Effect of high-dose 1.25 dihydroxyvitamin D 3 on remyelination in the cuprizone model AGNES E. NYSTAD, 1,2 STIG WERGELAND, 1,2 LAGE AKSNES, 3,4 KJELL-MORTEN MYHR, 1,2 LARS BØ 1,2 and ØIVIND TORKILDSEN 1,2 1 Department of Neurology, Norwegian Multiple Sclerosis Competence Centre, Haukeland University Hospital, Bergen; 2 Kristian Gerhard Jebsen MS Research Centre, Department of Clinical Medicine, University of Bergen, Bergen; 3 Department of Clinical Science, University of Bergen, Bergen; and 4 Department of Pediatrics, Haukeland University Hospital, Bergen, Norway Nystad AE, Wergeland S, Aksnes L, Myhr KM, Bø L, Torkildsen Ø. Effect of high-dose 1.25 dihydroxyvitamin D 3 on remyelination in the cuprizone model. APMIS 2014. Vitamin D supplementation is increasingly recommended to patients with multiple sclerosis (MS). To study the effect of high-dose vitamin D on remyelination, female C57Bl/6 mice were demyelinated with dietary 0.2% cuprizone for 7 weeks. The mice received intraperitoneal injections of 1.25-dihydroxyvitamin D 3 (calcitriol) or placebo (vehicle) injec- tions twice a week, from week 6, throughout week 9. Mice that received calcitriol had initially increased demyelination (p = 0.021), astrocytosis (p = 0.043), and microglia activation. However, levels of astrocytosis and microglia activation dropped below those of the placebo group during the remyelination phase. There was a significant increase in myelina- tion in the calcitriol group throughout the remyelination phase (p = 0.041), while the remyelination in the placebo group was not significant (p = 0.317). After 3 weeks of remyelination, the calcitriol group had more myelin than the placebo group (p = 0.001). The calcitriol group had a higher density of NOGO-A positive cells throughout the remyeli- nation phase, and the number of NOGO-A positive cells was significantly higher in the calcitriol group at one week of remyelination (p = 0.019). There were no significant differences in extent of T-lymphocyte infiltration. High-dose calcit- riol seems to be safe regarding remyelination. Our results indicate that this treatment could actually promote the repair process, possibly through a stimulating effect on oligodendrocyte maturation and astrocyte activation. The potential of calcitriol to stimulate the remyelination process should be investigated further in functional studies. Key words: Multiple sclerosis; cuprizone; calcitriol; vitamin D; remyelination. Correspondence: Øivind Torkildsen, Department of Neurology, Haukeland University Hospital, N-5021 Bergen, Nor- way. e-mail: oivind.torkildsen@gmail.com Multiple Sclerosis (MS) is an immune-mediated disease of the central nervous system (CNS). The cause of the disease is unknown, but it seems to evolve as a result of a complex interplay between genetic and environmental risk factors (1, 2). Vita- min D deficiency is emerging as one of the most promising environmental risk factor candidates (3). Studies have repeatedly found associations between vitamin D levels and disease activity (4, 5), and sev- eral authors have suggested that supplementation of vitamin D should be recommended to MS patients (6–8). It has, however, been found that high-dose vitamin D supplements does not have any therapeu- tic advantages compared with low-dose supplements, and that high-dose vitamin D treatment might even have negative effects (9, 10). The cuprizone model is a toxic non-T cell-depen- dent model of CNS demyelination in mice. The cop- per chelator cuprizone mainly targets mature oligodendrocytes. Oligodendrocyte precursor cells and other glial cells are not or only marginally affected (11). Cuprizone induces oligodendrocyte death with subsequent myelin disruption, and microglia- and macrophage activation. Astroglial activation occurs in the early stages of demyelination (12, 13). The model demonstrates acute demyelina- tion with subsequent spontaneous remyelination within days after termination of cuprizone exposure (14–16). We have previously used the cuprizone model to study how vitamin D 3 affects demyelination (17). Received 25 October 2013. Accepted 10 March 2014 1 APMIS © 2014 APMIS. Published by John Wiley & Sons Ltd. DOI 10.1111/apm.12281