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Panax notoginseng saponins inhibit the viability of mouse melanoma B16-F10 cells by regulating macrophage polarization via JAK2/STAT3 pathway
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Abstract:
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 cocultured 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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