Cell-to-cell interaction prevents cell death in cultured neonatal rat ventricular myocytes.

OBJECTIVES Loss of cardiac cells and the anatomical or functional remodeling of intercellular coupling occur under several pathological conditions. We have assessed the significance of intercellular coupling for cell death. METHODS AND RESULTS Ventricular cells obtained from 1 day old Wistar rats were cultured. Apoptosis was detected by nick-end labeling. Cells were plated at low and high cell density (3x10(4)/ml and 12x10(4)/ml, respectively). Cultured myocytes died spontaneously by apoptosis in a time dependent manner. The increase of the apoptotic cell population in a culture with high cell density on day 4 (1+/-1.2%, n=4) was significantly lower than that in a culture with low cell density (20+/-5.5%, n=4). The progression of apoptosis in the culture of low cell density was prevented in part after application of the medium extract from the culture of high cell density; the apoptotic cell population on day 6 decreased from 57+/-8.0% (n=4) to 36+/-3.8% (n=4). Treatment of the cultured myocytes at high cell density with antisense oligonucleotide for connexin43 (Cx43) for 24 h on day 2 resulted in a significant decrease in Cx43 expression as judged by Western blot, dye transfer and immunocytochemistry using mouse monoclonal antibody for Cx43. In association with the down-regulation of Cx43, the progress of apoptosis was accelerated; the apoptotic cell population on day 5 in the antisense-treated cultures (27+/-5.7%, n=4) was significantly higher than the sense-treated cultures (5+/-1.1%, n=4). The effect of Cx43 antisense treatment to promote apoptosis was not reversed by application of high cell-density culture medium. CONCLUSIONS These findings suggest that cell-cell communication through gap junction formation and some humoral factors play important roles in the survival of cultured myocytes.