健脾消癌方通过调控NLRP3/Caspase-1/GSDMD信号通路对大肠癌细胞焦亡的影响OA

Objective It is to investigate the effect of prescription for fortifying spleen and dispersing cancer on the py-roptosis of colorectal cancer cells through the regulation of the NOD-like receptor thermal protein domain associated protein 3(NLRP3)/Caspase-1/gasdermin D(GSDMD)signaling pathway.Methods The prescription for fortifying spleen and dispersing cancer was decocted and concentrated to 1.00 g/mL,then diluted with PRMI1640 medium containing 10%fetal bovine serum to prepare drug-containing serum solutions with drug concentrations of 0.00 g/mL,0.25 g/mL,0.50 g/mL,and1.00 g/mL;Concurrently,recombinant human NLRP3(rhNLRP3)was taken and diluted with PRMI1640 medium containing 10%fetal bovine serum to prepare drug-containing serum solution with a concentration of 10 μmol/mL.Human colorectal cancer HCT116 cells were used as the research subjects and divided into8 groups:which were0.00 g/mL herbal decoction group,0.25 g/mL herbal decoction group,0.50 g/mL herbal decoction group,1.00 g/mL herbal decoction group,10 μmol/mL rhNLRP3 group,0.25 g/mL herbal decoction+10 μmol/mL rhNLRP3 group,0.50 g/mL herbal de-coction+10 μmol/mL rhNLRP3 group,1.00 g/mL herbal decoction+10 μmol/mL rhNLRP3 group.The cell proliferation rate of each group was detected by CCK-8 method,the colony formation ability of the cells was assessed by colony formation assay,the positive protein expressions of cleaved-Caspase1 and GSDMD were detected by immunofluorescence chemistry,and the levels of interleukin-18(IL-18)and interleukin-1β(IL-1β)in cell culture supernatant were measured by ELISA.Results Compared with the0.00 g/mL herbal decoction group,the cell proliferation rates and mean optical density values of GSDMD positive expression were significantly decreased in the 0.25 g/mL herbal decoction group,0.50 g/mL herbal decoc-tion group,1.00 g/mL herbal decoction group,10 μmol/mL rhNLRP3 group,0.25 g/mL herbal decoction+10 μmol/mL rhNLRP3 group,0.50 g/mL herbal decoction+10 μmol/mL rhNLRP3 group,1.00 g/mL herbal decoction+10 μmol/mL rhNLRP3 group(all P<0.05),the numbers of cell colonies were significantly reduced(all P<0.05),and the mean op-tical density values of cleaved Caspase-1 positive expression,as well as the levels of IL-1β and IL-18 in the cells were sig-nificantly increased(all P<0.05).The cell proliferation rate and the mean optical density of GSDMD positive expression in cells were decreased as the decoction concentration increased in each simple herbal decoction group,while the numbers of cell colonies were also decreased as the decoction concentration increased,the mean optical density values of cleaved Caspase-1 positive expression and the levels of IL-1β and IL-18 in the cells were all decreased as the concentration of the herbal decoction increased,all pairwise comparisons showed statistically significant differences(all P<0.05).Compared with the 10 μmol/mL rhNLRP3 group,the cell proliferation rates and mean optical density values of GSDMD positive ex-pression in the cells were obviously decreased,and the numbers of cell colonies were significantly reduced in each herbal decoction+rhNLRP3 group(all P<0.05),while the mean optical density values of cleaved Caspase-1 positive expression and the levels of IL-1β and IL-18 in the cells were all significantly increased(all P<0.05);furthermore,the cell prolifer-ation rates,numbers of cell colonies,mean optical density values of GSDMD and cleaved Caspase-1 positive expressions,and the levels of IL-1β and IL-18 in the cells of each herbal decoction+rhNLRP3 group were all increased or decreased with increasing of decoction concentration,and all pairwise comparisons showed statistically significant differences(all P<0.05).Conclusion Prescription for fortifying spleen and dispersing cancer can inhibit the proliferation and promote the py-roptosis of colorectal cancer HCT116 cells through the activation of the NLRP3/caspase-1/GSDMD signaling pathway to re-lease large amounts of IL-1β and IL-18.