Application of Hypoxia to Establish Epithelial-Mesenchymal Transition Models inRenal Cell Carcinoma Cell Lines

Abstract

Purpose: To establish epithelial-mesenchymal transition (EMT) models in renal cell carcinoma cell lines by CoCl2-induced hypoxia. Materials and methods: The renal cell carcinoma cell lines A498 and 786-O were used in the experiment and CoCl2 was used to simulate hypoxia. The cancer cells were cultured with different concentrations of CoCl2. Morphology and cyto-activity changes were detected to obtain the optimal concentration of CoCl2 for simulating hypoxia. After CoCl2 treatment, the cells were subjected to Western blot analysis to test the expression of HIF-1α and the changes of EMT-related molecules (E-cadherin, fibronectin). Results: Cell conjunctions of CoCl2-treated groups were loose and scattered compared to the control. The effect of CoCl2 on A498 cell viability was not distinct at a low dosage, but when the concentration of CoCl2 reached 250 mM, cell activity gradually declined. In contrast, CoCl2 induced 786-O cell proliferation in the range of 50 mM- 200 mM, but it inhibited cell growth at dosages higher than 200 mM. The expression of E-cadherin was significantly down-regulated, and fibronectin was up-regulated in both A498 and 786-O cell lines under CoCl2-simulated hypoxia in comparison with normoxic conditions (P<0.01). Conclusions: EMT models of the renal cell carcinoma cell lines were successfully established by CoCl2-induced hypoxia. The models will help us further study the mechanisms of EMT and investigate novel therapeutic targets to inhibit tumor invasion and metastasis.