NNOs And LncRNAs Are Invovled In The Protective Effect Of Myocardial Ischemic Postconditioning

Part one nNOS is Involved in Myocardial Ischemic Postconditioning by Alleviating the Opening of mPTPCardiomyocytes are severely deficient in nutrients and oxygen and the production of ROS and free radicals will increase,which all result in irreversible necrosis and injury of myocardial tissue during long-term ischemia.After long-term ischemia,the process can also cause a continuous damage to the myocardium even if myocardial tissue blood supply is recovery(myocardial ischemia and reperfusion),which is known as myocardial ischemia reperfusion injury.Previous studies have confirmed that Ischemic Postconditioning(IPostC)can significantly improve the myocardial injury induced by ischemia and reperfusion.IPostC refers to several circulations of transient ischemia and reperfusion before reperfusion.IPostC can activate the Reperfusion Injury Salvage Kinases(RISK)and other endogenous myocardial protective mechanism,which can significantly reduce the infarct size and protect the cardiac tissue.But the molecular mechanism involved in the process of IPostC is complex,it has not been clear,yet to be further improved.Previous studies have found that endogenous nitric oxide and nitric oxide synthases(including endothelial nitric oxide synthase,neuronal nitric oxide synthase and induced nitric oxide synthase)play an important protective role in the process of myocardial ischemia.Neuronal nitric oxide synthase(nNOS)has been concerned about the effect of protecting cardiac cells under different pathological conditions.In our previous study,we also found that IPostC can reduce mitochondrial oxidative stress and maintain the integrity of mitochondrial structure to improve cardiac function.However,selective nNOS inhibitors L-VNIO can significantly attenuated the protective effect of IPostC,prompting that nNOS may achieve the effect of protecting myocardial cells by protecting mitochondrial function in IPostC process.The irreversible opening of a large number of mitochondria permeability transition pores(mPTP)is one of the characteristics of cell death,and VDAC 1,important component of mPTP,is the most abundant protein expression in the outer membrane of mitochondria.Therefore,in this study,we also discussed how nNOS affect the opening of mPTP and the activity and expression of VDAC1.The objective,methods,results and conclusions of this study are as follows.Objective:To investigate the protective effect of nNOS on mitochondria and the effect of nNOS on the opening of mPTP and the expression and activity of VDAC1 in the process of IPostC.Methods:Ischemia reperfusion models were established using isolated hearts from mice with langendorff perfusion system.Cardiomyocytes hypoxia and reoxygenation(H/R)models were established with a hypoxia tank.using an in vitro hypoxia tank to make myocardial hypoxia/reoxygenation model to observe the change of cell morphology.The infarct sizt were detected by TTC Kit.Mitochondrial membrane potential was determined by JC-1 as a fluorescent signal using a flow cytometer.The opening of mPTP was determined by Calcein-AM as a fluorescent signals using fluorescence microscopy.The expression and activity of nNOS were detected by immunofluorescence technique.the expression and activity of VDAC1 were detected by immunofluorescence technique when adding or not adding nNOS selective inhibitors of L-VNIO.Results:nNOS participate in reducing mice cardiac infarct size;During ischemic postconditioning,the expression of nNOS increased;mitochondrial membrane polarization and the opening of mPTP were improved,and when combined use of nNOS inhibitor L-VNIO,the effect were weakened;ischemic postconditioning also reduced the expression of VDAC1,combined use of nNOS inhibitor L-VNIO weakened the the degree of decrease.Conclusion:nNOS is involved in ischemic postconditioning.nNOS participates in the protection of impaired myocardium and play a protective role in reducing mitochondrial membrane potential depolarization and improving opening of mPTP opening.This protective effect may be achieved by regulating the expression of VDAC1.Part two LncRNA Expression Character Related to Cardiac Ischemic Reperfusion InjuryIschemic reperfusion injury(IRI)contributes to morbidity and mortality worldwide,which resulted in poor outcome of myocardial infarction(MI)patients.Ischemic post-conditioning(IPostC),consisting of one or several brief periods of ischemia and reperfusion,generates powerful protection against ischemia-reperfusion injury.During the past decades,the mechanism of IPostC initiation and development has been explored in depth,while it’s still not clear.As rising stars in post-genome time,IncRNAs were found to play critical roles in kinds of cardiovascular diseases.However,there is little information about the systematic analysis of IRI-related lncRNA expression signature.In this study,we used microarray to analyze the lncRNA expression characters of ischemic postconditioning(IPostc)(corresponding to IRI),and found that there were 22 lncRNAs up-regulated and 12 lncRNAs down-regulated significantly.Subsequently,we identified that the dys-regulated lncRNAs participated in kinds of important biological progression though Gene ontology(GO)and Pathway analysis.Finally we validated lncRNA AK144818,ENSMUST00000156637,ENSMUST00000118342,ENSMUST000001 18149,uc008ane.1,ENSMUST00000164933,ENSMUST00000162347,AK050713,ENSMUST00000125121 and ENSMUST00000120587 should be related to the initiation and development of ischemic postconditioning(IPostC)This study might be helpful to understand the initiation and development mechanism of ischemic postconditioning(IPostC)comprehensively and probably afford the potential biomarkers for diagnosis or therapy targets for treat of IRI.The purpose,methods,results and conclusions of this study are as follows.Objective:To screen and verify LncRNA differentially expressed between IRI and IPostC organizations,initially revealed potential mechanisms in IPostC organization.Methods:(1)We were taken IRI and IPostC organization 3 parts,the use of chip technology LncRNA expression were detected IRI and IPostC LncRNAs organization to change and having more than 2 times statistically significant(P<0.05)as the criterion,screened LncRNAs differentially expressed;(2)Integrated NCBI RefSeq,UCSC,RNAdb,LncRNAs and other database resources,LncRNA differentially expressed preliminary biological Gene Ontology,Pathways and other informatics analysis;(3)expand the samples,preclude the use of real-time quantitative polymerase chain reaction(qRT-PCR)technique to verify the results of gene chip.Results:(1)the chip and the raw data were normalized between the two groups,LncRNAs differential expression of 4140,which raised the LncRNAs2292 strip,strip down the LncRNAs1848;(2)GO(Gene Ontology)analysis showed:cellular component,(CC):LncRNA these differentially expressed mainly in the extracellular space,extracellular domain and intracellular;biological process(BP):Its LncRNA mainly involved in regulation of immune process,stress reaction;molecular functions(MF):The main having a protein binding binding,zinc ion binding and other functions.Pathways analysis showed:These LncRNA mainly involved in cytokine-cytokine receptor interaction,malaria,African trypanosomiasis,TNF signaling pathway,rheumatoid arthritis,chemokine signaling pathway,dilated cardiomyopathy and so on(3)qRT-PCR was found in IPostC organizations and 34 had LncRNAs significantly differentially expressed,including 22 upregulated LncRNAs,12 were down LncRNAs.Conclusion:Compared with IRI organizations IPostC organizations exists differentially expressed LncRNAs,these significant changes may be IPostC organizations LncNRAs biomarkers and potential therapeutic targets.