The regenerative inadequacy of the injured myocardium leads to adverse remodeling, cardiac dysfunction, and heart disease. Stem cellreplacement of damaged myocardium faces major challenges such as inappropriate differentiation, cellular uncoupling, scar formation, and accelerated apoptosis of transplanted cells. These challenges can be met by engineering an in vitro system for delivering stem cells capable of cardiac differentiation, tissue integration, and resistance to oxidative stress. In this study, we describe the formation of three-dimensional (3D) cell aggregates (''cardiospheres'') by putative stem cells isolated from adult dog myocardium using poly-L-ornithine. De novo formation of cardiospheres in growth factor-containing medium occurred over a period of 2-3 weeks, but accelerated to 2-3 days when seeded on poly-Lornithine. Older cardiospheres developed foci of ''beating'' cells upon co-culture with rat neonatal cardiomyocytes. Cardiospheres contained cells that exhibited characteristics of undifferentiated cells; differentiating cardiomyocytes with organized contractile machinery; and vascular cells capable of forming ''vessel-like'' networks. They exhibited strong resistance to elevated concentrations of hydrogen peroxide in culture and survived subcutaneous injections without undergoing neoplastic transformation up to 3 weeks post-transplantation. These findings suggest that cardiospheres are potentially useful for delivering functional stem cells to the damaged heart. J. Cell. Biochem. 105: 612-623, 2008. ß 2008 KEY WORDS: CARDIOMYOCYTES; CARDIOSPHERES; DIFFERENTIATION; ENDOTHELIAL CELLS; GROWTH FACTORS; OXIDATIVE STRESS; POLY-L-ORNITHINE; SMOOTH MUSCLE CELLS; SPHEROIDS; STEM CELLS T he progressive loss of cardiomyocytes coupled with the poor regenerative capacity of myocardial tissue following acute ischemia or oxidative stress result in the proliferation of fibroblasts, scarring, decreased cardiac performance, compensatory hypertrophy, and ultimately, lead to heart failure [Claycomb, 1992;Frey and Olson, 2003]. Advances in medicine and technology have led to new strategies in cardiac repair involving transplantation or infusion of neonatal cardiomyocytes [Muller-Ehmsen et al., 2002], skeletal myoblasts [Taylor et al., 1998;Suzuki et al., 2001], embryonic stem cells [Min et al., 2002], endothelial cells and/or progenitors [Condorelli et al., 2001;Murasawa et al., 2005], adult mesenchymal cells [Gojo et al., 2003;Makkar et al., 2005], or bone marrow-derived cells [Orlic et al., 2001;Strauer et al., 2002;Beeres et al., 2006]. Promising results have been obtained in many cases showing improvement in cardiac performance [Kehat et al., 2004], however, studies demonstrating development of a fully functional syncytium between the grafted cells and the recipient myocardium are limited and short-term. Grafted cells often exhibit abnormal action Journal of Cellular Biochemistry 105:612-623 (2008)
Journal of Cellular Biochemistry
ARTICLE
612Abbreviations used: 3D, three-dimensional; FGF-2, fibro...
