| Objectivecoronary artery disease is common in the elderly,which can gradually develop into myocardial ischemia,and a series of treatments for coronary recanalization are needed.Myocardial ischemia / reperfusion(I/R)injury refers to the injury of cells and myocardial tissue in myocardial reperfusion after coronary artery recanalization,which impairs cardiac function.There are a lot of research results on the mechanism of myocardial ischemia-reperfusion injury(MIRI),among which inflammation is one of the more important mechanisms.Nevertheless,there are still deficiencies in the study of the pathophysiological mechanism of the disease,which greatly limits the progress of clinical treatment of MIRI.Therefore,it is necessary to further explore the mechanism of disease occurrence and progression in order to promote the optimization of MIRI treatment.With the large increase of surgical patients,the number of patients at risk of MIRI also increases.We expect to use drugs during operation to reduce the occurrence of MIRI.Sevoflurane is an inhalation anesthetic.Studies have shown that Sevoflurane can reduce the occurrence and development of MIRI by inhibiting inflammation.In this study,we observed the effect of Sevoflurane on myocardial function in patients with myocardial ischemia.In addition,the injury model of cardiomyocyte hypoxia/reoxygenation(H/R)was constructed with primary rat ventricular myocytes(NRVM)to simulate the occurrence of inflammation and pyroptosis of cardiomyocytes in the H/R.Then NLRP3 inhibitors and agonists were used to observe the effects on the changes of H / R injury of cardiomyocytes,so as to determine the inflammation and pyroptosis of cardiomyocytes.Further verify the observation in the rat experiment,and then explore the molecular mechanism of Sevoflurane in anti NRVM H / R injury and rat MIRI.Methods117 patients undergoing abdominal surgery under general anesthesia were recruited in this study.87 patients with a history of myocardial ischemia were divided into Sevoflurane group and propofol group.30 patients without a history of myocardial ischemia were used as the control group.The clinical data of patients,the changes of blood pressure,heart rate,rate pressure product(RPP)and pressure rate ratio(PRQ)during anesthesia and cardiac related complications after anesthesia were recorded.In addition,heart type fatty acid binding protein(H-FABP),ischemia modified albumin(IMA),IL-1β and IL-18 were measured at different time points during perioperative anesthesia.The NRVM H / R injury model was established.The 1.8%,2.4% and 3.6%Sevoflurane groups were pretreated with different concentrations of Sevoflurane for 45 minutes,and then treated with hypoxia for 3 hours and reoxygenation for 1hour.After treatment,the m RNA expressions of P2X7 and NLRP3 were detected by quantitative PCR,and the expressions of the following proteins were detected by Western blot: P2X7,NLRP3,apoptosis associated speck like protein(ASC),cysteine aspartic acid specific protein-1(caspase-1),pyroporin-d(gsdmd)Bcl-2associated X protein(Bax)and B-cell lymphoma-2(Bcl-2).To reflect the occurrence of myocardial inflammation and pyroptosis after NRVM H / R.The rat MIRI model was established.After treatment with Sevoflurane at different concentrations,ischemia for 45 minutes and reperfusion for 3 hours.To detect the cardiac function of rats under ultrasound.TTC / Evans blue double staining was used to detect the myocardial infarction area after MIRI in rats.Detected IL-1β,IL-18,malondialdehyde(MDA),superoxide dismutase(SOD),lactate dehydrogenase(LDH),creatine kinase(CK)and creatine kinase isoenzymes(CK-MB)to reflect the degree of cardiomyocyte injury.The expression levels of inflammatory factors and signal pathway factors in myocardial tissue were detected by HE staining,immunofluorescence,DHE staining and WB.The role of P2X7-NLRP3 signaling pathway in Sevoflurane reducing MIRI damage was clarified by using NLRP3 inhibitors and agonists.ResultsPart I Sevoflurane alleviates myocardial injury in patients with myocardial ischemia1.The blood pressure and heart rate in Sevoflurane and propofol groups were lower than those before anesthesia.The inhibitory effect of propofol on blood pressure was more obvious.RPP<12000 and PRQ>1.0 were achieved in all groups,but PRQ was close to 1.0 in the propofol group,which may be related to the inhibitory effect of propofol on blood pressure.Compared with the control group,the incidence of new ECG ST-T abnormalities was decreased at 24 h and48h after anesthesia in the Sevoflurane group.Among the three groups,the incidence of unstable angina pectoris 30 days after anesthesia was highest in the propofol group.2.IMA,H-FABP,IL-1β and IL-18 increased in Sevoflurane group and propofol group at 0h,8h and 24 h after anesthesia,but the increase was more obvious in propofol group,indicating that Sevoflurane,also a general anesthesia drug,had a better effect on myocardial injury and inflammation inhibition.Part II effects of Sevoflurane on NRVM H / R injury and MIRI in rats3.After MIRI,LVEF and LVFS decreased significantly,myocardial infarction area and myocardial enzyme release increased.After Sevoflurane treatment,LVEF and LVFS were improved,myocardial infarction and injury indexes were also decreased,and the effect of 3.6% Sevoflurane group had the best effect.4.Myocardial apoptosis and oxidative stress were significantly increased after MIRI in rats.Sevoflurane treatment significantly reduced the occurrence of apoptosis and oxidative stress,indicating that Sevoflurane has a certain myocardial protection effect.Part III Sevoflurane inhibits cardiomyocyte inflammation and apoptosis through P2X7-NLRP3 signaling pathway1.After MIRI,the number of inflammatory cells in myocardium increased,the expression of NLRP3 and GSDMD increased,the levels of IL-1β and IL-18 in serum increased,and inflammatory cells and cytokines decreased in Sevoflurane group.2.The protein expression of P2X7,NLRP3,ASC and Caspase-1 decreased in Sevoflurane group,and the protein expression of GSDMD increased after MIRI in rats,indicating apoptosis was increased,but Sevoflurane decreased apoptosis.After NRVM H/R,m RNA expression of P2X7 and NLRP3,protein expression of P2X7,NLRP3,ASC,Caspase-1 and GSDMD increased.Different concentrations of Sevoflurane decreased the expression of P2X7 and NLRP3 m RNA,P2X7,NLRP3,ASC and Caspase-1 proteins.3.After NRVM was hypoxic and reoxygenated,the co-localization of NLRP3 and ASC was observed by confocal laser microscopy.It was observed that NLRP3 and ASC was co-located in the cytoplasm and formed spots.4.The NLRP3 inhibitor MCC950 100 n M inhibits the expression of NLRP3,caspase-1 and GSDMD,and reduces the secretion of IL-1β and IL-18 and the release of LDH.The NLRP3 agonist Nigericin 10 u M increased the expression of P2X7,NLRP3,caspase-1 and GSDMD proteins,and increased the secretion and release of IL-1β,IL-18 and LDH.Conclusion1.Sevoflurane has myocardial protection in patients with myocardial ischemia during operation.2.Sevoflurane inhibits cardiomyocyte inflammation and apoptosis through p2X7-NLRP3 signaling pathway in MIRI. |
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