Study On The Mechanism Of Silica-activated Macrophages Promoting EMT In Lung Cancer Cells By Inducing IL-6 Autocrine

Objective: To investigate whether silica-treated macrophages can promote the EMT of lung cancer cells through IL-6,and the specific mechanism of promoting EMT of lung cancer cells,so as to provide a new idea and theoretical basis for global occupational health problems caused by Silica exposure.Methods:1)The effect of Silica-induced macrophages on lung cancer cell migration was verified by scratch and Transwell experiments.The expression levels of EMT markers in lung cancer cells were determined by Western blot and real time quantitative PCR(RTQPCR).2)The expression levels of inflammatory factors in macrophages and lung cancer cells were detected by RT-QPCR,and the key cytokines promoting EMT in silica-induced macrophages in lung cancer cells were searched.Western blot and Enzyme Linked immunosorbent assay(ELISA)were used to verify the key cytokines.3)To further explore the mechanism of autocrine of IL-6 in lung cancer cells,the expression level of p-NFκB in lung cancer cells was determined by Western blot and immunofluorescence,and the promoting effect of p-NFκB on IL-6 was verified by ELISA and Western blot experiments.4)Western blot was used to detect the expression levels of p-PI3 K,p-AKT,p-Raf and p-ERK,and AKT and ERK inhibitors were used to verify the regulation of PI3K/AKT and Raf/ERK signaling pathways on EMT of lung cancer cells.5)The animal model of silicosis complicated with lung cancer was established in C57BL/6 mice,and the effects of IL-6R inhibitor on tumor were investigated by HE staining and immunofluorescence.Results:1)Silica treated macrophages promoted the expression and migration of EMT markers in lung cancer cells.2)Silica promoted IL-6 secretion in macrophages and induced IL-6 autosecretion in lung cancer cells.After IL-6R Ab was used to inhibit the binding of IL-6 to lung cancer cells,silica-induced macrophages were found to promote the reversal of EMT in lung cancer cells.3)Silica-induced macrophages promoted STAT3 phosphorylation and NFκB entry into the nucleus of lung cancer cells.After NFκB inhibitor blocked NFκB entry into the nucleus,IL-6 autocrine and EMT marker expression were inhibited in lung cancer cells.4)Silica-induced macrophages promoted the phosphorylation of PI3K/AKT and Raf/ERK in lung cancer cells,and AKT and ERK inhibitors could reverse the promotion of Silica-induced macrophages on EMT in lung cancer cells.5)Compared with the control group,the tumor tissue volume of silicosis combined with lung cancer mice increased,and the expression level of IL-6 increased.IL-6R inhibitor significantly inhibited the expression of IL-6 in tumor tissues of silicosis combined with lung cancer mice,and the tumor tissues of IL-6R inhibitor group were smaller than those of silicosis combined with lung cancer group.Conclusion: Silica-treated macrophages can induce IL-6 autocrine in lung cancer cells,and IL-6 autocrine can promote EMT in lung cancer cells by activating PI3K/AKT and Raf/ERK signaling pathways.These results provide a new theoretical basis for the study of the mechanism of Silica exposure promoting lung cancer,and provide a new strategy for the prevention and treatment of global occupational health problems caused by Silica exposure.Figure eighteen Table one Reference fifty-four...