hhlim对心肌肥大的影响及其作用机制探讨

hhlim Regulate Cardiac Hypertrophy In vitro

hhlim是从胎儿心脏中新近分离和克隆得到的与心脏发生相关的基因 ,其表达产物作为转录因子参与多种基因的转录调控和细胞的发育与分化过程 .用细胞转染方法将外源性hhlim基因导入原代心肌细胞 ,发现该基因强制性表达可使心肌细胞体积明显增大 .RT PCR和蛋白质印迹结果表明 ,hhlim促心肌细胞肥大与诱导α 肌动蛋白 (α actin)过表达及重新启动胚胎期表达基因脑钠肽 (BNP)表达有关 .用可表达hhlim反义RNA的真核表达载体转染心肌细胞后 ,致心肌细胞肥大因子ET 1对BNP和α actin表达的诱导受到显著抑制 .这些结果表明 ,ET 1促进BNP和α actin表达及引发心肌肥大的效应可能由hhlim所介导 ,提示hhlim表达与心肌细胞肥大的启动有关 .单独或共转染转录因子hhlim、Nkx2 5、GATA 4表达质粒和BNP转录调控区指导的报告基因结果显示 ,hhlim强制性表达不仅能直接激活BNP基因表达 ,而且与NKx2 5具有协同作用 .结果表明 ,hhlim可以通过直接或与Nkx2

hhlim is a new heart-related gene cloned from human embryonic heart whose product participates in transcriptional regulation and cell development as a kind of transcriptional factor. Over expression of hhlim gene using a recombinant plasmid was sufficient to induce a greater than 2.49 fold increase in cardiac myocyte area compared with that untransfected with hhlim. RT-PCR and Western blot testified that transfection of hhlim into the cardiac myocyte could induce skeletal a-actin over expression and trigger the expression of embryonic related gene-BNP, which is related to the cardiac hypertrophy. Antisense hhlim expression plasmid was constructed for analyze of hypertrophy in cultured neonatal cardiomyocytes. Endothelin-1 can induce cardiomyocyte hypertrophy. Cardiac myocyte treated by ET-1 was transfected with antisense hhlim palsmid. Western blot and RT-PCR analysis demonstraned that antisense hhlim restrained the increased cell surface area induced by ET-1, or increases expression of a-actin and BNP. Individual expression vectors for hhlim, Nkx2.5 and GATA-4 could enhance BNP reporter gene expression in cardiac myocytes. Cotransfection of hhlim and Nkx2.5 produced additive luciferase expression. The results demonstrate that hhlim protein is capable of initiating the hypertrophic response in cultured cardiac myocyte by activing BNP gene expression directly and indirectly.