Vitamin D Status and Supplementation in Clinically Isolated Syndrome: Implications for Disease Progression and Neuroimaging Outcomes. A Systematic Review

ABSTRACT Clinically Isolated Syndrome (CIS) represents the first neurological episode of inflammation and demyelination in the central nervous system and often precedes multiple sclerosis (MS). Vitamin D deficiency is common in CIS and has been implicated as a potential environmental risk factor influencing disease progression. Through its neuroprotective and immunomodulatory effects, vitamin D may affect lesion development, relapse frequency, and disability outcomes. However, findings from randomized controlled trials (RCTs) on vitamin D supplementation in CIS remain inconsistent. This systematic review evaluated RCT evidence on the effects of vitamin D supplementation compared with placebo or standard care in adults with CIS. Primary outcomes included gadolinium-enhancing and T2-weighted MRI lesions, relapse rate, Expanded Disability Status Scale (EDSS) progression, and conversion to clinically definite MS (CDMS). Four RCTs met the inclusion criteria. Two trials demonstrated a reduction in gadolinium-enhancing lesions in the vitamin D group, while others showed no significant difference. For T2 lesions, one trial reported fewer new lesions, but pooled continuous data revealed no statistically significant effect. Relapse rates, EDSS progression, and composite MRI activity were largely unchanged between groups. Subgroup analyses indicated potential benefit among patients with severe baseline vitamin D deficiency (<30 nmol/L), whereas achieved serum vitamin D levels after supplementation showed no consistent correlation with MS progression. Overall, current RCT evidence does not consistently support vitamin D supplementation as an effective intervention for CIS, underscoring the need for large, standardized trials to clarify its therapeutic role in delaying conversion to multiple sclerosis. Vitamin D Status and Supplementation in Clinically Isolated Syndrome: Implications for Disease Progression and Neuroimaging Outcomes. A Systematic Review Running Title: Impact of Vitamin D on Clinically Isolated Syndrome Outcomes Yashfeen Amjad (1), Menahil Faheem Siddiqui (2), Muhammad Owais (3), Salwa Bashir (4), Adan Ali (5), Muhammad Huzaifa Sabir (6), Faraz Ahmed (7), Azeen Asghar (8), Wajiha Altaf (9), Asma Azam (10) . Affiliations: 1,4,5,8,9 Allama Iqbal Medical College, Lahore, Pakistan. 2,10 Karachi Medical and Dental College, Karachi, Pakistan. 3,6 Faisalabad Medical University, Faisalabad, Pakistan. 7 Tahir Heart Institute, Chenab Nagar, Pakistan. Authors and Affiliations 1. Yashfeen Amjad, Allama Iqbal Medical College, Lahore, Pakistan Email: [email protected] ORCID: 0009-0003-7333-5418 2. Menahil Faheem Siddiqui Karachi Medical and Dental College, North Nazimabad, Karachi 74700, Pakistan Email: [email protected] ORCID: 0009-0002-5484-4543 3. Muhammad Owais Faisalabad Medical University, Faisalabad, Pakistan Email: [email protected] ORCID: 0009-0009-5639-5682 4. Salwa Bashir Allama Iqbal Medical College, Lahore, Pakistan Email: [email protected] 5. Adan Ali Allama Iqbal Medical College, Lahore, Pakistan Email: [email protected] 6. Muhammad Huzaifa Sabir Faisalabad Medical University, Faisalabad, Pakistan . Email: [email protected] ORCID: 0009-0004-7796-6361 7. Faraz Ahmed Chenab Nagar, Pakistan Email: [email protected] ORCID: 0009-0001-1860-7148 8. Azeen Asghar Allama Iqbal Medical College, Lahore, Pakistan Email: [email protected] 9. Wajiha Altaf Allama Iqbal Medical College, Lahore, Pakistan Email: [email protected] 10. Asma Azam Karachi Medical and Dental College, North Nazimabad, Karachi 74700, Pakistan Email: [email protected] ORCID: 0009-0001-3592-7493 Corresponding Author Menahil Faheem Siddiqui Karachi Medical and Dental College, North Nazimabad, Karachi 74700, Pakistan Email: [email protected] Phone: +92 332 5395050 ORCID: 0009-0002-5484-4543 Prospero Registration Details: Date: 8 th August 2025 Registration Number: CRD420251021810 Ethics Approval and Consent to Participate: Not applicable. Consent for Publication: Not applicable. Competing Interests: The authors declare no potential conflicts of interest concerning the research, authorship, and/or publication of this article. Funding: The authors received no extramural funding for this study. Authors’ Contributions: All authors contributed substantially to study conception, design, data acquisition, analysis, and interpretation. All participated in drafting or revising the manuscript, approved the final version for publication, and agree to be accountable for all aspects of the work. Availability of Data and Materials: Data supporting this study are available within the article. Extracted datasets are available from the corresponding author upon reasonable request. Acknowledgments: None to declare.

Clinically Isolated Syndrome (CIS) represents the first neurological episode of inflammation and demyelination in the central nervous system and often precedes multiple sclerosis (MS). Vitamin D deficiency is common in CIS and has been implicated as a potential environmental risk factor influencing disease progression. Through its neuroprotective and immunomodulatory effects, vitamin D may affect lesion development, relapse frequency, and disability outcomes. However, findings from randomized controlled trials (RCTs) on vitamin D supplementation in CIS remain inconsistent. This systematic review evaluated RCT evidence on the effects of vitamin D supplementation compared with placebo or standard care in adults with CIS. Primary outcomes included gadolinium-enhancing and T2-weighted MRI lesions, relapse rate, Expanded Disability Status Scale (EDSS) progression, and conversion to clinically definite MS (CDMS). Four RCTs met the inclusion criteria. Two trials demonstrated a reduction in gadolinium-enhancing lesions in the vitamin D group, while others showed no significant difference. For T2 lesions, one trial reported fewer new lesions, but pooled continuous data revealed no statistically significant effect. Relapse rates, EDSS progression, and composite MRI activity were largely unchanged between groups. Subgroup analyses indicated potential benefit among patients with severe baseline vitamin D deficiency (<30 nmol/L), whereas achieved serum vitamin D levels after supplementation showed no consistent correlation with MS progression. Overall, current RCT evidence does not consistently support vitamin D supplementation as an effective intervention for CIS, underscoring the need for large, standardized trials to clarify its therapeutic role in delaying conversion to multiple sclerosis.

Clinically Isolated Syndrome, Vitamin D, Multiple Sclerosis, MRI lesions, Relapse, Disability progression, Systematic review.

Clinically Isolated Syndrome (CIS) is the first episode of neurological symptoms caused by inflammation and demyelination in the central nervous system (CNS) that lasts at least 24 hours and does not occur with infection or fever, in individuals without prior diagnoses of multiple sclerosis (MS) [1]. CIS typically affects young adults between 20 and 40 years of age and commonly presents with optic neuritis, focal supratentorial deficits, brainstem or cerebellar dysfunction, or partial transverse myelitis [2]. Vitamin D deficiency, defined as serum 25-hydroxyvitamin D levels below 50 nmol/L, impacts approximately 58% of the global population and is particularly prevalent among those diagnosed with CIS [3]. Vitamin D has become a well-recognized environmental risk factor for multiple sclerosis (MS), particularly among CIS patients whose levels fall well below the protective threshold [4]. Vitamin D plays a neuroprotective and immunomodulatory role through receptors located throughout brain regions including the hippocampus and cortex, where it regulates neurotrophic factors and reduces neuroinflammation [5]. Vitamin D deficiency affects approximately 68% of South Asian adults, with women being most heavily affected compared to men at 76% versus 51% respectively. Pakistan shows the highest prevalence among all South Asian countries at 73% due to sun avoidance practices and cultural norms [6]. Around 52% of patients diagnosed with CIS had vitamin D deficiency and those in the lowest range had more than a threefold greater risk of progressing to MS (4). CIS such as unilateral optic neuritis, brainstem dysfunction, or partial myelitis typically represent the early presentation in approximately 85% of patients who eventually develop relapsing and remitting multiple sclerosis (MS). These initial neurological events may serve as important diagnostic clues, serving as early indications of MS [7]. Brain MRI at the time of CIS presentation serves as a powerful prognostic tool, with longitudinal studies showing conversion rates to clinically definite MS (CDMS) of 56% to 82% in patients with detectable MRI lesions, compared to just 20% in those with normal imaging. Notably, 60% to 80% of CIS cases exhibiting visible brain lesions on MRI progress to CDMS which emphasizing the critical importance of neuroimaging in risk stratification and early diagnosis [7,8]. MS once considered rare in Pakistan, now affects approximately 10 individuals per 100,000 with the Global Burden of Disease study reporting 29,780 prevalent cases in 2016 and female patients predominate 1.6:1, which conforms with global and regional trends [9]. Existing research surrounding vitamin D supplementation and CIS shows varying findings, with various clinical trials yielding different conclusions. While studies such as D-Lay MS trial [10] may demonstrate potential benefits in terms of reduced disease activity, others, such as O’Connell [11] study have failed to demonstrate clinical or radiological improvements; moreover trials such as EVIDIMS [12] did not show substantial therapeutic effects, and Derakhshandi’s [13] study suggest a possible delay in MS conversion among vitamin D deficient patients, while Butzkueven’s [14] findings did not support any benefit in high-risk CIS cases, further underscoring its complexity and variability associated with its intervention. Due to its importance, no comprehensive systematic review on vitamin D supplementation for people in CIS stage has yet been conducted. This gap in literature underscores the necessity of conducting such a review that synthesizes RCT evidence focused on clarifying vitamin D’s role in preventing conversion from CIS to clinically definite multiple sclerosis (CDMS). This systematic review aims to provide important information by focusing on CIS as an early intervention point, this systematic review seeks to offer valuable insight into how vitamin D impacts MS conversion rates, lesion progression rates, relapse rates and disability ratings using Expanded Disability Status Scale (EDSS). This research could prove especially relevant in regions with both MS prevalence and vitamin D deficiency coexisting, offering important implications for public health strategies as well as preventive care among individuals at risk of MS. Addressing gaps will ultimately result in more effective management protocols as well as better patient outcomes overall for patients living with CIS.

The systematic review was conducted according to the PRISMA 2020 (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. Its protocol was registered in PROSPERO under the registration number Data Sources and Search Strategy An extensive and thorough literature search was carried out on the following databases: PubMed, Google Scholar, MEDLINE and Cochrane Library. The search included the following keywords and Medical Subject Headings (MeSH): (”Vitamin D”[Mesh] OR ”Cholecalciferol”[Mesh]) AND (Clinically Isolated Syndrome OR ”Demyelinating Diseases”[Mesh] OR CIS) AND (MRI OR ”Magnetic Resonance Imaging”[Mesh]). No restrictions regarding race, place, sex and language were applied. Moreover, we conducted a manual search of the reference lists of the included research, as well as related meta-analyses and review articles, to identify relevant studies. Study Selection The eligibility criteria were randomized controlled trials (RCTs) with a target population of adults (≥18 years) diagnosed with Clinically Isolated Syndrome (CIS) and suggestive of Multiple Sclerosis (MS) including first demyelinating events. The intervention was oral vitamin D3 (cholecalciferol) supplementation while the comparator was placebo. Patients were excluded if they were not diagnosed with CIS, were vitamin D metabolism, suffered from renal dysfunction, were on treatment with specific medications, were unable to undergo MRI or adhere to protocols, or had definite Multiple Sclerosis at baseline. Studies other than RCTs such as case reports, opinion articles, narrative reviews, meta-analyses or systematic reviews, editorials or comments, animal studies, in-vitro studies and grey literature were also excluded. The study selection criteria is shown in Table 1. Table 1. Study Selection Criteria. Data Extraction At first, duplicate studies were removed from the list through Rayyan Software. All reviewers then carefully assessed the remaining articles, and only those that met the previously stated eligibility criteria were included. All the articles were initially screened based on the title and abstract, followed by full text. A third reviewer was consulted to resolve any disagreements regarding the results. A Microsoft Excel Spreadsheet was created online using data from the completed RCTs for baseline characteristics and outcomes. Following are the baseline parameters: age, gender, BMI (kg/m 2 ), baseline serum vitamin D, number of T2 and gadolinium enhancing lesions, infratentorial lesions, smoking status and steroids given at baseline. The following primary and secondary outcomes were included: Primary Outcomes MRI activity, T2 lesions. Secondary Outcomes Gadolinium enhancing lesions, Conversion to clinically definite multiple sclerosis (CDMS), Change in Expanded Disability Status Scale (EDSS) score, Relapse rate, Disease activity, Serum Vitamin D levels. Quality Assessment One author assessed the literature quality and bias of all four clinical trials using the Revised Cochrane Risk of Bias tool for randomized control trials (ROB-2). Studies were analysed for the randomization process, deviations from intended interventions, measurement of outcome, missing outcome data and selective reporting. The risk of bias was classified as unclear, high or low. Grading of Recommendation, Assessment, Development and Evaluation (GRADEpro) was used to evaluate the studies according to international methodology. ROB Assessment All 4 RCT studies included in the review were assessed for bias risk using the Revised Cochrane Risk of Bias tool for randomized control trials (ROB-2). The results shown that all studies have a low risk of bias for the randomization process. However, two studies exhibited a high risk of bias due to deviations from the intended interventions and missing outcome data, resulting in an overall high risk of bias (low quality) for these studies. One study demonstrated a low risk of bias across most domains but raised some concerns regarding the selection of reported result and the remaining one study was rated as low risk of bias across all domains, resulting in an overall low risk of bias (high quality). The detailed result was presented in figure; The detailed results are presented in Figure 1. Data Synthesis We were unable to perform a meta-analysis of all primary outcomes due to the unavailability of complete and consistent data in the included studies. While MRI activity and T2 lesions were reported in multiple trials, significant variations in study designs, outcome definitions, and data reporting methods limited the feasibility of statistical pooling. Moreover, differences in patient populations, heterogeneous dosing regimens and different follow-up times further constrained direct comparisons. Given these limitations, we conducted a systematic review of randomized controlled trials (RCTs) to qualitatively synthesize the available evidence on effect of vitamin D supplementation on disease progression and MRI in patients with Clinically Isolated Syndrome (CIS). However, forest plots were generated for the visual display of safety outcomes by using Review Manager (version 5.4, Copenhagen: Nordic Cochrane Center, The Cochrane Collaboration, 2014). These outcomes were displayed as risk ratios (RR) with a 95% confidence interval using the random effects model.

Study Selection By comprehensive literature search, we retrieved 241 articles across multiple databases. 27 duplicate articles were removed before screening, resulting in 211 unique articles which were screened for relevance based on titles and abstracts. Of these, 197 articles were excluded due to not meeting the predefined inclusion criteria. The remaining 14 articles were subjected to full text assessment for eligibility. No reports were excluded due to retrieval issues. Following full text evaluation, 5 studies were excluded for wrong intervention, 2 for inappropriate publication, 2 for wrong population and 1 for irrelevant outcome measures. Ultimately, 4 studies met all eligibility criteria and were included in the final qualitative synthesis. (The detailed PRISMA flow chart is in figure). Study Characteristics The four studies included in this systematic review were randomized controlled trials (RCTs), published between 2013 and 2025, and conducted in Iran, Belgium, Ireland, and France. A total of 599 participants were enrolled, with individual study sample sizes ranging from 29 to 309 patients. The follow-up duration varied between 12 and 24 months. Participants were predominantly aged 20–50 years and included both male and female patients, although the proportion of females was higher in most studies. Baseline characteristics reported across studies are given in Table. OUTCOMES Gadolinium enhancing lesions Three out of the four studies included in this systematic review reported outcomes relative to gadolinium (GD)-enhancing lesions after vitamin D supplementation in CIS patients. When compared to placebo, vitamin D proved to be effective in reducing the occurrence of new lesions as reported by Derakhshandi et al. (vitamin D = 0 ± 0; placebo = 1.36 ± 2.24) after 50,000 IU/week supplementation for 12 months.Thouvenot et al. also reported fewer events in the vitamin D group (29/156) compared to placebo (50/147) after 20,000 IU/2weeks for 24 months. On the contrary, O’ Connell et al. found no significant difference between the two arms on follow-up MRI (10,000 IU = 0; 5,000 IU = 0.1 ± 0.3; placebo = 0.1 ± 0.4). T2-Lesions All four studies documented the effect of vitamin D supplementation on T2-weighted lesion counts in patients with CIS. Since reporting formats varied, two studies, O’Connell et al. and Derakhshandi et al., that provided continuous outcome data as mean and standard deviation (SD), were quantitatively synthesized. The pooled mean difference (MD) using a random-effects model was −0.67 (95% CI: −2.86 to 1.51, p = 0.55), indicating statistically insignificant difference between vitamin D and placebo in the measured outcome. After combining the two vitamin D intervention arms (5000 UI/day and 10,000 UI/day), O’Connell reported a mean difference of 0.27 (95% CI: −0.56 to 1.10), while Derakhshandi et al. reported a mean difference of −1.99 (95% CI: −3.98 to 0.00). Diversity between studies was suggested by the significant heterogeneity (I2 = 76%) and the statistical significance (p = 0.04) of the Chi2 test. The overall Z-test (Z = 0.60, p = 0.55) and the wide confidence intervals indicate considerable uncertainty in the true effect estimate. Thouvenot et al., based on new enhancing lesion occurrence, found that such lesions were observed less frequently in the vitamin D group than in placebo (72/156 [46.2%] vs 87/147 [59.2%]), corresponding to a hazard ratio of 0.61 (95% CI: 0.44–0.84, p = 0.003). In contrast, Butzkueven et al. reported that the number of baseline cerebral MRI T2 lesions was not associated with conversion to clinically definite or radiological MS (p = 0.08), without providing a relative effect measure for the vitamin D versus placebo comparison. A Forest plot is shown in Figure 2 . Relapse Figure shows a forest plot evaluating the difference in the effects of vitamin D supplementation versus placebo on relapse risk in patients with clinically isolated syndrome (CIS). The pooled RR using a random-effects model is 0.78 (95% CI: 0.44–1.38, p = 0.39), indicating no statistically significant difference in relapse risk between vitamin D and placebo. The heterogeneity analysis shows low variability (I² = 4%), and the Chi² test (p = 0.37) does not indicate significant heterogeneity. The wide confidence intervals and the pooled estimate crossing the null line (RR = 1) indicate considerable uncertainty in the true effect size. With an overall Z-test of 0.86 (p = 0.39), these findings do not provide statistically enough evidence that vitamin D supplementation reduces recurrent risk compared with placebo. A Forest plot is shown in Figure 3 . EDSS Progression of disability was checked using the Expanded Disability Status Scale (EDSS) in the Thouvenot trial. The annual change in EDSS was −0.17 (SE = 0.05) in the vitamin D group and −0.14 (SE = 0.06) in the placebo group. However, the difference was not significant, and the findings do not support a measurable effect of vitamin D supplementation on disability. MRI activity Three studies reported overall MRI-based outcomes beyond individual lesion counts. Two of these studies, Thouvenot et al. and O’Connell et al., explicitly reported MRI activity as a composite imaging outcome. In O’Connell et al., the two vitamin D dosage arms (10,000 IU/day and 5,000 IU/day) were combined for analysis. Pooled analysis of these two studies using a random-effects model yielded an RR of 0.88 (95% CI: 0.74–1.05, p =0 .17), indicating no statistically significant difference in MRI activity between vitamin D supplementation and control. Thouvenot et al. reported a RR of 0.87 (95% CI: 0.73–1.05), while O’Connell et al., in which the two vitamin D intervention arms were combined for analysis, reported a RR of 1.23 (95% CI: 0.47–3.19). The heterogeneity analysis shows no observed variability (I² = 0%), suggesting consistency between studies, and the Chi² test (p = .49) further supports this finding. With an overall Z-test of 1.45 (p = .15), the results do not provide statistically significant evidence that vitamin D supplementation reduces MRI activity compared with placebo.The third study, Butzkueven, reported radiological conversion to MS, defined as conversion from CIS to MS based on MRI criteria [10,000 IU/day = 21 (42%), 5000 IU/day = 24 (47%), placebo = 17 (34%)]. This was included under the MRI activity outcome due to its reliance on imaging. A Forest plot is shown in Figure 4 . Effect of Vitamin D levels and Supplementation on Disease Progression The results of two of the studies highlighted the role of vitamin D levels, both at baseline and attained serum levels after supplementation in modulating disease activity in CIS patients. O’Connell et al., after adjusting for baseline serum 25(OH)D level, and comparing CIS patients with a serum 25(OH)D level of 100 nmol/l following supplementation recorded a freedom rate of 50% (7/14) versus 67% (8/12),although the difference was not statistically significant (p = 0.4). This shows that the increased vitamin D levels after supplementation are associated with decreased progression to multiple sclerosis. The second study, Thouvenot et al., demonstrated the significance of baseline serum vitamin D deficiency and reported that patients with severe deficiency, defined by levels <30 nmol/L, observed the greatest benefit of supplementation. Analysis revealed a significant interaction between treatment effect and baseline vitamin D status, with severely deficient patients responding more favorably (HR, 0.33 vs 0.78; P value for interaction = .03) In contrast to the findings of Thouvenot et al, Butzkueven et al. did not find evidence of a protective effect of supplementation at any dose level in reducing the risk of conversion to MS [1000 IU/day 0.79 (0.44, 1.41), 5000 IU/day 1.15 (0.67, 1.99), 10 000 IU/day 1.06 (0.61, 1.83)]. There was no overall effect of the four treatments, (P = 0.60.) All these outcomes are summarized in Table 2. Table 2. Outcomes Reported in Included Studies.

In patients with clinically isolated syndrome (CIS) which is the first demyelinating event which may progress to Multiple sclerosis (MS), vitamin D deficiency was frequently observed. Lesion formation, relapse risk, and disability progression in CIS might be influenced by vitamin D owing to its role in neuroprotection [15] and immune regulation [16] as suggested by prior research. Vitamin D’s possible function in the delay or prevention of CIS progression is underscored by several immunological and neurobiological mechanisms. It enhances regulatory T-cell function and suppresses pro-inflammatory responses by Th1 and Th17 cells involved in demyelination, and by vitamin D receptors in the hippocampus and cortex, it modulates neurotrophic factors and restricts oxidative damage. Moreover, epidemiological data also indicate that vitamin D sufficiency can lower the risk for EBV reactivation as a suspected precipitant of demyelinating events [17], and experimental models point towards the fact that it is increasing the stability of blood-brain barrier, which can curtail immune cell entry into the CNS. Collectively, these mechanisms offer a biologically coherent explanation for supplementation among high-risk CIS individuals, especially those with extreme deficiency. However, results from randomized controlled trials are inconsistent. Some trials indicate benefits seen in radiological studies, while others report no notable clinical impact. The rationale of this systematic review is to clarify these uncertainties and improve understanding of vitamin D’s possible role in CIS. Prior systematic reviews were based on the efficacy of Vitamin D on Multiple Sclerosis (MS) outcomes like relapse rate, magnetic resonance imaging activity, and disability development. Langlois and Denimal (2023) performed a systematic review to estimate the impact of vitamin D supplementation on clinical and imaging findings in patients with multiple sclerosis (MS) [18]. They examined 19 trials and concluded that, while vitamin D supplements had no effect on relapse frequency or disability worsening (based on the Expanded Disability Status Scale), it was associated with a reduction in new MRI lesions. The results indicate that vitamin D supplementation may have an effect on disease activity at the subclinical level and that further studies are needed to explore its potential as a treatment for MS. Conversely, our review specifically addresses Clinically Isolated Syndrome (CIS) the first detectable stage of CNS demyelination. CIS is an important therapeutic window in which modifiable risk factors may be able to change the course towards MS. All the included studies reported heterogenous data with inconsistent dosage of vitamin D supplementation although the intervention and control group were similar i.e. Vitamin D supplemented group and placebo group. Additionally, Vitamin D was supplemented orally. Table: Study Characteristics: Intervention, Control, and Duration. The findings from this systematic review state that patients with CIS after providing supplementation of vitamin D showed positive outcomes in reducing Gadolinium (GD) enhancing lesions and occurrence of new lesions when compared to placebo. Although variation of vitamin D supplementation was seen, 50,000 IU/week showed reduced effects when used for 12 months and 100,000 IU/2weeks when used for 24 months. One study also found no difference between 10,000IU/day and 20,000IU/day supplements when compared with a placebo. Similarly, the outcome of T2 lesions varied in all studies included. One study, which combined 5,000 IU/ day with 10,000 IU/day and two other studies which directly compared Vitamin D supplementation with placebo, did not show statistically significant results in reducing these lesions. However, one study showed a decrease in the occurrence of T2 lesions in the intervention group as compared to the placebo. The outcome of relapse risk of CIS in patients when compared with placebo did not show statistically significant evidence, as the heterogeneity test showed a low score and wide confidence intervals showed uncertain effect size; therefore, placebo and vitamin D supplementation did not show relative positive outcomes. Additionally, one study discussed the disability progression outcome; the annual data reported did not show a significant difference in disability progression between placebo and vitamin D supplementation. Hence, disability progression was not influenced by Vitamin D supplementation. Furthermore, the disease progression of CIS to Multiple sclerosis is found to be influenced by Vitamin D supplementation. Three of the included studies showed that when vitamin D administered in patients with baseline deficiency showed positive outcomes for reducing disease progression to MS after supplementation of vitamin D. Whereas, one of the included studying showed contradicting results as all doses (1000IU/day, 5000IU/day, 10,000IU/day and 20,000IU/day) did not show reduce overall outcomes in conversion to MS. Despite of these positive outcomes results remain inconsistent due to lack of statistically significant data reported owing to the heterogeneity of data reported in all studies. Lastly, the MRI-based outcomes were discussed by combining two supplementations of vitamin D (5000 IU/day and 10,000 IU/day) compared with placebo in one study, which showed no statistically significant difference between the intervention and control group (placebo group). Another included study, which used 20,000 IU/day, also showed no significant difference between the groups. The third study, although it does not explicitly discuss MRI-based outcomes, showed conversion of CIS to Multiple Sclerosis (MS). This study included 5000 IU/day and 10,000 IU/day as the intervention and a placebo as the control group. The strengths of this study included vitamin D supplementation as 1000 IU, 5000 IU, 10,000 IU and 20,000 IU per day. Due to the heterogeneity of the data reported across the studies a meta-analysis was not performed but comparative data analysis was done among the outcomes which reported similar data and units. Additionally, vitamin D supplementation was compared with CIS but the studies that reported conversion of CIS to MS data was also considered. Furthermore, no specific age and population were considered so this review provides diverse collective outcomes and a broad analysis was done. Particularly in those regions where CIS is prevalent and vitamin D deficiency is common; the findings of this review have real relevance for public health as well as clinical treatment. Although overall data on vitamin D supplementation are unclear, the advantages observed in very deficient individuals emphasize the need for regular screening of serum vitamin D levels in CIS patients so that more focused and individualized treatments may be applied. Knowing that usual dosage regimens and long-term efficacy are still unknown, doctors may start treating deficiency early in the course of the disease to maximize neuroprotection and maybe lower the risk of acquiring clinically proven multiple sclerosis. At a public health level, these results highlight a requirement for awareness campaigns, prevention programs, and legislation supporting adequate vitamin D consumption among sensitive populations, therefore complementing MS prevention efforts, lowering the disease burden, and improving long-term neurological outcomes among susceptible groups.

In conclusion, it is still up for consideration if vitamin D supplementation has any significant role in Clinically Isolated Syndrome. Conflicting results from various dose regimens, outcome measures, and follow-up lengths in evidence from recent randomized trials show limitations in both quality and quantity. While this contradiction makes it challenging to reach definitive conclusions, extensive multicenter studies with consistent clinical and radiological outcomes, longer follow-up, and careful examination of effect modifiers including baseline vitamin D levels, heredity, and environmental exposures will be essential. Vitamin D should be seen as a safe and helpful complement to general management rather than a proven disease-modifying intervention in CIS until such evidence becomes available.

One of the major limitations is that there are a limited number of studies eligible for inclusion, which limits the external validity of the results. The studies included were highly heterogeneous in intervention, dosages of vitamin D, outcome measures, and follow-up times, which made quantitative synthesis challenging and created uncertainty in pooled effect estimates. Extra variation emerged from variations in baseline vitamin D status, sample sizes, and geographic populations, making direct comparisons between studies difficult. Future research Future studies should center on resolving the contradictions and inconsistencies discovered in the current vitamin D supplementation data for CIS. Longer follow-up periods, well-defined clinical and radiological results, and standardized dose regimens from large, multicenter randomized controlled trials will help to define the efficacy and best treatment plans. Moreover, in order to identify the subgroups most likely to profit, more study on how baseline vitamin D levels interact with genetic predisposition and environmental exposures is required. Furthermore, providing mechanistic understanding of how vitamin D influences the course of the disease, studies looking at indicators of neuroinflammation and neurodegeneration could be a beneficial insight in more understanding of the progression of disease. These will help guide customized treatment plans and give more weight to the evidence base while also informing future clinical guidelines for MS prevention and CIS treatment.

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Authors Metrics & Citations Metrics Article Usage 181views 103downloads Citations Download citation Yashfeen Amjad, Menahil Faheem Siddiqui, Muhammad Younas, et al. Vitamin D Status and Supplementation in Clinically Isolated Syndrome: Implications for Disease Progression and Neuroimaging Outcomes. A Systematic Review. Authorea. 24 October 2025. DOI: https://doi.org/10.22541/au.176128044.44919305/v1 DOI: https://doi.org/10.22541/au.176128044.44919305/v1 If you have the appropriate software installed, you can download article citation data to the citation manager of your choice. Simply select your manager software from the list below and click Download. For more information or tips please see 'Downloading to a citation manager' in the Help menu.