Study of monocyte chemotactic protein-4 mediating p38 MAPK to regulate pyroptosis of nasal mucosa epithelial cells in chronic sinusitis
Objective To investigate the effects of monocyte chemotactic protein-4(MCP-4)on pyroptosis and p38 mitogen-activated protein kinase(p38 MAPK)activation in human nasal epithelial cells(HNEpCs).Methods Forty patients diagnosed with chronic rhinosinusitis(CRS)were selected as the case group,and another 40 healthy individuals were selected as the normal group.Enzyme-linked immunosorbent assay(ELISA)was used to detect the levels of MCP-4 in serum samples from both groups.Receiver operator characteristic(ROC)curve analysis was performed to assess the diagnostic efficacy of serum MCP-4 levels for CRS.HNEpCs were divided into six groups:control group,5 mg/L lipopolysaccharide(LPS)model group,LPS+siRNA negative control(si-Con)group,LPS+siRNA MCP-4(si-MCP-4)group,LPS+pcDNA3.1-vector group,and LPS+pcDNA3.1-MCP-4 group.Cell viability was assessed using the cell counting kit-8(CCK-8),and cell proliferation activity was detected by 5-Ethynyl-2'-deoxyuridine(EdU)staining.Western blot was used to detect the expression levels of phosphorylated p38 MAPK(p-p38 MAPK),Gasdermin D(GSDMD),NOD-like receptor family,pyrin domain containing 3(NLRP3),apoptosis-associated speck-like protein(ASC),and caspase-1.ELISA was performed to measure the levels of tumor necrosis factor-α(TNF-α),interleukin-18(IL-18),and IL-1β in cell supernatants.The Transwell chamber was utilized to assess the effects of MCP-4 on the migratory capacity of HNEpCs.HNEpCs were divided into four groups:control group,10 ng/ml MCP-4 group,and 20 ng/ml MCP-4 group.Results Compared with the normal group,the level of MCP-4 in the serum of patients in the case group was significantly upregulated.ROC curve analysis showed that the area under the curve(AUC)for serum MCP-4 levels in the diagnosis of CRS was 0.921.In vitro experiments demonstrated that LPS stimulation reduced the survival rate and proliferation ability of HNEpCs,increased the expression of pyroptosis proteins,and upregulated p-p38 MAPK levels,and increased the release of TNF-α,IL-1β,and IL-18(P<0.01).Compared with the LPS model group,the LPS+si-MCP-4 group showed increased survival rate and proliferation ability of HNEpCs,decreased expression levels of p-p38 MAPK,pyroptosis proteins and reduced levels of TNF-α,IL-1β,and IL-18(P<0.01).In contrast,compared with the LPS model group,the LPS+pcDNA3.1-MCP-4 group exhibited further decreased survival rate and proliferation ability of HNEpCs,increased expression levels of p-p38 MAPK,pyroptosis proteins,and elevated levels of inflammatory cytokines TNF-α,IL-1β,and IL-18(P<0.01).Compared with the control group,there was no significant increase in the migration ability of HNEpCs in the 10 ng/mL MCP-4 and 20 ng/ml MCP-4 groups.Conclusion Inhibiting the expression of MCP-4 can block the activation of p38 MAPK,reducing pyroptosis and the release of inflammatory cytokines in HNEpCs.
万方数据
维普
