| Objective: To observe the effects of massage on pain behavior indexes(spontaneous pain,heat pain,gait),cell morphology changes in the right L4 and 5 dorsal root ganglion and Purinergic receptor P2 X ligang-gated ion channel 3(P2X3)and Protein Kinase C(PKC)protein expression levels in the right L4 and 5 dorsal root ganglion of chronic constriction injury(CCI)model rats,to explore the effect of PKC-P2X3 pathway in DRG on peripheral analgesia in regulating massage intervention of neuropathic pain and enrich the theoretical system of massage analgesia.Methods: 64 healthy male male SD(Sprague Dawley)rats were randomly divided into the normal group,the sham group,the model group and the massage group according to the random number table method,with 16 rats in each group.CCI model was established in model group and massage group,the rats in the massage group were intervened by massage on the fourth day after operation,once a day,once for 15 minutes,for 18 consecutive days,the rest groups only fixed the same as the massage group,without massage.Spontaneous pain scoring criteria,hot water tail-flick test were used respectively to spin experiment,Catwalk gait analysis system observation group rats in front of the building,building 3,7,14,21 days after the spontaneous pain,thermal hyperalgesia and the parameters change of the gait,and within 21 days after the model preparation,remove groups of rats on the right side of L4-5 DRG,the western blot and immunohistochemical technology measure P2X3,PKC protein expression,hematoxylin-eosin staining is used to observe L4-5 DRG cells morphological changes.Results:(1)There were no statistically significant differences among spontaneous pain score,thermal withdrawal latency and Catwalk gait parameters before modeling,and the differences were comparable(P>0.05).(2)After establishing of the model,compared with the rats in the normal group and the sham group,the rats in the model group and the massage group showed the hind limb protection phenomenon such as tail slapping and licking,while the right posterior lower limb did not touch the floor and the load bearing decreased when walking,the spontaneous pain score increased,and the thermal withdrawal latency decreased,the difference is statistically significant(P<0.01);Changes in CatWalk gait analysis system parameters:stand,printarea,meanintensity and maxcontactarea,decreased significantly,while the swing increased,the duration time remains unchanged,and the footfall pattern is still dominated by alternating pattern,the difference is statistically significant(P<0.01).(3)After the massage intervention,the spontaneous pain score of the massage group decreased and the thermal withdrawal latency rose,the difference is statistically significant(P<0.01);Changes in CatWalk gait parameters:stand,printarea,meanintensity,and maxcontactarea began to rise and gradually approached the normal baseline level,swing begins to decrease,with statistically significant differences compared with the model group,the difference is statistically significant(P<0.01).(4)Western blot results:compared with the normal group and the sham group,the expression of P2X3 and PKC protein in L4 and 5 dorsal root ganglion increased in model group and massage group,while the P2X3 and PKC expression level in the massage group was lower than than in the model group,the difference is statistically significant(P<0.01).(5)Immunohistochemical results:compared with the normal group and the sham group,the expression of P2X3 and PKC protein in L4 and 5 dorsal root ganglion increased in model group and massage group,while P2X3 and PKC expression level in the massage group was lower than that in the model group,the difference is statistically significant(P<0.05).(6)Hematoxylin-eosin staining results:the DRG neurons in the normal group and the sham group were large and round,with clear nucleoli and uniform cytoplasm staining.The neurons in the model group and the massage group showed partial degeneration and necrosis,with uneven cytoplasmic staining and unclear nucleolus boundaries.Conclusion: Massage not only can reduce the degree of spontaneous pain and thermal hyperalgesia caused by sciatic nerve injury effectively,but also can improve the abnormal gait caused by modeling,promote the recovery of gait after nerve injury,and reduce the expression level of PKC and P2X3 protein in L4 and 5 dorsal root ganglion of rats in massage group.The results showed that PKC-P2X3 pathway in DRG was involved in the mechanism of peripheral analgesia of massage intervention on neuropathic pain,the possible mechanism is to inhibit the protein expression levels of PKC and P2X3 in DRG,thereby weakening the further upward transmission of pain signals mediated by the PKC and P2X3. |
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