Development of novel therapeutic strategy for heart failure by improving Ca2+ transport function of sarcoplasmic reticulum

1)激活SERCA2基因转录的新药物我们已经筛选了8万多个化合物，从中筛选出9个能够激活H9C2心肌细胞SERCA2基因转录的化合物。将这些化合物应用于主动脉缩窄所致的心力衰竭大鼠，观察这些化合物对SERCA2mRNA水平激活的影响。其中一种化合物使SERCA2的mRNA水平增加了20%。然而，由于在体内改善心力衰竭的效果还不够高，我们现在正在进一步搜索化合物文库，以寻找更有效的SERCA2基因治疗。将SERCA2基因导入心肌梗死心衰大鼠冠状动脉，观察SERCA2基因导入对心衰大鼠心功能的影响。基因转移7天后，转染SERCA2的细胞出现…信号短轴缩短率从16.5%增加到26%，而心脏收缩的非功能蛋白GFP基因转移对缩短率没有明显改善(从16%增加到19%)。这些变化与SERCA2基因表达水平和蛋白水平的上调相一致，提示慢病毒SERCA2基因转移系统作为治疗心力衰竭的手段是可行的。3)磷蛋白消融方法的建立利用HVJ膜将针对磷蛋白基因编码区的双链和21核苷酸序列导入新生大鼠心肌细胞。磷蛋白siRNA对靶基因的作用具有高度的基因特异性，其基因表达水平仅为对照组的10%。重要的是，钙吸收动力学发生了改变，使效率提高了38%。在过氧化氢诱导的心脏衰竭模型中，磷蛋白RNAi的这些有益作用也得到了证实。我们的研究表明，钙转运蛋白的基因调控在治疗心力衰竭方面有一定的疗效。较少

1)New pharmacological agent which activates the transcription of the SERCA2 geneWe have screened over 80,000 chemical compounds and picked up 9 compounds that activate the transcription of the SERCA2 gene in H9C2 cardiac cells. These compounds are applied to rats under heart failure induced by aortic banding and the effect of these compounds on the activation of the SERCA2 mRNA levels examined. One of these compounds increased the SERCA2 mRNA level by 20%. However, because the effect is not sufficiently high to improve heart failure in vivo, we are now further searching the compound library to find out more potent one.2)SERCA2 gene therapyWe have developed lentiviral gene transfer system bearing human SERCA2 cDNA. We introduced the gene construct into rat coronary artery of failing heart induced by myocardial infarction and examined the effect of SERCA2 gene introduction on the cardiac function of failing heart. Seven days after gene transfer, SERCA2-transfected demonstrated the signif … More icant increase of the fractional shortening from 16.5% to 26%, whereas the GFP, a non-functioning protein for contraction of heart, gene transfer did not significantly improve the fractional shortening (from 16 to 19%). These changed were corresponded with the upregulation of the SERCA2 mRNA and protein levels, which suggests the feasibility of the lentiviral SERCA2 gene transfer system as a therapeutic modality of heart failure.3)Development of phospholamban ablation methodDouble strand 21 ribonucleotide sequence specific for coding region of phospholamban gene was introduced into rat neonatal cardiac myocytes using HVJ envelope. The effect of phospholamban siRNA was highly gene specific for target mRNA and reduced its mRNA level to 10% of control group. Importantly, Ca2+ uptake kinetics was shifted to increase the efficiency by 38%. These beneficial effect of phospholamban RNAi was also demonstrated in the hydrogen peroxide-induced failing heart model. Decreased Ca2+ uptake was restored in the phospholamban ablation group.Our research suggests that genetic modulation of Ca2+ transporting protein has a therapeutic benefit in the treatment of heart failure. Less