The Role Of MiR-126 In Cardiac Myocytes Apoptosis And Vascular Endothelium Inflammation Induced By Hypoxia-Acidosis

BACKGROUDHypoxia-ischemia is a common pathophysiological process in cardiovascular diseases such as myocardial infarction and heart failure,which can cause disruption of myocardial cell energy metabolism and oxidative stress,leading to mitochondrial dysfunction and cell apoptosis.In addition,hypoxia-ischemia can cause dysfunction of vascular endothelial cells and inflammatory responses,which can lead to vascular damage and decreased contractile function.Therefore,the impact of hypoxia and acidosis on the cardiovascular system is a complex process involving multiple cell types and biological processes.A deeper understanding of the mechanisms underlying the effects of hypoxia-ischemia on the cardiovascular system can provide new insights and strategies for the prevention and treatment of cardiovascular diseases.MicroRNA play a regulatory role by binding to the 3’ untranslated region(3’UTR)of target genes,leading to the degradation of target messenger RNA or inhibition of target gene translation.MiR-126 is a highly expressed miRNA in vascular endothelial cells,which plays a critical role in maintaining endothelial cell integrity,regulating vascular development,preventing atherosclerosis,and regulating vascular function.Although miR-126 has been shown to play an important role in many cardiovascular diseases,its role in hypoxia-ischemia is not fully understood.OBJECTIVETo investigate the role of miR-126 in hypoxia-ischemia induced cardiac myocytes apoptosis and vascular endothelial inflammation,with the aim of providing new experimental evidence for a deeper understanding of the occurrence and development of cardiovascular diseases.RESEARCH CONTENTSSection 1:Regulatory Mechanisms of miR-126 in Myocardial Infarction:A Bioinformatics Analysis Based on Target Gene Pathways.Using NCBI’s GEO database to screen datasets,the GSE24591 dataset was selected for miRNA expression analysis.Differential analysis and plotting were performed using Sangerbox.miRNA-126-5p target gene prediction was performed using miRWalk.GO and KEGG pathway enrichment were performed using the clusterProfiler R package.The StringDB database was used to search for target gene interaction networks,and the interaction network of target genes was visualized using Cytoscape.Section 2:The role of miR-126 in hypoxia-acidosis induced apoptosis of cardiac myocytes.(1)Mouse myocardial cells were cultured under hypoxia-acidosis conditions,and the expression level of miR-126 was detected by RT-qPCR.The severity of apoptosis was evaluated using the TUNEL assay,and the levels of inflammatory factors TNF-α,IL-6,and IL-8 were detected by Western blot analysis.(2)Myocardial cells were transfected with antimiR-126 and then cultured under hypoxia-acidosis conditions.The expression of miR-126 was detected by RT-qPCR,the degree of apoptosis was evaluated using the TUNEL assay,and the levels of inflammatory factors TNF-α,IL-6,and IL-8 were detected by Western blot analysis.(3)Myocardial cells were transfected with antimiR-126 and then cultured under varying degrees of hypoxia-acidosis conditions.The levels of inflammatory factors TNF-α,IL-6,and IL-8 were detected by Western blot analysis.(4)Myocardial cells were transfected with antimiR-126 and then cultured under hypoxia-acidosis conditions.The expression of p-p38,p-JNK,and Bcl-2 was detected by Western blot analysis.Section 3:The role of miR-126 in hypoxia-acidosis induced inflammation of vascular endothelium(1)Endothelial cells were cultured under hypoxic conditions,and the levels of miR-126 and HMGB1 RNA were measured by RT-qPCR,while the protein expression level of HMGB1 was assessed by Western blot analysis.(2)Endothelial cells were subjected to hypoxic conditions,and the levels of NADPH,TNF-α,p-Akt,and Bcl-2 were measured by Western blot,while the level of ROS in cells was detected using the DCFH-DA probe.(3)After transfection with premiR-126 or antimiR-126,endothelial cells were subjected to hypoxic conditions.The expression of miR-126 and HMGB1 RNA was determined by RT-qPCR,while the level of HMGB1 protein was measured by Western blot analysis.(4)A dual luciferase reporter gene vector was constructed containing the wild-type or mutant HMGB1 gene 3’UTR seed sequence,and co-transfected with miR-126-5p to assess luciferase activity.(5)After transfection with premiR-126 or antimiR-126,endothelial cells were subjected to hypoxic conditions,and the levels of NADPH,TNF-α,p-Akt,and Bcl-2 were measured by Western blot analysis,while the level of ROS in cells was detected using the DCFH-DA probe.RESULTS1.Using the GEO database and Sangerbox analysis tools,it was found that miR126 was significantly upregulated in myocardial infarction tissues.The results of bioinformatics analysis showed that miR-126 could regulate the MAPK signaling pathway and had a regulatory effect on the target gene HMGB1.2.Under hypoxia-acidosis conditions,the expression of miR-126 in cardiac myocytes increased,and apoptosis and expression of inflammatory factors also significantly increased.After transfection with antimiR-126,the apoptosis and expression of inflammatory factors induced by hypoxia-acidosis in cardiac myocytes were reduced.3.Under hypoxia-acidosis conditions,the phosphorylation levels of p38 and JNK in cardiac myocytes were enhanced,and the expression of Bcl-2 was decreased.After transfection with antimiR-126,the phosphorylation levels of p38 and JNK were weakened,and the expression of the anti-apoptotic factor Bcl-2 was increased in cardiac myocytes.4.Under hypoxia-acidosis conditions,transfection of endothelial cells with antimiR-126 leads to increased expression of HMGB1 and upregulation of ROS,NADPH,and TNF-α expression,enhanced phosphorylation of Akt,and a significant increase in the expression of Bcl-2.5.Under hypoxia-acidosis conditions,miR-126 in endothelial cells promotes the degradation and suppresses the translation of HMGB1 gene by binding to the 3’UTR of the HMGB1 gene,resulting in decreased expression of HMGB1.CONCLUSIONS1.The expression of miR-126 is significantly upregulated in myocardial infarction tissues.Bioinformatic analysis results suggest that miR-126 can regulate the MAPK signaling pathway and has a regulatory effect on the target gene HMGB1.2.Under conditions of hypoxia-acidosis,miR-126 triggers an apoptotic response by activating the p38MAPK/JNK signaling pathway and suppressing the expression of Bcl-2 in cardiac myocytes.3.Under conditions of hypoxia-acidosis,miR-126 downregulates the inflammatory response by inhibiting the expression of the HMGB1 gene in cultured vascular endothelium cells.