类风湿关节炎患者外周血Th17细胞/调节性T细胞变化及其临床意义

目的 通过检测类风湿关节炎(RA)患者外周血,Th17细胞和调节性T细胞的变化,探讨其在RA发病机制中的作用及其临床意义.方法 采用流式细胞术(FCM)检测57例RA患者和32名健康对照者外周血单个核细胞(PBMCs)中IL-17+CD4+T细胞(Th17细胞)与Foxp3+CD25+CD4+T细胞(调节性T细胞)占CD4+T细胞的百分比,分析其与RA临床及实验室指标的关系.统计学处理采用t检验和方差分析,相关分析采用Pearson直线相关分析.结果 RA组Th17细胞占CD4+T细胞的百分率较健康对照组明显升高[(4.2±2.2)%和(2.3±1.4)%,P=0.000],调节性T细胞降低[(3.9±1.6)%和(7.0±2.2)%,P=0.000],Th17细胞/涮节性T细胞比值显著升高(1.15±0.62和O.34±0.17,P=0.000).RA早期组与非早期组外周血Thl7细胞、调节性T细胞及,Th17调节性T细胞比值比较,差异无统计学意义(P均>0.05).相关性分析显示RA患者Th17细胞及Th17细胞/调节性T细胞比值与关节压痛数、患者疼痛模拟视觉评分(VAS)、RA28关节疾病活动评分(DAS)28评分等病情活动指标呈正相关(P均<0.05),而调节性T细胞与上述病情活动指标无明显相关性(P均>0.05).单因素回归分析显示Th17细胞、Th17细胞/调节性T细胞比值、DAS28评分及病程对RA患者骨侵蚀有显著影响(P均<0.05).经抗风湿药物治疗后,RA患者Th17细胞阙节性T细胞比值降低.结论 RA患者Th17细胞明显增多,调节性T细胞减少,Th17细胞/调节性T细胞比例明显增高.可能是RA发病的重要原因,通过抗风湿药物治疗可调节Th17细胞/调节性T细胞亚群平衡.

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Objective To analyze the level of Th17 and Treg cells in the peripheral blood of patients with rheumatoid arthritis(RA),and to investigate its role in the pathogenesis of RA and the clinical significance.Methods Flow cytometry(FCM)was used to analyze the ratio of interleukin(IL)-17+CD4+T (Thl7)cells and Foxp3+CD25+CD4+T(Treg)cells in CD4+T cells from the peripheral blood of 57 RA patients and 32 normal controls.T-test and Chi-square test were used for inter-group comparison and Pearson's linear analysis was used for correlation analysis.Resuits Compared with normal controls.the level of both IL-17+CD4+T cells and the ratio of Th17/Treg in RA patients increased significantly.while the level of Foxp3+CD4+CD25+T cells decreased markedly[(4.2±2.2)%vs(2.3±1.4)%,P=0.000;1.15±0.62 I)5 0.34±0.17,P=0.000;(3.9±1.6)%vs(7.0±2.2)%,P=0.000].Compared with early RA(persistent for 2 years or less)patients,the levels of Th17,Treg and the ratio of Th17/Treg in chronic RA(duration for more than 2 years)patients didn't markedly changed(P>0.05).The level of Th17 cells and the ratio of Th17/Treg was directlycorrelated with disease activity parameter (including tender ioint counts,visual analog seale of patients,disease activitv score in 28 joints,etc).Regression analysis discovered that risk factors of bone erosion were the level of Th17 cells,the ratio of Th17/Treg,disease activity score in 28 joints and disease duration.Antirheumatic drugs could decreascthe ratio of Th 17/Treg.Conclusions Treg cells is decreased in RA patients while Th7 cells is increased in patients with RA.Th17/Treg ratio goes up significantly as well.Change of T celt subsets,especially Th17 and Treg cells are important for the pathogenesis of RA.Th17 and Treg cells could aggrevate disease activity and bone destruction throughout the whole disease process.Anti-rheumatic medications is effective by regulating Th17/Treg subset balance.     

系统性红斑狼疮患儿外周血辅助性T17细胞表达的初步研究

目的 观察系统性红斑狼疮(SLE)患儿外周血辅助性T17(Th17)细胞表达变化,探讨Th17细胞及其相关的细胞因子在SLE患儿发病中的作用.方法 以15名健康儿童为健康对照组,流式细胞术(FCM)检测25例SLE患儿(SLE组)外周血Th17细胞百分率,用酶联免疫吸附试验(ELISA)法检测外周血血浆白细胞介素(IL)-17、IL-21水平,分析Th17细胞与SLE活动度之间的关系.统计学处理采用t检验和Spearman相关性检验.结果 与健康对照组相比,SLE患儿外周血中CD3+CD8-IL-17+T细胞的频率[(1.24±0.64)%,(0.59±0.21)%],CD3+CD8-IL-21+T细胞的频率[(1.5±0.6)%,(0.8±0.4)%]明显增高,P均＜0.01;血浆中IL-17,IL-21水平明显升高(P＜0.01).CD3+CD8-IL-17+T细胞频率与疾病活动度相关(r=0.732,P＜0.01),CD3+CD8-IL-21+T细胞频率与疾病活动度无相关性(r=-0.002,P＞0.05).结论 Th17细胞及其相关细胞因子参与儿童SLE发病及病情发展.

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Objective To investigate the Th17 cell expression in peripheral blood of childron with systemic lupus erythematosus (SLE) and explore the role of Th17 cells and the cytokines in the pathogenesis of SLE. Methods Twenty-five children with SLE were enrolled and 15 healthy children were controls. Flow cytometry (FCM) was employed to detect the expression of Th17 cells in peripheral blood of SLE children (SLE group, n=25), and IL-17, IL-21 levels in plasma were detected by ELISA. Two-independent sample t-test and Spearmen's test were used for correlation analysis. Results Compared with that of the control, the frequencies of CD3+CD8-IL-17+T[(1.24±0.64)% vs (0.59±0.21)%], CD3+CD8-IL-21+T cells[(1.5±0.6)%vs (0.8±0.4)% ] increased significantly in SLE patients (P＜0.01) and the plasma concentrations of IL-17, IL-21 were significantly higher (P＜0.01). The SLE activity was positively correlated with the frequencies of CD3+CD8-IL-17+T cells, but not with CD3+CD8-IL-21+T cells. Conclusion Th17 cells and the related cytokinesplay an important role in the pathogenesis of childhood SLE.     

调节性T细胞和辅助性T细胞17的平衡变化在慢性乙型肝炎病毒感染中的作用

目的 研究调节性T细胞(Treg)和辅助性T细胞17(Th17)的平衡变化在慢性HBV感染中的作用.方法 应用ELISA法和流式细胞术分别对34例慢性乙型肝炎(CHB)患者、20例HBV相关慢加急性肝功能衰竭(ACHBLF)患者和20例健康对照者外周血中Treg和Th17分化相关细胞因子及外周血Th17和Treg细胞水平进行检测.计数资料应用Fisher's确切概率法,计量资料应用单因素方差分析和Tukey's多重比较检验.结果 ACHBLF组Th17分化相关因子IL-1β为(3.97±2.85) pg/mL,IL-6为(12.75±8.87)pg/mL,IL-21为(360.0±335.7)pg/mL,比健康对照组的IL-1β[(1.87±0.94)pg/mL,q=4.559,P＜0.01)、IL-6[(5.28±o.72)pg/mL,q=7.309,P＜0.01)和IL-21[(46.68±20.17) pg/mL,q=6.946,P＜0.01)均明显上调.ACHBLF组外周血Th17细胞比例明显高于健康对照组(q=3.972,P＜0.05).与健康对照组和ACHBLF组相比,CHB组Treg细胞分化相关因子TGF-β明显升高(q=4.536、5.323,均P＜0.01),外周血Treg比例也明显升高.ACHBLF组Th17细胞效应因子IL-17A水平最高,ACHBLF患者外周血Th17细胞比例与血清TBil水平呈正相关(γ=0.74,P＜0.01).结论 慢性HBV感染中,宿主免疫存在Th17和Treg失衡,ACHBLF组以Th17细胞活动为主,CHB组以Treg细胞活动为主.     

流式细胞术分析外周血辅助性T17细胞在强直性脊柱炎患者外周血的表达及其意义

目的 检测强直性脊柱炎(AS)患者外周血辅助性T17(Th17)细胞的表达,探讨其在AS发病机制中的作用.方法 用流式细胞术检测20例AS患者和15名健康对照者外周血Th17[白细胞介素(IL)-17]、Th1[干扰素(IFN)-γ]、Th2(IL-4)细胞及人类白细胞抗原(HLA)-B27,同时检测红细胞沉降率(ESR)、C反应蛋白(CRp)等,并分析Th17细胞与Th1、Th2、HLA-B27及AS活动性指标的相关性.采用t检验和Spearman相关分析.结果 AS患者外周血Th17细胞[(2.6±0.8)%]高于对照组[(1.1±0.4)%](P＜0.01);Th1(IFN-γ)细胞[(3.9±0.8)%]低于对照组[(5.1±1.3)%](P＜0.05);Th2(IL-4)细胞[(4.1±1.6)%]比对照组[(3.1±1.4)%]升高,但差异无统计学意义(P＞0.05);Th17细胞与HLA-B27百分数及与HLA-B27平均荧光强度、ESR、CRP无相关性(P＞0.05);但是,Th17细胞表达水平有随HLA-B27百分数及HLA-B27平均荧光强度增加升高的趋势.结论 AS患者外周血Th1细胞减少,而Th17细胞增加,AS患者体内存在Th细胞的失衡,Th17比例变化是反映AS整个复杂免疫反应异常的一个重要环节.

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Objective To detect the expression of T helper 17 (Th17) cells in the peripheral blood of patient with ankylosing spondylitis (AS),and discuss its role in thc pathogenesis of AS.Methods Twenty AS patients and fifteen healthy controls were enrolled in the study.Th17 (IL-17),Th1 (IFN-γ),Th2 (IL-4)cells and HLA-B27 of their peripheral blood were analyzed by flow cytometry,at the same time,the levels of ESR and CRP were also measured in order to analyze thc relation of Th17 and HLA-B27,ESR as well as the CRP level.The statistical analysis was carried out with single-sample t-test and Speraman's correlation test.Results The level of Th17 cells was significantly higher in the perpipheral blood of AS patients[(2.6±0.8 )%] than those in healthy controls[ (1.1±0.4)% ] (P＜0.01).The level of Th 1 cells was significantly lower in the peripheral blood of AS patients[(3.9±0.8)%] than those in healthy controls[(5.1±1.3)%] (P＜0.01)and the level of Th2 cells was not different in the peripheral blood of AS patients[(4.1±1.6)%] when compared to healthy controls[ (3.1±1.4)% ] (P＞0.05).Th 17 cells was not significantly correlated with the percentage and mean fluorescent intensity of HLA-B27,ESR,CRP(P＞0.05); but there was a tendency that increased expression level of Th17 cells was associated with elevated percentage and mean fluorescent intensity of HLA-B27.Conclusion The level of Th1 cells is decreased,but Th17 is increased in the peripheral blood of AS patients.Th cells are imbalance in AS,patients.The change of Th17 cells may be an important part of the pathogenesis of AS.     

羟氯喹抑制紫外线诱导的系统性红斑狼疮CD4+T细胞白细胞介素-10和干扰素-γ的表达

目的 研究紫外线对系统性红斑狼疮(SLE)CD4+T细胞因子的影响和羟氯喹的抑制作用.方法 选择SLE 30例,健康对照10名.磁珠分选SLE患者和健康人的CD4+T细胞,紫外线311 nm窄谱中波紫外线暴露,加入羟氯喹共培养,酶联免疫吸附试验(ELISA)检测培养上清白细胞介素(IL)-10和干扰素-γ的表达水平.采用t检验进行统计学分析.结果 SLE患者CD4+T细胞IL-10表达高于健康对照[(27±4)和(18±3) pg/ml,P=0.011];经45、100 mJ/cm2紫外线暴露后,SLE活动患者CD4+T细胞IL-10表达升高[(27±4)和(77±42) pg/ml,(40±18)和(77±42) pg/ml,P=0.022,P=0.048],经100 mJ/cm2紫外线暴露后,活动患者CD4+T细胞IL-10表达高于稳定患者[(77±42)和(24±4)pg/ml,P=0.029];羟氯喹降低SLE活动患者CD4+T细胞IL-10和干扰素-γ表达[(2.6±4.0)和(17.9±2.3)pg/ml,P=0.018,P=-0.017)];羟氯喹降低经45,100 mJ/cm2紫外线暴露后SLE活动患者T细胞IL-10表达[(40±18)和(22±6)pg/ml,(77±42)和(21±5) pg/ml,P=0.037,P=0.04];羟氯喹降低经100 mJ/cm2紫外线暴露的SLE活动和稳定患者T细胞干扰素-γ表达[(18±3)和(13±14) pg/ml,(19±7)和(12±5) pg/ml,P=0.013,P=0.049].结论 紫外线加重SLE患者体内Th1/Th2细胞因子的比例失衡;羟氯喹抑制了紫外线诱发SLE患者干扰素-γ和IL-10的表达.

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Objective To explore the role of hydroxychloroquine (HCQ) in ultraviolet B (UVB)- induced expression of interleukin (IL)-10 and interferon (IFN)-γ from CD4+T cells in patients with systemic lupus erythematosus (SLE). Methods Thirty patients with SLE and 10 healthy controls were enrolled in the study. CD4+ T cells were isolated using magnetic beads from SLE patients and healthy controls. HCQ was added in culture media before and after irradiation with UVB 311 nm narrow band ultraviolet B (NB-UVB). The levels of IL-10 and IFN-γ in the supernatant were detected with enzyme-linked immunosorbent (ELISA). Comparisons between groups were performed by t-test. Results The level of IL-10 was higher in SLE patients [(27±4) pg/ml] than that in healthy controls [(18±3) pg/ml, P=0.011]. After exposure of CD4+T cells to UVB in 45 or 100 mJ/cm2 dosages, the level of IL-10 was increased significantly in patients with active disease (P=0.022, P=0.048). After exposure of CD4+T cells to UVB in 100 mJ/cm2 dosages, the levels of IL-10 was higher in patients with active disease [(77±42) pg/ml] than patients with stable disease [(24± 4) pg/ml, P=0.029]. When CD4+ T cell were cultured with HCQ, IL-10 and IFN-γ levels in patients with active disease [(2.6±4.0), (17.5±2.3) pg/ml] were decreased significantly (P=0.018, P=0.017). HCQ reversed UVB-induced IL-10 expression in active SLE patients after exposure of CD4+T cells to UVB in 45 or 100 mJ/cm2 dosages (P=0.037, P=0.04). HCQ also reversed UVB-induced IFN-7 expression in active SLE patients and stable SLE patients after exposure to CD4+T cells with UVB in 100 mJ/cm2 dosages (P=0.013, P= 0.049). Conclusion UVB can aggravate the imbalance of Th1 and Th2 cytokines. HCQ inhibits UVB-induced IL-10 and IFN-7 expression of CD4+T cells in patients with SLE, especially in patients with active disease.     

脐带间充质干细胞对系统性红斑狼疮患者Th17细胞的调节

目的 探讨脐带间充质干细胞(UC-MSCs)体内外对系统性红斑狼疮(SLE)患者CD3+CD8-IL17A+T淋巴细胞及相关细胞因子的调节作用.方法 UC-MSCs移植治疗14例SLE患者,观察移植前后的临床表现及实验窜指标的变化.采用流式细胞术检测各时间点患者外周血CD3+D8-IL17A+T细胞百分率,酶联免疫吸附试验(ELISA)测定血浆白细胞介素(IL)-6、转化生长因子(TGF)-β、IL-17A、IL-22表达水平.10例SLE患者外周血单个核细胞(PBMC)分别与UC-MSCs按不同比例体外共培养72 h,流式细胞术检测PBMC中CD3+CD8-IL17A+T细胞百分率.采用配对t检验和独立样本t检验.结果 UC-MSCs移植后患者疾病活动指数评分(SLEDAI)在3个月(7.8±1.2,t=2.19)及6个月(6.9±0.9,t=4.2)均显著低于移植前(10.4±0.9,P<0.05);尿蛋白定量(24 h)移植后6个月显著下降[(1489±260)与(2454±322)mg,t=2.6,P<0.05];血清白蛋白水平在移植后1、3、6个月均显著高于移植前(P<0.05),补体C3移植后持续升高.UC-MSCs移植后1个月及6个月,患者外周血CD3+CD8-IL17A+T淋巴细胞百分率显著下降(P<0.05).UC-MSCs与SLE患者PBMC共培养可显著下调CD3+CD8-IL17A+T淋巴细胞百分率(P<0.05),但无剂量依赖性.UC-MSCs移植后患者血浆IL-6、TGF-β、IL-17A、IL-22水平与移植前比,差异无统计学意义(P>0.05).结论 UC-MSCs移植治疗SLE有效,移植后SLE患者PBMC中CD3+CD8-IL17A+T淋巴细胞水平显著下降.

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Objective To investigate the regulatory effects of umbilical cord-derived mesenchymal stem cells (UC -MSCs) on Th17 cells and related cytokines in patients with systemic lupus erythematosus (SLE). Methods Human UC-MSCs were isolated and expanded and infused into fourteen SLE patients. Clinical changes were evaluated before and after transplantation by SLE disease activity index (SLEDAI), 24-hour urine protein, serum albumin and complement C3. The percentages of CD3 +CD8-IL17A + T cells in peripheral blood were detected by flow cytometry. Concentrations of plasma IL-6, TGF-β, IL-17A, IL-22were determined by enzyme-linked immunosorbent assay (ELISA). UC-MSCs and peripheral blood mononindependent samples t-test. Results SLEDAI scores decreased significantly at 3 month (7.8±1.2, t=2.19) and 6 month (6.9±0.9, t=4.2) after UC-MSCs transplantation than pre-transplantation level (10.4±0.9, P< 0.05). Twenty-four-hour proteinuria decreased significantly 6 months after MSCs infusion [(1489±260) mg vs (2454±322) mg, t=2.6, P<0.05]. Meanwhile, serum albumin and complement C3 levels had increased significantly since 1 month after transplantation (P<0.05). The percentages of peripheral blood CD3+CD8-IL17A+T cells decreased obviously in 1 week, 1 month and 6 months after UC-MSCs transplantation (all P<0.05). The coculture of UC-MSCs with PBMC from SLE patients resulted in a statistically significant reduction of CD3+CD8-IL17A+T cells percentage in PBMC (P<0.05), but was not in a dose dependent manner. No change of plasma IL+6, TGF-β, IL-17A and IL-22 levels was observed after UC-MSCs transplantation (P>0.05).Conclusion UC-MSCs transplantation down-regulates the percentages of CD3+CD8-IL17A+T cells in SLE patients, which may be one of the mechanisms for its therapeutic effect in refractory SLE.     

系统性红斑狼疮患儿外周血辅助性T17细胞表达的初步研究

Objective To investigate the Th17 cell expression in peripheral blood of childron with systemic lupus erythematosus (SLE) and explore the role of Th17 cells and the cytokines in the pathogenesis of SLE. Methods Twenty-five children with SLE were enrolled and 15 healthy children were controls. Flow cytometry (FCM) was employed to detect the expression of Th17 cells in peripheral blood of SLE children (SLE group, n=25), and IL-17, IL-21 levels in plasma were detected by ELISA. Two-independent sample t-test and Spearmen's test were used for correlation analysis. Results Compared with that of the control, the frequencies of CD3+CD8-IL-17+T[(1.24±0.64)% vs (0.59±0.21)%], CD3+CD8-IL-21+T cells[(1.5±0.6)%vs (0.8±0.4)% ] increased significantly in SLE patients (P＜0.01) and the plasma concentrations of IL-17, IL-21 were significantly higher (P＜0.01). The SLE activity was positively correlated with the frequencies of CD3+CD8-IL-17+T cells, but not with CD3+CD8-IL-21+T cells. Conclusion Th17 cells and the related cytokinesplay an important role in the pathogenesis of childhood SLE.     

非小细胞肺癌患者外周血调节性T细胞和辅助性T细胞17的平衡表达特征

目的 通过检测辅助性T(Th)细胞17、调节性T细胞(regulatory T cell,Treg)相关细胞因子及特异性转录因子维A酸相关孤核受体(RORγt)和叉头转录因子3(forkhead box P3,FoxP3) mRNA的表达变化,探讨非小细胞肺癌患者外周血Treg和Th17的平衡表达特征.方法 57例非小细胞肺癌患者作为研究对象,25名健康人作为对照.通过荧光定量PCR方法检测外周血单个核细胞(PBMC)中RORγt和FoxP3 mRNA的表达.应用酶联免疫吸附方法(ELISA)检测患者外周血转化生长因子β(transforming gowth factor-β,TGF-β)和Th17相关细胞因子白介素17 A(IL-17A)、IL-6的表达.结果 Ⅳ期非小细胞肺癌患者PBMC的RORγt mRNA水平[(11.30±4.07)×10-4]高于健康组[(7.50±2.75)×10-4]及Ⅰ～Ⅲ期非小细胞肺癌患者[(5.18±2.10)×10-4](q值分别为7.53、6.82,P值均＜0.01).Ⅳ期患者血浆TGF-β水平[(69.52±10.04) pg/ml]明显高于I～Ⅲ期非小细胞肺癌患者[(42.24±16.64)pg/ml]和健康对照组[(24.51±10.96)pg/ml](q值分别为3.91、3.87,P值均＜0.05).血浆IL-6、IL-17A各组间差异无统计学意义[非小细胞肺癌Ⅳ期组、Ⅰ～Ⅲ期组、健康对照组血浆IL-17A含量分别为(25.20±4.58)pg/ml、(18.58±5.92)pg/ml、(18.93±5.24)pg/ml];[非小细胞肺癌Ⅳ期组、Ⅰ～Ⅲ期组、健康对照组血浆IL-6含量(pg/ml)分别为(39.53±11.30) pg/ml]、(42.41±12.38)pg/ml、(38.15±12.09)pg/ml(血浆IL-17A组间两两比较q值分别为1.21、1.09、1.12;血浆IL-6组间两两比较q值分别为1.19、1.76、1.06,P值均＞0.05).结论 晚期非小细胞肺癌中RORγt mRNA表达增强、外周血TGF-β表达增高,Th17以及Treg细胞可能同时参与了非小细胞肺癌的肿瘤远处转移过程,从而影响肿瘤进程.     

CD4+白细胞介素-17+辅助性T细胞对香烟暴露小鼠肺部炎症及肺气肿的作用

目的 观察香烟烟雾暴露小鼠肺实质中CD4+白细胞介素(IL)-17+辅助性T细胞(Th17)数量及活性的表达,探讨其在香烟暴露小鼠肺部CD4+ γ-干扰素+(Th1)炎症及肺气肿中的作用及相关机制.方法 将40只雄性Balb/c小鼠按随机数字表法分为4组:对照12周(C12)组、对照24周(C24)组、烟雾暴露12周(S12)组、烟雾暴露24周(S24)组,每组10只.香烟烟雾暴露法建立小鼠肺气肿模型.HE染色观察小鼠肺气肿的改变,计算平均内衬间隔和肺泡破坏指数(DI);流式细胞术检测小鼠肺实质中CD4+IL-17+T(Th17)细胞、CD4+γ-干扰素+T(Th1)细胞、CD4+IL-17+γ-干扰素+T(Th17/Th1)细胞、CD8+γ-干扰素+T(Tc1)细胞、CD8+IL-21R+细胞及CD4+IL-17+IL-21+细胞比例;荧光定量PCR法检测小鼠肺实质中维甲酸相关孤独受体(RORγt)和IL-17的mRNA表达,并分析这些指标的相互关系.结果 S12组和S24组的平均内衬间隔[(39±4)μm和(47±7)μm]和DI(39.1±1.6和45.2±3.1)明显高于C12组[(32±4)μm和28.2±1.6]和C24组[(33±3)μm和28.9±2.1],且以S24组的增高更为明显,差异均有统计学意义(t值为4.378～15.188,均P＜0.05);S12组和S24组Th17细胞比例[(3.3±1.1)%和(7.2±2.2)%]均明显高于C12组和C24组[(1.8±0.8)%和(2.0±0.6)%];S12组和S24组RORγtmRNA表达量[(25±4)和(35±3)]及IL-17的mRNA表达量[(26±3)和(36±3)]亦明显高于C12组[(10±5)和(13±5)]和C24组[(11±7)和(8±6)],以S24组增高更为明显,差异均有统计学意义(P＜0.05);S12组和S24组Th1细胞比例[(10.0±3.7)%和(26.2 ±6.0)%]、Th17/Th1细胞比例[(0.61±0.30)%和(1.82±0.52)%]及Tc1细胞比例[(17.0±4.5)%和(26.8±8.5)%]均明显高于C12组[(3.8±1.7)%、(0.27±0.17)%和(4.8±1.9)%]和C24组[(4.2±1.3)%、(0.28±0.11)%和(5.2±1.0)%],以S24组增高更为明显,差异均有统计学意义(P＜0.05);S12组和S24组小鼠Th17细胞与Th1、Tc1细胞比例、平均内衬间隔、DI值均呈显著正相关(r值为0.519～0.797,均P＜0.01);Th17/Th1细胞比例与平均内衬间隔、DI值呈显著正相关(r值分别为0.742和0.802,均P＜0.01);S12组和S24组CD4+IL-17+IL-21+细胞比例[(0.19±0.04)和(0.55±0.24)]明显高于C12组和C24组[(0.07±0.03)和(0.08±0.03)],S24组增高更为明显,差异均有统计学意义(P＜0.05).S12组和S24组的CD8+IL-21R+细胞比例[(2.94±1.26)和(4.12±2.26)]高于C12组和C24组[(1.22±0.31)和(1.34±0.18)](P＞0.05);S12组及S24组小鼠CD4+IL-17+IL-21+细胞比例与Th1、Tc1细胞比例、平均内衬间隔和DI值均呈显著正相关(r值为0.694～0.754,均P＜0.05);S12及S24组小鼠CD8+IL-21R+细胞比例与平均内衬间隔和DI呈显著正相关(r值分别为0.516和0.725均P＜0.05).结论 香烟暴露导致肺气肿小鼠肺内Th17细胞数量及活性上调,并随烟雾暴露时间延长而增强;Th17细胞通过IL-21及IL-21R在肺部Th1/Tc1炎症中起重要促进作用;这对探讨COPD肺部炎症和肺气肿发生机制以及新的治疗靶点具有重要意义.

Abstract：

Objective To evaluate the expression and the role of Th 17 in cigarette smoke-induced lung inflammation and emphysema in mice.Methods Forty male BALB/c mice were randomly divided into 4 groups, including a control group C12, a control group C24, a smoke-exposure 12 week group (S12) and a smoke-exposure 24 week group S24 (n = 10 each).Morphological changes were evaluated by mean linear intercepts and destructive index (DI).The proportion of CD4+ IL-17 + Th17, CD4+ IFN-γ+ Th1, CD4+ IL-17 +IFN-γ+ T( Th17/ Th1 ), CD8+ IFN-γ+ Tc1, CD8+ IL-21R + and CD4+ IL-17 + IL-21 + T cells in lungs of mice was determined by flow cytometry.The mRNA expressions of RORγt and IL-17 were evaluated by real-time PCR.Results Mean linear intercepts and DI were significantly higher in S12 and S24 groups [(39 ± 4)μm, (47 ±7) μm], (39.1 ± 1.6, 45.2 ±3.1 ) as compared to C12[(32 ±4) μm,28.2 ± 1.6] and C24groups [(33 ± 3 ) μm ,28.9 ± 2.1], all P ＜ O.05.The percentage of Th17 of S12 and S24 groups [(3.3 ±1.1 )%, (7.2 ±2.2)%] was significantly increased as compared with that of C12 and C24 groups [( 1.8± 0.8) %, (2.0 ± 0.6) %], all P ＜ 0.05.The mRNA levels of RORγt [( 25 ± 4), ( 35 ± 3 )] and IL-17 [(26 ± 3), (36 ± 3 )] in S12 and S24 groups were higher than in C12 [(10 ± 5 ), (13 ± 5 )] and C24 groups [( 11 ± 7 ), (8 ± 6)], all P ＜ 0.05.The percentage of Th 1, Th17/Th1 and Tc1 cells of S12 and S24 groups [(10.0 ±3.7)%, (26.2 ±6.0)%], [(0.61 ±0.30)%, (1.82 ±0.52)%], [(17.0±4.5 ) %, ( 26.8 ± 8.5 ) %] was significantly increased as compared with that of C12 [( 3.8 ± 1.7 ) %,(0.27±0.17)%, (4.8 ±1.9)%] and C24 groups [(4.2±1.3)%, (0.28±0.11)%, (5.2±1.0)%], all P＜0.05.Moreover, the frequency of Th17 cells had a positive correlation with Th1, Tc1 cells and emphysematous lesions ( r =0.519 - 0.797, all P ＜ 0.01 ).In addition, a positive correlation between Th17/Th1 cells and emphysematous lesions was also found (r =0.742, 0.802, all P ＜0.01 ).The percentage of CD4+ IL-17+ IL-21 +T cells was significantly increased in S12 and S24 groups [(0.19 ±0.04) %, (0.55 ± 0.24) %] compared to controls [(0.07 ± 0.03 ) %, (0.08 ± 0.03 ) %], all P ＜ 0.05.Meanwhile, as compared with that of the controls [( 1.22 ± 0.31 ), ( 1.34 ± 0.18 )], the percentage of CD8+ IL-21 R + T cells was also increased in SI 2 and S24 groups [( 2.94 ± 1.26 ), (4.12 ± 2.26 )], but there were no differences among smoke-exposure groups ( P ＞0.05 ).The frequency of CD4+ IL-17 + IL-21 + T cells had a positive correlation with Th 1, Tc1 cells and emphysematous lesions (r = 0.694 -0.754, all P ＜0.05).And the frequency of CD8+ IL-21R+ T cells also had a positive correlation with emphysematous lesions ( r = 0.516, 0.725, all P ＜ 0.05).Conclusions Cigarette smoke increased the expression and the activity of Th17 in mice.Th17 may play a potential (active) role in the development of lung inflammation through IL-21/IL-21R pathway.     

系统性红斑狼疮患者外周血Th17细胞与调节性T细胞的变化

目的 探讨外周血辅助性T细胞17(Th17)与调节性T细胞在系统性红斑狼疮(SLE)发生、发展中的变化.方法 选取32例SLE活动期患者作为SLE活动组、30例SLE稳定期患者作为SLE稳定组及25名健康者作为对照组,采用流式细胞术及定量聚合酶链反应(PCR)的方法,分别从蛋白质水平与mRNA水平检测其外周血CD4+T细胞IL-17及FoxP3的表达.采用单因素方差分析进行统计学处理.结果 SLE活动组外周血CD4+T细胞IL-17蛋白质(1.01±0.22)％及mRNA(2.04±0.63)表达水平显著高于SLE稳定组(0.48±0.16)％、(1.12±0.34)及健康对照组(0.41±0.12)％、1(P＜0.01),但SLE稳定组与健康者对照组差异无统计学意义(P＞0.05).SLE活动组外周血CD4+T细胞FoxP3蛋白质(2.36±0.54)％及mRNA(0.42±0.16)表达水平显著低于SLE稳定组(4.34±0.95)％、(0.87±0.28)及健康对照组(5.09±1.17)％、 1(P＜0.01),SLE稳定组又低于健康对照组(P＜0.05).结论 SLE患者外周血中可能存在Th17/调节性T细胞的失衡,且失衡程度可能与病情活动性相关.     

系统性红斑狼疮患者外周血白细胞介素-17与辅助T细胞17的检测及其意义

目的 研究系统性红斑狼疮(SLE)患者外周血白细胞介素(IL)-17蛋白和mRNA水平、辅助T细胞(Th17)细胞表达比例,探讨其临床意义.方法 用酶联免疫吸附试验检测SLE患者及对照者血浆中IL-17的蛋白水平;采用实时荧光定量反转录-聚合酶链式反应(RT-PCR)检测2组外周血中IL-17 mRNA表达水平;运用流式细胞术检测SLE患者外周血单个核细胞(PBMCs)中Th17细胞比例,进一步分析IL-17/Th17细胞与SLE临床实验室指标的相关性.计量资料组间比较采用t检验,相关性分析采用Spearman秩和相关分析.结果 SLE组患者血浆IL-17含量明显高于健康对照组,SLE组患者外周血IL-17 mRNA表达水平[(28.3±11.7)×10-5]高于对照组[(9.8±2.2)×10-5](P均＜0.01).SLE患者PBMCs中Th17细胞比例(2.5±1.5)%及IL-17荧光强度(1937±1022)显著高于对照组[(1.5±0.7)%,(1245±413)],且SLE活动期患者Th17细胞百分比高于非活动组,SLE肾病组Th17细胞比例较无肾病组明显升高(P均＜0.05).SLE患者血浆IL-17水平、Th17细胞数与SLE疾病活动性指数(SLEDAI)呈正相关(r=0.681,P＜0.01;r=0.426,P=0.034).结论 SLE患者体内IL-17蛋白分泌及基因表达水平存在明显异常,外周血Th17细胞比例亦显著升高,且与疾病活动性有明显相关,提示IL-17/Th17细胞可能在SLE发病中起着重要作用.     

系统性红斑狼疮患者外周血CD4+CD25highFoxp3+调节性T细胞数量和Foxp3mRNA的表达水平与疾病活动性的相关性

目的 研究系统性红斑狼疮(SEE)患者CD4+CD25highFoxp3+调节性T细胞的数量及其功能基因Foxp3 mRNA的表达水平与SLE疾病活动性和肾脏损伤的相关性.方法 采用四色流式细胞术以Foxp3-异硫氰酸荧光素(FITC )/CD25-藻红蛋白/CD4-多甲藻叶绿素蛋白(PerCP)/CD3-藻蓝蛋白7抗体组合检测40名健康对照者及42例SLE患者外周血CD4+CD25highFoxp3+调节性T细胞的数量,实时荧光定量聚合酶链反应(PCR)检测特异性转录因子Foxp3 mRNA的表达水平,并分析其与SLE患者疾病活动指数(SLEDAI)、补体C3及血清抗双链DNA(dsDNA)抗体的关系.统计学方法采用t检验和Spearman相关分析.结果 活动期SLE患者外周血CD4+CD25highFoxp3+调节性T细胞数量显著低于健康对照组[(4±3)％与(7±4)％,P＜0.05],稳定期与健康对照组差异无统计学意义(P＞0.05);活动期SLE患者外周血CD4+CD25highFoxp3+调节性T细胞数量及CD4+CD25highFoxp3+调节性T细胞/CD4+比值显著低于稳定期患者[(4±3)％,(9±6)％与(5±4)％,(10±6)％,P均＜0.05];活动期SLE患者外周血Foxp3 mRNA的表达水平明显低于稳定期和对照组(P＜0.01,P＜0.05);SLE患者并发肾病组外周血CD4+CD25highFoxp3+调节性T细胞数量及CD4+CD25highFoxp3+调节性T细胞/CD4+比值显著低于SLE非肾病组(P＜0.05).相关分析显示,SLE患者外周血CD4+CD25highFoxp3+调节性T细胞数量与SLEDAI呈负相关(r=-0.5782,P＜0.05);CD4+CD25highFoxp3+调节性T细胞/CD4+比值与SLEDAI呈负相关(r=-0.4913,P＜0.05),与补体C3呈正相关(r=0.3687,P＜0.05);SLE患者外周血CD4+CD25highFoxp3+调节性T细胞数量与Foxp3 mRNA的表达水平呈正相关(r=0.6142,P＜0.0l).结论 SLE患者外周血CD4+CD25highFoxp3+调节性T细胞和Foxp3 mRNA的变化可能是导致SLE疾病发生和发展的关键因素之一,与疾病的活动性有密切关系.     

系统性红斑狼疮患者体内Th17、Th1细胞及其相关因子的表达及意义

目的 研究系统性红斑狼疮（SLE）患者血浆IL-17、干扰素（IFN）-γ水平及外周血辅助性T细胞（Th）17和Th1细胞表达比例,探讨其临床意义.方法 用酶联免疫吸附法检测SLE患者及对照组血浆中IL-17、IFN-γ浓度;运用流式细胞术检测SLE患者外周血单个核细胞（PBMCs）中Th17、Th1细胞表达比例.进一步分析Th17/Th1细胞及其相关因子与SLE临床实验室指标的相关性.结果 SLE患者PBMCs中Th17细胞比例显著高于对照组,且SLE活动期患者Th17细胞百分比高于非活动期患者（P〈0.05）,而SLE患者Th1细胞比例与对照组无差异（P〉0.05）.SLE患者血浆IL-17水平、Th17细胞数与SLE疾病活动性指数呈正相关.结论 SLE患者体内IL-17水平、Th17细胞比例显著升高,且与疾病活动性明显相关,提示IL-17/Th17细胞在SLE发病机制中可能发挥重要作用.     

系统性红斑狼疮患者外周血中γδT细胞的变化及临床意义

目的 探讨γδT细胞在系统性红斑狼疮(SLE)患者外周血中的变化及临床意义.方法 流式细胞术检测42例SLE患者、20名健康对照组外周血中CD3+γδ+T细胞百分率及绝对数;用膜联蛋白-Ⅴ/碘化丙啶法检测6例活动期SLE患者及6名正常对照组外周血中CD3+T细胞及γδ+T细胞的凋亡率.酶联免疫吸附试验( ELISA)法测定42例SLE患者血清抗核抗体和抗双链DNA(dsDNA)抗体水平.统计学方法采用t检验及Pearson相关分析.结果 活动期SLE患者外周血中T细胞百分率[(3.0±1.8)％]较非活动期[(5.3±3.0)％]及健康对照组[(6.8±2.8)％]均显著下降(t值分别为-3.071和-5.913,P均＜o.01);活动期SLE患者γδT细胞绝对数[(1.7±1.6)×107/L]明显低于非活动期[(5.3±3.6) ×107/L](t=-3.292,P＜0.01),且两者均低于健康对照组[(10.1 ±5.0) ×107/L](t值分别为-7.247和-2.905,P均＜0.01).活动期SLE患者γδT细胞绝对数与SLE疾病活动指数(SLEDAI)评分呈负相关(r=-0.365,P=0.047).SLE患者外周血中γδT细胞百分率(r=-0.336,P=0.030)及绝对数(r=0.410,P=0.007)均与红细胞沉降率( ESR)呈负相关,γδT细胞绝对数与血红蛋白呈正相关(r=0.409,P=0.007).此外,SLE患者外周血中γδT细胞凋亡率显著高于健康对照组及SLE患者CD3+T细胞凋亡率(t值分别为2.886和2.952,P均＜0.05).结论 SLE患者外周血中γδT细胞数量下降,与病情活动相关;γδT细胞数量下降的机制可能与凋亡增加有关.