The Mechanism Of Macrophage Migration Inhibitory Factor In Warburg Effect Of Lung Cancer Cells

BackgroundLung cancer is the most common cancer and the leading cause of cancer-associated mortality in the world.It is estimated that the 5-year survival rate is<16%.However,the underlying molecular mechanisms of lung cancer pathogenesis are poorly understood.Therefore,further elucidation of the mechanism of lung cancer development and development is the key to lung cancer treatment.Most solid tumor cells can provide energy and related raw materials through glycolysis after hypoxia,but existing studies have also found that in aerobic environment,there are a lot of glycolysis in tumor cells,called Warburg effect(also called Oxygen glycolysis,an important mechanism is the activation of hypoxia-inducible factor 1?(HIF-1?)in tumor cells.Macrophage migration inhibitory factor(MIF)is a multifunctional pro-inflammatory cytokine that can be secreted by a variety of tissues and cells.Existing studies have shown that MIF expression is significantly increased in a variety of solid tumors,and can regulate tumor cell proliferation,apoptosis,drug resistance and the like.Studies have confirmed that MIF overexpression can promote lung tumor growth and progression.However,does MIF affect the proliferation of lung cancer cells by regulating the Warburg effect?It remains to be further elaborated.ObjectiveTo explore the role of MIF on the Warburg effect of lung cancer cells and related molecular mechanisms,and to elucidate the role of MIF-regulated Warburg effect in the proliferation of lung cancer cells.MethodsHuman lung cancer H358,H460,H524,H1650,H838,H1975 and A549 cell lines were used to examine the expression of MIF by real time-quantitative polymerase chain reaction and western blotting.Selecting the highest expression and minimum expression of MIF cell lines to complete the experimental study.The lentivirus was used to over-express MIF and the expression of MIF and hypoxia-inducible factor 1-?(HIF-1?)were knocked down by shRNA or siRNA.The proliferation of cell lines was detected by MTT assay to evaluate the effect of MIF and HIF-la on cell proliferation.Glucose uptake,adenosine 5'-triphosphate(ATP)production,the glycolytic rate and lactate production in these cell lines were used to examine the Warburg effect of MIF and HIF-1? in cells.In the highly expressed MIF cell line,the expression of HIF-1? was knocked down by transfection technology.The cell proliferation curve,glucose uptake rate,ATP production,glycolysis rate and lactate production were observed before and after transfection to evaluate whether MIF regulates Warburg effect through HIF-1?.BAY 11-7082(BAY)was used to inhibit the nuclear transcription of NF-?B,which the expression of HIF-lawas detected by immunofluorescence,which is to evaluate whether HIF-1?was regulated by NF-?B?Results1.Effect of MIF on proliferation of lung cancer cells.The expression of MIF in the lung cancer cell lines of 7 groups was detected by PCR and Western blot.The results showed that MIF was the highest in H358 cells and the least in H524 cells.MIF expression was silenced in H358 cell line and MIF was highly expressed in H524 cell line.The cell proliferation curve suggested that silencing of MIF significantly inhibited the proliferation of H358 cells.On the contrary,high expression of MIF could enhance the proliferation of H524 cells.2.Effect of MIF on Warburg effect of lung cancer cells.By comparing the glucose uptake rate,ATP production,glycolysis rate and lactic acid production of cell lines before and after MIF expression,it was suggested that high MIF expression significantly promoted the Warburg effect of H524 cells,while MIF expression silencing significantly inhibited the Warburg effect of H358 cells.These results indicate that MIF promotes the Warburg effect of lung cancer cells.3.Effect of MIF on the expression level of HIF-1? in lung cancer cells.The results of Western blot showed that the expression level of HIF-1? was significantly decreased in H358 cells after knockdown of MIF.In H524 cells,the expression level of HIF-1? was significantly increased after MIF overexpression.This experiment demonstrates that MIF can induce the expression of HIF-1?.4.Role of HIF-1? in MIF-induced lung cancer cell proliferation and Warburgeffect.The proliferation ability of H358 cells after interference with HIF-1? expression was significantly lower than that of the control group,and the Warburg effect was also weaker than that of the control group.In H524 cells overexpressing MIF,MIF overexpression-induced cell proliferation and Warburg effect were also inhibited by siRNA interfering with HIF-1? expression.The above experiments show that MIF promotes lung cancer proliferation and Warburg effect by increasing HIF-1?expression.5.The role of NF-?B in the regulation of HIF-1? expression by MIF.Inhibition of NF-?B pathway activation by BAY significantly reduced HIF-la expression in H358 cells.In H524 cell line,high expression of MIF up-regulated the expression of HIF-1?,whereas inhibition of NF-?B pathway activation by BAY significantly inhibited MIF-induced upregulation of HIF-1?.It was shown that MIF regulates HIF-1? by activating the NF-?B pathway.Conclusions1.MIF can promote the Warburg effect of lung cancer cells,thereby inducing the proliferation of lung cancer cells;2.MIF induces the Warburg effect of lung cancer cells by promoting the expression of HIF-la;3.MIF promotes HIF-la expression by activating NF-?B pathwayMIF is a key component in lung cancer progression through promoting the Warburg effect,and that the novel MIF/NF-KB/HIF-la axis may prove to be useful for the development of new strategies for treating patients with lung cancer.