Up-regulation of hypoxia inducible factor-1α by cobalt chloride correlates with proliferation and apoptosis in PC-2 cells

PC-2 cells were cultured with different concentration (50-200 μmol/L) of CoCl2 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-quantitive RT-PCR and Western blot analysis. Apoptosis of PC-2 cells were demonstrated by flow cytometry with Annexin V-FITC/PI double staining.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 CoCl2. RT-PCR and Western blot analysis indicated that hypoxia could up-regulate the expression of HIF-1α on both mRNA and protein levels.Hypoxic microenvironment stimulated by CoCl2 could effectively induce apoptosis and influence cell proliferation in PC-2 cells, the mechanism could be related to up-expression of HIF-1α.Hypoxia is one of the most important pathological characteristics of solid tumor which is the result of imbalance between tumor cell proliferation and blood supply [1]. As solid tumor growing, its center becomes a hypoxic area because of lacking blood and oxygen. The hypoxic status of various solid tumor has been recognized as an important determinant for the outcome of anti-cancer therapies in a number of tumors [2].Hypoxia-inducible factor-1 (HIF-1) was found in the 1992 when Semenza [3] researched the expression of erythropoietin gene induced by hypoxia. Human HIF-1 has been depurated and isolated, it is a heterodimeric transcription factor composed of oxygen-dependent HIF-1α and constitutively expressed HIF-1β subu