ABSTRACT. To understand the mechanism underlying the sudden animal death caused by acute heart failure during heat stress, the relationships among the heat-induced pathological changes and apoptosis and the variations in the levels of protective Hsp90α and its mRNA in the heat-stressed primary myocardial cells of neonatal rats in vitro were studied by cytopathological observation, immunoblotting, RT-PCR, and analysis of the related enzymes. After a period of adaptive cell culture, the myocardial cells were immediately exposed to heat stress at 42°C for 10, 20, 40, 60, 120, 240, 360, and 480 min. Levels of creatine kinase increased from the beginning of heat stress, and the cells exposed to heat stress showed acute cellular Hsp90α, heat stress and myocardial cells lesions characterized by vacuolar degeneration and necrosis after 40 min of heat stress, suggesting that the myocardial cells in vitro were obviously stressed and damaged by higher temperature. The levels of cleaved caspase-3 and cytochrome C, which were related to apoptosis, increased significantly after 40 min of heat stress while the Hsp90α protein level significantly decreased. In contrast, after 6 h of exposure to heat stress, the levels of cleaved caspase-3 and cytochrome C decreased while those of Hsp90α significantly increased, suggesting that early depletion of Hsp90α coincides with a high rate of necrosis and apoptosis in heat-stressed myocardial cells, while the Hsp90α level in surviving cells increases again with significantly less apoptosis after 6 h of heat stress. These findings also indicate that apoptosis of myocardial cells occurs through the activation of the cytochrome C and caspase-3 pathway. The cell repair capacity of Hsp90α is overstrained in the early phase of heat treatment and needs some hours to stabilize. As a result, in the primary myocardial cells in vitro, Hsp90α shows protective activity against damage at the end period of the heat exposure.

ABSTRACT. We investigated and described the kinetics of heat shock protein (Hsp) 110 expression and distribution in rat primary myocardial cells exposed to heat stress in vitro. After incubation at 37°C for 72 h, myocardial cells were heat stressed at 42°C for 0, 10, 20, 40, 60, 120, 240, 360, and 480 min. Significant increases in aspartate transaminase, lactate dehydrogenase, and creatine kinase enzymatic activities in the myocardial cell culture media were observed during heat stress, suggesting that the integrity of the myocardial cells was altered. Immunocytochemical analysis revealed that the expressed Hsp110 was constitutively localized in the cytoplasm and in the nuclei ©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 11 (4): 4728-4738 (2012) Hsp110 expression changes in vitro 4729 in small amounts characterized by a granular pattern. Nuclear Hsp110 levels increased significantly after 240 min of heat stress compared with levels in the control. The overall levels of Hsp110 expression increased significantly after 20 min. After 240 min, Hsp110 levels were approximately 1.2-fold higher than those in the control. Increasing levels of hsp110 messenger RNA detected using real-time quantitative polymerase chain reaction were observed after 20 min of heat stress, and the levels peaked with a 10-fold increase after 240 min of heat stress. These results indicate that the expression of Hsp110 in primary myocardial cells in vitro is sensitive to hyperthermic stress and that Hsp110 is involved in the potential acquisition of thermotolerance after heat stress. Therefore, Hsp110 might play a fundamental role in opposing and alleviating heat-induced damage caused by hyperthermic stress in primary myocardial cells.

ABSTRACT. The objective of this study was to investigate the mechanism of heat shock protein 90 alpha (Hsp90α) protection against heart damage resulting from heat stress by detecting Hsp90α mRNA, Hsp90α protein, protein localization, and cell damage in primary myocardial cells of neonatal rats in response to heat stress in vitro. The cells were heat-stressed at 42°C in an incubator with 95% air and 5% CO 2 for different periods. Levels of Hsp90α, protein localization, enzymes, and cytopathological lesions were detected using Western blot, immunocytochemistry enzymatic assays, and cytopathological techniques. Aspartate aminotransferase, lactate Heat shock, Hsp90α and primary myocardial cells dehydrogenase, and creatine kinase enzyme levels were elevated during heat stress, and acute cellular lesions that were characterized by vacuolar degeneration and necrosis were observed. Hsp90α levels decreased between 10 and 60 min of heat stress and increased after 360 and 480 min, while Hsp90α mRNA decreased after 360 min. These results indicate that heat stress might induce irreversible damage in certain myocardial cells. The elevated Hsp90α level at the end of heat stress and its positive signal in the cytoplasm of myocardial cells after heat stress could be associated with its protective role. Additionally, the consumption of Hsp90α exceeded its production in the first period of treatment.