BMSCs Promote The Neurological Recovery Of SCI Rats Via The Mediation Of The Differentiation Of ENSCs And The Inflammatory Process

Object: To investigate the effect of BMSC-CM on the neurological regeneration and immune system in spinal cord injured rats.Methods: In vitro,the differentiation of NSCs was detected by immunofluorescence after 7 days co-culutre with BMSC-CM.In vivo,by using the tissueimmunofluorescence and Western blot,the expression of neurons and astrocytes was detected.In addition,ELISA,WB,He staining and BBB score were used to detect the effects of bmsc-cm on the expression of inflammatory factors,neuronal apoptosis,cavity size and neurological function.Resutles: In vitro,BMSC-CM was able to promote the differentiation of NSCs to into neurons and reduce their differentiation into astrocytes.In vivo,the treatment of BMSC-CM to SCI rats reduced the glial scar around the injured cavity and promoted,the regeneration of neurons and the sprouting of nerve fibers into the scar.In addition,by the downregulation of pro-inflammatory factors and upregulation of anti-inflammatory factors,BMSC-CM reduced the local neuronal apoptosis,cavity size and improving the neurological functional recovery.Conclusion: BMSC-CM was able to promote the improvement of neural function after spinal cord injury through the mediation of inflammation and the differentiation of endogenous neural stem cells.Object: To investigate the effect of BMSC-CM on the BMP4/Smad 1/5/8 signaling pathway in NSCsMethods: In vitro,the differentiation of NSCs was detected by immunofluorescence after 7 days co-culutre in the presence of BMP4 with or without BMSC-CM,and by using WB,the expression of p-Smad 1/5/8 was analysized.In vivo,WB was used to detect the expression of both BMP4 and p-Smad 1/5/8 in the injured lesion at different time points following the onset of SCI.Resutles: In vitro,the proportion of astrocytes were significantly increased by the addition of BMP4 to NSCs,while this BMP4-induced effects on the differentiation of NSCs were abolished by the treatment of BMSC-CM.WB anaylsis showed that the p-Smad 1/5/8 expression was downregulated by the treatment of BMSC-CM to NSCs in the presence of BMP4.In vivo,both of the BMP and p-Smad 1/5/8 expression was reduced by the treatment of BMSC-CM,compare to the rats without the BMSC-CM treatment.Conclusion: BMSC-CM was able to mediate the differentiation of NSCs via inhibiting the BMP4/Smad 1/5/8 sigaling pathway.Object: To evaluate The role of hepatocyte growth factor in mesenchymal stem cell-induced recovery in spinal cord injured rats.Methods: A conditioned medium(collected from the MSCs)and hepatocyte growth factor(HGF)were used to test the effects on the differentiation of neural stem cells(NSCS)in the presence of BMP4 with or without a c-Met antibody.In SCI rats,Western blot,ELISA,immunohistochemistry,and hematoxylin-eosin staining were used to investigate the biological effects of MSC-conditioned medium and HGF on nerve cell regeneration and inflammation with or without the pre-treatment using a c-Met antibody.In addition,the possible molecular mechanism(cross-talk between HGF/c-Met and the BMP/Smad 1/5/8 signaling pathway)was also detected by Western blot both in vivo and in vitro.Resutles: The conditioned medium from bone marrow-derived MSCs(BMSCs)was able to promote the NSC differentiation into neurons in vitro and the neurite outgrowth in the scar boundary of SCI rats by inhibiting the BMP/Smad signaling pathway as well as reduces the secondary damage through the modulation of the inflammatory process.The supplementation of HGF showed similar biological effects to those of BMSC-CM,whereas a functional blocking of the c-Met antibody or HGF knockdown in BMSCs significantly reversed the functional improvement mediated by the BMSC-CM.Conclusion: The MSC-associated biological effects on the recovery of SCI rats mainly depend on the secretion of HGF.Object: To investigate the effects of BMSC-CM on the Smad 6 expression and its role in the mediation of the differentiation of NSCs and in SCI rats.Method: Conditioned medium(CM)was collected from bone marrow MSCs(BMSCs)isolated from rats with SCIs,and its effect on the regulation of Smad 6 expression was tested in vitro(in NSCs)and in vivo(SCI rats).Western blot analysis and immunohistochemistry staining were used to investigate the proportion of neurons and astrocytes in vitro and in vivo.BBB scores were used to assess the neurological outcome of SCI rats at different time points.Results: BMSC-CM could upregulate Smad 6 expression in vitro.BMSC-CM-induced upregulation was suppressed by pre-treatment with the TGF-β type I receptor kinase inhibitor SB431542.BMSC-CM was able to promote the differentiation of NSCs to neurons;Smad 6 knockdown in NSCs partly weakened this effect on neural differentiation.In vivo,Smad 6 expression in the later phase of injury was closely associated with BMSC-CM treatment.Conclusion: BMSC-CM can upregulate Smad 6 expression by the secretion of TGF-β.It promotes the differentiation of NSCs into neurons,partly through upregulation of Smad 6.