A rapid and strong apoptotic process is triggered by hyperosmotic stress in cultured rat cardiac myocytes

In all cell types, the maintenance of normal cell volume is an essential ho meostatic function. Relatively little is known about the induction of apopt osis by hyperosmotic stress and its molecular mechanism in terminally diffe rentiated cardiac myocytes. We compared the apoptotic response of cultured neonatal rat cardiomyoctes to hyperosmotic stress by sorbitol (SOR) with th ose induced by doxorubicin (Doxo) or angiotensin II (Ang II). We also exami ned the apoptotic-signaling pathway stimulated by the hyperosmotic stress. Apoptosis was assessed by the observation of: (1) cell viability, (2) DNA f ragmentation detected by the TUNEL method and by agarose gel electrophoresi s, and (3) poly(ADP-ribose)polymerase (PARP) degradation, and Bcl-X-s and B cl-X-L levels by Western blot analysis. Exposure of cardiomyocytes to 0.3 M SOR for 24 h resulted in decreased cell viability and increased generation of oligosomal DNA fragments (2.5-fold of controls). At this time, 83+/-5% of SOR-treated myocytes were TUNEL-positive (vs 23.7+/-6.8% in controls, P< 0.01). PARP levels also decreased by approximately 42% when cardiac myocyte s were exposed to SOR. Hyperosmotic stress induced a more rapid and stronge r apoptotic response in cardiomyocytes than Doxo or Ang II. In addition, SO R increased 3.2-fold Bcl-X-s proapoptotic protein without changes in Bcl-X- L antiapoptotic protein levels and in the p53-transactivating activity. Tak en together, these results strongly suggest that hyperosmotic stress trigge rs cardiac myocyte apoptosis in a p53-independent manner. being earlier and stronger than apoptosis induced by Doxo and Ang II.