[Show abstract][Hide abstract]ABSTRACT: Phenolic compounds EGCG [(-)-epigallocatechin-3-gallate], resveratrol (3,4',5-trihydroxy-trans-stilbene) and capsaicin (trans-8-methyl-N-vanillyl-6-nonenamide) are worth investigating for clinical application in cancer prevention and chemotherapy. Hypoxia-induced drug resistance is a major obstacle in the development of effective cancer chemotherapy. Therefore, we examined whether drug resistance to these phenolic compounds is acquired by hypoxia.
Hep3B hepatoma, Caki-1 renal carcinoma, SK-N-MC neuroblastoma, and HEK293 cell lines were cultured under normoxic or hypoxic conditions. Drug sensitivities to the phenolic compounds and expression of hypoxia-inducible factor-1alpha (HIF-1alpha) and the multidrug resistance genes were examined in these cell lines.
Drug resistance was acquired 24 h after hypoxia and subsided 8 h after reoxygenation. Protein synthesis inhibitors abolished this drug resistance. A transfection study demonstrated that HIF-1alpha enhanced this hypoxia-induced resistance and that its dominant-negative isoform suppressed resistance acquisition. However, MDR1 and MRP1, which provide multidrug resistance to conventional anticancer agents, were not induced by hypoxia.
These results suggest that HIF-1alpha-dependent gene expression participates in the cellular process of the hypoxia-induced resistance to phenolic compounds.
[Show abstract][Hide abstract]ABSTRACT: Atrial natriuretic peptide (ANP) is a cardiac peptide, the transcription of which is up-regulated in the ischaemic ventricle. However, the molecular mechanism of ANP induction is unclear. This study demonstrated that ANP mRNA expression in rat ventricular myocardium is induced in an early phase of ischaemia, preceded by hypoxia-inducible factor-1 (HIF-1) alpha expression. The ANP gene was also induced by hypoxia or HIF-1 inducers such as CoCl2 and desferrioxamine in H9c2 and neonatal cardiomyocytes. The 2307 bp 5'-flanking region of the rat ANP gene was cloned and fused to the luciferase gene. Evidence of the promoter activity was only apparent in the myocytes and was induced by hypoxia and HIF-1 inducers. The overexpression of HIF-1alpha markedly enhanced ANP promoter activity, and a dominant-negative isoform completely suppressed it. We demonstrated that the promoter regions are essential for hypoxic ANP induction. One promoter region, containing the HIF-1-binding sequence, is regulated directly by HIF-1. The other region is also activated by HIF-1 despite having no HIF-1-binding sequence. These results suggest that HIF-1 enhances the transactivation of the ANP gene in hypoxic myocytes, implying that stimulation of the ANP promoter by HIF-1 may in fact be responsible for the induction of the ANP gene in ischaemic ventricular myocardium.