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Hypoxia-derived oxidative stress mediates epigenetic repression of PKC? gene in foetal rat hearts.

ABSTRACT: Hypoxia causes protein kinase C epsilon (PKC?) gene repression in foetal hearts, resulting in heightened cardiac susceptibility to ischaemic injury in offspring. We tested the hypothesis that hypoxia inducible factor 1 (HIF-1) and/or reactive oxygen species (ROS) mediate hypoxia-induced PKC? gene repression.Hypoxia induced in vivo to pregnant rats, ex vivo to isolated foetal rat hearts, and in vitro in the rat embryonic ventricular myocyte cell line H9c2 resulted in a comparable decrease in PKC? protein and mRNA abundance in foetal hearts and H9c2 cells, which was associated with a significant increase in CpG methylation of the SP1-binding sites at the PKC? promoter. In H9c2 cells and foetal hearts, hypoxia caused nuclear accumulation of HIF-1?, which was inhibited by 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole and 2-methoxy estradiol. The HIF-1? inhibitors had no significant effect on hypoxia-induced PKC? mRNA repression. Hypoxia produced a time-dependent increase in ROS production in H9c2 cells and foetal hearts that was blocked by ROS scavengers N-acetyl-cysteine or tempol. In accordance, N-acetyl-cysteine and tempol, but not apocynin, inhibited the hypoxic effect and restored PKC? protein and mRNA expression to the control values in foetal hearts and H9c2 cells. The ROS scavengers blocked hypoxia-induced CpG methylation of the SP1-binding sites, restored SP1 binding to the PKC? promoter, and abrogated the hypoxia-induced increase in the susceptibility of the heart to ischaemic injury in offspring.The results demonstrate that hypoxia induces epigenetic repression of the PKC? gene through a NADPH oxidase-independent ROS-mediated pathway in the foetal heart, leading to heightened heart vulnerability to ischaemic injury in offspring.

SUBMITTER: Patterson AJ

PROVIDER: S-EPMC3258654 | biostudies-other | 2012 Feb

REPOSITORIES: biostudies-other

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Hypoxia-derived oxidative stress mediates epigenetic repression of PKCε gene in foetal rat hearts.

Patterson Andrew J AJ Xiao Daliao D Xiong Fuxia F Dixon Brandon B Zhang Lubo L

Cardiovascular research 20111202 2

<h4>Aims</h4>Hypoxia causes protein kinase C epsilon (PKCε) gene repression in foetal hearts, resulting in heightened cardiac susceptibility to ischaemic injury in offspring. We tested the hypothesis that hypoxia inducible factor 1 (HIF-1) and/or reactive oxygen species (ROS) mediate hypoxia-induced PKCε gene repression.<h4>Methods and results</h4>Hypoxia induced in vivo to pregnant rats, ex vivo to isolated foetal rat hearts, and in vitro in the rat embryonic ventricular myocyte cell line H9c2 ...[more]

PMID: 22139554

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Project description:Epidemiological studies demonstrate a clear association of adverse intrauterine environment with an increased risk of ischemic heart disease in adulthood. Hypoxia is a common stress to the fetus and results in decreased protein kinase C epsilon (PKCepsilon) expression in the heart and increased cardiac vulnerability to ischemia and reperfusion injury in adult offspring in rats.The present study tested the hypothesis that fetal hypoxia-induced methylation of cytosine-phosphate-guanine dinucleotides at the PKCepsilon promoter is repressive and contributes to PKCepsilon gene repression in the heart of adult offspring.Hypoxic treatment of pregnant rats from days 15 to 21 of gestation resulted in significant decreases in PKCepsilon protein and mRNA in fetal hearts. Similar results were obtained in ex vivo hypoxic treatment of isolated fetal hearts and rat embryonic ventricular myocyte cell line H9c2. Increased methylation of PKCepsilon promoter at SP1 binding sites, -346 and -268, were demonstrated in both fetal hearts of maternal hypoxia and H9c2 cells treated with 1% O(2) for 24 hours. Whereas hypoxia had no significant effect on the binding affinity of SP1 to the unmethylated sites in H9c2 cells, hearts of fetuses and adult offspring, methylation of both SP1 sites reduced SP1 binding. The addition of 5-aza-2'-deoxycytidine blocked the hypoxia-induced increase in methylation of both SP1 binding sites and restored PKCepsilon mRNA and protein to the control levels. In hearts of both fetuses and adult offspring, hypoxia-induced methylation of SP1 sites was significantly greater in males than in females, and decreased PKCepsilon mRNA was seen only in males. In fetal hearts, there was significantly higher abundance of estrogen receptor alpha and beta isoforms in females than in males. Both estrogen receptor alpha and beta interacted with the SP1 binding sites in the fetal heart, which may explain the sex differences in SP1 methylation in the fetal heart. Additionally, selective activation of PKCepsilon restored the hypoxia-induced cardiac vulnerability to ischemic injury in offspring.The findings demonstrate a direct effect of hypoxia on epigenetic modification of DNA methylation and programming of cardiac PKCepsilon gene repression in a sex-dependent manner, linking fetal hypoxia and pathophysiological consequences in the hearts of adult offspring.

Project description:Senescence is characterized by permanent cell-cycle arrest despite continued viability and metabolic activity, in conjunction with the secretion of a complex mixture of extracellular proteins and soluble factors known as the senescence-associated secretory phenotype (SASP). Cellular senescence has been shown to prevent the proliferation of potentially tumorigenic cells, and is thus generally considered a tumor suppressive process. However, some SASP components may act as pro-tumorigenic mediators on premalignant cells in the microenvironment. A limited number of studies indicated that protein kinase C (PKC) has a role in senescence, with different isoforms having opposing effects. It is therefore important to elucidate the functional role of specific PKCs in senescence. Here we show that PKC?, an epithelial specific and anti-apoptotic kinase, promotes senescence induced by oxidative stress and DNA damage. We further demonstrate that PKC? promotes senescence through its ability to upregulate the expression of the cell cycle inhibitors p21(Cip1) and p27(Kip1) and enhance transcription and secretion of interleukin-6 (IL-6). Moreover, we demonstrate that PKC? creates a positive loop for reinforcing senescence by increasing the transcription of both IL-6 and IL-6 receptor, whereas the expression of IL-8 is specifically suppressed by PKC?. Thus, the presence/absence of PKC? modulates major components of SASP. Furthermore, we show that the human polymorphic variant of PKC?, 374I, that exhibits higher kinase activity in comparison to WT-374V, is also more effective in IL-6 secretion, p21(Cip1) expression and the promotion of senescence, further supporting a role for PKC? in senescence. As there is now considerable interest in senescence activation/elimination to control tumor progression, it is first crucial to reveal the molecular regulators of senescence. This will improve our ability to develop new strategies to harness senescence as a potential cancer therapy in the future.

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Hypoxia-derived oxidative stress mediates epigenetic repression of PKC? gene in foetal rat hearts.

Publications

Hypoxia-derived oxidative stress mediates epigenetic repression of PKCε gene in foetal rat hearts.

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