S100A8 Promotes Inflammation Via Toll-like Receptor 4 After Experimental Traumatic Brain Injury

Objective: S100 calcium-binding protein A8(S100A8),a member of the S100 family of proteins,is also known as macrophage-related protein 8,which is involved in various pathological processes in the central nervous system post traumatic brain injury(TBI),plays a critical role in inducing inflammatory cytokines.Accumulating evidence has indicated that Toll-like receptor 4(TLR4)is involved in inflammatory responses post TBI.The present study was designed to analyze the hypothesis that S100A8 is the key molecule that induces inflammation via TLR4 in TBI.Methods: In the present animal study,the weight-drop TBI model was used and implemented on mice that were randomly assigned to one of the six groups: Sham,NS,S100A8,S100A8+TAK-242 groups,TBI,and TBI+TAK-242 groups.S100A8 was injected into the lateral ventricle of the brain of mice in the S100A8 and S100A8+TAK-242 groups.In the S100A8+TAK-242 and TBI+TAK-242 groups,at half an hour prior to the intracerebroventricular administration of S100A8 or TBI,mice were intraperitoneally injected with TAK-242 that acts as a selective inhibitor of TLR4.In order to evaluate the association between S100A8 and TLR4,Western blot,immunofluorescence,enzymelinked immunosorbent assay(ELISA),and Nissl staining were employed.Simultaneously,the neurological score and brain water content were also assessed.In addition to the above mentioned techniques,in the in vitro analysis,BV-2 murine microglial cells were stimulated with lipopolysaccharide(LPS)or S100A8 recombinant protein,with or without TAK-242 pre-treatment.Hence,the expressions of the functional proteins were subsequently detected by Western blot or ELISA.Results: The levels of S100A8 protein and pro-inflammatory cytokines were significantly elevated after TBI.The neurological scores were reduced in non-TBI animals with remarkable severe brain edema after the intracerebroventricular administration of S100A8.Furthermore,the TLR4,p-p65 and myeloid differentiation factor 88(My D88)levels were elevated after the administration of S100A8 or TBI,which were blocked by TAK-242 pre-injection.Meanwhile,in the in vitro analysis,S100A8 or LPS treatment,upregulated the expression of p-p65 and My D88 in cultured BV-2microglial cells,whereas TAK-242 pre-treatment suppressed these upregulations.Conclusion: The present study demonstrates that the TLR4-My D88 pathway is activated by S100A8,which is essential for the development of inflammation in the brain after TBI.