The Role Of Tlr4 In The Recovery Of Sciatic Nerve Injury In C3H/HeJ Mice

Objective The TLR4 mutant mice C3H/He J are used to build the model of sciatic nerve injury.Our goals are to identify short-stage and long-stage changes after sciatic nerve injury,mainly by checking the expression changes of inflammation factors in the short-stage and the differences in the recovery of the injured sciatic nerve in the long-stage.Further explore the relationship between TLR4 signaling pathway and sciatic nerve injury.The experimental theory basis was provided for the clinical treatment of peripheral nerve injury.Methods To investigate the correlation between TLR4 and recovery after sciatic nerve injury,the model of sciatic nerve injury was conducted using TLR4-mutated mice(C3H/He J)and wild mice(C3H/He N).The experiment divide into C3H/He J injury group(He J),C3H/He N injury group(He N)and C3H/He N control group(control).The injury of sciatic nerve operate under the condition of sterile environment.The short-stage and longstage change after sciatic nerve injury are detected.Real-time quantitative PCR(RT-q PCR)is applied to detect the dynamic expressions of Interleukin-1?(IL-1?),Interleukin-6(IL-6),Tumor necrosis factor-?(TNF-?),Monocyte chemotactic protein 1(MCP-1),Nlrp3,Nlrp6 and caspase1 in post-operative 6h,12 h,24h and 48 h.Western blot is used to find the protein expression of p75 neurotrophin receptor(P75NTR),growth-associated protein-43(GAP-43),Nlrp3,Nlrp6 and caspase1 in 12 h and 24 h postoperatively.After 1week,2weeks,4weeks and 8weeks of sciatic nerve injury in mice,the pathological variations were observed by hematoxylin-eosin staining(HE staining),protein expressions of GAP-43,p75 NTR,Nlrp3,Nlrp6 and caspase1 were detected by immunohistochemistry.Sciatic nerve function index(SFI)evaluate motor function recovery after sciatic nerve injury.Results The result of short-stage: q PCR showed that compared with control group,the expression of IL-1?,IL-6,TNF-? and MCP-1 were significantly increased after 6h,12 h and 24 h of sciatic nerve injury in He N group(P < 0.01),and these changes of gene expression in He J group is not significant.Contrast with TNF-? between He N and He J group mice showed significant difference again(P < 0.01).According to the results of 12 h or 24 h after sciatic nerve injury,the p75 NTR protein expression in He J mice significantly increased using Western blot,compared with the He N group(P < 0.001).Meanwhile,the caspase1 expression was high,although lower Nlrp3 expression and different Nlrp6 expression in the He N and He J Group.The long-stage difference mainly concerned the recovery of injured sciatic nerve,so HE staining,immunochemistry and sciatic functional index(SFI)were used.There were more neutrophils and macrophages in the He N Group at 1week,2weeks,4weeks and 8weeks after sciatic nerve injury,showing that the intense inflammation happened in He N Group by HE staining and the higher expression of GAP-43 and p75 NTR showed in He J Group at the same detected time.The results of immunohistochemistry in Nlrp3,Nlrp6 and caspase1 showed that,Nlrp3 and caspase1 did not have much difference,but Nlrp6 in the He J Group express higher than the He N Group mice.SFI results show that,compared with the He N group,SFI score of He J Group in the postoperative period is relatively high and the nerve recovery is better.Conclusion We can conclude that sciatic nerve injury model is successfully built in mutant mice,and the study revealed that TLR4 can speed up the sciatic nerve repair by interfering inflammatory factors.The changes in the expression of Nlrp6,which are related to the TLR4 mutation,may influence recovery of the injured sciatic nerve.Further studies will be conducted to confirm these results.The experimental theory basis was provided for the clinical treatment of peripheral nerve injury.