Inhibition of nuclear transcription factor E2-related factor 2 pathway regulates pancreatic cancer cell metastasis and immune escape factor expression under high glucose
Objective:To investigate effect of inhibiting nuclear transcription factor-E2 related factor 2(Nrf2)pathway on metastasis and secretion of immune escape factors of pancreatic cancer cells under high glucose state.Methods:Human pancreatic cancer cell line Panc-1 was cultured,and treated with 5.5,10,25 and 50 mmol/L glucose.Cell proliferation rate was detected by MTT at 12 h,24 h and 48 h,respectively,and at 24 h,changes of intracellular Nrf2 expression were detected by RT-qPCR and Western blot;experimental groups included control group,high glucose(HG)group,Nrf2 inhibitor ML385+high glucose(ML385+HG)group,MTT was used to detect cell proliferation rate,cell colony formation assay was used to detect number of colonies formed,Tran-swell was used to detect number of cell migration and invasion,in vitro scratch assay was used to detect cell wound healing,ELISA was used to determine contents of vascular endothelial growth factor(VEGF),IFN-γ,transforming growth factor-β1(TGF-β1)and IL-6,distribution of intracellular Nrf2 was observed by immunofluorescence staining,Western blot was used to detect protein expres-sions of Nrf2 and heme oxygenase-1(HO-1)in cells.Results:Compared with 5.5 mmol/L glucose group,proliferation rate of Panc-1 cells was increased when treated with 10,25 and 50 mmol/L glucose for 12 h and 24 h,expressions of Nrf2 mRNA and protein were increased(P<0.05);compared with control group,proliferation rate of cells in HG group was increased,number of colonies formed was increased,numbers of migration and invasion were also increased,and rate of wound healing was increased,contents of VEGF,IFN-γ,TGF-β1 and IL-6 in cell culture supernatant were increased,Nrf2 fluorescence staining was significantly enhanced,expres-sion of Nrf2 in nucleus was increased,and protein expressions of Nrf2 and HO-1 were up-regulated(P<0.05);compared with HG group,cell proliferation rate in ML385+HG group was decreased,number of colonies formed,numbers of migration and invasion were also decreased,wound healing rate was decreased,VEGF,IFN-γ,TGF-β1 and IL-6 contents in cell culture supernatant were also decreased,intracellular Nrf2 fluorescent staining was weaken,and protein expressions of Nrf2 and HO-1 were down-regulated(P<0.05).Conclusion:Nrf2 is highly expressed in pancreatic cancer cells under high glucose state,inhibiting Nrf2 can inhibit prolifera-tion,migration and invasion of pancreatic cancer cells promoted by high glucose,and reduce secretion of immune escape factors.
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