摘要
目的:观察IL-36在溃疡性结肠炎(UC)中的表达,分析IL-36受体拮抗剂(IL-36Ra)对UC患者单核细胞功能的调控作用。方法:入组2021年4月至2023年1月在新乡医学院第一附属医院就诊的UC患者72例和对照者20例,分离血浆和外周血单个核细胞(PBMCs),分离UC组织和正常组织中组织浸润淋巴细胞(TILs),纯化PBMCs和TILs中的单核细胞。ELISA检测血浆中IL-36α、IL-36β、IL-36γ、IL-36Ra水平,实时定量PCR检测组织中IL-36和单核细胞中IL-36受体亚基mRNA表达。使用重组IL-36Ra刺激单核细胞,与CaCO_(2)细胞共培养,检测CaCO_(2)细胞死亡比例,ELISA检测上清中TNF-α、颗粒酶B、颗粒酶H水平,流式细胞术检测单核细胞中Fas配体(FasL)和TNF相关凋亡诱导配体(TRAIL)平均荧光强度(MFI)。结果:UC组和对照组血浆IL-36α、IL-36β、IL-36γ水平差异无统计学意义(P>0.05),UC组血浆IL-36Ra水平低于对照组[(87.33±11.95)pg/ml vs(100.81±15.62)pg/ml,t=4.162,P<0.001],UC组织和正常组织IL-36α、IL-36β、IL-36γmRNA表达差异无统计学意义(P>0.05),UC组织中IL-36Ra mRNA表达低于正常组织(1.00±0.23 vs 1.36±0.21,t=5.500,P<0.001)。UC组和对照组、UC组织和正常组织单核细胞中IL-36受体亚基mRNA表达差异均无统计学意义(P>0.05)。UC组外周血单核细胞诱导CaCO_(2)细胞死亡比例高于对照组[(14.45±3.33)%vs(10.92±2.87)%,t=2.965,P=0.005],UC组织纯化的单核细胞诱导CaCO_(2)细胞死亡比例高于正常组织[(17.41±4.19)%vs(14.31±3.02)%,t=2.327,P=0.027],上清中TNF-α、颗粒酶B、颗粒酶H水平均升高(P<0.05),但UC组和对照组、UC组织和正常组织FasL和TRAIL MFI差异均无统计学意义(P>0.05)。重组IL-36Ra刺激UC患者外周血和病变组织纯化的单核细胞后,CaCO_(2)细胞死亡比例降低(P<0.05),上清中TNF-α、颗粒酶B、颗粒酶H水平均降低(P<0.05),但FasL和TRAIL MFI在无刺激和经IL-36Ra刺激间差异无统计学意义(P>0.05)。结论:IL-36Ra可能通过减少TNF-α和颗粒酶分泌抑制UC患者单核细胞杀伤功能。
Objective:To investigate expression of IL-36 in ulcerative colitis(UC),and to assess regulatory activity of IL-36 receptor antagonist(IL-36Ra)to monocytes function in UC patients.Methods:Seventy-two UC patients and twenty controls were enrolled between April 2021 and January 2023 in the First Affiliated Hospital of Xinxiang Medical University.Plasma and peripheral blood mononuclear cells(PBMCs)were isolated.Tissue infiltrating lymphocytes(TILs)were isolated from UC tissue and normal tis-sue.Monocytes from PBMCs and TILs were purified.Plasma IL-36α,IL-36β,IL-36γand IL-36Ra levels were measured by ELISA.mRNA levels of IL-36 in tissue and IL-36 receptor subunits in monocytes were semi-quantified by real-time PCR.Purified monocytes were stimulated with recombinant IL-36Ra and co-culture with CaCO_(2) cells.Percentage of CaCO_(2) cell death was measured.Serum TNF-α,granzyme B and granzyme H levels were measured by ELISA.Mean fluorescence intensity(MFI)of Fas ligand(FasL)and TNF-related apoptosis-inducing ligand(TRAIL)in monocytes was measured by flow cytometry.Results:There were no significant differences of plasma IL-36α,IL-36βor IL-36γlevels between UC group and control group(P>0.05).Plasma IL-36Ra levels were lower in UC group compared with control group[(87.33±11.95)pg/ml vs(100.81±15.62)pg/ml,t=4.162,P<0.001].There were no significant differences of IL-36α,IL-36βor IL-36γmRNA levels between UC tissue and normal tissue(P>0.05).IL-36Ra mRNA level was remarkably reduced in UC tissue compared with normal tissue(1.00±0.23 vs 1.36±0.21,t=5.500,P<0.001).There were no signifi-cant differences of IL-36 receptor subunits mRNA relative levels between UC group and control group,as well as between UC tissue and normal tissue(P>0.05).Peripheral monocytes-induced CaCO_(2) cell death was higher in UC group compared with control group[(14.45±3.33)%vs(10.92±2.87)%,t=2.965,P=0.005].Monocyes purified from UC tissue also mediated increased CaCO_(2) cell death compared with those from normal tissue[(17.41±4.19)%vs(14.31±3.02)%,t=2.327,P=0.027].TNF-α,granzyme B and granzyme H levels in supernatants were significantly elevated(P<0.05).However,there were no remarkable differences of FasL or TRAIL MFI in monocytes between UC group and control group,as well as between UC tissue and normal tissue(P>0.05).Recombi-nant IL-36Ra stimulation reduced CaCO_(2) cell death was decreased in both monocytes purified from peripheral blood and UC tissue in UC patients(P<0.05),TNF-α,granzyme B and granzyme H levels in supernatants were also down-regulated(P<0.05).There were no remarkable differences of FasL or TRAIL MFI in monocytes between cells with and without IL-36Ra stimulation(P>0.05).Conclu-sion:IL-36Ra may suppress monocytes cytotoxicity through down-regulation of TNF-αand granzyme secretions in UC patients.
作者
董戴源
郭晓鹤
薛耀峰
DONG Daiyuan;GUO Xiaohe;XUE Yaofeng(Department of Gastroenterology WardⅡ,the First Affiliated Hospital of Xinxiang Medical University,Weihui 453100,China)
出处
《中国免疫学杂志》
北大核心
2025年第8期1989-1996,共8页
Chinese Journal of Immunology
基金
2017年度新乡医学院第一附属医院青年基金项目(QN-2017-B019)。
