The Study Of Icariin Enhances Remyelination Process After Acute Demyelination Induced By Cuprizone Exposure

ObjectiveMultiple sclerosis(MS)is an autoimmune disease with inflammatory demyelinations in the central nervous system(CNS),which is characterized by classic relapsing remitting pattern.Pathological changes,including inflammation,demyelination,and axon injury will appear since the onset of MS.During the remission course,pathology are still progressive and cumulative,thus the remission period is a key time point for the regeneration of the myelin sheath.Treated patients with drugs during the remission period may improve adequate levels of remyelination and further benefit MS patients.As an effective monomer component extracted from traditional Chinese medicine Epimedium,Icariin(ICA)has extensive pharmacological effects,known as neuroprotective drug to against oxidative-stress induced neurodegeneration and to improve the memory deficiency.In order to explore the effects of ICA on de-or remyelination process and the underlying mechanisms,the C57BL/6J mice in the present study were fed with rodent chows containing cuprizone(CPZ)for 5 weeks to induce acute demyelination.We detected the effects of ICA on the pathological changes of NF200-positive axons,APC+/Olig2+ mature oligodendrocytes and neuron-derived neurotrophic factor such as nerve growth factor(NGF)in the brain.Our findings suggest that ICA is a protective drug in the CNS demyelinating disorders such as MS and provides new clue for the development of MS therapeutic drugs.Methods1.Female C57BL/6 mice aged six-week old(weighing between 12 to 17 g)were fed with cuprizone(CPZ,0.2%w/w)for 5 weeks to induce acute demyelination and oligodendrocytes degeneration,after which CPZ was withdrawn to allow recovery.Icariin(ICA,6.25,12.5 and 25 mg/kg/day),vehicle(0.5%sodium carboxymethyl cellulose solution)or distilled water was administrated orally to mice for 1 week after CPZ withdrawal.The body weights of each mice were recorded every week.2.Brain tissue was taken at the point of 5 weeks’ CPZ exposure and 1 week after regeneration.Luxol-fast blue(LFB),immunohistochemical or immunofluorescence staining was used to detect morphological and pathological changes such as myelin basic protein(MBP)-positive myelin,NF200-positive axons,APC+/Olig2+ mature oligodendrocytes and neuron-derived nerve growth factor(NGF).Results3.C57BL/6J mice fed with CPZ diet showed weight loss.Since CPZ diet was withdrawn,the body weight of mice was restored.The impact of ICA on body weight is not significant.Feeding CPZ diet could successfully induce extensive demyelination in the corpus callosum region of C57BL/6J mice,which is suggested by LFB staining.Normal myelinated neural fibers were stained blue in LFB stained brain sections,however,blue staining was rarely seen in the central part of corpus callosum(CC)either at levels 0.74 mm or-2.18 mm bregma when mice were exposed to CPZ for 5 weeks.4.Demyelination and remyelination were also determined by myelin basic protein(MBP)immunohistochemistry and axon changes were detected by NF200 immunofluorescence.After 1 week recovery following CPZ withdrawal,the MBP immunostaining in the central part of CC reappeared and seemed to similar to that of normal mice in ICA treatment groups with dose-dependent manner.And the decrease of NF-200 positive axons both in the corpus callosum and frontal cortex induced by CPZ diet was reversed and further enhanced by ICA.5.In this study,it was found that CPZ could induce the apoptosis of Olig2+/APC+ mature oligodendrocytes(myelin-forming cells)in the frontal cortex of C57BL/6J mice,and ICA could promote the regeneration of mature oligodendrocytes during demyelination.6.The expression of nerve growth factor(NGF)in neurons of the frontal cortex was increased after CPZ diet,and ICA could increase the expression of nerve growth factor.The effect of ICA on promoting remyelination may be to improve neuronal expression of nerve growth factors.ConclusionAnimal experiments showed that ICA could promote regeneration of myelin and axons loss.The mechanisms may be the promotion of mature oligodendrocytes and the increase of neurotrophic factors such as nerve growth factor.Our results suggest that ICA may deserve to be a candidate of potential drug for the treatment of neurodegenerative diseases including MS.