Cheng Wen-Pin, Wang Bao-Wei, Lo Huey-Ming, Shyu Kou-Gi
Department of Medical Education and Research, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan.
Division of Cardiology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan; School of Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan.
PLoS One. 2015 Apr 21;10(4):e0123235. doi: 10.1371/journal.pone.0123235. eCollection 2015.
The expression of TRB3 (tribbles 3), an apoptosis regulated gene, increases during endoplasmic reticulum (ER) stress. How mechanical stress affects the regulation of TRB3 in cardiomyocytes during apoptosis is not fully understood. An in vivo model of aorta-caval shunt in adult rats demonstrated the increased TRB3 protein expression in the myocardium. The tumor necrosis factor-alpha (TNF-α) antagonist etanercept reversed the TRB3 protein expression and cardiomyocyte apoptosis induced by AV shunt. An in vitro model of cyclic stretch in neonatal rats was also used to investigate TRB3 expression. We hypothesized that cardiomyocyte apoptosis induced by cyclic stretch is TRB3 dependent. Neonatal rat cardiomyocytes grown on a flexible membrane base were stretched by vacuum to 20% of maximum elongation, at 60 cycles/min. Cyclic stretch significantly increased TRB3 protein and mRNA expression. Addition of c-jun N-terminal kinase (JNK) inhibitor SP600125, TNF-α antibody and etanercept 30 min before stretch reversed the induction of TRB3 protein induced by stretch. Cyclic stretch induced the DNA-binding activity of growth arrest and DNA damaged inducible gene-153 (GADD153) by electrophoretic mobility shift assay. SP600125, JNK siRNA, TNF-α antibody and etanercept abolished the binding activity induced by stretch. TRB3 promoter activity was enhanced by stretch and TRB3-mut plasmid, SP600125, TNF-α antibody and etanercept attenuated TRB3 promoter activity induced by stretch. Exogenous administration of TNF-α recombinant protein to the non-stretched cardiomyocytes increased TRB3 protein expression similar to that seen after stretch. Cyclic stretch induced cardiomyocyte apoptosis is inhibited by TRB3 siRNA and etanercept. The stretch-induced TRB3 is mediated by TNF-α、JNK and GADD153 pathway. These results indicate that TRB3 plays an important role in stretch-induced cardiomyocyte apoptosis.
TRB3（ Tribbles 3）是一种凋亡调节基因，其表达在内质网（ER）应激期间会增加。机械应激如何在凋亡过程中影响心肌细胞中TRB3的调节尚未完全清楚。成年大鼠主动脉-腔静脉分流的体内模型显示心肌中TRB3蛋白表达增加。肿瘤坏死因子-α（TNF-α）拮抗剂依那西普可逆转AV分流诱导的TRB3蛋白表达和心肌细胞凋亡。还使用新生大鼠的体外循环拉伸模型来研究TRB3的表达。我们假设循环拉伸诱导的心肌细胞凋亡是TRB3依赖性的。在柔性膜基质上生长的新生大鼠心肌细胞通过真空拉伸至最大伸长率的20%，频率为60次/分钟。循环拉伸显著增加了TRB3蛋白和mRNA表达。在拉伸前30分钟添加c-jun N末端激酶（JNK）抑制剂SP600125、TNF-α抗体和依那西普可逆转拉伸诱导的TRB3蛋白的诱导。循环拉伸通过电泳迁移率变动分析诱导生长停滞和DNA损伤诱导基因-153（GADD153）的DNA结合活性。SP600125、JNK siRNA、TNF-α抗体和依那西普消除了拉伸诱导的结合活性。TRB3启动子活性通过拉伸增强，TRB3突变质粒、SP600125、TNF-α抗体和依那西普减弱了拉伸诱导的TRB3启动子活性。向未拉伸的心肌细胞外源性给予TNF-α重组蛋白可增加TRB3蛋白表达，类似于拉伸后所见。TRB3 siRNA和依那西普可抑制循环拉伸诱导的心肌细胞凋亡。拉伸诱导的TRB3由TNF-α、JNK和GADD153途径介导。这些结果表明TRB3在拉伸诱导的心肌细胞凋亡中起重要作用。
