Long-term primary cultures of human adult atrial cardiac myocytes: Cell viability, structural properties and BNP secretion in vitro

Abstract

BackgroundHuman adult cardiomyocytes (CM) have been used in short-term cultures for in vitro studies of the adult myocardium. However, little information is available regarding human adult CMs cultured for long term (>2 weeks). MethodsHuman adult CMs were isolated from atrial specimens of 43 patients undergoing cardiopulmonary bypass surgery. Cell viability, cytoskeletal properties, intercellular junctional mediators and responsiveness to extracellular stimuli were monitored in CM cultures for 8 weeks. ResultsAbsolute numbers of CMs decreased through the first 2 weeks, with substantially lower rates of cell loss thereafter. Apoptosis predominated over necrosis as the principal mode of cell death, affecting 4.1±1.6% of freshly dissociated cells, that declined in culture (3.6±1.0% week 1, 1.3±0.5% week 2). CMs maintained rod-shaped morphology and cross-striated expression pattern of sarcomeric proteins desmin and β-myosin heavy chain for the first 4 weeks. Levels of desmin remained stable on first 3 weeks, but declined thereafter. CMs expressed cardiac-specific adherence molecule N-cadherin throughout the culture duration, indicating conserved contractile potential. CMs remained functional early in culture, as indicated by BNP secretion, with maximal levels on 1st week that declined gradually by week 4. Cell responsiveness to metabolic stresses (serum deprivation) was detected, inducing an early (6 h) 1.8-fold increase in levels of BNP. ConclusionLong-term cultured human adult CMs maintain morphological integrity, adult-type cytoskeletal protein expression, cell–cell communication potential and functionality for 3–4 weeks in vitro.