CAJ | 학술논문

目的评价吗啡预处理对心力衰竭大鼠离体心肌细胞缺氧复氧时miRNA表达的影响.方法清洁级健康成年雄性SD大鼠,体重200～ 220 g,制备大鼠心力衰竭模型并分离心室肌细胞接种于24孔板或60mm培养皿,采用随机数字表法分为3组(n=16):对照组(C组)、缺氧复氧组(H/R组)和吗啡预处理组(MP组).C组正常培养,H/R组和MP组细胞通入5％CO2-95％N2缺氧90 min后用含10％新生牛血清的高糖DMEM培养120 min.M组细胞缺氧前即刻采用吗啡培养液(吗啡终浓度为0.3μmol/L)培养10 min,再换为无吗啡培养液培养30 min进行预处理.于复氧120m in时各组随机取8孔细胞,采用台盼蓝染色法检测细胞活力和乳酸脱氢酶(LDH)活性.各组余8孔细胞提取总RNA行miRNA芯片检测,筛选各组差异表达的miRNA.结果与C组比较,H/R组和MP组心肌细胞活力降低,LDH活性升高,miR-6216和let-7e-5p上调,miR-133b-5p下调(P＜0.05);与H/R组比较,MP组心肌细胞活力升高,LDH活性降低,miR-133b-5p上调,miR-6216和let-7e-5p下调(P＜0.05).结论吗啡预处理通过调控miR-133b-5 P、miR-6216和let-7e-5p等miRNA表达减轻心力衰竭大鼠离体心肌细胞缺氧复氧损伤.
목적 평개마배예처리대심력쇠갈대서리체심기세포결양복양시miRNA표체적영향.방법 청길급건강성년웅성SD대서,체중200～ 220 g,제비대서심력쇠갈모형병분리심실기세포접충우24공판혹60mm배양명,채용수궤수자표법분위3조(n=16):대조조(C조)、결양복양조(H/R조)화마배예처리조(MP조).C조정상배양,H/R조화MP조세포통입5％CO2-95％N2결양90 min후용함10％신생우혈청적고당DMEM배양120 min.M조세포결양전즉각채용마배배양액(마배종농도위0.3μmol/L)배양10 min,재환위무마배배양액배양30 min진행예처리.우복양120m in시각조수궤취8공세포,채용태반람염색법검측세포활력화유산탈경매(LDH)활성.각조여8공세포제취총RNA행miRNA심편검측,사선각조차이표체적miRNA.결과 여C조비교,H/R조화MP조심기세포활력강저,LDH활성승고,miR-6216화let-7e-5p상조,miR-133b-5p하조(P＜0.05);여H/R조비교,MP조심기세포활력승고,LDH활성강저,miR-133b-5p상조,miR-6216화let-7e-5p하조(P＜0.05).결론 마배예처리통과조공miR-133b-5 P、miR-6216화let-7e-5p등miRNA표체감경심력쇠갈대서리체심기세포결양복양손상.
Objective To evaluate the effects of morphine preconditioning on the expression of microRNAs (miRNAs) during hypoxia-reoxygenation (H/R) in isolated cardiomyocytes in rats with heart failure.Methods Healthy adult male Sprague Dawley rats,w eighing 200-220 g,were used in this study.Adriamycin 2.0 mg/kg was injected once a week for 6 weeks via the tail vein to induce heart failure.The cardiomyocytes were isolated from the failing hearts of rats and seeded in 24-well plates or in 60 mm diameter dishes.The cells were then randomly divided into 3 groups (n =16 each) using a random number table:control group (group C); group H/R;morphine preconditioning group (group MP).The cells were cultured in normal culture atmosphere in group C.After being exposed to hypoxic air (5％ CO2-95％ N2) for 90 min,the cells were returned to the high-glucose DMEM supplemented with 10％ newborn bovine serum and were then cultured for 120 min in H/R and MP groups.In group M,the cells were cultured in morphine culture medium (final concentration of morphine 0.3 μmol/L) for 10 min and then were returned to the culture medium without morphine and cultured for 30 min immediately before hypoxia.At 120 min of reoxygenation,the cells of 8 wells in each group were chosen to detect the cell viability and lactate dehydrogenase (LDH) activity (by Typan blue staining).All the RNAs were extracted from the cardiomyocytes of the left 8 wells in each group and subjected to miRNA microarray to screen differentially expressed miRNAs.Results The cell viability was significantly lower,the activity of LDH was higher,the expression of miR-6216 and let7e-5p was higher,and the expression of miR-133b-5p was lower in H/R and MP groups than in group C (P ＜ 0.05).Compared with H/R group,the cell viability was significantly increased,the activity of LDH was decreased,the expression of miR-133b-5p was up-regulated,and the expression of miR-6216 and let-7e-5p was down-regulated in MP group (P ＜ 0.05).Conclusion Morphine preconditioning reduces H/R injury to isolated cardiomyocytes in rats with heart failure through regulating the expression of miRNAs such as miR133b-5p,miR-6216 and let-7e-5p.