Regulating Embryo Cardiomyocyte Cell Division - Implications for Gene Therapy

Fischman, Donald

Abstract

Two growth factor pathways play major roles in the regulation of cardiac myocyte proliferation in the embryo. These are the fibroblast growth factors (FGF) and TGF-B factors and their cognate receptors (FGFR and TGF-BR). Our group (mima et al; 1995) has demonstrated that FGF signaling is essential for the in vivo proliferation of cardiac myocytes in early embryonic hearts (week-1 of the embryonic chick development). By week-2 of development myocyte growth becomes FGF-independent, and this coincides with a prolongation of myocyte cell cycle kinetics. Studies by Sporn and colleagues (letterio et al; 1994) have shown that genetic knockouts of TGF-B reception in the mouse cause a dramatic thickening of the myocardial wall, suggesting that TGF-B signaling plays a negative control on myocyte proliferation. Cell culture studies also support this conclusion. Thus, regulation of myocyte cell division involves positive (FGF) and negative (TGF-B) pathways. We suggest that future attempts to control myocyte proliferation in the adult will require an understanding of these two signaling pathways and how they are regulated pre- and postnatally. We plan three sets of experiments using chicken embryos. (1) explore the role of FGF signaling in regulating the proliferation of cardia myocytes in the tubular stage embryonic chick heart. This will involve; (a) the up- and down-regulation of FGFR1 in cardiac myocytes between days E3 and E7 of embryonic development using replication-defective retroviral constructs; and (b) the co-expression of FGFR1 plus the FGF ligand in myocytes using a dicistronic retroviral vector. (2) examine the negative control of myocyte cell division by TGF-B using dominant-negative TGF-B receptor constructs to disrupt this signaling pathway. (3) Using replication-defective adenoviral vectors, examine the role of TGF-B and FGF signaling in myocytes which are withdrawing from the cell cycle during the week-2 of embryonic chick development. The chick embryo was selected as a model system because: a) early morphogeneisis of the heart in chick embryos is essentially identical to that in mouse, rat or man; b) cell cycl regulation in the chicken is no differnet from that in higher mammals;c) the decline in myocyte proliferation during chick heart development mirrors that of man; d) it offers many experimental advantages for transgene insertion and manipilation of cardiac precursors by microsurgery.