The Study On The Involvement Of Adipose-derived Mesenchymal Stem Cells In The Regulation Of Post-spinal Cord Injury Pain Through The Modulation Of Microglial Phenotypes

Lesions in the central nervous system such as the dorsal root of the spinal cord and the spinal cord caused by mechanical trauma are clinically classified as spinal cord injury(SCI).Even after timely surgical treatment,patients will still have neurogenic inflammation,which will lead to adverse reactions such as neurogenic pain,which seriously affects the physical and mental health of patients.Studies have shown that microglial activation after SCI releases inflammatory factors,participates in the neuroinflammatory process,and is one of the causes of hyperalgesia after spinal cord injury.Mesenchymal stem cells have a certain role in the treatment of spinal cord injury and avoid convalescent pain,but the role of adipose-derived Mesenchymal Stem Cells(Ad-MSCs)in pain relief of spinal cord injury has not been reported.Therefore,in this study,adipose mesenchymal stem cells were injected intrathecal into the injury site of rats with spinal cord injury models,and the mechanism of adipose mesenchymal stem cells regulating the phenotype of microglia participating in the regulation of pain in spinal cord injury was explored through microglial activation markers and inflammation-related factors,which provided a new theoretical basis for clinical treatment.1.METHODS(1)Analyze the related genes of spinal cord injury and stem cells through bioinformatics;(2)Identification of adipose mesenchymal stem cells activation markers by flow cytometry;(3)Identification of microglia activation markers by flow cytometry;(4)BBB scoring method and pain threshold detection method were used to detect the recovery of motor ability and change of pain threshold in rats with spinal cord injury;(5)HE staining and histochemical staining were used to detect the morphology of microglia;(6)Western blotting and ELISA were used to detect the expression of inflammation-related molecules.2.RESULT(1)There are 85 genes related to spinal cord injury and stem cells.We select the top 20 genes with the highest correlation for mapping,and select NF-κB、TNF-α and IL-1β as the focus of follow-up research;(2)The extracted Ad-MSCs expressed CD44 and CD90 at a high level and CD45 and CD103 at a low level,which conformed to the characteristics of stem cells and had good ability of adipogenesis and osteogenesis;(3)After the injection of stem cells,the microglia activation marker CDllb was identified by flow cytometry,which showed that compared with the spinal cord injury group,the microglia activation was decreased;(4)The BBB score of rats in the group injected with Ad-MSCs and stem cell supernatant rose,and the 50%threshold rose to a certain extent,it indicated that the exercise ability of rats recovered and the pain sensitivity was relieved after treatment;(5)HE staining results showed that injection of stem cells could effectively maintain the normal morphology of spinal cord tissue and maintain the activity of neurons.Immunohistochemical staining results showed that injection of stem cells could effectively inhibit the activation of microglia;(6)Western blotting detection of inflammatory factor NF-κB、TNF-α and IL-1βexpression,the result showed that after injection of stem cells,the expression of NFκB、TNF-α and IL-1β were down;Elisa detects inflammatory factor TNF-α、IL-6and IL-1β,the results showed that after injection of stem cells,the expression of TNF-α、IL-6 and IL-1β were down.3.CONCLUSION(1)The results of the behavior,histochemistry and related inflammatory factors of SCI model rats showed that the animal model used in the experiment was successful,and SCI rats exhibited neurogenic pain,and the possible mechanism was the activation of microglia in the spinal cord after injury,releasing inflammatory factors,and then mediating the inflammatory response,resulting in the phenomenon of neurogenic pain;(2)After injection of Ad-MSCs in SCI model rats,Ad-MSCs may effectively alleviate neurogenic pain in rats with spinal cord injury by inhibiting microglial activation,thereby inhibiting the inflammatory response mediated by activated microglia;Injection of stem cell supernatant can also alleviate the pain response after SCI to some extent,but its effect is weaker than that of stem cell injection.