| ObjectiveMyocardial ischemia-reperfusion injury refers to the more serious structural and functional changes of myocardial tissue in patients with coronary heart disease after percutaneous coronary intervention.Electroacupuncture preconditioning can alleviate the symptoms,but the mechanisms remain unclear cause the limitation of acupuncture to prevent and treat of Myocardial ischemia-reperfusion injury in clinical application.Related studies have shown that ventricular reconstruction happened in the process of myocardial ischemia-reperfusion injury,including myocardial remodeling and sympathetic remodeling.The studytaking the myocardial ischemia-reperfusion model rats as the research object,according to the theory of "preventive treatment of disease" of guidance from acupuncture,using the immunohistochemical and western blot to observe the effect of electroacupuncture preconditioning on myocardial remodeling and sympathetic remodeling,in order to explore the treatment and prevention mechanisms of the electroacupuncture preconditioning with "Combined Biao and Ben Acupoints " on myocardial ischemia-reperfusion injury,and to provide experimental basis for acupuncture to prevent andalleviate myocardial ischemia-reperfusion injury and to give full play to the advantages of "Combined Biao and Ben Acupoints Treatment".Methods1.40 SPF male Wistar rats grade at 3 months old were randomly divided into 4 groups with 10 rats in each group after adaptive feeding for 1 week.sham operation group(SO group),myocardial ischemia/ reperfusion injury group(MI/RI group),electroacupuncture preconditioning with Neiguan group(MI/RI+PC6group),and electroacupuncture preconditioning with Biao-Ben Acupoints group(myocardial ischemia reperfusion group with acupuncture pretreatment on Neiguan,Zusanli and Guanyuan,MI/RI+AP group).2.All rats were bound at the same time once a day for 20 min for 7 days.The rats in SO group and MI/RI group were bound without acupuncture,while the rats in MI/RI+PC6 and MI/RI+AP were pretreated with electroacupuncture.The rats in MI/RI+PC6 group were treated by acupuncture at the bilateral Neiguan(PC6)point,meanwhile another needle was inserted subcutaneously at0.3cm nearby the Neiguan(PC6)point as auxiliary electrode which would form a pair of electrodes with Neiguan(PC6)point.The rats in MI/RI+AP group were treated by acupuncture at the bilateral Neiguan(PC6),Zusanli(ST36)and Guanyuan(CV4)point,and a pair of electrodes were formed by the Neiguan(PC6)and Zusanli(ST36)acupoints at the same side,meanwhile another needle was inserted subcutaneously nearby the Guanyuan(CV4)point to 0.3cm as auxiliary electrode which would form a pair of electrodes with Guanyuan(CV4)point.Setting the han's electroacupuncture therapeutic apparatusas continuous wave,frequency of 2 Hz,intensity of 1 m A.The treatment would last for 20 min once a day for 7 days.3.At the end of 7 day's acupuncture preconditioning treatment,except for the rats in SO group,all rats were underwent thoracotomy,next caused myocardial ischemia of left ventricular anterior wall for 20 min by using suture to block the anterior descending branch of left coronary artery at 2 mm below the root of left atrial appendage and pulmonary artery,and then opened the suture to restore blood supply and reperfusion for 40 min,resulting in MI/RI model.Using BL-420 biological function experiment system to recorded the change of ST voltage after 20 min ligation and 40 min reperfusion.The elevation of ST voltage higher than 0.2 m V would mark myocardial ischemia,and the decrease of ST voltage would mark reperfusion.4.The mortality of rats in each group were observed during the modeling.The chemiluminescence was used to examine the content of the ROS in the MI/RI tissue,and the biochemistry was used to examine the activity of the serum GSH-PX.The Masson staining was used to observe the morphological distribution of myocardial collagen fibers and calculate the CVF.The expression of CTGF and the distribution of TH positive nerve fibers in the MI/RI tissues were detected by immunohistochemistry,while GAP43 protein in the MI/RI area was detected by Western blot.Results1.(1)Mortality rate:compared with SO group,the mortality rate of MI/RI group was significantly higher(P<0.01);compared with MI/RI group,the mortality rate of MI/RI+PC6 group and MI/RI+AP group was lower,but the difference was not statisticallysignificant(P>0.05).(2)ST segment voltage change of ECG:compared with SO group,MI/RI group was significantly higher after 20 min ligation and 40 min reperfusion(P<0.01);compared with MI/RI group,MI/RI+PC6 group and MI/RI+AP group were significantly lower after 20 min ligation and 40 min reperfusion(P<0.01);compared with MI/RI+PC6 group,MI/RI+AP group was lower after 20 min ligation and40 min reperfusion(P<0.05).(3)Markers of oxidative stress:compared with SO group,the content of ROS in MI/RI group was significantly higher(P<0.01),and the activity of GSH-PX was significantly lower(P<0.01);compared with MI/RI group,the content of ROS in MI/RI+PC6 group and MI/RI+AP group were lower(P<0.05),and the activity of GSH-PX were higher(P<0.05);compared with MI/RI+PC6group,the content of ROS in MI/RI+AP group was lower(P<0.05),the activity of GSH-PX was higher(P<0.05).2.(1)Expression of CTGF in myocardial tissue:compared with SO group,the expression of CTGF in MI/RI group was significantly increased(P<0.01);compared with MI/RI group,the expression of CTGF in MI/RI+PC6 group was decreased(P<0.05)and was significantly decreased in MI/RI+AP group(P<0.01);compared with MI/RI+PC6 group,the expression of CTGF in MI/RI+AP group was decreased(P<0.05).(2)Morphological comparison of myocardial tissue:after Masson staining of myocardial tissue,compared with SO group,the myocardial was obviously damaged in MI/RI group,the myofilaments arranged in disorder,and the collagen fibers were thickened and fibrosis were obvious;compared with MI/RI group,the myofilaments MI/RI+PC6 group arranged in order,only a few hyperplastic collagen fibers,partial fibrosis;compared with MI/RI+PC6 group,the pathological changes of myocardial tissue inMI/RI+AP group were further alleviated.(3)The ratio of CVF:compared with SO group,the ratio of CVF in MI/RI group was significantly increased(P<0.01);compared with MI/RI group,the ratio of CVF in MI/RI+PC6 group was decreased(P<0.05)and was significantly decreased in MI/RI+AP group(P<0.01);compared with MI/RI+PC6 group,the ratio of CVF in MI/RI+AP group was decreased(P<0.05).3.(1)The density of TH positive nerve fiber: compared with SO group,the density of TH positive nerve fiber in MI/RI group was significantly higher(P<0.01);compared with MI/RI group,the density of TH positive nerve fiber in MI/RI+PC6 group was lower(P<0.05)and was significantly lower in MI/RI+AP group(P<0.01);compared with MI/RI+PC6 group,the density of TH positive nerve fiber in MI/RI+AP group was lower(P<0.05).(2)The ratio of gray value of GAP43 protein:compared with SO group,the expression of GAP43 in MI/RI group was significantly increased(P<0.01);compared with MI/RI group,the expression of CTGF in MI/RI+PC6 group was decreased(P<0.05)and was significantly decreased in MI/RI+AP group(P<0.01);compared with MI/RI+PC6 group,the expression of CTGF in MI/RI+AP group was decreased(P<0.05).Conclusion1.The effect of oxidative stress and ST segment voltage of ECG:electroacupuncture preconditioning can promote the decrease of ST segment voltage of ECG,decrease the content of ROS in myocardial tissue and increase the activity of GSH-PX in serum to inhibit oxidative stress,so that effectively protect MI/RI model rats.The effect of electroacupuncture preconditioning with "Combined Biao and Ben Acupoints " is better than Neiguan single acupoint.2.The effect of myocardial remodeling:electroacupuncture preconditioning can inhibit the expression of CTGF,reduce the ratio of CVF and decrease the content of collagen fiber,improve the morphology of myocardial tissue,alleviate the pathological changes of myocardial fibrosis,so as to effectively protect the myocardial.The effect of electroacupuncture preconditioning with "Combined Biao and Ben Acupoints " is better than Neiguan single acupoint.3.The effect of sympathetic remodeling:electroacupuncture preconditioning can inhibit the expression of GAP43,reduce the springing of sympathetic nerve,improve the density of TH positive nerve fibers,decrease the secondary electrophysiological changes to prevent MI/RI.The effect of electroacupuncture preconditioning with "Combined Biao and Ben Acupoints" is better than Neiguan single acupoint. |
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