ABSTRACT
This study examined whether insulin-stimulated hypoxia-inducible factor 1α(HIF-1α)expression plays a crucial role in promoting the proliferative vitality and invasive capability in human pancreatic cancer cells.PANC-1 cells were divided into three groups: Control group,insulin group and insulin+YC-1(a pharmacological inhibitor of HIF-1α)group in terms of different treatments.Cells in the insulin group or insulin+YC-1 group were treated with insulin(0.1,1,10 and 100 nmol/L)alone or combined with 3-(5'-hydroxymethyl-2'-furyl)-1-benzyl indazole(YC-1,0.1,1,10 and 100μmol/L).HIF-1α mRNA and protein expression in PANC-1 cells was determined by real-time RT-PCR and Western blotting respectively.Cell proliferation and invasion were measured by using growth curve and invasion assay,respectively.Western blot analysis demonstrated that insulin dose-dependently increased the HIF-1α protein expression,and YC-1 could dose-dependently block this effect.However,neither insulin nor YC-1 altered HIF-1α mRNA levels in PANC-1 cells.Moreover,insulin could enhance the proliferation and invasion of PANC-1 cells,while YC-1 could weaken this effect.It was concluded that the malignant proliferation and local invasion of pancreatic cancer cells may be related to high-insulin microenvironment.The tumor biological behavior change resulting from high-insulin microenvironment may be associated with the increased expression of HIF-1αprotein.
ABSTRACT
Summary: The effect of target-directed regulation of the uncoupling protein-2 (UCP-2) gene expression on the ischemia-reperfusion injury of hepatocytes under different conditions was investigated. The expression plasmid and RNAi plasmid targeting UCP-2 gene were constructed and transfected into normal hepatocytes and fatty liver cells, respectively. The expression of UCP-2 mRNA was detected by real time PCR. The cells were divided into normal cell group (NCG), group of normal cells transfected with empty vector (EVNCG), group of normal cells transfected with expression plasmid (EPNCG), fatty liver cell group (FCG) and group of fatty liver cells transfected with RNAi plasmid (RPFCG). The ischemia-reperfusion model in vitro was established. One, 6, 12 and 24h after reperfusion, Annexin V/PI flow cytometry was used to measure cell necrosis rate, apoptosis rate and survival rate. Simultaneously, the intracellular ATP, ROS and MDA levels were determined. The resuits showed that 1, 6, 12 and 24h after ischemia-reperfusion, the intracellular ROS, MDA and ATPlevels and cell survival rate in EPNCG were significantly lower, and cell necrosis rate significantly higher than in NCG and EVNCG, but there was no significant difference in apoptosis rate among NCG, EVNCG and EPNCG (P005). Six, 12 and 24h after reperfusion there was no significant difference in ROS, MDA levels and apoptosis rate between FCG and RPFCG (P0.05), but the ATP level and survival rate of cells in RPFCG were higher than in FCG (P<0.05). It was concluded that down-regulation of the UCP-2 gene expression in steatotic hepatocytes could alleviate the ischemia-reperfusion injury of liver cells.