| Objective:Multiple sclerosis (MS) is an inflammatory demyelination disease of the central nervous system (CNS) that causes relapsing and progressive neurological impairment. Experimental autoimmune encephalomyelitis (EAE), an autoimmune disease, characterised by myelinolysis and perivascular cuffings, is an ideal animal model for MS.There has been some reports that the neural stem cell marker expression is increasing when the brain and spinal injury, but there is few reports about that in relapsing-remitting EAE rat.Oxidative stress plays an important role in the pathogenesis of MS. Alpha-lipoic acid has attracted wide attention as a strong natural anti-oxidants. It mainly plays its antioxidant activity through scavenging free radicals, chelating metal ions, and regenerating other antioxidants. We found that alpha-lipoic acid can significantly reduce the morbidity of EAE, but weather it can promote the neural stem cell proliferation or not is unclear.In the research, we want to know the neural stem cell proliferation and its significance in EAE rat by detecting the induction of Nestin and PCNA. We administrate alpha-lipoic acid to the EAE rat in order to know its impact about the neural stem cell proliferation, thus to provide new ideas and experimental evidence to the treatment of MS.Methods:A total of 36 adult healthy female Wistar rats weighing between 180g and 200g were divided randomly into three groups: normal control group(6 rats),other rats were immunized subcutaneously in the four foot pads and backside by fresh guinea pig spinal cord homogenate (GPSCH) and complete Freund′s adjuvant(CFA). After the first paroxysm, divided randomly out 6 rats as disease group, others divided into EAE group and alpha-lipoic acid (LA group, 100mg. Kg-1. d-1), and this two groups were divided into 7 day group and 14 day group, separately.Clinical signs of EAE were assessed the mean of twice daily by two observations. Scores were assigned on the basis of the following symptoms: 1. tail weakness; 2. tail weakness plus limb asthenia; 3. mild limb paralysis; 4. severe limb paralysis; 5. moribund/dead. LA group were injected intra- peritioneal resectively LA100mg.Kg-1.d-1. This treatment was started on the first day of EAE outbreak and continued daily for 7 days.Rats were sacrificed after anesthesia with intraperitoneal injection. Tissue of the spinal cord was fixed with 4% formalin, then the tissue was embedded in paraffin and sectioned at 6μm thickness. The sections were stained with immunehistochemistry staining of Nestin and PCNA. At last, the results were analyzed with microscopy.Results:1 Nestin expression in the spinal cord: The number of positive cells in EAE disease stage is increased. In the remitted stage, it is decreased. In relapsing stage, it is increased again. The number of positive cells in spinal cord of rats in EAE disease group is higher than Relapsing group (P<0.05). In the remitted stage, there is no statistical significance between LA group and EAE group (P>0.05). In the relapsed stage, the number of positive cells of LA group is lower than that of EAE group (P<0.05). In the study, we find that there are Nestin positive cells similar with neuron or astrocytes.2 PCNA expression in the spinal cord: The number of positive cells in EAE disease stage is increased. In the remitted stage, it is decreased. In relapsing stage, it is increased again. The number of positive cells in spinal cord of rats in EAE disease stage is more than that in Relapsing stage (P<0.05). In the remitted stage, there is no statistical significance between LA group and EAE group (P>0.05). In the relapsed stage, the number of positive cells of LA group is lower than that of EAE group (P<0.05). And the positive cells are mainly distributed in the department of inflammatory cell infiltration.Conclusions:1 alpha-lipoic acid can protect the EAE rats from the severity of the disease, but it has no significant effect on the endogenous neural stem cell proliferation.2 The degree of cell proliferation is closely related to the development of EAE disease; alpha-Lipoic acid can reduce cell proliferation by reducing the degree of inflammation.
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