Endothelial exposure to hypoxia induces Egr‐1 expression involving PKCα‐mediated Ras/Raf‐1/ERK1/2 pathway

Abstract

Hypoxia induces endothelial dysfunction that results in a series of cardiovascular injuries. Early growth response-1 (Egr-1) has been indicated as a common theme in vascular injury. Here we demonstrates that in bovine aortic endothelial cells (ECs) subjected to hypoxia (PO(2) approximately 23 mmHg), rapidly increased Egr-1 mRNA expression which peaked within 30 min and decreased afterwards. Treatment of ECs with PD98059, a specific inhibitor to mitogen-activated protein kinase (MAPK/ERK), inhibited this hypoxia-induced Egr-1 expression. The involvement of ERK pathway was further substantiated by the inhibition of Egr-1 promoter activities when ECs were co-transfected with a dominant negative mutant of Ras (RasN17), Raf-1 (Raf 301), or a catalytically inactive mutant of ERK2 (mERK). In addition, the hypoxia-induced transcriptional activity of Elk-1, an ERK substrate, was abolished by administration of PD98059. Addition of calphostin C, a protein kinase C (PKC) inhibitor, completely blocked the hypoxia-augmented Egr-1 expression. The likewise occurred while exposing ECs to D609 to inhibit phospholipase C and BAPTA/AM to chelate intracellular calcium. Hypoxia to ECs increased ERK phosphorylation within 10 min and which was abolished by administration of PD98095, calphostin C, and BAPTA/AM. Hypoxia triggered a transient translocation of PKCalpha from cytosol to membrane fraction concurrent with the association of PKCalpha to Raf-1. Involvement of PKCalpha in mediating ERK activation was further confirmed by the inhibition of ERK and the subsequent Egr-1 gene induction with antisense oligonucleotides to PKCalpha. These results indicate that ECs under hypoxia induce Egr-1 expression and this induction requires calcium, phospholipase C activation, and PKCalpha-mediated Ras/Raf-1/ERK1/2 signaling pathway. Our finding support the importance of specific PKC isozyme linked to MAPK pathway in the regulation of endothelial responses to hypoxia.

Regulation of apoA-I gene expression

“…These results are consistent with the concept that upregulation of apoA-I expression is effected by estrogen-mediated activation of the MAP kinase pathway. PKC has been implicated in the regulation of MAP kinase activation (32). However, in our experiments, inhibition of PKC did not have an effect on apoA-I activation by estrogen and genistein.…”

Histamine Induces Egr-1 Expression in Human Aortic Endothelial Cells via the H1 Receptor-mediated Protein Kinase Cδ-dependent ERK Activation Pathway

“…Our results also reveal that the histamine-triggered pathway leading to Egr-1 expression is distinct from the hypoxia-induced pathway, in which activation of PKC␣ or PKC␤ leads to Egr-1 expression in endothelial cells and in a monocyte-like cell line (35,36). These results indicate that these specific PKC isoforms serve as the central mediators for Egr-1 expression in various cell types and in response to different stimuli.…”

Inhibition of PKCα induces a PKCδ-dependent apoptotic program in salivary epithelial cells

“…While this may indicate that PKCa normally functions to suppress ERK expression and activity in parotid C5 cells, this seems unlikely given a variety of reports which demonstrate that PKCa, as well as other PKC isoforms, activate ERK. [53][54][55] A more likely explanation is that activation of ERK1/ 2 and its increased expression is a component of the apoptotic pathway induced by the loss of PKCa activity, and does not reflect loss of a normal physiologic function of PKCa. Interestingly, PKCd has also been shown to be a positive regulator of ERK, 50,55,56 and expression of a dominant negative PKCd construct blocked the activation of ERK in cells induced to undergo apoptosis by treatment with UV light.…”