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Novel Therapy for Heart Failure of cardiomyopathic Hamster with Gene Transfection of Hepatocyte Growth Factor

肝细胞生长因子基因转染治疗心肌病仓鼠心力衰竭的新疗法

基本信息

批准号：
11670729
负责人：
KOMAMURA Kazuo
金额：
$ 2.43万
依托单位：
Research Institute, National Cardiovascular Center
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

1) We integrated rat Hepatocyte growth factor CDNA (rHGF) into PCAGGS plasmid vector, which has β-actin promoter and affinity to muscle cells (PCAGGS-rHGF). Likewise, we made PCAGGS-human HGR We could confirm expression of rHGF alone, and used this forthe experiment.2) Ultrasound alone did not enhance transfection of rHGF. Sonoporation wrth microbubble made rat myocardial culture cells excrete HGF into medium.3) We tried direct injection of genes into the heart of cardiomyopathic hamster. Reporter gene LacZ was found to be transfected in the myocaraium. There was no excretion of HGF protein into peripheral blood.4) In vivo electroporation of LacZ and interleukin 5 into skeletal musde of hamster successfully transfected these genes. We employed this method, 6 pulses of 100V 50 ms, as a standard in vivo transfection method.5) We tried in vivo electroponation of rHGF into tibialis anterior musde of cardiomyopathic hamsterTO-2 using the dose of 200, 400 and 800 μg. Dose of 800 μg. resulted into maximum blood concentration of HGF. This dose with standard in vivo transfection method turned out to be a sustained elevation of HGF concentration over 5 ng/mL. Cardiac function estimated by echo-Doppler improved in HGF-treated hamsters compared with normal control and placebo given hamsters. Myocardial fibrosis was suppressed and capillary density was increased in HGF-treated hamsters compared with placebo-given hamsters. Increase in MMP-1 activity might be attributable to one of possible mechanisms of beneficial effects of HGF treatment for heart failure in cardiomyopathic hamsters (submitted).

1)将大鼠肝细胞生长因子cDNA（rHGF）整合到具有β-actin启动子和肌细胞亲和性的PCAGGS质粒载体（PCAGS-rHGF）中。同样，我们制备了PCAGGS-人HGR，我们可以证实单独的rHGF表达，并将其用于实验。2）单独的超声不能增强rHGF的转染。微泡声穿孔法使大鼠心肌细胞分泌HGF。3）心肌病仓鼠心脏直接注射基因。报告基因LacZ被转染到心肌细胞中。4）将LacZ和IL-5基因在体内电穿孔转染仓鼠骨骼肌，成功地转染了两种基因。5）将rHGF以200、400和800 μg的剂量电穿孔到心肌病仓鼠TO-2的胫骨前肌中。剂量为800 μg。导致HGF的最大血液浓度。使用标准体内转染方法的该剂量被证明是HGF浓度超过5 ng/mL的持续升高。与正常对照组和安慰剂组仓鼠相比，HGF治疗组仓鼠的心脏功能通过超声多普勒评估得到改善。与给予安慰剂的仓鼠相比，HGF治疗的仓鼠心肌纤维化受到抑制，毛细血管密度增加。MMP-1活性的增加可能归因于HGF治疗心肌病仓鼠心力衰竭的有益作用的可能机制之一（已提交）。