双歧杆菌对哮喘儿童DC表达CD86和HLA-DR的影响

目的研究双歧杆菌对过敏性哮喘儿童外周血单个核细胞来源的树突状细胞（DC）表面表达CD86和HLA-DR的影响。方法从12例过敏性哮喘儿童和10例对照组儿童的外周血单个核细胞诱导生成未成熟DC,分别经双歧杆菌或细菌脂多糖（LPS）处理,用流式细胞仪检测各组DC表面CD86和HLA-DR分子表达。结果双歧杆菌刺激后,哮喘儿童DC表面CD86表达明显增高（P〈0．05）,HLA-DR表达无明显变化（P〉0．05）,对照组儿童CD86和HLA-DR表达无明显影响;LPS刺激可明显增加哮喘儿童和对照组儿童CD86和HLA-DR的表达。结论过敏性哮喘儿童DC表面CD86的表达可能存在缺陷,双歧杆菌能适度上调其表达,在DC的成熟过程中可能起调节作用。     

过敏性哮喘小鼠树突状细胞负荷Der p2后改变及与T细胞增殖分化的关系研究

目的:探讨哮喘小鼠与正常小鼠骨髓源性树突状细胞(DC)负荷Der p2抗原后表达表面分子(CD11c、CD86)和细胞因子(IL-10、IL-12p70)的差异及其对Th1和Th2型细胞因子平衡的影响,进一步研究过敏性哮喘发生中DC的可能作用。方法:分别从哮喘组和对照组提取骨髓培养DC,第五天负荷Der p2,24小时后吹打收集细胞,观察DC形态,用流式细胞仪检测孵育后细胞表面CD11c、CD86表达。并留取负荷Der f2前后培养上清,ELISA法检测IL-10及IL-12p70含量。同时以DC:反应细胞比例为1:10混合培养,72 h后ELISA法检测混合培养上清中IL-4、IL-5、IFN-γ的水平。结果:1负荷Der p2后,哮喘组CD86、CD11c表达比对照组高,分别为(t=11,P0.05)(t=4.9,P0.05),差异有统计学意义;2在细胞因子分泌方面,Der p2负荷前,两组DC均能分泌IL-10与IL-12p70,IL-10水平哮喘组高(t=9.5,P0.05),而IL-12p70水平对照组高(P0.05);负荷Der p2后,对照组IL-10、IL-12p7分泌量比负荷前明显增加(P0.05),哮喘组无明显差异(P0.05);3在DC刺激同种T细胞因子分泌方面,负荷Der p2后哮喘组DC刺激T细胞分泌IL-4、IL-5分泌能力明显增强(P0.05),而刺激INF-γ能力降低(P0.05)。结论:DC在过敏性哮喘中起着重要作用,异常DC通过增加CD86、CD11c的表达和减少IL-10及IL-12的合成,致使T细胞向Th2细胞优势分化。     

双歧杆菌WPG对正常和哮喘模型小鼠骨髓来源树突状细胞分泌细胞因子的影响

目的研究双歧杆菌完整肽聚糖(WPG)对正常和哮喘模型小鼠骨髓来源树突状细胞(DC)分泌细胞因子的影响,探讨双歧杆菌WPG在防治过敏性疾病中的免疫调节作用。方法分离正常及哮喘模型小鼠骨髓单个核细胞,诱导生成未成熟DC,在培养的第7天将细胞分为WPG组、阳性组,其中WPG组每毫升培养液中加入双歧杆菌WPG 5μg,阳性组每毫升培养液中加入脂多糖(LPS)1μg,ELISA法检测DC培养上清中IL-12及IL-10的水平。结果正常小鼠WPG组DC分泌的IL-12、IL-10的量低于阳性组,哮喘小鼠WPG组DC分泌IL-12、IL-10的量高于阳性组,差异均具有统计学意义(P0.05);哮喘小鼠WPG组、阳性组DC分泌IL-12、IL-10的量分别低于正常小鼠WPG组、阳性组,差异均具有统计学意义(P0.05)。结论双歧杆菌WPG对具有功能缺陷的哮喘小鼠DC功能有一定调节作用,但其调节作用有一定限度,不能使细胞功能完全恢复到正常时的水平。     

支气管哮喘病儿外周血树突状细胞功能变化

目的:观察支气管哮喘病儿外周血单个核细胞(PBMC)来源树突状细胞(DC)功能变化。方法:以18例健康儿童为对照,选择16例支气管哮喘发作期病儿为研究对象,分离PBMC并经rhGM-CSF诱生成熟DC。采用流式细胞仪(FACS)检测DC表面共刺激分子CD80(B7-1)、CD86(B7-2)和CD83的表达率;ELISA法检测培养上清液中IL-10和IL-12的变化。结果:①哮喘组DC表面CD86的表达率明显高于健康对照组(t=2.27,P<0.05),CD80、CD83的表达率与健康对照组比较均无显著性差异(t=1.17,1.34;P>0.05)。②哮喘组DC分泌IL-10、IL-12水平均明显低于健康对照组(t’=3.31,3.39;P<0.01)。③哮喘组DC分泌IL-10与IL-12成正相关(r=0.740,P<0.01),而健康对照组IL-10与IL-12无相关性(r=0.232,P>0.05)。结论:支气管哮喘病儿DC存在功能缺陷,主要表现在CD86表达升高、IL-10、IL-12分泌减少。     

乳杆菌对牛乳β-乳球蛋白致敏小鼠淋巴细胞分泌Th1/Th2型细胞因子和抗体的体外影响

为了分析乳杆菌对致敏小鼠脾淋巴细胞分泌Th1/Th2细胞因子及抗体的体外影响,用牛乳β-乳球蛋白腹腔注射BALB/c小鼠建立过敏症模型,造模成功后,分离致敏小鼠的脾淋巴细胞并与4种活/死乳杆菌(107 CFU/mL)体外共同孵育,ELISA法检测细胞上清液中细胞因子(IL-12、IFN-γ和IL-4)和抗体(总IgE、β-Lg特异性IgE和总IgG)含量。4种活/死乳杆菌均可体外调节致敏小鼠脾淋巴细胞分泌细胞因子和抗体的水平,特别是热致死的发酵乳杆菌和嗜酸乳杆菌可提高淋巴细胞IL-12和IFN-γ的分泌,抑制IL-4的分泌,使其IFN-γ/IL-4比值(代表Th1/Th2细胞平衡)高于活菌,与空白对照组比较差异显著(P<0.05)。同时,这两株热致死菌还可显著下调细胞上清液中总IgE、特异性IgE和总IgG抗体的浓度(P<0.05)。试验结果表明乳杆菌可提高牛乳β-乳球蛋白致敏小鼠脾淋巴细胞的IFN-γ/IL-4比值,进而纠正Th2占优势的Th1/Th2失衡,下调抗体分泌量,且具有菌株特异性。     

白细胞介素18

白细胞介素-18(IL-18)是新发现的细胞因子,具有多种生物学功能.IL-18能促进外周血单个核细胞产生干扰素-γ(IFN-γ)、IL-2、粒细胞巨噬细胞集落刺激因子(GM-CSF)等细胞因子,增强天然杀伤细胞(NK细胞)的细胞毒作用.IL-18结构上与IL-1相似,而功能更接近IL-12,IL-18与IL-12均能诱导Th1细胞产生IFN-γ,存在协同效应,但它们的作用途径不同.IL-18在抗感染抗肿瘤等方面有着潜在的应用前景,并与自身免疫性疾病的发病密切相关.     

乙型肝炎肝硬化患者肠道菌群变化与血清细胞因子水平的相关性

目的探讨乙型肝炎肝硬化患者肠道菌群变化与血清干扰素-α(IFN-α)、白细胞介素6(IL-6)、白细胞介素1β(IL-1β)水平的相关性。方法选择2017年12月至2019年12月我院收治的105例乙型肝炎肝硬化患者为A组,50例乙型肝炎患者为B组,同期于我院体检健康者50例为C组。比较3组患者粪便标本中菌群分布情况,同时比较3组患者血清IFN-α、IL-6、IL-1β水平,不同程度肠道菌群失调乙型肝炎肝硬化患者血清IFN-α、IL-6、IL-1β水平。采用Pearson相关分析乙型肝炎肝硬化患者肠道菌群与血清IFN-α、IL-6、IL-1β的相关性。结果 A组患者肠道乳杆菌、双歧杆菌数量显著低于B组,而肠杆菌、肠球菌及血清IFN-α、IL-6、IL-1β水平显著高于B组(均P0.05)。B组患者肠道乳杆菌、双歧杆菌数量显著低于C组,而肠杆菌、肠球菌及血清IFN-α、IL-6、IL-1β水平显著高于C组(均P0.05)。Ⅱ度菌群失调乙型肝炎肝硬化患者血清IFN-α、IL-6、IL-1β水平显著高于I度菌群失调者(均P0.05)。Ⅲ度菌群失调乙型肝炎肝硬化患者血清IFN-α、IL-6、IL-1β水平显著高于Ⅱ度菌群失调者(均P0.05)。乙型肝炎肝硬化患者肠道肠杆菌、肠球菌数量与血清IFN-α、IL-6、IL-1β水平呈正相关,而其肠道乳杆菌、双歧杆菌数量与血清IFN-α、IL-6、IL-1β水平呈负相关(均P0.05)。结论乙型肝炎肝硬化患者存在明显的肠道菌群失调,同时其外周血IFN-α、IL-6、IL-1β水平呈现上升趋势。肠道菌群与血清IFN-α、IL-6、IL-1β可能协同参与了乙型肝炎肝硬化的发生及发展。     

五种益生菌菌株对脐血单个核细胞免疫作用的实验研究

目的比较不同剂量的5种不同益生菌菌株的免疫调节作用,为选择适当的菌株进行治疗提供依据。方法取7例健康孕妇的脐血分离出的脐血单个核细胞(cord blood monocular cells,CBMC),分别与长双歧杆菌6-1株、婴儿双歧杆菌CGMCC313-1株、嗜酸乳杆菌YIT2004株、粪链球菌YIT0072株和酪酸梭状芽胞杆菌CGMCC313-2株,以菌和CBMC比例2∶1(低)、20∶1(中)和200∶1(高)共培养24~36 h,同时设阴性对照(PBS)和阳性对照(脂多糖,LPS)。然后采用流式细胞仪检测各组CBMC表面CD4、CD25分子表达情况,用ELISA方法检测培养上清中IL-10、IL-12、IL-4、TGF-β1和IFN-γ的水平。结果 (1)与阴性对照(PBS)组相比,除200∶1比例的酪酸梭状芽胞杆菌CGMCC313-2株能够显著提高CBMC表达CD4CD25(10.45±3.16 vs 5.84±2.32,P=0.009)以外,其余益生菌菌株对表达CD4CD25差异均无统计学意义(P0.05)。(2)长双歧杆菌6-1株在中高剂量下,能够刺激CBMC产生IL-10和IFN-γ,对产生IL-12无明显影响。(3)婴儿型双歧杆菌CGMCC313-1株在各个剂量下均能够刺激CBMC产生IL-10,对IL-12和IFN-γ产生无明显影响。(4)酪酸梭状芽胞杆菌CGMCC313-2株在中高剂量下,能够刺激CBMC产生IL-10,对IL-12和IFN-γ产生无明显影响。(5)粪链球菌YIT0072株在低中剂量下,能够刺激CBMC产生IL-10、IL-12和IFN-γ,而高剂量则无影响。(6)嗜酸乳杆菌YIT2004株在中高剂量下,能够刺激CBMC产生IL-10,在中剂量下,能够刺激CBMC产生IFN-γ,对IL-12无影响。(7)在本研究中均未能检测出IL-4和TGF-β1。结论在目前国内使用的益生菌菌株中,仅酪酸梭状芽胞杆菌CGMCC313-2株能够显著提高CBMC表达CD4CD25。5种菌株均能够刺激CBMC产生抗炎症因子IL-10;长双歧杆菌6-1株、粪链球菌YIT0072株和嗜酸乳杆菌YIT2004株能够刺激CBMC产生Th1型细胞因子INF-γ,仅粪链球菌YIT0072株能够刺激CBMC产生IL-12。各个菌株在不同的剂量下,具有不同作用。提示在应用益生菌治疗免疫等相关性疾病时,应该考虑不同菌株对免疫细胞的不同作用机制。     

白细胞介素21对SHIV感染的恒河猴外周血CD8~＋ T细胞分泌γ干扰素的影响

目的研究白细胞介素21（interleukin 21,IL-21）对SHIV感染CD8＋T细胞分泌干扰素γ（interferon-γ,IFN-γ）的影响。方法从SHIV/恒河猴模型外周血中分选出CD8＋T细胞,加入IL-21诱导培养,应用ELISA方法检测细胞培养上清液中IFN-γ浓度,RT-PCR方法检测细胞中IFN-γmRNA的表达水平,流式细胞术检测分泌IFN-γ的CD8＋T细胞所占的百分比。结果 10 ng/mL IL-21明显促进CD8＋T细胞分泌IFN-γ（P〈0.05）,IFN-γmRNA的表达明显升高,4 h为刺激CD8＋T细胞胞内IFN-γ合成的最佳时间。结论 IL-21对CD8＋T细胞分泌IFN-γ有促进作用。     

蜂毒肽对成熟树突细胞诱导T细胞分化的免疫调节的影响

通过蜂毒肽(Melittin,MT)干预脂多糖(Lipopolysaccharide,LPS)刺激树突细胞(Dendritic Cell,DC)与淋巴细胞共培养,以探究MT对Th1/Th2分化的影响。用MTS检测LPS、MT对DC与淋巴细胞共培养活性;流式细胞术检测MT对LPS刺激DC与淋巴细胞共培养中Th1、Th2比例;ELISA检测LPS刺激DC与淋巴细胞共培养中细胞上清液中IL-4、INF-γ浓度。结果显示MT、LPS在一定浓度下刺激DC与淋巴细胞共培养的活性;在成熟的DC诱导下,MT能上调Th1/Th2比例、Th1百分率(P0.05)及IFN-γ浓度(P0.01);对IL-4浓度、Th2百分率无影响(P0.05)。本研究说明MT通过上调Th1细胞比例,IFN-γ浓度,促进Th1/Th2细胞向Th1方向分化途径对T细胞起免疫调节作用。     

IL-32gamma induces the maturation of dendritic cells with Th1- and Th17-polarizing ability through enhanced IL-12 and IL-6 production

IL-32, a newly described multifunctional cytokine, has been associated with a variety of inflammatory diseases, including rheumatoid arthritis, vasculitis, and Crohn's disease. In this study, we investigated the immunomodulatory effects of IL-32γ on bone marrow-derived dendritic cell (DC)-driven Th responses and analyzed the underlying signaling events. IL-32γ-treated DCs exhibited upregulated expression of cell-surface molecules and proinflammatory cytokines associated with DC maturation and activation. In particular, IL-32γ treatment significantly increased production of IL-12 and IL-6 in DCs, which are known as Th1- and Th17-polarizing cytokines, respectively. This increased production was inhibited by the addition of specific inhibitors of the activities of phospholipase C (PLC), JNK, and NF-κB. IL-32γ treatment increased the phosphorylation of JNK and the degradation of both IκBα and IκBβ in DCs, as well as NF-κB binding activity to the κB site. The PLC inhibitor suppressed NF-κB DNA binding activity and JNK phosphorylation increased by IL-32γ treatment, thereby indicating that IL-32γ induced IL-12 and IL-6 production in DCs via a PLC/JNK/NF-κB signaling pathway. Importantly, IL-32γ-stimulated DCs significantly induced both Th1 and Th17 responses when cocultured with CD4(+) T cells. The addition of a neutralizing anti-IL-12 mAb abolished the secretion of IFN-γ in a dose-dependent manner; additionally, the blockage of IL-1β and IL-6, but not of IL-21 or IL-23p19, profoundly inhibited IL-32γ-induced IL-17 production. These results demonstrated that IL-32γ could effectively induce the maturation and activation of immature DCs, leading to enhanced Th1 and Th17 responses as the result of increased IL-12 and IL-6 production in DCs.     

Effect of short-term interferon-β treatment on cytokines in multiple sclerosis: significant modulation of IL-17 and IL-23

Therapeutic effect of interferon-β (IFN-β) treatment has been associated with modulation of the balance between Th1, Th17, Th2 and regulatory T (Treg) cells, whereas the impact of disease modifying drugs on Th9-immunity in multiple sclerosis (MS) has not been studied. To investigate the short-term effects of IFN-β treatment on cytokines in MS, we determined serum levels of IL-17, IL-23, IL-10, IL-4, IFN-γ, IL-9 and TGF-β in relapsing remitting MS patients before and 2 months after IFN-β treatment by ELISA. MS patients showed increased IL-17, IL-23 and IL-4 levels and decreased IL-9 levels as compared to healthy controls. IFN-β treatment only reduced IL-17 and IL-23 levels, whereas the levels of other cytokines remained unchanged. IFN-β treatment appears to exert its earliest therapeutic effect on Th17-immunity. The influence of IL-9 on MS pathogenesis needs to be further studied.     

Th17 cells induce colitis and promote Th1 cell responses through IL-17 induction of innate IL-12 and IL-23 production

Both Th1 and Th17 cells have been implicated in the pathogenesis of inflammatory bowel disease and experimental colitis. However, the complex relationship between Th1 and Th17 cells and their relative contributions to the pathogenesis of inflammatory bowel disease have not been completely analyzed. Although it has been recently shown that Th17 cells can convert into Th1 cells, the underlying in vivo mechanisms and the role of Th1 cells converted from Th17 cells in the pathogenesis of colitis are still largely unknown. In this study, we report that Th17 cells from CBir1 TCR transgenic mice, which are specific for an immunodominant microbiota Ag, are more potent than Th1 cells in the induction of colitis, as Th17 cells induced severe colitis, whereas Th1 cells induced mild colitis when transferred into TCRβxδ(-/-) mice. High levels of IL-12 and IL-23 and substantial numbers of IFN-γ(+) Th1 cells emerged in the colons of Th17 cell recipients. Administration of anti-IL-17 mAb abrogated Th17 cell-induced colitis development, blocked colonic IL-12 and IL-23 production, and inhibited IFN-γ(+) Th1 cell induction. IL-17 promoted dendritic cell production of IL-12 and IL-23. Furthermore, conditioned media from colonic tissues of colitic Th17 cell recipients induced IFN-γ production by Th17 cells, which was inhibited by blockade of IL-12 and IL-23. Collectively, these data indicate that Th17 cells convert to Th1 cells through IL-17 induction of mucosal innate IL-12 and IL-23 production.     

TGF-β is necessary for induction of IL-23R and Th17 differentiation by IL-6 and IL-23

TGF-β and IL-6 induce Th17 differentiation, and IL-23 is required for expansion and maintenance of Th17 cells. Recently, it was shown that IL-6 up-regulates IL-23R mRNA in naive CD4⁺ T cells and therefore IL-6 and IL-23 synergistically promote Th17 differentiation. However, the molecular mechanism whereby IL-6 and IL-23 induce Th17 differentiation and the relevance to TGF-β remain unknown. Here, we found that IL-6 up-regulated IL-23R mRNA expression, and IL-6 and IL-23 synergistically augmented its protein expression. The combination induced Th17 differentiation, and TGF-β1 further enhanced it. IL-6 augmented endogenous TGF-β1 mRNA expression, whereas the amount of TGF-β produced was not enough to induce Th17 differentiation by IL-6 alone. However, unexpectedly, the up-regulation of IL-23R and induction of Th17 differentiation by IL-6 and IL-23 were almost completely inhibited by anti-TGF-β. These results suggest that the induction of IL-23R and Th17 differentiation by IL-6 and IL-23 is mediated through endogenously produced TGF-β.     

双歧杆菌及其完整肽聚糖对脐血来源树突状细胞分泌IL-12的影响

目的探讨两歧双歧杆菌和不同剂量双歧杆菌的完整肽聚糖（WPG）对脐血来源树突状细胞（DC）分泌IL-12的影响。方法以双歧杆菌全菌（量）和不同剂量双歧杆菌WPG（1-8μg/ml）与脐血来源树突状细胞共培养,用ELISA的方法测定培养上清中IL-12的量。结果双歧杆菌和其WPG（1-6μg/ml）与树突状细胞共培养后,树突状细胞分泌的IL-12的量显著高于阴性对照组（P〈0.01）,当WPG量为1-5μg/ml时树突状细胞分泌的IL-12的量呈剂量依赖性,其中WPG量为5μg/ml时作用最为显著,WPG量为6μg/ml时分泌IL-12量减少,WPG量为8μg/ml时,分泌的IL-12量与阴性对照组差异无显著性（P〉0.05）。结论两歧双歧杆菌及其WPG能够刺激脐血来源的树突状细胞IL-12分泌;双歧杆菌WPG的免疫刺激作用呈一定的量效关系     

IL-23/Th17轴在变应性哮喘中的研究进展

变应性哮喘是一种由辅助性T细胞（T helper cell,Th cell）调节的慢性炎症性疾病。Th1/Th2的失衡一直被认为是变应性哮喘的发病机制,Th2细胞及其分泌的细胞因子白介素4（interleukin 4,IL-4）、IL-5以及IL-13在变应性哮喘特异性症状的发病中发挥重要作用。最近研究发现Th17细胞及其分泌的IL-17参与变应性哮喘的发展过程,IL-23在Th17细胞维持生存和功能成熟中发挥重要作用,并参与抗原诱导的气道炎症反应。该文对目前IL-23/Th17轴在变应性气道炎症反应中的研究进展作一综述。     

Increased expression and activation of IL-12-induced Stat4 signaling in the mucosa of ulcerative colitis patients

Inflammatory bowel diseases (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC), are chronic inflammatory diseases with unsolved pathogenesis. Imbalanced Th1/Th2 may play a role in the sustained inflammation of IBD. In China, CD is rare but the incidence of UC has been rising steadily in the last two decades. We investigated the expression of IL-12 (p40) and IFN-γ, and the activational state of Stat4 signaling in mucosal tissues at the site of disease from 30 active UC patients in comparison with 30 healthy controls. RT-PCR analyses revealed increased mRNA expression of IL-12 (p40) but not IFN-γ in UC patients. Western blot analyses discovered, for the first time, increased levels of constitutive Stat4 in the cytoplasm and phosphorylated Stat4 in the nucleus of mucosal cells from UC patients. We conclude that a heightened, perhaps persistent, activational state of IL-12/Stat4, and/or IL-23/Stat4 signaling may be present in active Chinese UC patients, and possibly involved in chronic inflammation in UC.     

Th 细胞因子IL-17、IFN-γ、IL-4 在狼疮性肾炎中的表达及意义

目的:探讨T辅助细胞(Th)相关细胞因子在狼疮性肾炎发病中的免疫机制作用。方法:64例系统性红斑狼疮患者和28例健康体检者作为对照,采用酶联免疫吸附测定法(ELISA法)检测所有受试者血清IL-17、IFN-γ、IL-4水平,并对其与SLEDAI、SDI、24小时尿蛋白量相关性进行研究。结果:狼疮性肾炎组血清IL-17水平显著高于狼疮无肾炎组和健康对照组(P<0.001),狼疮性肾炎组血清IFN-γ水平显著高于狼疮无肾炎组(P<0.05)和健康对照组(P<0.01),血清IL-4水平在狼疮性肾炎组、狼疮无肾炎组均显著高于健康对照组(P<0.01)。狼疮性肾炎组IFN-γ/IL-4比值显著高于狼疮无肾炎组(P<0.01)和健康对照组(P<0.05);狼疮无肾炎组IFN-γ/IL-4比值显著低于健康对照组(P<0.01)。SLE患者血清IFN-γ表达水平与SLEDAI积分呈正相关(r=0.402,P<0.05),血清IL-17、IL-4表达水平与SLEDAI、SDI、抗ds-DNA抗体、C3、24小时尿蛋白量均无相关性。结论:狼疮性肾炎患者外周血中IL-17、IFN-γ、IL-4等促炎细胞因子均有不同程度升高促起炎症发生及组织损伤,参与了狼疮性肾炎的免疫发病过程。