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The Expression And Mechanism Of Pyroptosis Related Factors IL-1β,IL-18,GSDMD,GSDMD-N In Psoriasis

Posted on:2022-10-07Degree:MasterType:Thesis
Country:ChinaCandidate:L Y WangFull Text:PDF
GTID:2504306344463254Subject:Master of Clinical Medicine
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Objectives:To detect the expression levels of serum interleukin-1 β(IL-1β),interleukin-18(IL-18)and the expression of GSDMD and GSDMD-N in skin lesions of patients with psoriasis vulgaris,to observe the proliferation of HaCaT cells induced by lipopolysaccharide(LPS)and its effect on the expression of IL-1β and GSDMD in HaCaT cells,and to explore the role of pyroptosis signal pathway in the pathogenesis of psoriasis.Methods:From April 2020 to January 2021,Serum of 20 patients with psoriasis vulgaris and lesions of 10 patients with psoriasis vulgaris diagnosed in the Department of Dermatology of Subei people’s Hospital were collected and enrolled into PSO group,Serum of 20 healthy persons in health check-up center and skin samples of 10 patients in plastic surgery were collected and and enrolled into CON group.The levels of serum IL-18 and IL-1β in PSO group and CON group were detected by enzyme linked immunosorbent assay(ELISA),and the expression of IL-1β mRNA in the serum of PSO group and CON group were detected by real-time fluorescence quantitative PCR(qRT-PCR).The expression of GSDMD and GSDMD-N terminal in skin tissue of PSO group and CON group were detected by Western Blot.HaCaT cells were treated with LPS,and the effects of different concentrations of LPS(0.1 ug/ml,lug/ml,10ug/ml)on the proliferation of HaCaT cells were detected by CCK-8 method.The expression of IL-1β in HaCaT cells culture supernatant induced by LPS was detected by enzyme-linked immunosorbent assay(ELISA).The expression levels of IL-1β and GSDMD mRNA in HaCaT cells induced by LPS were detected by real-time fluorescence quantitative PCR(qRT-PCR).Result:1.Serum IL-18 and IL-1β expression levels:ELISA enzyme-linked immunosorbent assay showed that IL-18 in the PSO group was 226.59(199.91-412.56)pg/ml compared with 73.89(62.53-86.76)pg/ml in the CON group,and the difference was statistically significant(P<0.0001).The serum IL-1β(58.20±10.51)pg/ml in the PSO group(58.20±10.51)pg/ml was significantly higher than that in the CON group(12.33±1.79)pg/ml,and the difference was statistically significant(P<0.0001).2.The expression level of IL-1β mRNA in serum:the results of qRT-PCR showed that the expression level of IL-1β mRNA in serum of PSO group patients was significantly higher than that of patients in CON group,and the difference was statistically significant(P<0.0001).3.The expression of GSDMD and GSDMD-N protein in skin lesions:Western blotting assay showed that the expression level of GSDMD protein in skin lesions of PSO group patients was significantly higher than that in CON group(P<0.01),and the expression level of GSDMD-N terminal protein in PSO group was also higher than that in CON group(P<0.0001).4.The effect of LPS on cell proliferation:the results of cell proliferation by CCK-8 assay showed that the proliferation ability of HaCaT cells stimulated by different concentrations(0.1ug/ml,1ug/ml,10ug/ml LPS)was increased compared with the untreated group,and 0.1ug/ml and lug/ml LPS promoted cell proliferation,the difference was statistically significant(P<0.001,P<0.01).LPS 10ug/ml stimulated cell proliferation,but there was no statistical significance.Therefore,the concentrations of 0.1ug/ml and lug/ml LPS were selected for following experiments.5.IL-1β expression level in cell culture supernatant:Compared with the untreated group(12.71±0.68)pg/ml,IL-1β expression level in 0.1ug/ml LPS group(24.52 ±1.93)pg/ml,lug/ml LPS group(16.56±1.29)pg/ml increased,the differences were statistically significant(P<0.0001).6.Expression level of IL-1β and GSDMDmRNA:qRT-PCR results showed that the relative expression of IL-1β mRNA in HaCaT cells stimulated by 0.1ug/mlLPS and lug/ml LPS was significantly higher than that in untreated group(P<0.0001,P<0.001).The relative expression of GSDMD mRNA in HaCaT cells stimulated by 0.1ug/ml LPS was significantly higher than that in the untreated group(P<0.001),but the relative expression of GSDMD mRNA in HaCaT cells stimulated by lug/ml LPS was not significantly different from that in the untreated group.Conclusion:The levels of serum IL-18 and IL-1β were increased in patients with psoriasis vulgaris,and the expression of GSDMD and GSDMD-N in psoriatic lesions was also increased.Compared with the control group,the proliferation ability of HaCaT cells induced by LPS was increased,the secretion of IL-1β was increased,and the expression levels of IL-1β and GSDMD mRNA were increased.The ability of cell proliferation induced by LPS 0.1ug/ml was the strongest,and the expression of IL-1β and GSDMD mRNA was the highest.These results above suggested that the pyroptosis related cytokines may play certain roles in the pathogenesis of psoriasis.
Keywords/Search Tags:Psoriasis, Pyroptosis, IL-1β, IL-18, GSDMD, GSDMD-N
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