The Study Of Spinal M1/M2 Microglia Polarization Is Involved In The Neuropathic Pain Of SNI Rats

Objective: The purpose of this study is to explore whether spinal M1/M2 microglia polarization is involved in the neuropathic pain induced by spared nerve injury(SNI),so as to further explore the role of neuroinflammation in neuropathic pain and its mechanism,and to provide scientific basis and guidance for the treatment of neuropathic pain.Methods:The rats were randomly divided into control group(Blank group),sham operation group(Sham group),SNI operation group(SNI group).The neuropathic pain model of rat SNI(sciatic nerve spared nerve injury)was constructed.The Von Frey hair was used to detect the mechanical allodynia changes at 1d,3d,7d,14 d and 21 d after operation;Immunofluorescence and Western blot were used to observe the activation of microglia in spinal dorsal horn of SNI rats with neuropathic pain;Immunofluorescence was used to observe the expression of polarization states of M1 and M2 microglia in the spinal dorsal horn of rats with neuropathic pain;The expression levels of pro-inflammatory factors(TNF-α,IL-1β)and anti-inflammatory factors(IL-10)in the spinal cord of neuropathic rats were detected by Western blot.Results:The neuropathic pain model of SNI in rats was successfully constructed.Compared with sham group,the mechanical withdrawalthreshold of SNI group decreased 1 day,3 days,7 days,14 days and 21 days after the operation(P < 0.05).Among them,the pain field was reduced to the lowest on the 14 th day after SNI;Microglia in spinal dorsal horn were activated significantly after SNI nerve injury in rats,and the expression of microglia marker Iba-1 was increased significantly(P <0.05),especially on the 14 th day after SNI.Immunofluorescence results showed that microglia activation in spinal dorsal horn 14 d after surgery:the number of microglia of the SNI group was significantly higher than Sham group,which was significantly different from the sham operation group(P < 0.05,n = 5),and the morphology of the microglia changed,the cell body became hypertrophy,the protrusion shortened,and it changed from a branch to an amoeba;The expression of pro-inflammatory M1 microglia phenotypes marker CD86 in lumbar spinal cord of rats 14 days after SNI was significantly higher than that of sham-operated rats(P <0.05);on the contrary,the expression of anti-inflammatory M2 microglia phenotypes marker CD206 in spinal cord of rats in SNI group was significantly lower than that in sham-operated rats(p < 0.05);The pro-inflammatory cytokines IL-1β and TNF-α in the lumbar spinal cord of SNI group were significantly higher than those in Sham group(P <0.05).The expression of anti-inflammatory cytokines IL-10 protein in spinal cord after SNI was changed.It was found that the expression of anti-inflammatory cytokines IL-10 was significantly up-regulated on day1 after SNI(P < 0.05).However,the expression of the anti-inflammatory cytokines IL-10 in the spinal dorsal horn was not significantly different from that in the Sham group at 3 days after SNI surgery.At the later time points(rats at 7,14,21 days after SNI surgery),IL-10 expression was significantly lower than that of the Sham group(P < 0.05).Conclusion: Spinal microglia can exhibit two different polarization states of M1 and M2 sin neuropathic pain of SNI rats,in which the proinflammatory effect of M1 microglia mediates the formation of neuropathic pain,while the anti-inflammatory effect of M2 microglia was significantly attenuated during the neuropathic pain,and neuropathic pain was associated with an increased proportion of M1/M2 microglia.