The Study Of The Interaction Between Microglia And Astrocyte In The Micro-environment Of Traumatic Brain Injury

Part One Characteristics of microglia and astrocytes' responses in traumatic brain injuryObjective: To study the response characteristics of microglia and astrocytes and the expression of related proteins in traumatic brain injury(TBI)environment.Methods: The model of traumatic brain injury in mice was established.The morphological and quantitative changes of microglia and astrocytes were detected by immunofluorescence,protein imprinting and immunohistochemistry.The proteins of TLR4/MyD88/NF-?B as well as HMGB1,IL-1? in tissues were also detected.Results: After TBI,there were obvious edema,neuronal apoptosis,BBB damage in the brain tissue lesion area,significant proliferation of microglia and astrocytes,and the morphology of microglia changed from long-branched to short-branched amoeba-like.After TBI,TLR4/My D88/NF-?B signaling pathway was activated significantly,and HMGB1,IL-1?and so on.The expression of subunit was significantly up-regulated.TBI in vitro model showed that inhibiting HMGB1 could significantly inhibit the pro-inflammatory reaction between microglia and astrocytes.Conclusion: After TBI,microglia and astrocytes show significant proliferation and activation,accompanied by significant activation of TLR4/My D88/NF-?B signaling pathway and significant up-regulation of molecular expression of HMGB1 and IL-1?.HMGB1 is an important factor in the interaction between microglia and astrocytes.Part Two The proinflammatory interactions between microglia and astrocytes in vitro and the relative mechanismsObjective: To investigate the proinflammatory interactions and the relative mechanism between microglia and astrocytes in vitro.Methods: Primary microglia and astrocytes stimulated by LPS were used as in vitro research models.The interaction between them was explored by conditioned medium.The activation of related signaling pathways was studied by immunofluorescence,protein imprinting and real-time quantitative PCR.Results: The TLR4/My D88/NF-?B signaling pathway related protein as well as inflammatory cytokines as IL1-? and i NOS were up-regulated both in microglia and astrocyte,associated with robust microglia proliferation.After administration of TAK242 and BAY117082,expression of TLR4,My D88 and phosphorylated NF-?B decreased as well as IL1 b and i NOS.Additionally,when using conditioned medium from microglia and astrocyte stimulated by LPS to activate astrocytes and microglia,expression of both IL-1? and i NOS was found to significantly increase in comparison with LPS stimulation alone.Such effects can be inhibited by TAK242 and BAY117082 respectively.Conclusion: LPS can mimic TBI in vitro and activate microglia and astrocytes to produce proinflammatory responses via the TLR4/NF-?B pathway,besides,microglia and astrocytes can enhance the pro-inflammation effects in astrocytes and microglia respectively through TLR4/NF-?B pathwayPart three Effect and mechanism of HMGB1 inhibitor on nerve function and BBB repair in TBI miceObjective: To study the interaction between microglia and astrocytes mediated by HMGB1 in TBI microenvironment.Methods: TBI mice model was established.HMGB1 inhibitors EP and GA were used to inhibit the expression of HMGB1.Immunofluorescence,Western blotting and real-time quantitative PCR were used to study the activation of related signaling pathways.Results: Under the action of TBI inhibitors EP and GA,the damage of nerve function and BBB in mice decreased significantly,and the number of pro-inflammatory microglia and astrocytes decreased significantly.Conclusion: HMGB1 plays an important role in TBI microenvironment.Microglia and astrocytes produce inflammatory reaction through HMGB1,which aggravates the brain injury after TBI.Inhibiting the expression of HMGB1 can promote the early recovery of nervous function and reduce BBB injury in TBI mice.