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

1)激活SERCA2基因转录的新药理学试剂我们筛选了超过80，000种化合物，并挑选了9种激活H9C2心脏细胞中SERCA2基因转录的化合物。将这些化合物应用于由主动脉缩窄诱导的心力衰竭的大鼠，并检查这些化合物对SERCA2 mRNA水平的活化的作用。其中一种化合物使SERCA2 mRNA水平增加了20%。然而，由于其在体内的效果不足以改善心力衰竭，我们正在进一步寻找化合物库，以找到更有效的化合物。2）SERCA 2基因治疗我们开发了携带人SERCA 2 cDNA的慢病毒基因转移系统。我们将该基因导入心肌梗死诱导的心力衰竭大鼠冠状动脉，观察SERCA 2基因导入对心力衰竭大鼠心功能的影响。基因转移后7天，SERCA2转染组显示出显著的细胞凋亡， ...更多信息 缩短分数从16.5%显著增加到26%，而GFP（心脏收缩的非功能性蛋白质）基因转移没有显著改善缩短分数（从16%到19%）。这些变化与SERCA 2基因表达和蛋白水平的上调相一致，提示慢病毒SERCA 2基因转移系统作为心力衰竭治疗手段的可行性。3）受磷蛋白消融方法的建立利用HVJ包膜将受磷蛋白基因编码区特异性的双链21核苷酸序列导入新生大鼠心肌细胞。受磷蛋白siRNA的作用对靶mRNA具有高度基因特异性，并将其mRNA水平降低至对照组的10%。重要的是，Ca2+吸收动力学发生了变化，使效率提高了38%。受磷蛋白RNAi的这些有益作用也在过氧化氢诱导的心力衰竭模型中得到证实。受磷蛋白消融组的Ca2+摄取恢复，我们的研究表明，Ca2+转运蛋白的遗传调节在心力衰竭的治疗中具有治疗益处。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