hydroxyvitamin D levels and CRP positivity were independently associated with hypertension in south Indian patients. Serum 25-hydroxyvitamin D deficiency was associated with a 1.5-times higher risk, positive CRP was associated with a 1.4-times higher risk, and abnormal ALT was associated with a 1.2-times higher risk for hypertension. Larger multicenter randomized control studies are required to determine the potential role of these findings in hypertension management to decrease the public heath burden.