NLRP3 inflammasome activation involved in LPS and coal tar pitch extract-induced malignant transformation of human bronchial epithelial cells.

Abstract

Inflammatory microenvironment has been found as a new characteristic of cancer; however, the mechanisms of inflammation-related lung cancer remain unclear. To explore the role of NLRP3 inflammsome activation in inflammation-related lung carcinogenesis, a cell model was set up. Human bronchial epithelial cells (BEAS-2B) were stimulated with 1 μg/mL lipopolysaccharide (LPS) for 24 hours, and then treated with 2.4 μg/mL coal tar pitch extract (CTPE) for 24 hours, after removal of LPS and CTPE, the cells were numbered passage 1 and were passaged and treated in this way until passage 30, which was called LPS + CTPE group. DMSO and Saline were used as vehicle controls. Malignant transformation of cells in passage 30 was evaluated by morphological change, platelet clone formation assay, and tumor formation in nude mice. The mRNA levels of NLRP3 and IL-1β were detected by real time-PCR. The combination of NLRP3 and caspase-1 were determined using immunofluorescence and confocal. The protein expression of NLRP3, cleaved caspase-1(p10), and cleaved IL-1β was detected using Western blot. It was shown that CTPE, LPS + CTPE-stimulated BEAS-2B cells of passage 30 changed a lot morphologically. The clone formation rates, the rates of positive cells of NLRP3 and caspase-1 combination, the mRNA levels of NLRP3 and IL-1β, the protein expression of NLRP3, cleaved caspase-1(p10) and cleaved IL-1β of cells exposed with CTPE and LPS + CTPE at passage 30 were significantly increased compared to vehicle controls. Furthermore, the ability of tumor formation in nude mice, the rates of clone formation and positive cells, mRNA and protein levels of NLRP3 inflammasome activation-related factors in LPS + CTPE-induced cells were all higher than those in cells stimulated with CTPE alone. In conclusion, the cell model of inflammation-related lung cancer is set up successfully, and NLRP3 inflammasome activation may be involved in the malignant transformation of BEAS-2B cells which induced by CTPE alone or LPS combined with CTPE.