The expression and clinical significance of hypoxia-induced factor-1α in gallbladder carcinoma tissues and its role on metformin-suppressed metastasis in GBC-SD cells

Abstract

Objective To study the expression and the clinical significance of hypoxia-induced factor-1α (HIF-1α ) in gallbladder cancer tissues, and the role and mechanism of HIF-1α in metformin-suppressed metastasis in gallbladder carcinoma GBC-SD cells. Methods 24 specimens of gallbladder cancer tissues and 5 specimens of chronic cholecystitis were collected from the First Affiliated Hospital of Zhengzhou University between June 2016 and February 2017. Immunohistochemistry and qPCR were used to detect the expression of HIF-1α in gallbladder cancer tissues, in adjacent non-cancer tissues and in chronic cholecystitis, and the clinical significance was analyzed. The model of metastasis was induced by hypoxia; the wound healing assay and the Transwell assay were used to detect the ability of cell metastasis; the expressions of HIF-1α and VEGF in gallbladder carcinoma GBC-SD cells were detected by western blotting assay and immunofluorescence. Results The expression of HIF-1α in gallbladder cancer tissues was higher than the adjacent non-cancer tissues and in chronic cholecystitis. The expression of HIF-1α was correlated with lymph node metastasis and TNM staging in gallbladder cancer tissues (P<0.05). The wound healing rate after 48 h in the negative control group and in the treatment with hypoxia group (1%O2) in GBC-SD cells were (46.5±4.8)% and (67.3±4.0)%, respectively. The Transwell data showed that the numbers of metastasis after 24 h in the negative control group and in the treatment with hypoxia group GBC-SD cells were (147.4±11.7) and (234.4±17.7), respectively. When compared with the negative control group, treatment with hypoxia significantly increased the ability of metastasis and up-regulated the expression of HIF-1α and VEGF in GBC-SD cells (P<0.05). The wound healing rate after 48 h in the negative control group, the metformin group, the hypoxia group and the metformin and hypoxia group in GBC-SD cells were (40.6±7.1)%, (16.4±9.4)%, (69.5±4.0)% and (22.4±7.4)%, respectively. The Transwell data showed that the numbers of metastasis after 24 h in the negative control group, the metformin group, the hypoxia group and the metformin and hypoxia group in GBC-SD cells were (148.4±6.9), (90.0±8.4), (185.8±10.2) and (113.4±8.6), respectively. When compared with the hypoxia group, treatment with metformin and hypoxia significantly decreased the ability of metastasis and down-regulated the expression of HIF-1α and VEGF in GBC-SD cells (P<0.05). The wound healing rate after 48 h in the negative control group, the 2MeoE2 group, the hypoxia group, the 2MeoE2 and hypoxia group in GBC-SD cells were (43.4±4.4)%, (25.9±9.0)%, (63.3±2.2)%, (46.2±4.5)%, respectively. The Transwell data showed that the numbers of metastasis after 24 h in the negative control group, the 2MeoE2 group, the hypoxia group, the 2MeoE2 and hypoxia group in GBC-SD cells were (144.2±12.6), (80.2±7.7), (203.8±7.0), (124.0±5.2), respectively. When compared with the hypoxia group, treatment with HIF-1α inhibitor 2MeoE2 and hypoxia significantly decreased the ability of metastasis and down-regulated the expression of HIF-1α and VEGF in GBC-SD cells (P<0.05). Conclusions The expression of HIF-1α was correlated with lymph node metastasis and TNM staging in gallbladder cancer tissues. Treatment with hypoxia significantly increased the expression of HIF-1α and VEGF and promoted metastasis of GBC-SD cells, while treatment with metformin decreased the ability of metastasis induced by hypoxia via inhibiting the HIF-1α/VEGF pathway in GBC-SD cells. Key words: Gallbladder carcinoma; Hypoxia-induced factor-1α; Metformin; Hypoxia; Metastasis