Background: The exact mechanism of the effects of hypoxia on the proliferation and apoptosis in carcinoma cells is still conflicting. This study investigated the variation of hypoxia-inducible factor-1α(HIF-1α) expression and the apoptosis effect of hypoxia stimulated by cobalt chloride (CoCl(2)) in pancreatic cancer PC-2 cells.
Methods: PC-2 cells were cultured with different concentration (50-200 μmol/L) of CoCl(2) after 24-120 hours to simulate hypoxia in vitro. The proliferation of PC-2 cells was examined by MTT assay. The cellular morphology of PC-2 cells were observed by light inverted microscope and transmission electron microscope(EM). The expression of HIF-1α on mRNA and protein level was measured by semi-quantitative RT-PCR and Western blot analysis. Apoptosis of PC-2 cells were demonstrated by flow cytometry with Annexin V-FITC/PI double staining.
Results: MTT assay showed that the proliferation of PC-2 cells were stimulated in the first 72 h, while after treated over 72 h, a dose- dependent inhibition of cell growth could be observed. By using transmission electron microscope, swollen chondrosomes, accumulated chromatin under the nuclear membrane and apoptosis bodies were observed. Flow cytometer(FCM) analysis showed the apoptosis rate was correlated with the dosage of CoCl(2). RT-PCR and Western blot analysis indicated that hypoxia could up-regulate the expression of HIF-1α on both mRNA and protein levels.
Conclusion: Hypoxic microenvironment stimulated by CoCl(2) could effectively induce apoptosis and influence cell proliferation in PC-2 cells, the mechanism could be related to up-expression of HIF-1α.