Abstract 4596: Restoring hypoxia-inducible factor-1α in MCF-7 breast cancer cells in vitro by preventing the excessive degradation induced by high oxygen unmasks strong estradiol induction of vascular endothelial growth factor expression
Authors: Molitoris Kristin H. , Brodie Angela , Koos Robert D. 
Published: February 2011

Abstract
Vascular endothelial growth factor (VEGF) is the most potent inducer of increased microvascular permeability known and the key mediator of angiogenesis, pivotal processes in development, reproduction, and tumor growth. VEGF expression is rapidly and strongly induced in normal uterine endometrial epithelial cells in vivo by 17β-estradiol (E2; Kazi et al. 2009 Biol Reprod 81:378). Recent studies indicate that E2-induced epithelial cell proliferation in the mammary gland is also dependent on VEGF. VEGF is also strongly up-regulated by E2 in rat mammary tumors in vivo (Nakamura et al. 1996 Endocrinology 137:5589). By contrast, numerous studies have found that E2 has little or no effect on VEGF expression by breast cancer cells in vitro. We have recently shown that the E2 induction of VEGF is mediated by both estrogen receptor α (ERα) and hypoxia-inducible factor 1α (HIF-1α; Kazi et al. 2005 Mol Endocrinol 19:2006; Molitoris et al. 2009 Endocrinology 150:5405). Cultured cells, however, usually lack HIF-1α due to excessive degradation caused by the abnormally high oxygen level (20%) to which they are exposed. To test the hypothesis that restoring HIF-1α would restore the ability of E2 to strongly induce VEGF expression in breast cancer cells, we treated MCF-7 cells with cobalt chloride (CoCl2; 12.5-100 µM), which prevents oxygen-induced HIF-1α degradation. HIF-1α was very low to undetectable in untreated MCF-7 cells grown in 20% O2, but increased in a dose-dependent manner in response to CoCl2. Although high doses of CoCl2 alone significantly stimulated VEGF mRNA expression, 12.5 µM did not do so. A modest increase in VEGF expression was also seen when MCF-7 cells were treated with E2 alone (∼ 2.5-fold). When cells were co-treated with E2 and 12.5 µM CoCl2, however, a much larger increase in VEGF expression (∼ 6-fold), similar to that induced by E2 in vivo, was seen. This finding is consistent with the hypothesis that HIF-1α is required for maximal E2 induction of VEGF expression in carcinoma cells. Interestingly, when cells were grown for 4 days before treatment, high levels of HIF-1α protein were present in untreated cells. The tendency for MCF-7 cells to express HIF-1α as they become crowded in vitro, even in the presence of high oxygen levels, may explain the ability of E2 alone to induce some VEGF expression in vitro as observed here and in a few previous studies. In conclusion, these results confirm that HIF-1 plays an essential role in E2-induced VEGF expression. The rapid induction of VEGF expression by E2 and the increased microvascular permeability that results may play a critical role in tumor growth by exposing cancer cells to circulating IGF-1, which synergizes with E2 to induce mitosis [Supported by NICHD SCCPIR U54 HD36207 and R01 HD047275].
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4596.