The role of vitamin D in MS pathogenesis is currently at the focus of multidisciplinary research initiatives. Prior research evaluated the role of vitamin D deficiency as a risk factor for development of MS and as a modifier of its clinical course  as well as of common symptoms of PwMS, such as pain and depression [19, 20]. The interaction between IFN-β and vitamin D in terms of their combined efficacy was also previously studied both clinically [8, 9, 11] and in an animal model , with yet conflicting results.
In the current work we focused on a different aspect of the vitamin D – IFN-β interaction, namely: the effect on FLS, a bothering, sometimes debilitating, adverse-effect of IFN-β treatment. We hypothesized that vitamin D may ameliorate these symptoms, by modulating a post-injection cytokine surge. In the present randomized, double-blind study, vitamin D supplementation did not affect IFN-β-induced FLS. Futility analysis showed that this conclusion was unlikely to change with recruitment of more patients.
The desired 25-OH-D level for the general population is derived from epidemiologic studies of the relationship between this level and the risk for osteoporosis. The controversy in translating the data to practice is reflected in conflicting recommendations of current guidelines. While the American institution of medicine recommends a target of 50 nmol/l , the American society of endocrinology defines levels below 75 nmol/l as insufficient and below 50 nmol/l as deficient . Additionally, since some findings suggest that high dose of vitamin D might have a direct toxic effect , the optimal level range for vitamin D has not yet reached consensus.
Our data show that the high dosage regimen is more effective in achieving 25-OH-D serum level above 75 nmol/l in MS patients. After 6 months, only 25% of the low dose vitamin D group reached serum levels of 75nmo/l or higher, compared to 100% of the high dose group. By the end of the study only 13% of those treated with 800 IU/d compared to 60% of those treated with 4,370 IU/d reached the 75 nmo/l target.
Our patients’ average EDSS score was about 3.5, implying that most of them were not housebound and could have been influenced by outdoor sun exposure. Therefore, the observed decline in 25-OH-D from 6 months to 12 months in both groups can be partly attributed to winter conditions at the 12 month visit. Annual variation in 25-OH-D levels, with decreased levels at winter, has previously been reported in Israel . All patients confirmed their adherence to the study regimen in every phone conversation as well as in each clinic visit. However, attrition in compliance can’t be ruled out and might have contributed to the decreased 25-OH-D levels after 12 months in both dosage groups.
As of today, only few, small scale randomized controlled trials of vitamin D supplementation have been completed, and with conflicting results. Burton et al.  randomized 49 MS patient to no supplementation or escalating vitamin D doses up to 40,000 IU/day over 28 weeks followed by 10,000 IU/day for 12 weeks, thence down-titrated to 0 IU/d until the end of 52 weeks of follow up. Treatment group patients appeared to have fewer relapses and a persistent reduction in T-cell proliferation compared to controls. In another study, 23 patients were randomized to 6000 IU/d vitamin D or placebo. After 6 months follow up no significant differences were detected in various MRI endpoints, and there were more relapses in the high dose group .
Recently, Soilu-Hanninen et al.  reported a 1 year, double blind, placebo controlled, randomized study in 66 MS patients with vitamin D3 as an add-on treatment to IFN-β. Patients in the vitamin D group showed improved MRI outcomes, with no significant differences in clinical outcomes. Similarly, a 96-week randomized controlled trial of 68 patients found that supplementation with 20,000 IU vitamin D(3) weekly did not result in beneficial effects on clinical MS-related outcomes .
The present study is akin to prior trials in duration and sample size. We found no trend of clinical improvement regarding disease activity in PwMS after vitamin D supplementation; however MRI outcomes were not included in this study. Since patients participating in our study were all but one under long-term IFN-β treatment, disease activity as assessed by relapse rate, relapse numbers and EDSS change was as expected relatively low. The study was therefore underpowered to assess the effect of vitamin D on MS activity, and larger scale trials and meta-analysis of available data, including MRI parameters, are needed to clarify the benefit of vitamin D on MS disease activity and/or its symptoms.
IL-17 is a pro-inflammatory cytokine produced by Th-17 cells, and has been advocated to play a role in MS pathogenesis . IL-17 has been found to be highly expressed in MS lesions, and Th17 cells are abundant in active MS lesions . Vitamin D has been shown to decrease Th-17 cell activity in animal models of MS [31, 32], and to down regulate IL-17 production in Th17 cells isolated from MS patients . IFN-β was also shown to inhibit Th17 cell development, and this effect is considered to be one of the mechanisms of actions of IFN-β in the treatment of MS .
Since both vitamin D and IFN-β have been shown to down regulate IL-17 production, we premised that vitamin D supplementation would decrease IL-17 serum levels in our IFN-β treated patients. Unexpectedly, a significant increase in serum IL-17 was found in the low dose group of this study, while non-homogenous trends in IL-17 levels were noted in the high dose group. Due to ethical considerations, this study did not include a pure placebo group, without any kind of vitamin D supplementation. Therefore, it is impossible to determine from this data, whether the demonstrated increased IL17 was due to the low dose supplementation, or the consequence of some other unrecognized confounder. Since accumulating data point towards both vitamin D and IFN-β as inhibitors of Th17, it’s more probable that the increased IL-17 in the low dose group in this study was not due to vitamin D. Notably, baseline measurements were carried out at winter while 3 months tests were done during spring. Seasonal variation in immune cell subsets and cytokines has been previously described, and could have contributed to the variation in serum cytokines in this study [35, 36].
The trend towards decreased IL-17 secretion in an increasing proportion of patients on higher pharmacological doses of vitamin D is in agreement with earlier studies which demonstrated that high dose vitamin D decreases Th-17 cell activity in animal models of MS [31, 32] as well as in PwMS . Beneficial effects of vitamin D increase may be undetectable in IFN-β treated patients, because of the strong immunomodulatory effect of this cytokine , however our results indicate that the add on of large doses of vit D could be beneficial. The question remains whether further increasing vitamin D dose, above 4,370 IU per day, would entail significant IL17 down-regulation in a greater proportion of MS patients.
Furthermore, IL-17 data must be interpreted with caution as serum IL-17 is not an established biomarker of MS disease activity. IL17 serum levels before treatment with IFN−β, as well as 3 months after treatment, could not be correlated to disease activity parameters in a cohort of 118 MS patients , and IL‑17 levels showed a trend towards being higher in MS patients with inactive disease compared to those with active disease in another patient cohort of 169 patients .
No change in serum levels of other cytokines was noted in this work, as well as in a previous study , assessing the impact of vit D supplementation.
Notably, PTH levels decreased significantly in the high dose vitamin D treated group. This change in PTH level was found also in healthy people after vitamin D supplementation . The role of PTH as a potential immune-modulator has been debated. Both B and T lymphocytes express PTH receptors .While some studies imply that PTH may be anti-inflammatory  other studies found that PTH is in fact increased during MS relapse ; still others found no correlation at all between PTH levels and immune cells function . Nevertheless, PTH is a well known stimulator of bone resorption. Its decrement in our patients who received the high dose of vitamin D may imply an advantage, though yet to be confirmed, in terms of bone health and prevention of osteoporosis, which is particularly common and debilitating in PwMS .