| Heat shock related cellular damage and death is of major scientific and clinical interest. In the present study we have established that subjecting rat cardiac myoblasts (H9c2 cells) to 46°C for 1 hour (lethal heat shock) resulted in optimum cell injury as determined by LDH release. Pretreatment of H9c2 cells for 24 hours with 17β estradiol or dehydroepiandrosterone, significantly protects myoblasts against subsequent lethal heat shock exposure in a concentration dependent manner with maximum protection obtained at 1 μM 17β estradiol and 5 μM dehydroepiandrosterone. Using Western blotting, it was observed that 17β estradiol or dehydroepiandrosterone pretreated, followed by lethal heat shock exposed cells had significantly higher levels of inducible heat shock protein 70 (Hsp70), as well as inducible nitric oxide synthase (iNOS) levels compared to lethal heat shock exposed cells. In contrast, lethal heat shock exposed cells had significantly higher levels of total cellular; both cytoplasmic plus nuclear, glucocorticoid receptors (GR) levels compared to 17β estradiol or dehydroepiandrosterone protected cells. Immunofluorescence technique using confocal microscopy, revealed intense nuclear localization of glucocorticoid receptor (GR) and Hsp 70 in lethal heat shock exposed H9c2 cells while 17β estradiol or dehydroepiandrosterone protected cells had primarily extranuclear localization of GR and hsp 70. The glucocorticoid receptor antagonist RU 486 protected H9c2 cells against heat shock induced cell injury indicating that protecting effect is mediated through GR. Selective iNOS inhibitor, S-methylisothiourea, partially reversed 17β estradiol and dehydroepiandrosterone mediated protective effect suggesting that iNOS is involved in the protective effect. Selective and potent mitochondrial K(ATP) channel blocker, 5-HD, was unable to block the 17β estradiol and dehydroepiandrosterone protective effects indicating that protective effects is independent of mitochondrial K (ATP) channel. It is concluded that both 17β estradiol and dehydroepiandrosterone (i) protect H9c2 cells against lethal heat shock insult injury through modulation of expression of GR, Hsp 70 and iNOS and (ii) may have potential clinical beneficial effects against malignant hyperthermia as well as heat stroke, endotoxin shock and acute phase response-induced cardiac injuries. |
