Background Liraglutide is glucagon-like peptide-1 receptor agonist for treating patients with type 2 diabetes mellitus. Our previous studies have demonstrated that liraglutide protects cardiac function through improvi...Background Liraglutide is glucagon-like peptide-1 receptor agonist for treating patients with type 2 diabetes mellitus. Our previous studies have demonstrated that liraglutide protects cardiac function through improving endothelial function in patients with acute myocardial infarction undergoing percutaneous coronary intervention. The present study will investigate whether liraglntide can perform direct protective effects on cardiomyocytes against reperfusion injury. Methods In vitro experiments were performed using H9C2 cells and neonatal rat ventricular cadiomyocytes undergoing simulative hypoxia/reoxygenation （H/R） induction. Cardiomyocytes apoptosis was detected by fluorescence TUNEL. Mitochondrial membrane potential （AWm） and intracellular reactive oxygen species （ROS） was assessed by JC-1 and DHE, respectively. Fura-2/AM was used to measure intracellular Ca2＋ concentration and calcium transient. Immtmofluorescence staining was used to assess the expression level of sarcoplasmic reticulum Ca2＋-ATPase （SERCA2a）. In vivo experiments, myocardial apoptosis and expression of SERCA2a were detected by colorimetric TUNEL and by immunofluorescence staining, respectively. Results In vitro liraglutide inhibited cardiomyotes apoptosis against H/R. △mψ of cardiomyocytes was higher in liraglntide group than H/R group. H/R increased ROS production in H9C2 cells which was attenuated by liraglutide. Liraglutide significantly lowered Ca2＋ overload and improved calcium transient compared with H/R group, lmmunofluorescence staining results showed liraglutide promoted SERCA2a expression which was decreased in H/R group. In ischemia/reperfusion rat hearts, apoptosis was significantly attenuated and SERCA2a expression was increased by liraglutide compared with H/R group. Conclusions Liraglutide can directly protect cardiomyocytes against reperfusion injury which is possibly through modulation of intracellular calcium homeostasis.展开更多
Objective.To investigate the protective effects of melatonin(MT )on cardiomyocytes against the ox-idative injury induced by H 2 O 2 .Methods.Ventricular myocytes were isolated from neonatal rats and cultured for3～5da...Objective.To investigate the protective effects of melatonin(MT )on cardiomyocytes against the ox-idative injury induced by H 2 O 2 .Methods.Ventricular myocytes were isolated from neonatal rats and cultured for3～5days.Cells were divided into4groups:control group,H 2 O 2 (100μmol/L H 2 O 2 )group,MT group(100μmol/L MT )and MT preconditioning group(100μmol/L MT+100μmol/L H 2 O 2 ).The fluorescent probe,DCFH?DA,was used to detect intracellular levels of reactive oxygen species(ROS)and another fluorescent probe,Fluo-3?AM,was used to detect[Ca 2+ ] i by using a laser scanning confocal microscopy(LSCM).The malondi-aldehyde(MDA)content in cardiomyocytes was determined by measuring thiobarbituric acid?reactive sub-stances to monitor lipid peroxidation.The activity of cytoplasmic enzyme lactate dehydrogenase(LDH)that was released into the culture media was assayed to indicate alternation in the integrity of the cellular membrane.Trypan blue exclusion was used to detect the cell viability.Results.Compared with the control group,intracellular ROS,[Ca 2+ ] i ,MDA content ,LDH leakage and cell death were significantly elevated when cells were treated by100μmol/L H 2 O 2 for60minutes(P<0.01).However,those changes were significantly attenuated in MT preconditioning group.Conclusion.MT has very good antioxidant effect and can protect the cardiomyocytes against H 2 O 2 -in-duced injury.展开更多
基金This work is supported by grants from National Natural Science Foundation of China （No. 81102079） and China Postdoctoral Science Foundation （No. 201003776）. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. The authors report no conflict of interest. The authors are responsible for the content and writing of the paper.
文摘Background Liraglutide is glucagon-like peptide-1 receptor agonist for treating patients with type 2 diabetes mellitus. Our previous studies have demonstrated that liraglutide protects cardiac function through improving endothelial function in patients with acute myocardial infarction undergoing percutaneous coronary intervention. The present study will investigate whether liraglntide can perform direct protective effects on cardiomyocytes against reperfusion injury. Methods In vitro experiments were performed using H9C2 cells and neonatal rat ventricular cadiomyocytes undergoing simulative hypoxia/reoxygenation （H/R） induction. Cardiomyocytes apoptosis was detected by fluorescence TUNEL. Mitochondrial membrane potential （AWm） and intracellular reactive oxygen species （ROS） was assessed by JC-1 and DHE, respectively. Fura-2/AM was used to measure intracellular Ca2＋ concentration and calcium transient. Immtmofluorescence staining was used to assess the expression level of sarcoplasmic reticulum Ca2＋-ATPase （SERCA2a）. In vivo experiments, myocardial apoptosis and expression of SERCA2a were detected by colorimetric TUNEL and by immunofluorescence staining, respectively. Results In vitro liraglutide inhibited cardiomyotes apoptosis against H/R. △mψ of cardiomyocytes was higher in liraglntide group than H/R group. H/R increased ROS production in H9C2 cells which was attenuated by liraglutide. Liraglutide significantly lowered Ca2＋ overload and improved calcium transient compared with H/R group, lmmunofluorescence staining results showed liraglutide promoted SERCA2a expression which was decreased in H/R group. In ischemia/reperfusion rat hearts, apoptosis was significantly attenuated and SERCA2a expression was increased by liraglutide compared with H/R group. Conclusions Liraglutide can directly protect cardiomyocytes against reperfusion injury which is possibly through modulation of intracellular calcium homeostasis.
文摘Objective.To investigate the protective effects of melatonin(MT )on cardiomyocytes against the ox-idative injury induced by H 2 O 2 .Methods.Ventricular myocytes were isolated from neonatal rats and cultured for3～5days.Cells were divided into4groups:control group,H 2 O 2 (100μmol/L H 2 O 2 )group,MT group(100μmol/L MT )and MT preconditioning group(100μmol/L MT+100μmol/L H 2 O 2 ).The fluorescent probe,DCFH?DA,was used to detect intracellular levels of reactive oxygen species(ROS)and another fluorescent probe,Fluo-3?AM,was used to detect[Ca 2+ ] i by using a laser scanning confocal microscopy(LSCM).The malondi-aldehyde(MDA)content in cardiomyocytes was determined by measuring thiobarbituric acid?reactive sub-stances to monitor lipid peroxidation.The activity of cytoplasmic enzyme lactate dehydrogenase(LDH)that was released into the culture media was assayed to indicate alternation in the integrity of the cellular membrane.Trypan blue exclusion was used to detect the cell viability.Results.Compared with the control group,intracellular ROS,[Ca 2+ ] i ,MDA content ,LDH leakage and cell death were significantly elevated when cells were treated by100μmol/L H 2 O 2 for60minutes(P<0.01).However,those changes were significantly attenuated in MT preconditioning group.Conclusion.MT has very good antioxidant effect and can protect the cardiomyocytes against H 2 O 2 -in-duced injury.
文摘目的研究表明桑白皮多糖具有抗氧化作用,而其对心肌细胞氧化损伤的影响尚不清楚,文章旨在探讨桑白皮多糖对缺氧/复氧诱导的心肌细胞损伤的作用及分子机制。方法H9C2细胞缺氧培养6 h后再复氧6 h,作为缺氧/复氧(H/R)组,不做任何处理的细胞作为正常对照组,用0.1、0.2、0.4μg/mL的桑白皮多糖预处理H9C2细胞,而后进行缺氧/复氧处理,作为H/R+桑白皮多糖低、中、高剂量组;将pcDNA、pcDNA-circDLGAP4转染至H9C2细胞后进行缺氧/复氧处理,记为H/R+pcDNA组、H/R+pcDNA-circDLGAP4组;将si-circDLGAP4转染至H9C2细胞后用0.4μg/mL桑白皮多糖处理,再进行缺氧/复氧处理,记为H/R+桑白皮多糖高剂量+si-circDLGAP4组。细胞计数试剂盒8(CCK-8)、流式细胞术分别检测以上各组细胞活性和凋亡率;丙二醛(MDA)、超氧化物歧化酶(SOD)和乳酸脱氢酶(LDH)试剂盒分别检测MDA含量、SOD活性和LDH活性;实时荧光定量PCR(RT-qPCR)检测各组细胞circDLGAP4和miR-320的表达水平。双荧光素酶报告实验检测circDLGAP4和miR-320的靶向关系。结果与正常对照组相比,H/R组H9C2细胞活性降低,凋亡率升高,Bax表达水平升高,Bcl-2表达水平降低(P<0.05);与H/R组相比,H/R+桑白皮多糖低、中、高剂量组H9C2细胞活性升高,凋亡率降低,Bax表达水平降低,Bcl-2表达水平升高,呈剂量依赖性(P<0.05)。与正常对照组H9C2细胞MDA[(18.35±1.11)nmL/mL]、LDH(26.81±1.11)U/L,SOD活性[(65.53±2.75)U/L]相比,H/R组H9C2细胞中MDA含量[(95.02±3.67)nmL/mL]及LDH活性[(133.97±5.30)U/L]升高,SOD活性[(11.92±0.63)U/L]降低(P<0.05);与H/R组相比,H/R+桑白皮多糖低、中、高剂量组H9C2细胞中MDA含量[(71.43±3.00)、(53.62±1.69)、(35.81±1.36)nmL/mL]及LDH活性[(106.43±4.00)、(84.50±2.68)、(58.51±1.94)U/L]降低,SOD活性[(24.60±1.14)、(42.37±2.26)、(58.27±2.61)U/L]升高,呈剂量依赖性(P<0.05)。与正常对照组相比,H/R组H9C2细胞中circDLGAP4表达水平降低,miR-320表达水平升高(P<0.05);与H/R组相比,H/R+桑白皮多糖低、中、高剂量组H9C2细胞中circDLGAP4表达水平升高,miR-320表达水平降低,呈剂量依赖性(P<0.05)。与pcDNA组相比,pcDNA-circDLGAP41、2、3组circDLGAP4表达水平升高。与H/R+pcDNA组相比,H/R+pcDNA-circDLGAP4组circDLGAP4表达水平升高,miR-320表达水平降低,MDA含量、LDH活性降低,SOD活性升高;H9C2细胞活性升高,H9C2细胞凋亡率及Bax表达水平降低,而Bcl-2表达水平升高(P<0.05)。与H/R+桑白皮多糖高剂量组相比,H/R+桑白皮多糖高剂量+si-circDLGAP4组circDLGAP4表达水平降低,miR-320表达水平升高,H9C2细胞中MDA含量、LDH活性升高,SOD活性降低,H9C2细胞活性降低,H9C2细胞凋亡率及Bax表达水平升高,而Bcl-2表达水平降低(P<0.05)。circDLGAP4和miR-320的互补序列。wt-circDLGAP4与miR-320共转染的细胞荧光素酶活性较与miR-NC共转染的降低(0.96±0.08 vs 0.35±0.03,P<0.05)。结论桑白皮多糖通过调控circDLGAP4/miR-320对缺氧/复氧诱导的心肌细胞损伤起保护作用。
