Mechanism Of Acupuncture Regulating RhoA/ROCK Signaling Pathway To Improve The Safety Of Thrombolysis In Cerebral Infarction And Clinical Effect Observation

Objective:(1)To investigate the effect of acupuncture on improving the safety of intravenous thrombolysis of rt-PA and to explore the mechanism of acupuncture regulating RhoA/ROCK signaling pathway in cerebral infarction model rats.(2)To clinically verify and observe the effect of acupuncture on improving the safety of intravenous thrombolysis of rt-PA in patients with cerebral infarction,in order to win more treatment opportunities for the majority of patients with cerebral infarction and provide new ideas and methods for improving the safety of thrombolysis in cerebral infarction.Methods:1.Experimental study:Experiment 1.Study on the effect of acupuncture on improving the safety of intravenous rt-PA thrombolysis in cerebral infarction model rats:SD rats were randomly divided into 4 groups:sham group,model group,4.5h rt-PA group,and acupuncture+4.5h rt-PA group.The cerebral infarction rat model was established by modified autothromboembolization method.Thrombolysis in the 4.5h rt-PA group and the acupuncture+4.5h rt-PA group were given intravenous rt-PA thrombolysis 4.5h after the success of the cerebral infarction model.Acupuncture+4.5h rt-PA group received Xingnao Kaiqiao acupuncture intervention immediately after 4.5h intravenous rt-PA therapy,and the acupuncture was kept for 30min,once a day,7 times as a course of treatment,for a course of treatment;the sham group and model group were injected with normal saline 4.5h after successful cerebral infarction model.The sham group,model group and 4.5h rt-PA group were given the same fixation only.Observe the effects of acupuncture on neurobehavioral score,percentage of cerebral infarct volume,content of hemoglobin,content of EB and percentage of cerebral water content in each group.Experiment 2.The mechanism of RhoA/ROCK signaling pathway in improving the safety of thrombolysis in rats with cerebral infarction model by acupuncture.Experiment 2 used the same model,grouping and intervention method as in Experiment 1.Western blot was used to detect RhoA/ROCK signaling pathway indicators(RhoA,ROCK2,MLC)and the expression of BBB(blood-brain barrier)structure-related proteins(Claudin5,ZO-1,Occludin).Real-time PCR was used to detect the mRNA expressions of RhoA and ROCK2,and immunofluorescence was used to detect the protein expressions of ZO-1,Claudin5 and MMP9 protein.2.Clinical observation:80 patients with cerebral infarction who received intravenous thrombolysis with rt-PA were randomly divided into observation group and control group,40 cases in each group.The control group was only given intravenous rt-PA thrombolysis within the time window of 4.5h,while the observation group was immediately given Xingnao Kaiqiao acupuncture treatment after intravenous rt-PA thrombolysis,and the needle was kept for 30min,once a day,7 times as a course of treatment,for 2 courses of continuous treatment,1 day interval between the two courses.Total response rate,NIHSS score,incidence of symptomatic intracranial hemorrhage(sICH),incidence of adverse events,sICH related indicators(TC,LDL-C,MPV,PLT,D-dimer,fibrinogen,CRP),and compliance were compared between the two groups.Results:1.Experimental study.Experiment 1.Study on the effect of acupuncture on improving the safety of intravenous rt-PA thrombolysis in cerebral infarction model rats(1)Neurobehavioral score:Compared with the model group,the neurobehavioral score decreased in the 4.5h rt-PA group and the acupuncture+4.5h rt-PA group(P<0.05).Compared with 4.5h rt-PA group,the neurobehavioral score decreased in the acupuncture+ 4.5h rt-PA group(P<0.01).(2)The percentage of cerebral infarction volume:The percentage of cerebral infarction volume in the 4.5h rt-PA group and the acupuncture+4.5h rt-PA group were lower than that in the model group(P<0.01).Compared with the 4.5h rt-PA group,the infarct volume in the acupuncture+4.5h rt-PA group was smaller(P<0.05).(3)BBB permeability:Compared with the model group,the content of EB increased in 4.5h rt-PA group and decreased in acupuncture+4.5h rt-PA group(P<0.01).Compared with 4.5h rt-PA group,the content of EB in the acupuncture+4.5h rt-PA group decreased(P<0.01).(4)Brain water content:Compared with the model group,the brain water content decreased in the 4.5h rt-PA group and the acupuncture+4.5h rt-PA group(P<0.01).The brain water content in the acupuncture+4.5h rt-PA group was lower than that in the 4.5h rt-PA group(P<0.05).(5)Brain hemorrhagic transformation:Compared with the model group,the content of hemoglobin in the 4.5h rt-PA group increased(P<0.01),but there was no significant difference in the acupuncture+ 4.5h rt-PA group(P>0.05).The content of hemoglobin in the acupuncture+4.5h rt-PA group was lower than that in the 4.5h rt-PA group(P<0.01).Experiment 2.The mechanism of RhoA/ROCK signaling pathway in improving the safety of thrombolysis in rats with cerebral infarction model by acupuncture.(1)RhoA/ROCK signal pathway index:Compared with the model group,the protein and mRNA expressions of RhoA and ROCK2 in 4.5h rt-PA group were increased(P<0.01),the protein and mRNA expressions of RhoA and ROCK2 were decreased in the acupuncture+4.5h rt-PA group(P<0.01).Compared with the 4.5h rt-PA group,the protein and mRNA expressions of RhoA and ROCK2 in the acupuncture+4.5h rt-PA group were decreased(P<0.01).Compared with the model group,the expression of MLC protein increased in the 4.5h rt-PA group(P<0.05),the expression of MLC protein in acupuncture+4.5h rt-PA group decreased(P<0.05).Compared with the 4.5h rt-PA group,the expression of MLC protein in the acupuncture+4.5h rt-PA group decreased(P<0.01).(2)MMP9 protein expression:Compared with the model group,the expression of MMP9 protein increased in the 4.5h rt-PA group(P<0.05),and decreased in acupuncture+4.5h rt-PA group(P<0.05).Compared with the 4.5h rt-PA group,the expression of MMP9 protein decreased in the acupuncture+4.5h rt-PA group(P<0.01).(3)Expression of ZO-1,Claudin5 and Occludin protein:Compared with the model group,the expression levels of ZO-1,Claudin5 and Occludin protein decreased in the 4.5h rt-PA group(P<0.05,P<0.05,P<0.01),increased in the acupuncture+4.5h rt-PA group(P<0.05,P<0.05,P<0.01).Compared with the 4.5h rt-PA group,the expression of ZO-1,Claudin5 and Occludin protein were increased in the acupuncture+4.5h rt-PA group(P<0.01).2.Clinical observation(1)Comparison of total effective rate between the two groups after treatmentThe total effective rate was 95%in the observation group and 88.57%in the control group,the observation group was higher than the control group.(2)Comparison of NIHSS scores between the two groups after treatmentAfter treatment,the NIHSS scores in the observation group and the control group were lower than those before treatment(P<0.05),and the observation group was lower than the control group(P<0.05).(3)Comparison of the incidence of sICH between the two groupsThe incidence of sICH in the observation group was 0%,and that in the control group was 10%,the observation group was lower than the control group(P<0.05).(4)Comparison of the incidence of adverse events between the two groupsThe incidence of adverse events was 0%in the observation group and 12.5%in the control group(P<0.05),the observation group was lower than the control group(P<0.05).(5)Comparison of sICH related indicators between the two groupsAfter treatment,the levels of LDL-C,MPV and CRP in the observation group and control group were lower than those before treatment.After treatment,the level of D-dimer in the observation group was lower than that before treatment(P<0,05).After treatment,the levels of TC,PLT and fibrinogen in the observation group were higher than those in the control group,while the levels of D-dimer and CRP in the observation group were lower than those in the control group(P<0.05).(6)Comparison of compliance between the two groupsClinical observation was completed in both groups,and the compliance was 100%(P>0.05).Conclusion:1.Acupuncture can improve the neurological function,reduce the volume of cerebellar infarction,reduce the degree of cerebral edema,reduce the exacerbation of BBB destruction and reduce the occurrence of HT after thrombolysis in rats with cerebral infarction,and achieve the effect of improving the efficacy and safety of thrombolysis.2.Acupuncture can effectively protect BBB by inhibiting RhoA/ROCK signaling pathway,so as to improve the safety of intravenous rt-PA thrombolysis in rats with cerebral infarction.3.Prompt acupuncture intervention can improve the efficacy of rt-PA intravenous thrombolysis in patients with cerebral infarction,reduce the incidence of sICH,and improve the safety of thrombolysis.It provides a new idea and method for improving the safety of thrombolysis in cerebral infarction.