Hypoxia Attenuates the Expression of E-Cadherin via Up-Regulation of SNAIL in Ovarian Carcinoma Cells

Abstract

Since ovarian carcinoma cells detach from the primary lesion and metastasize via peritoneal dissemination, we hypothesized that these cells are exposed to hypoxia, which may affect cell attachment and invasiveness. To address this hypothesis, we first examined in vivo the immunohistochemical expression of hypoxia-inducible factor-1␣ (HIF-1␣) and its topological correlation with E-cadherin expression in ovarian carcinomas. We then examined in vitro the effect of hypoxia on the mRNA and protein expressions of E-cadherin using two ovarian cancer cell lines, SKOV3 and OVCAR3, and normal ovarian surface epithelial (OSE) cells. In addition, hypoxia-induced change in the expression of SNAIL, a transcriptional factor repressing E-cadherin expression, was also analyzed. Finally, we examined the facilitation of invasiveness of ovarian cancer cells under hypoxia using Matrigel invasion assay. Immunohistochemically, nuclear localization of HIF-1␣ was observed in 32 of the 76 (42%) carcinomas studied, and showed a topological correlation with loss of E-cadherin expression. Northern blotting, real-time PCR and Western blotting demonstrated that E-cadherin expression was remarkably decreased under hypoxia in both SKOV3 and OVCAR3 cells, but not in normal OSE cells. mRNA expression of SNAIL was increased under hypoxia in both ovarian cancer cell lines. Invasion assay revealed that hypoxia increases the invasiveness of ovarian cancer cells. Accordingly, the present study demonstrated that hypoxia induces down-regulation of E-cadherin in ovarian carcinoma cells, via up-regulation of the transcriptional repressor SNAIL. These findings suggest that hypoxia plays an important role in the change in intercellular attachment, which may be involved in the initiation of tumor progression of ovarian cancer cells.

Metabolic reprogramming of the tumor

“…First, HIF cooperates with oncogenic MYC to foster OXPHOS-uncoupled glycolysis (Kim et al, 2007). Second, it provokes E-cadherin loss in carcinoma cells (Imai et al, 2003), summoning a cell biological program, the epithelial-mesenchymal transition (EMT), whereby a tightly concerted alteration in gene expression leads to the transformation of epithelial cells into pseudomesenchymal cells (Thiery and Sleeman, 2006). This endows cancer cells with novel properties, such as anchorage-independent growth, and enhanced motility Figure 3 Proposed roles of cancer metabolism in the expression of cancer hallmarks (Hanahan and Weinberg, 2011).…”

“…As in tumor growth, hypoxia induces HIF-1, as a pleiotropic master switch of the cell biological processes responsible for tumor invasion. Its activity leads to constitutive lactate production (Kim et al, 2007), angiogenesis (Ikeda et al, 1995;Mandriota and Pepper, 1998), metalloproteinase expression (Pouyssegur et al, 2006) and EMT induction (Imai et al, 2003). From the point of view of the cancer cell, moving to other sites when microenvironment conditions become detrimental makes sense.…”

Mouse Snail Is a Target Gene for HIF

“…Activity of this enhancer is associated with enrichment of H3 Lys (4) dimethylation and H3 acetylation of both the enhancer and promoter. Although hypoxia has been implicated in upregulation of Snail (19)(20)(21), no functional hypoxia-responsive element has been identified in the snail promoter.…”

“…Imai and colleagues (19) reported that mRNA expression of Snail was increased under hypoxia in ovarian cancer cell lines. In addition, they demonstrated that hypoxia downregulated E-cadherin in these cells.…”

“…GSK-3 is an endogenous inhibitor of Snail transcription (17) and GSK-3b mediated phosphorylation of Snail has been shown to facilitate its proteasomal degradation (18). Low oxygen induces Snail transcription (19)(20)(21)(22) and both the canonical HIF (22,23) and Notch (24,25) pathways have been implicated in hypoxia-induced EMT. However, Snail is yet to be identified as a direct HIF target gene.…”