Panax notoginseng saponins inhibit the viability of mouse melanoma B16-F10 cells by regulating macrophage polarization via JAK2/STAT3 pathway
Objective:To investigate the effect of total panax notoginseng saponin (PNS) on the survival of mouse melanoma B16-F10 cells by regulating macrophage polarization through JAK2/STAT3 pathway. Methods:B16-F10 cells and macrophage RAW264.7 were cultured regularly. The effects of different concentrations of PNS on the survival rate of RAW264.7 or B16-F10 cells were detected by MTT assay. The experiment was divided into the following groups:blank group (B16-F10 cells only),control group (B16-F10 cells co-cultured with RAW264.7 cells),PNS groups of various concentrations (B16-F10 cells co-cultured with RAW 264.7 cells,treated with 50,100,200 μg/mL PNS),and PNS+colivelin[B16-F10 cells co-cultured with RAW264.7 cells,200 μg/mL PNS,0.5 μmol/L colivelin (JAK2/STAT3 pathway activator)]group. MTT assay and flow cytometry were applied to detect the survival rate and apoptosis rate of co-cultured cells in each group,and the morphological changes of macrophages were observed under a microscope. ELISA was applied to detect the levels of cytokines TNF-α,IL-6 and IL-1β in the supernatant. qPCR was applied to detect the mRNA expression of macrophage polarization-related genes,inducible nitric oxide synthase (iNOS),IL-12,CD206,and arginase-1 (Arg-1). Western blotting was applied to detect the phosphorylation levels of JAK2 and STAT3 proteins pathway in cells. Results:PNS of different concentrations did not significantly affect the viability of RAW264.7 cells or B16-F10 cells cultured alone (all P>0.05). Compared with the control group,PNS significantly promoted cell apoptosis,protein levels of IL-6,TNF-α,IL-1β,and mRNA levels of IL-12 and iNOS in a concentration-dependent manner (all P<0.05);additionally,PNS reduced the survival rate of co-cultured cells and the phosphorylation levels of JAK2 and STAT3 proteins (all P<0.05). These effects of PNS on co-cultured cells were partially inhibited by colivelin. Conclusion:PNS inhibits the viability of mouse melanoma B16-F10 cells by promoting the polarization of M1 macrophages via inhibiting JAK2/STAT3 pathway.
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