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目的:探讨不同浓度肿瘤坏死因子-α(TNF-α)对培养的乳鼠心肌细胞活力、蛋白合成、分泌AngⅡ及AT1受体表达的影响。方法:体外培养的SD乳鼠心肌细胞随机分为对照组和不同浓度TNF-α(20、40、60、80、100μg/L)干预组,用BCA法测定心肌细胞蛋白合成总量,MTT比色法和LDH检测反映心肌细胞的活力,ELISA法检测心肌细胞培养液中AngⅡ的含量,免疫细胞化学染色法检测心肌细胞膜AT1受体表达的变化。结果:TNF-α浓度依赖性地增强乳鼠心肌细胞活力、增加蛋白合成,20、40、60、80μg/L TNF-α组细胞活力较对照组分别增加1.21(P<0.05)、1.42、1.51和1.73倍(均P<0.01),蛋白合成分别增加27.8%(P<0.05)、38.9%、46%和66.7%(均P<0.01),而LDH含量差异无显著性。100μg/L TNF-α组与对照组比较,心肌细胞活力降低18.5%(P<0.01)、蛋白合成降低18.3%(P<0.01)及心肌LDH生成增加1.48倍(P<0.01)。TNF-α浓度依赖性地增加乳鼠心肌细胞AngⅡ分泌,与对照组比较分别增加0.5、1.1、1.6、3和3.6倍(均P<0.01)。TNF-α还具有诱导AT1受体的表达的作用。结论:TNF-α可促进心肌细胞内源性AngⅡ产生,引起心肌细胞活力、蛋白合成改变,AT1受体表达上调,可能介导了心肌肥大、心肌受损等心肌改建的病理生理过程。
Objective: To investigate the effects of tumor necrosis factor-α (TNF-α) on the viability, protein synthesis, AngⅡ and AT1 receptor expression of cultured neonatal rat cardiomyocytes. Methods: Cardiomyocytes from neonatal SD rats were randomly divided into control group and intervention group with different concentrations of TNF-α (20, 40, 60, 80 and 100 μg / L). Cardiomyocyte protein synthesis, The colorimetric method and LDH test reflected the viability of cardiomyocytes, the content of AngⅡ in cardiomyocyte culture medium was detected by ELISA, the expression of AT1 receptor in myocardial cell membrane was detected by immunocytochemical staining. Results: Compared with control group, the viability of TNF-α group increased by 1.42 (P <0.05), 1.42 and 1.51 And 1.73 folds (all P <0.01). The protein synthesis increased by 27.8% (P <0.05), 38.9%, 46% and 66.7% respectively (all P <0.01), while the LDH content had no significant difference. The viability of myocardial cells was decreased by 18.5% (P <0.01), the protein synthesis decreased by 18.3% (P <0.01) and the myocardial LDH production was increased by 1.48 times (P <0.01) compared with the control group. TNF-α increased AngⅡ secretion in neonatal rat cardiocytes in a concentration-dependent manner, which were increased by 0.5, 1.1, 1.6, 3 and 3.6 times (all P <0.01) compared with the control group. TNF-α also has the effect of inducing AT1 receptor expression. CONCLUSION: TNF-α can promote the production of endogenous AngⅡ in cardiomyocytes, induce the changes of myocardial cell viability, protein synthesis and up-regulation of AT1 receptor, which may be involved in the pathophysiological process of myocardial remodeling.