本論文研究的目的，在探討大鼠心肌梗塞(myocardial infarction)動物模式下，注射感染肝生長因子(hepatocyte growth factor)的胚胎心肌細胞(H9c2)，對心臟功能及動脈形態的影響。

　　以縫線將大鼠心臟冠狀動脈左下降枝(left antirior descending coronary artery)血流給予阻斷，造成急性冠狀心肌梗塞(acute myocardial infarciton)。心肌梗塞14天之後，老鼠隨機分成五組，分別給予細胞培養基質(medium)(50μl)、腺病毒(adenovirus carrying LacZ)(5x105 Pfu)、腺病毒攜帶肝細胞因子(adenovirus carrying HGF)(1x109 )、胚胎心肌細胞(5x105 )以及胚胎心肌細胞感染肝生長因子(H9c2 cell carrying HGF)(5x105 )。實驗結果發現：1) 與細胞培養基質組比較，在胚胎心肌細胞感染肝生長因子和腺病毒攜帶肝細胞因子兩組中，心臟纖維組織(scar)有明顯的減少；2) 與細胞培養基質組相比，在胚胎心肌細胞感染肝生長因子和腺病毒攜帶肝細胞因子兩組中，心肌等容舒張能力(diastolic dp/dt; isovolumetric relaxation)有顯著的增加；3) 與細胞培養基質組相比，胚胎心肌細胞感染肝生長因子組中，直徑大於100μm的動脈數目沒有變化; 4) 和細胞培養基質組相比，腺病毒攜帶肝細胞因子組中，動脈外膜纖維組織(adventitia)層與平滑肌層的面積比有明顯增加。本論文証實了注射感染肝生長因子(hepatocyte growth factor)的胚胎心肌細胞(H9c2)，能減少大鼠心肌梗塞所造成的心臟纖維組織的形成，並增加心肌舒張功能。

　Backgroud- The purpose of this study was to evaluate the effects of embryonic cardiomyocytes mediated-HGF gene therapy on ventricular function of infarcted hearts, and arterial development.

　Methods and Resutls-In rats, left anterior descending coronary artery ligation was operated to create MI. Two weeks after ligation, rats were randomly divided into five groups: (1) the medium group for control (injection of 50μm culture medium, n=3), (2) the Ad-LacZ group (injection of 5*10^5 pfu Ad.LacZ, n=3), (3) the HGF group (injection of 1*10^9 pfu Ad.HGF, n=3), (4) the cell group (1*10^5 H9c2 cells, n=3), and (5) the C-HGF group (1*10^5 H9c2 cells infected with Ad.HGF, n=3). Scar formation was reduced after either the HGF group or the C-HGF group compared to the medium group. Cardiac functions determinated by Millar demonstrated that end-diastolic pressure was significantly decreased in two treatment groups when compared with the medium group. Further, the minimal dp/dt was significantly increased in the C-HGF group compared to the medium group. In vascular system, the arterial density was no changed among groups. The ratio of adventitia area to smooth muscle cell area was increased in the HGF group compared to the medium group.

　Conclusions-The present study proves that the application of H9c2 cell-mediated HGF gene therapy affected cardiac function by reducing scar expansion and improving cardiac relaxation. The intervention of this combine therapy may contribute to the prevention of heart failure from a chronic MI.

Reference
Alexander, M.Y., K.A. Webster, P.H. McDonald, and H.M. Prentice. 1999. Gene transfer and models of gene therapy for the myocardium. Clin Exp Pharmacol Physiol. 26:661-8.
Birchmeier, C., and E. Gherardi. 1998. Developmental roles of HGF/SF and its receptor, the c-Met tyrosine kinase. Trends Cell Biol. 8:404-10.
Erlebacher, J.A., J.L. Weiss, M.L. Weisfeldt, and B.H. Bulkley. 1984. Early dilation of the infarcted segment in acute transmural myocardial infarction: role of infarct expansion in acute left ventricular enlargement. J Am Coll Cardiol. 4:201-8.
French, B.A., W. Mazur, R.S. Geske, and R. Bolli. 1994. Direct in vivo gene transfer into porcine myocardium using replication-deficient adenoviral vectors. Circulation. 90:2414-24.
Gal, D., L. Weir, G. Leclerc, J.G. Pickering, J. Hogan, and J.M. Isner. 1993. Direct myocardial transfection in two animal models. Evaluation of parameters affecting gene expression and percutaneous gene delivery. Lab Invest. 68:18-25.
Guzman, R.J., P. Lemarchand, R.G. Crystal, S.E. Epstein, and T. Finkel. 1993. Efficient gene transfer into myocardium by direct injection of adenovirus vectors. Circ Res. 73:1202-7.
Helisch, A., and W. Schaper. 2003. Arteriogenesis: the development and growth of collateral arteries. Microcirculation. 10:83-97.
Jayasankar, V., Y.J. Woo, T.J. Pirolli, L.T. Bish, M.F. Berry, J. Burdick, T.J. Gardner, and H.L. Sweeney. 2005. Induction of angiogenesis and inhibition of apoptosis by hepatocyte growth factor effectively treats postischemic heart failure. J Card Surg. 20:93-101.
Kass-Eisler, A., E. Falck-Pedersen, M. Alvira, J. Rivera, P.M. Buttrick, B.A. Wittenberg, L. Cipriani, and L.A. Leinwand. 1993. Quantitative determination of adenovirus-mediated gene delivery to rat cardiac myocytes in vitro and in vivo. Proc Natl Acad Sci U S A. 90:11498-502.
Koh, G.Y., S.J. Kim, M.G. Klug, K. Park, M.H. Soonpaa, and L.J. Field. 1995. Targeted expression of transforming growth factor-beta 1 in intracardiac grafts promotes vascular endothelial cell DNA synthesis. J Clin Invest. 95:114-21.
Li, Y., G. Takemura, K. Kosai, K. Yuge, S. Nagano, M. Esaki, K. Goto, T. Takahashi, K. Hayakawa, M. Koda, Y. Kawase, R. Maruyama, H. Okada, S. Minatoguchi, H. Mizuguchi, T. Fujiwara, and H. Fujiwara. 2003. Postinfarction treatment with an adenoviral vector expressing hepatocyte growth factor relieves chronic left ventricular remodeling and dysfunction in mice. Circulation. 107:2499-506.
Lin, H., M.S. Parmacek, G. Morle, S. Bolling, and J.M. Leiden. 1990. Expression of recombinant genes in myocardium in vivo after direct injection of DNA. Circulation. 82:2217-21.
Liu, Y., K. Rajur, E. Tolbert, and L.D. Dworkin. 2000. Endogenous hepatocyte growth factor ameliorates chronic renal injury by activating matrix degradation pathways. Kidney Int. 58:2028-43.
Markkanen, J.E., T.T. Rissanen, A. Kivela, and S. Yla-Herttuala. 2005. Growth factor-induced therapeutic angiogenesis and arteriogenesis in the heart--gene therapy. Cardiovasc Res. 65:656-64.
Menard, C., S. Pupier, D. Mornet, M. Kitzmann, J. Nargeot, and P. Lory. 1999. Modulation of L-type calcium channel expression during retinoic acid-induced differentiation of H9C2 cardiac cells. J Biol Chem. 274:29063-70.
Miyagawa, S., Y. Sawa, S. Taketani, N. Kawaguchi, T. Nakamura, N. Matsuura, and H. Matsuda. 2002. Myocardial regeneration therapy for heart failure: hepatocyte growth factor enhances the effect of cellular cardiomyoplasty. Circulation. 105:2556-61.
Murry, C.E., R.W. Wiseman, S.M. Schwartz, and S.D. Hauschka. 1996. Skeletal myoblast transplantation for repair of myocardial necrosis. J Clin Invest. 98:2512-23.
Nakamura, T., S. Mizuno, K. Matsumoto, Y. Sawa, and H. Matsuda. 2000. Myocardial protection from ischemia/reperfusion injury by endogenous and exogenous HGF. J Clin Invest. 106:1511-9.
Nakamura, T., T. Nishizawa, M. Hagiya, T. Seki, M. Shimonishi, A. Sugimura, K. Tashiro, and S. Shimizu. 1989. Molecular cloning and expression of human hepatocyte growth factor. Nature. 342:440-3.
Nakamura, Y., R. Morishita, J. Higaki, I. Kida, M. Aoki, A. Moriguchi, K. Yamada, S. Hayashi, Y. Yo, K. Matsumoto, and et al. 1995. Expression of local hepatocyte growth factor system in vascular tissues. Biochem Biophys Res Commun. 215:483-8.
Pfeffer, M.A., and E. Braunwald. 1990. Ventricular remodeling after myocardial infarction. Experimental observations and clinical implications. Circulation. 81:1161-72.
Pouzet, B., J.T. Vilquin, A.A. Hagege, M. Scorsin, E. Messas, M. Fiszman, K. Schwartz, and P. Menasche. 2000. Intramyocardial transplantation of autologous myoblasts: can tissue processing be optimized? Circulation. 102:III210-5.
Rappolee, D.A., A. Iyer, and Y. Patel. 1996. Hepatocyte growth factor and its receptor are expressed in cardiac myocytes during early cardiogenesis. Circ Res. 78:1028-36.
Raya, T.E., M. Gaballa, P. Anderson, and S. Goldman. 1997. Left ventricular function and remodeling after myocardial infarction in aging rats. Am J Physiol. 273:H2652-8.
Reinecke, H., G.H. MacDonald, S.D. Hauschka, and C.E. Murry. 2000. Electromechanical coupling between skeletal and cardiac muscle. Implications for infarct repair. J Cell Biol. 149:731-40.
Smits, P.C. 2004. Myocardial repair with autologous skeletal myoblasts: a review of the clinical studies and problems. Minerva Cardioangiol. 52:525-35.
Sutton, M.G., and N. Sharpe. 2000. Left ventricular remodeling after myocardial infarction: pathophysiology and therapy. Circulation. 101:2981-8.
Suzuki, K., B. Murtuza, R.T. Smolenski, I.A. Sammut, N. Suzuki, Y. Kaneda, and M.H. Yacoub. 2001. Cell transplantation for the treatment of acute myocardial infarction using vascular endothelial growth factor-expressing skeletal myoblasts. Circulation. 104:I207-12.
Taylor, D.A., B.Z. Atkins, P. Hungspreugs, T.R. Jones, M.C. Reedy, K.A. Hutcheson, D.D. Glower, and W.E. Kraus. 1998. Regenerating functional myocardium: improved performance after skeletal myoblast transplantation. Nat Med. 4:929-33.
Van Belle, E., B. Witzenbichler, D. Chen, M. Silver, L. Chang, R. Schwall, and J.M. Isner. 1998. Potentiated angiogenic effect of scatter factor/hepatocyte growth factor via induction of vascular endothelial growth factor: the case for paracrine amplification of angiogenesis. Circulation. 97:381-90.
von Degenfeld, G., A. Banfi, M.L. Springer, and H.M. Blau. 2003. Myoblast-mediated gene transfer for therapeutic angiogenesis and arteriogenesis. Br J Pharmacol. 140:620-6.
Zhang, M., D. Methot, V. Poppa, Y. Fujio, K. Walsh, and C.E. Murry. 2001. Cardiomyocyte grafting for cardiac repair: graft cell death and anti-death strategies. J Mol Cell Cardiol. 33:907-21.
Zhang, Y.W., and G.F. Vande Woude. 2003. HGF/SF-met signaling in the control of branching morphogenesis and invasion. J Cell Biochem. 88:408-17.