Analyses of ischemia-induced signals in cardiac myocytes

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The objective of this project was to elucidate functional significance of hypoxia-inducible factor 1α (HIF-1α), a member of the basic-helix-loop-helix-Per/ARNT/Sim (bHLH-PAS) protein family, in the ischemic-reperfused myocardium. Particularly, we analyzed expression of vascular endothelial growth factor (VEGF) and inducible nitric oxide synthase (iNOS), which are among target genes of HIF-1α, in cardiac myocytes.1) Analyses of VEGF expression in cardiac myocytes : Using cultured neonatal rat myocytes, we have observed that expression of VEGF was augmented by lipopolysaccaharide (LPS) (Sugishita Y, et al. Biochem Biophys Res Commun, 2000) or adenosine (via its A_2 receptor). On the other hand, we have analyzed expression of the chick VEGF in cultured 10-day-old chick embryonic ventricular myocytes (CEVM) (Sugishita Y, et al. J Mol Cell Caardiol, 2000).2) Analyses of the chick iNOS gene expression in CEVM : We have obtained the data showing that both NF-κB and Ets-1 are involved in the LPS-induced transactivation of the chick iNOS geen in CEVM (Takahashi T, et al. Mol Cell Biochem, submitted).3) Molecular cloning and expressional analyses of the chick HIF-1α cDNA in CEVM : We have cloned cDNAs encoding the chick HIF-1α from CEVM with RT-PCR and Race methods. We then examined expression of the chick HIF-1α mRNA species in various embryonic tissues. The deduced amino acid sequence of the chick HIF-1α cDNA showed 79% identity with that of the human HIF-1α cDNA.In contrast, sequence homology between the chick HIF-1α and endothelial PAS protein 1 (EPAS1), another member of the bHLH-PAS proteins, was only low (49%). HIF-1α mRNA was expressed abundantly in CEVM, but scarcely in the liver, which was quite different from expression pattem of EPAS1 mRNA.These data suggest that HIF-1α may be involved in embryonic cardiovascular development (Takahashi T, et al. Biochem Biophys Res Commun, 2001, in press).