Cell transplantation to myocadial infarction area

• Project/Area Number: 12470267
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Thoracic surgery
• Research Institution: Tohoku University
• Principal Investigator: SAI Sadahiro Tohoku University Hospital, Research Associate, 医学部/附属病院, 助手 (60312576)
• Co-Investigator(Kenkyū-buntansha): IGUCHI Atsushi Tohoku University, Graduate School of Medicine, Associate Professor, 大学院・医学系研究科, 助教授 (90222851)
• Project Period (FY): 2000 – 2001
• Project Status: Completed (Fiscal Year 2001)
• Budget Amount: ¥10,800,000 (Direct Cost: ¥10,800,000); Fiscal Year 2001: ¥900,000 (Direct Cost: ¥900,000); Fiscal Year 2000: ¥9,900,000 (Direct Cost: ¥9,900,000)
• Keywords: bioenginering / ES cell / cell transplantation / gene therapy / heart failure / myocardial infarction / myosin heavychain / promoter / 再生医療 / 心筋細胞移植 / ES細胞 / 細胞分化 / 心筋虚血 / 心筋梗塞 / 慢性心不全

Research Abstract

The purpose of the present study was to evaluate the application of cell transplantation into myocardium in a rodent infarction model. Embryonic stem (ES) cells were cultured and induced to differentiate inadvertently, and cells expressing myocardial phenotype revealed. We postulated that ES cells differentiating to cardiomyocytes expressed alpha-myosin heavy chain. We constructed an expression gene that consisted of alpha-myosin heavy chain promoter and cDNA encoding EGFP-IRES-zeocin and puromycin resistant gene, and this gene was transfected into ES cells. ES cells transfected with the gene were cultured in medium supplemented with zeoncin, and the transfected clones were determined by PCR. Cells were cultured in medium not supplemented with human leukemia inhibitory factor and differentiated to form embryoidbody. Some cells of the embryoidbody started beating spontaneously. EGFP expressing cells were existed but we could not detect enough number of the cells transfected with the gen e by flowcytometric analysis. We performed puromycin selection alternatively, but the selection was not effective to detect enough number of beating cells. Since the specific master genes for the differentiation to myocardial cells were not detected. To isolate the myocardial-phenotype cells from cultured deferentiated ES cells seemed to be a promising strategy for us. However, the transfected gene did not activated well in ES cells unexpectedly. The transfected gene seemed to be tuned off by unknown gene inactivation mechanism. It may be a methylation of transduced promoter gene to be speculated as one of the possible mechanism of the inactivation. To evaluate the effectiveness of the cell transplantation, we used cryo-injured model. Myocardium of Fisher rat were injured with a cryogenic probe and regional movement of the left ventricle were analyzed by using ultrasonic crystal probes. The accuracy of the measurement technique, however, was limited for the detection of reduction in the cardiac function.