IngentaConnect Spontaneous death of isolated adult rat cardiocytes in culture in...

Abstract:

Many isolated adult cardiocytes do not survive beyond the early days of culture, but why they die has not been defined. We examined the possibility of apoptosis as the mechanism of death in cultured atrial and ventricular rat cardiocytes. Calcium-tolerant cardiocytes isolated by enzymatic dissociation were cultured with a medium containing FBS. Nucleosomal DNA fragmentation was detected by electrophoresis of DNA extracted from the cardiocytes, by immunohistochemical in situ DNA nick-end labelling of single cells, and by enzyme immunoassay for in vitro quantification in cytoplasmic fraction. Electrophoresis on the 5th to 14th day of culture revealed the ladder appearance characteristic of internucleosomal DNA cleavage in apoptosis with a consistent single peak of increased cytoplasmic DNA fragments. After the 14th day, the cytoplasmic DNA fragments decreased, and the ladder appearance could no longer be detected by electrophoresis. Cardiocytes positive with nick-end labelling were seen by the 5th day, and then increased in number over the remaining days. These results indicate that many isolated cardiocytes die spontaneously by apoptosis within the first 2 weeks of culture, suggesting a possible signal dependence for survival of adult cardiocytes. In addition to chemical signal depletion in culture, other possible explanations for this apoptosis include the absence of an electric signal during culture, lack of contractile activity, and initial loss of intercellular connections.

Keywords: Apoptosis; enzyme immunoassay; gel electrophoresis; growth factor; immunohistochemistry

Document Type: Research article

Affiliations: 1: World Health Organization Cardiovascular Center; Departments of Medicine and Pathology, UK 2: Department of Physiology and Biophysics, University of Texas Medical Branch Galveston, USA

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Spontaneous death of isolated adult rat cardiocytes in culture in association with internucleosomal cleavage of genomic DNA