Impact of Two Regimens of Vitamin D Supplementation on Calcium - Vitamin D - PTH Axis of Schoolgirls of Delhi

 PII:S097475590900056-1

 

Objective: To determine the efficacy of supplementation with oral vitamin D3 (cholecalciferol) on bone mineral biochemical parameters of school-going girls.

Setting: Government school (government-aided) and Private school (fee paying) in Delhi.

Design: Randomized controlled trial.

Intervention: Cholecalciferol granules (60,000 IU) orally with water, either once in two months (two-monthly D3 group) or once a month (one-monthly D3 group) for one year.

Participants: 290 healthy schoolgirls (6-17 y), 124 from lower socioeconomic strata (LSES) (attending government schools) and 166 from upper socioeconomic strata (USES) (attending private schools).

Outcome measures: Serum 25(OH)D, calcium, phosphorus, parathyroid hormone, and alkaline phosphatase levels at 6 and 12 months after start of supplementation.

Results: At baseline, 93.7% schoolgirls were vitamin D deficient [25(OH)D<50 nmol/L]. While significant increase in serum calcium and decrease in alkaline phosphatase levels was noted in both groups with both interventions, PTH response was inconsistent. In LSES subjects, two-monthly D3 and one-monthly D3 supplementation resulted in a significant increase in serum 25(OH)D levels by 8.3 nmol/L and 11.0 nmol/L, respectively at 6 months (P<0.05). Similarly, the increase in the two intervention arms in USES subjects was 10.5 nmol/L and 16.0 nmol/L, respectively (P<0.05). In both groups, this increase in serum 25(OH)D levels persisted at 12 months (P<0.05). Despite supplementation with 60,000 IU of Vitamin D3 (monthly or two-monthly), only 47% were vitamin D sufficient at the end of one year.

Conclusions: 60,000 IU of cholecalciferol, monthly or two-monthly, resulted in a significant increase in serum 25(OH)D levels in vitamin D deficient schoolgirls.

Key words: Cholecalciferol, India, Supplementation, Schoolgirls, Vitamin D.

Adequate vitamin D status for optimum bone health has received increased recognition in recent years. Vitamin D insufficiency has been reported in healthy children, adolescents and adults worldwide(1), which has been attributed to low vitamin D intake and inadequate sunlight exposure(2,3). Rickets is seen in children with severe vitamin D deficiency, but previous studies have shown that even mild vitamin D insufficiency can have detrimental effects on bone mineral acquisition(4,5) and bone remodeling(6,7) in adolescence. Nutrition guidelines for optimizing bone health in children and adolescents have focused on calcium and exercise, but have neglected vitamin D(8,9). Furthermore, the optimal serum 25(OH)D concentrations for children and adolescents is still a subject of debate. While most professional bodies have recommended a daily vitamin D intake of 10mg(10), the Indian Council of Medical Research has made no recommendation for India, with the view that adequate sunlight exposure would provide the necessary daily vitamin D requirement(11). However, increasing awareness of wide prevalence of vitamin D deficiency in Asian Indian adults(12-17) and children(18-20) has prompted a renewed discussion on the desired vitamin D intake to ensure optimal bone health.

In the absence of results of vitamin D fortification studies, it is not possible to evaluate the merits of food fortification relative to the systemic use of vitamin D containing dietary supplements(21). While several intervention trials have evaluated the role of calcium on the bone health of growing children and adolescents(22,23), information on the effect of vitamin D supplementation remains limited. We planned this study with the primary objective to assess the impact of oral vitamin D₃ (cholecalciferol) supplementation for one year, on bone mineral biochemical parameters, in healthy school-going girls (6-17 years) from two different socioeconomic strata, residing in Delhi, India.

Methods

Subjects

Three hundred and fifty five healthy school girls, aged 6-17 years, who responded to a request to participate, were recruited from two schools of Delhi, which were located in geographic proximity to the principal investigator’s institution. Socioeconomic stratification of the subjects was based on the type of school attended. Girls studying in the government-aided school were considered to represent the lower socioeconomic strata (LSES, n=165), while those enrolled in the fee-paying private school represented the upper socioeconomic strata (USES, n=190). A convenient sample was selected because available information in literature was limited and was based on populations receiving food items fortified with vitamin D and with different baseline 25(OH)D levels. Children with systemic illness, endocrine disorders and drugs affecting bone mineral health were excluded. Each class in a school had 5 sections, and one section from each class was randomly selected. Of the 355 girls screened, 290 girls were enrolled for the intervention after excluding 65 girls. These 290 subjects underwent baseline assessment in summer months (July-August, 2006).

The study protocol was approved by the institutional ethics committee of the Institute of Nuclear Medicine and Allied Sciences (INMAS). A prior written consent for the study was taken from the school administration and from the parents. The details of the study design are shown in Fig 1.

Fig.1 Study design.

Data collection

Anthropometric measurements and dietary information were collected from the subjects at baseline. Standing height was recorded to the nearest 1mm without wearing shoes and by using a wall stadiometer. Weight was recorded to the nearest 0.1kg by using a clinical balance, with the subject wearing light clothing. Body mass index was calculated using the formula weight (in kg) / height (in m2). Every morning, the scale and stadiometer were calibrated with standard weight and height, respectively.

Blood samples were collected from subjects in the fasting state at 0800 hours without venostasis under basal conditions for estimation of serum calcium (Ca), phosphorus (P), alkaline phosphatase (ALP), 25-hydroxyvitamin D [25(OH)D], and immunoreactive parathyroid hormone (PTH) prior to initiating intervention. Serum calcium, phosphorus, and ALP were estimated on the same day, two aliquots were stored at –20ºC until PTH and 25(OH)D were estimated. Serum calcium (Randox Laboratory Ltd, Crumlin, UK) and phosphorus (Clonital; Ampli Medical SPA, Milan, Italy) were measured by colorimetric method and ALP by liquid kinetic method (Clonital; Ampli Medical SPA, Milan, Italy). Normal laboratory range for serum calcium was 2.02-2.60 mmol/L (8.10-10.40 mg/dL) and for serum phosphorus in adults was 0.81-1.55 mmol/L (2.5-4.8 mg/dL), according to the kit manufacturers. The reported upper limit of serum P in mid-childhood is 1.87 mmol/L (5.8 mg/dL)(28). Normal laboratory range for serum ALP at 37ºC was 100-275 IU/L in adults and 180-1200 IU/L in children before epiphyseal closure.

Supplementation was initiated in the winter (December, 2006) of the same year for all subjects and was carried out for a period of one year. The subjects were randomized to two intervention groups receiving either 60,000 IU of cholecalciferol granules (Calcirol, Cadila Pharmaceuticals, Ahmedabad) every two months (two-monthly D3 group; equivalent to 1000 IU/day) or every month (one-monthly D3 group; equivalent to 2000 IU/day), for a period of one year. These doses represent 25% and 50%, respectively of the lowest adult dose considered likely to cause adverse effects(29).

Statistical analysis was carried out using STATA 9.0 (College Station, Texas, U.S.A.). Baseline data are presented as mean (SD) or number (percentage) as appropriate. Effect of vitamin D3 supplementation (once in two months and once a month) on serum Ca, P, ALP, 25(OH)D and PTH was analyzed using generalized estimating equation (GEE). P values <0.05 were considered significant.

TABLE I



Baseline Anthropometric and Dietary Parameters of the Study Subjects [Mean(SD)]

LSES: Lower socioeconomic strata; USES: Upper socioeconomic strata; BMI: Body Mass Index; 

*Height and weight Z scores were calculated based on the reference values provided by Agarwal, et al.(24).

TABLE II



Effect of Vitamin D supplementation on Serum Calcium, Phosphorous, Alkaline Phosphatase, 

25(OH)D and Parathyroid Hormone

LSES: Lower socioeconomic strata; USES: Upper socioeconomic strata; Two-monthly D3 group: Vitamin D (60,000 IU) 

once in two months; One-monthly D3 group: Vitamin D (60,000IU) once a month; 

* = P<0.05 for baseline vs. 6 month;   # = P<0.05 for baseline vs. 12 month; $ = P<0.05 for 6 months vs. 12 month.

In the study population, 93.7% girls (97.5% vs. 90.9% in LSES and USES, respectively) were found to be vitamin D deficient at baseline. In LSES group, this prevalence declined from 98% to 74% to 38% in two-monthly D₃ group and from 97% to 69% to 28% in one-monthly D₃ group at 6 and 12 months post-supplementation, respectively. However, in USES strata, the percentage of vitamin D deficient girls decreased from 94% to 84% to 80% in two-monthly D3 group and 88% to 68% to 57% in one-monthly D₃ group at 6 and 12 months of supplementation, respectively.

As reviewed recently, most experts agree that in the absence of adequate sun exposure, children and adults need 800-1000 IU of vitamin D/day(1). Since data from several workers in India suggest a high prevalence of vitamin D deficiency(12-21), we considered evaluating two dose strengths; namely, the equivalent of 1000 units/day and 2000 units/day. Oral vitamin D3 supplementation with a dose equivalent to 1000-2000 units/day for 1 year was safe, and increased serum 25(OH)D concentrations significantly in both upper and lower socio-economic strata. However, despite the supplementation, only 47% of schoolgirls studied became vitamin D sufficient. The mode and frequency of administration of vitamin D₃, and the extent and possible chronicity of hypovitaminosis D in the study population may in part explain this observation.

Two important observations need to be highlighted. It is an established fact that serum 25(OH)D levels in volunteers in Delhi are lower when measured in winter as compared with summer(17). This anticipated decline in serum 25(OH) D levels during winter was overcome by the vitamin D supplementation, especially in the LSES group. Secondly, monthly supplementation (equivalent to 2000 IU D₃/day) was superior to two-monthly supplementation (equivalent to 1000 IU D₃/day), though this effect was significant only in the USES.

The superiority of a higher dose of vitamin D was also reported by Fuleihan, et al.(34) and Maalouf, et al.(36) comparing 200 IU vs. 2000 IU of vitamin D/day, and by Viljakainen, et al.(35), comparing 200 IU vs. 400 IU of vitamin D per day. Maalouf, et al.(36) reported that in children and adolescents with serum 25(OH)D concentration below 20 ng/mL (i.e. <50 nmol/L), a vitamin D₃ dose equivalent to 2000 IU/day resulted in desirable vitamin D levels. In a study assessing only winter time vitamin D₂ supplementation in Finnish girls, prevalence of hypovitaminosis D was significantly reduced in those receiving vitamin D(4). The supplementation had more effect on those with severe hypovitaminosis D than those with normal vitamin D levels at baseline. In a similar study assessing the impact of winter time supplementation of vitamin D3 in French male adolescents, Guillemant, et al.(31), reported that a monthly dose of 50,000 IU was adequate to prevent the decline in serum 25(OH)D levels usually observed in winter.

The increment in serum 25(OH)D levels in response to monthly D3 supplementation in the present study was significantly less than that reported by Maalouf, et al.(36), using a comparable dose of D₃. The possible reasons for the differences in outcome between the two studies could be as follows: use of oil containing vitamin D₃ preparation by Maalouf, et al.(36), in contrast to the direct ingestion of granulated vitamin D₃ followed by water in our study; weekly delivery versus monthly delivery of vitamin D₃; and, differences in the mean baseline calcium intake and severity of hypovitaminosis D in the study population.

Vitamin D supplementation resulted in a significant increase in serum calcium in both intervention arms (two-monthly and one-monthly D₃ groups) of both socioeconomic strata. This has also been reported by other workers(33,35). Similarly, both interventions resulted in a decline in ALP in both SES, with the maximum decline observed at 6 months in both groups. However, Maalouf, et al.(36) also showed a significant reduction in ALP at 12 months with both low (200 IU/day) and high (2000 IU/day) doses of vitamin D₃ but the reduction at 6 months was not significant. In contrast, Rajakumar, et al.(33) showed decline in ALP with a lower vitamin D dose of 400 IU/day, after one month of intervention.

Certain limitations of the present study need to be highlighted. Firstly, due to constraints in the study conditions, the cholecalciferol granules had to be given to the subjects with water instead of milk or an oily preparation. Secondly, due to lack of permission from the school authorities, pubertal staging could not be carried out. Thirdly, due to procedural delay, the intervention was initiated in the winter, while the baseline assessment was performed in summer months. Finally, since serum albumin was not measured, we have not provided corrected values of serum calcium. Thus, in the absence of food fortified with vitamin D, monthly supplementation providing the equivalent of 2000 IU of Vitamin D₃/day may be the preferred approach to combat the prevalence of hypovitaminosis D in Indian school girls.

40. Guillemant J, Cabrol S, Allemandou A, Peres G, Guillemant S. Vitamin D-dependent seasonal variation in growing male adolescents. Bone 1995; 17: 513-516.