| Background:Cerebral infarction,called cerebral ischemic stroke as well,is a most commonly type of cerebrovascular disease(CVD),couting for about 70% of all CVD. Neurologic defect appears duing to the ischemic necrosis of brain tissue after stroke.So the concerning about nerve regenerration after stroke is increasing daily.It was showed in the current studies that the restriction of neurite outgrowth can be mostly attributed to the over express of myelin inhibitors,including Nogo, myelin associated glycoprotein(MAG) and oligodendrocyte myelin glycoprotein(OMgp), which fuction through the Rho-ROCK pathway by combining Nogo receptor (NgR).So the Nogo or NgR became a new target for the nerve repairing.Animal experiments suggested that the absence of NgR allows a greater number of corticorubral fibers from the intact hemisphere to sprout into the bain stem and contralateral spinal cord denervated by the stroke ,which reached an . anatomically compensatory remodeling. The roles of the myelin inhibitors were foud in the experiment in vitro that the DRG neurites repelled by the CSPG,MAG and Nogo-A in the culture medium.Interestingly,co-culture of human MSC with DRG explanted reduced the inhibitory effects of the myelin mentioned above on neurite outgrowth. These findings suggested that MSC transplantation may promote axonal regeneration both by stimulating nerve growth via secreted factors and also by reducing the nerve-inhibitory effects of the extracellular molecules present.It is proved that there may be some mechanisms in MSCs transplantation after cerebral infarction:cells replacement; promoting proliferation of endogenous neural stem cells;secreting nutrition factors to promting angiogenesis,protecting penumbra neurons from death and promoting the neurites growth.Among these,whether the nerve regeneration promoted by MSCs works by reducing the express of NgR has not reported in current SCI papers. Therefor experoments were carried out in rats to study the effect of MSCs on the express of NgR and to explore the possibility of MSCs may work through the Nogo/NgR pathway.Objective: to study the effect of MSCs on the express of NgR and to explore the possibility of MSCs may promot neurites growth through the Nogo/NgR pathway.Materials and methods: MSCs were isolated and purified by the attachment culture method and cultured in specified culture medium.Cell phenotype were detected by flow cytometry, and Osteoblastic differentiation and neural differentiation were induced to comfirm the cells.Then rat stroke models were stablished by MCAO surgeris and two doses(3×106,or 6×106) of MSCs,labelled with Hoechst,were injected into the tail vein at 24th hour after stroke.Animals were sacrificed 1, 2, and 3 weeks after operation.All brains were processed for paraffin embedded and 9 sections,3μm thick,were cut for Nissl stain,to observe the perilesional neurons, and immunohistochemisty, to detect the expression of NgR, GFAP and ki67.Western Blotting was performed for the NgR expression.Results: 1.Higher purity of rMSCs could be achieved by the attachment culture method and cells can be induced to differentiate to neuron-like cells and osteoblast.2.Sensorimotor forelimg cortex(FL),primary and secondary somatosensory cortices(S1 and S2) of right side were damaged,resulting in fuction defect in moter and sensation of left forelimb in the rats after stroke compared with the sham-operated rats. The rats treated with MSCs showed significant improvement in NSS compared with MCAO rats,but there were no significant differences between the two groups treated with different doses of MSCs. 3.The NgR over-expressed in brain tissue,obviously on 1st week. 4. The perilesional neurons and the expression of ki67 around the lesion in the groups treated MSCs was higher than that in the MCAO group;the expression of ki67 in the ipsilateral SVZ in the former groups were higher than that in the latter groups,and the the expression of ki67 in the ipsilateral SVZ showed higher in the 6×106 MSCs groups than that in the 3×106 MSCs groups.5.The NgR expression,detected by immunohistochemisty and WB,in the ipsilateral primary and secondsry motor cortices, insular cortex and the contralateral primary motor cortice in groups treated with MSCs,represented a decrease,obviously in the group treated with 6×106 MSCs.Conclusions: The ipsilateral FL,S1 and S2 were damaged afer MCAO, resulting in fuction defect in moter and sensation of left forelimb in the rats after stroke, with NgR increased not in ipsilateral cortex but also contralateral cortex,obviously on 1st week.But the NgR showed a decrease in the groups treated with MSCs, especially in the 6×106 MSCs group, suggesting that efficacy may be dose-dependent.Meanshile the proliferation of neurons increased in the ipsilateral primary and secondsry motor cortices, insular cortex and the contralateral primary motor cortice with MSCs treated after stroke specially in the 6×106 MSCs group.These findings suggested that MSCs may promote nerve regeneration by reducing NgR expression afer transplantation. |
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