| Objective:Study on effect of electroacupuncture combined with rehabilitation training on middle cerebral artery occlusion(MCAO)model rat micro RNA-223-3p(Micror NA-223-3p)The influence of miR-223-3p 、 NLRP3,to explore the possible mechanism of action of electroacupuncture combined with rehabilitation training on pyrodeath of cells after cerebral ischemia.It provides a new therapeutic direction and experimental basis for the clinical treatment of cerebral ischemia.Method:Study on effect of electroacupuncture combined with rehabilitation training on middle cerebral artery occlusion(MCAO)model rat micro RNA-223-3p(Micror NA-223-3p)The effect of miR-223-3p 、 Nod-like receptor protein 3(NLRP3),to explore the possible mechanism of electroacupuncture combined with rehabilitation training on pyrodeath after cerebral ischemia,and to provide a new therapeutic direction and experimental basis for clinical treatment of cerebral ischemia.Method:1.45 SPF male SD rats were purchased and randomly divided into model group,electroacupuncture group(EA),rehabilitation group(RT),electroacupuncture combined rehabilitation group(EA+RT)and sham operation group,with 9 rats in each group.MCAO model was established in the sham operation group except for the right common carotid artery.2.Sham operation group and model group were not treated."Baihui","Dazhui" on the left side of the EA group were given electroacupuncture treatment 24 h after the establishment of the model,RT group was given treadmill training,and EA+RT group was given combined intervention with acupuncture and kang.Each treatment was performed for 20 minutes,once a day for 7 days.3.Longa score was observed 4h and 7d after the completion of modeling.The morphological changes of ischemic cortex were observed by HE staining.The concentrations of IL-1β and IL-18 in ischemic cortex were determined by enzyme-linked immunosorbent assay.The positive expressions of NLRP3 and Caspase-1 in the ischemic cortex were detected by immunofluorescence method.The expressions of GSDMD and miR-223-3p in ischemic cortex were detected by RT-q PCR.At the same time,the positive expression of NLRP3 and Caspase-1 and the concentration of IL-1β and IL-18 were interactive in factorial analysis.Results:1.Comparison of neurological impairment scores in each groupAt 4h after modeling,the neurological function scores of each group were between 1 and 3,and were significantly higher than those of the sham operation group(P<0.01),indicating that the model was successfully constructed.Compared with model group,the scores of EA and RT groups were decreased after 7 days(P<0.05),while the scores of EA+RT group were significantly decreased(P<0.01).There was no difference in scores among EA,RT and EA+RT groups(P>0.05).2.Morphological changes of ischemic brain tissue in each groupIn the sham operation group,the cell morphology was orderly,the nuclei were large,and the nucleoli were located in the center and obvious.In the model group,the ischemic cortical region had loose structure,blurred nuclear margin,irregular staining and morphology.Compared with the model group,the three treatment groups had regular and complete nuclei and clear nucleoli.In the EA+RT group,compared with the EA and RT groups,the cerebral ischemic area was improved more significantly,the cell morphology was more regular,and the number of nuclear deformed cells was significantly reduced.3.Comparison of IL-1β and IL-18 contents in ischemic cortical brain tissueCompared with sham operation group,the contents of IL-1β and IL-18 in model group were significant(P<0.01).Compared with model group,the contents of inflammatory cytokines IL-1β and IL-18 in EA,RT and EA+RT treatment groups were significantly decreased(P<0.01).Compared with EA group,the contents of IL-1β and IL-18 in EA+RT group were decreased(P<0.05).Compared with RT group,the contents of IL-1β and IL-18 in EA+RT group were also decreased(P<0.05).There were no differences in IL-1β and IL-18 contents between RT group and EA group(P>0.05).4.Comparison of NLRP3 and Caspase-1 expression in ischemic cortexCompared with sham operation group,the proportion of NLRP3 and Caspase-1 positive cells was significantly higher than that in model group(P<0.01).Compared with model group,the expressions of NLRP3 and Caspase-1 in EA,RT and EA+RT groups were significantly decreased(P<0.01).Compared with EA group,the positive expression level of immunofluorescence NLRP3 in EA+RT group was significantly decreased(P<0.05),and the expression level of Caspase-1 positive cells was significantly decreased(P<0.01).Compared with RT group,the immunofluorescence NLRP3 and Caspase-1 positive expression levels in EA+RT group were decreased significantly(P<0.01).The expressions of NLRP3 and Caspase-1 in EA group were not significantly different from those in RT group(P>0.05).5.m RNA expressions of miR-223-3p and GSDMD in the ischemic cortex of rats were detected by PCRCompared with sham operation group,the expression of miR-223-3p was significantly lower than that of model group(P<0.01),GSDMD was significantly increased(P<0.01).Compared with model group,the content of miR-223-3p in EA,RT and EA+RT was significantly increased(P<0.01),while the content of GSDMD was significantly decreased(P<0.05).Compared with EA group,the content of miR-223-3p in EA+RT group was significantly increased(P<0.05),while the content of GSDMD was significantly decreased(P<0.05).Compared with RT group,the expression of miR-223-3p in EA+RT group was also increased(P<0.05),while the content of GSDMD was also significantly decreased(P<0.05).Compared with RT group,there were no significant expressions of miR-223-3p and GSDMD genes in EA group(P>0.05).6.Factorial analysis of NLRP3 channel correlation factorsElectroacupuncture and rehabilitation training had significant effects on the positive expression of NLRP3 and Caspase-1 and the main effect of inflammatory cytokines IL-1βand IL-18(P<0.05).Electroacupuncture and rehabilitation training had synergistic interaction on the positive expression of NLRP3 and Caspase-1 as well as the inflammatory cytokines IL-1β and IL-18(P<0.05),and the four indexes were affected by positive synergistic interaction,resulting in a greater decrease in their levels than that of single factor.Conclusion:1.Electroacupuncture therapy and treadmill rehabilitation training can effectively improve the neurological defects of cerebral ischemia rats,improve the histopathological changes of ischemic brain,and promote the recovery of neurological function.Electroacupuncture and rehabilitation training have positive synergistic effect.2.Electroacupuncture combined with rehabilitation training can enhance the antiinflammatory and anti-pyrogenic ability by regulating the expression of miR-223-3p and NLRP3 related factors in the ischemic cortex of cerebral ischemia rats,which may play a role in inhibiting inflammatory injury and pyrogenic effect of nerve cells in cerebral ischemia rats,thus achieving neuroprotective effect. |
