T_H17型细胞在移植排斥反应中的研究进展

在不同细胞因子和转录因子作用下,初始CD4~+T淋巴细胞分化为效应性T淋巴细胞(T_H1型、T_H2型、T_H17型细胞)和调节性T淋巴细胞(Treg细胞).初始CD4~+T淋巴细胞在转化生长因子β(TGF-β)和白细胞介素6(IL-6)的共同诱导下分化为T_Hl7型细胞,分泌IL-17和IL-6等细胞因子,主要参与炎症反应和自身免疫性疾病等的发生.近年来发现,T_H17型细胞与移植排斥反应有关,现就其与排斥反应的关系综述如下.     

血必净促进内毒素/脂多糖刺激调节性T淋巴细胞凋亡并介导辅助性T淋巴细胞漂移的作用

目的 了解血必净注射液促进LPS刺激CD4+CD25+调节性T淋巴细胞(Treg细胞)凋亡过程及介导辅助性T淋巴细胞(Th)漂移的调节作用.方法 免疫磁珠法分选获得大鼠脾脏CD4+CD25+Treg细胞,分为常规培养对照组、抗CD3/CD28组、抗CD3/CD28+LPS组、抗CD3/CD28+血必净组和抗CD3/CD28+LPS+血必净组,培养3 d后应用流式细胞术检测Treg细胞凋亡率及叉头翼状螺旋转录因子3(Foxp3)表达.将CD4+CD25+Treg细胞与CD4+CD25+T淋巴细胞1:1培养,伴刀豆球蛋白A刺激68 h,检测上清液中Th1分泌的γ干扰素(IFN-γ)、Th2分泌的IL-4、Th17分泌的IL-17水平.结果 抗CD3/CD28+LPS+血必净组Treg细胞凋亡率为(45.1±2.7)%,明显高于抗CD3/CD28+LPS组[(29.4±1.6)%,P＜0.01];2组Foxp3平均荧光强度分别为95±9、140±18,差异有统计学意义(P＜0.01).同时,抗CD3/CD28+LPS+血必净组IFN-γ分泌水平显著高于抗CD3/CD28+LPS组(P＜0.01),IL-4则呈相反变化(P＜0.05),抗CD3/CD28+LPS+血必净组IFN-γ/IL-4较对照组升高(P＜0.01);抗CD3/CD28+血必净组IL-17分泌水平较抗CD3/CD28组明显下降(P＜0.05).结论 CD4+CD25+Treg细胞活化介导了Th1向Th2功能性极化;血必净对LPS诱导的T淋巴细胞免疫功能有重要调节作用,可促进CD4+CD25+Treg细胞凋亡并介导Th2向Th1漂移,从而缓解细胞免疫抑制状态.     

OX40信号调节抑制小鼠CD4+CD25+调节性T淋巴细胞Foxp3的表达

目的 探讨共刺激信号OX40对体外诱导的小鼠CD4+ CD25+适应性调节性T淋巴细胞(iTreg)的Foxp3表达的影响.方法 制备C57BL/6小鼠淋巴细胞悬液,经免疫磁珠法分选,获得CD4+ CD25-静息T淋巴细胞,与抗CD3单克隆抗体、抗CD28单克隆抗体、转化生长因子β1、白细胞介素2共孵育,诱导产生Foxp3+ iTreg.在此基础上,于培养体系中加入OX40激动型抗体及其对照抗体,利用流式细胞仪分析研究OX40信号刺激对iTreg Foxp3表达的影响.结果 C57BL/6小鼠淋巴结中CD4+ CD25+天然调节性T淋巴细胞(Treg)比例为(5.0±0.4)％,体外诱导培养的CD4+CD25+ Treg比例为(71.8±13.4)％,其中Foxp3阳性表达占(74.9±1.9)％.OX40激动型抗体组CD4+ CD25+ Treg细胞比例为(80.0±1.6)％,其中Foxp3表达水平为(59.2±0.7)％;OX40激动型抗体对照抗体组CD4+ CD25+ Treg细胞比例为(86.0±1.4)％,其中Foxp3表达水平为(70.0±0.8)％,两组间差异有统计学意义(P＜0.05).结论 静息T淋巴细胞可以在体外诱导培养获得高纯度iTreg;OX40信号刺激可以显著抑制CD25+ iTreg细胞Foxp3的表达.     

西罗莫司对T淋巴细胞分化发育的影响

西罗莫司(SRL)最初作为低毒性的抗真菌药物,于1977年被发现具有免疫抑制作用,1989年开始将SRL用于预防器官移植后排斥反应.由于SRL在预防排斥反应中的疗效显著,不良反应较少,现在已广泛应用于各种器官移植.SRL不仅能抑制T淋巴细胞的分化发育,而且能够诱导效应性T淋巴细胞转化为调节性T淋巴细胞(Treg细胞),在体内外能明显扩增CD4+CD25+Treg细胞.     

不同免疫抑制剂对肝移植受者外周血中调节性T淋巴细胞比例的影响

目的 探讨西罗莫司(SRL)和钙调磷酸酶抑制剂(CNI)对肝移植受者外周血中CD4+CD25high T淋巴细胞水平的影响.方法 排除肝移植远期移植肝功能异常的受者,将移植肝功能长期(超过2年)稳定的受者47例纳入研究,其中免疫抑制方案使用SRL者15例(SRL组),使用CNI(均为他克莫司)者32例(CNI组).以同期38名健康成人志愿者作为正常对照.使用流式细胞仪检测各组受试者外周血中单个核细胞CD4、CD25及Foxp3的表达水平,比较各组间外周血中CD4+CD25high调节性T淋巴细胞(Treg细胞)的差异.结果 与正常对照组相比,CNI组外周血淋巴细胞中CD4+ CD25high T淋巴细胞的比例显著减少(P<0.05),SRL组CD4+ CD25high T淋巴细胞的比例显著升高(P<0.05).SRL组、正常对照组和CNI组受试者外周血中CD4+ CD25high Foxp3+ Treg 细胞占CD4+ T淋巴细胞的比例依次降低,分别为1.88%(1.56%～2.60%)、1.15%(0.57%～1.48%)和0.84%(0.46%～1.45%),3组间两两比较,差异均有统计学意义(P<0.01或P<0.05).CD4+ CD25 high T淋巴细胞表达Foxp3的阳性率超过95%,CD4+ CD25 low T淋巴细胞表达Foxp3的阳性率低于20%,CD4+ CD25-T淋巴细胞不表达Foxp3.结论 SRL可促进肝移植受者外周血中Treg细胞水平的升高,而CNI可降低Treg细胞的水平.     

转化生长因子-β1差异性诱导CD4+和CD8+T细胞表达转录因子FOXP3的研究

目的 研究CD4+T细胞和CD8+T细胞被转化生长因子-β1(TGF-β1)诱导表达转录因子FOXP3的差异性.方法 给予CD4+T细胞和CD8+T细胞抗T细胞受体(TCR)刺激的同时,加入TGF-β1培养4d后,通过胞内细胞因子染色,分析二者被诱导表达FOXP3的情况.同时通过CFSE标记实验检测被TGF-β1诱导表达FOXP3的淋巴细胞的增殖情况;通过annexin V流式染色检测TGF-β1对于T淋巴细胞凋亡的影响.结果 相对于CD8+T淋巴细胞,TGF-β1主要诱导CD4+T淋巴细胞阳性表达FOXP3[外周血单个核细胞(PBMC):29.66±3.624比7.430±0.643;癌旁组织浸润淋巴细胞(NIL):31.74±2.612比8.637±1.146].被TGF-β1诱导表达FOXP3的淋巴细胞增殖活跃(94.39±1.179),受到活化刺激后依旧能够有效分泌干扰素-γ(IFN-γ)效应细胞因子(39.58±1.611).TGF-β1能够有效降低CD8+T淋巴细胞活化后的细胞凋亡率(25.39±2.158比9.320±0.3219).结论 与肝癌患者FOXP3表达阳性的淋巴细胞＞95％(97.15±0.3807,n=10)集中在CD4+T淋巴细胞的结果一致,TGF-β1优先选择性诱导CD4+T细胞表达FOXP3.与天然Treg(调节性T细胞)低增殖活性和低细胞因子分泌活性不同,被TGF β1阳性诱导表达FOXP3的T细胞依旧具有活跃的细胞增殖和分泌IFN γ效应细胞因子的功能.CD8+T淋巴细胞相对于CD4+T淋巴细胞,更易发生活化后细胞凋亡,而TGF-β1能够有效降低这种细胞凋亡率.     

T_H17型细胞的分化及调节在自身免疫性疾病和排斥反应中的作用

初始T淋巴细胞被树突状细胞活化后,在体内不同的微环境下可分化成为功能及表型不同的效应性T淋巴细胞和调节性T淋巴细胞(Treg细胞).当有白细胞介素12(IL-12)存在时,初始T淋巴细胞分化为T_H 1型细胞,主要分泌IL-12、γ干扰素(IFN-γ)及特异性表达转录因子T-bet;有IL-4存在时,初始T淋巴细胞分化为T_H2型细胞,主要分泌IL-4、IL-10,特异性表达转录因子GATA-3;在转化生长因子β(TGF-β)单独存在时,初始T淋巴细胞分化为Treg细胞,特异性表达Foxp3.     

石斛多糖对胶原诱导关节炎小鼠免疫活性的影响

目的 探究石斛多糖（DSP）对胶原蛋白诱导关节炎（CIA）小鼠体内辅助T（Th17）/调节性T（Treg）细胞比例平衡及免疫炎症的影响。方法 构建CIA小鼠，通过关节HE染色和后肢右爪形态观察进行模型鉴定。分别给予CIA小鼠100、200和300 mg/kg DSP灌胃处理将小鼠分为对照组，模型组，低、中、高浓度DSP组。观察小鼠足爪病变情况，HE染色观察小鼠右肢滑膜组织病理改变，流式细胞术检测小鼠脾脏中Th17细胞，Treg细胞比例和CD4+CD28?PD-1lowOX40+T细胞比例的变化，免疫组化法观察小鼠右肢膝关节中IL-7的表达水平，ELISA检测不同时间点（0、14、28、42 d）小鼠血清IL-6、IL-10及小鼠右肢膝关节IL-6和IL-10表达水平。结果 与对照组小鼠相比，模型组小鼠足爪病变加重，滑膜组织病理变化明显，Th17细胞比例上升，Treg细胞和CD4+CD28?PD-1lowOX40+T细胞比例下降，IL-7阳性表达增加，Foxp3阳性表达下降，血清和膝关节中IL-6水平增加，IL-10水平下降（均P<0.0.5）。与模型组相比，低、中、高浓度DSP组小鼠足爪病变减轻，滑膜组织病理变化改善，Th17细胞比例下降，Treg细胞和CD4+CD28?PD-1lowOX40+T细胞比例上升，IL-7阳性表达减少，Foxp3阳性表达增多，血清和膝关节中IL-6水平下降，IL-10水平增加（均P<0.0.5），且高浓度DSP作用效果最好。结论 DSP对CIA小鼠体内Th17/Treg细胞比例平衡具有调节作用，能抑制炎症反应，减轻病理改变。     

西罗莫司预处理减轻大鼠肝脏缺血再灌注损伤的作用及其机制

目的 探讨西罗莫司(SRL)预处理减轻大鼠肝脏缺血再灌注损伤的作用及其机制.方法 将SD大鼠随机分为4组,每组12只.假手术对照组:开腹后仅用生理盐水纱布覆盖切口60min,关腹;假手术SRL组:手术方式同假手术对照组;实验对照组:开腹后夹闭肝门静脉左支、肝动脉左支及左肝管,60 min后开放血管;实验SRL组:手术方式同实验对照组.两SRL组大鼠术前2周开始给予SRL 2 mg·kg~(-1)·d~(-1)灌胃,术前6 h加用1次.而两对照组同期仅给予等体积无菌生理盐水灌胃.术后24 h分别采集各组大鼠的血液和肝组织,检测血清丙氨酸转氨酶(ALT)和天冬氨酸转氨酶(AST)水平,光镜下观察肝组织的病理学变化,采用流式细胞术检测肝组织中CD4~+ CD25~+ T淋巴细胞占单个核细胞的比例.采用实时聚合酶链反应检测肝组织中Foxp3 mRNA的表达,采用酶联免疫吸附试验检测血清巾转化生长因子-β(TGF-β)和白细胞介素10(IL-10)的含量.结果 假手术对照组和假手术SRL组血清ALT和AST均处于较低水平,而实验SRL组和实验对照组明显升高,且实验SRL组低于实验对照组(P<0.05).假手术对照组和假手术SRL组肝组织结构正常,实验对照组可见明显的片状坏死,实验SRL组肝小叶结构基本完整,未见明显细胞坏死.假手术对照组、假手术SRL组、实验对照组和实验SRL组CD4~+ CD25~+ T淋巴细胞占单个核细胞的比例分别为(6.12±1.87)%、(22.36±6.75)%、(4.53±1.02)%和(13.29±3.16)%.假手术SRL组Foxp3 mRNA的相对表达量以及血清中TGF-β和IL-10的含量明显高于假手术对照组(P<0.05),实验SRL组明显高于实验对照组(P<0.05).结论 SRL可以减轻大鼠肝脏缺血再灌注损伤,其机制可能与SRL诱导体内CD4~+ CD25~+ Foxp3~+ 调节性T淋巴细胞的分化以及增加TGF-β和IL-10的分泌抑制炎症反应有关.     

HIV感染者CD4~+T细胞亚群变化的研究进展

人免疫缺陷病毒(HIV)侵入人体后主要侵犯CD4+T淋巴细胞,CD4+T细胞根据诱导因子和分泌因子的不同,主要分为辅助性T(Th)1细胞、Th2细胞、Th17细胞、调节性T(Treg)细胞等4类。本文针对HIV对CD4+T亚群变化的最新进展进行综述。     

Immunoregulatory Function of IL-27 and TGF-β1 in Cardiac Allograft Transplantation

BACKGROUND: Deciphering the mechanisms of tolerance represents a crucial aim of research in transplantation. We previously identified by DNA chip interleukin (IL)-27 p28 and transforming growth factor (TGF)-β1 as overexpressed in a model of rat cardiac allograft tolerance mediated by regulatory CD4CD25 T cells. The role of these two molecules on the control of the inflammatory response remains controversial. However, both are involved in the regulation of the T helper 17/Treg axis, suggesting their involvement in tolerance. METHODS: We analyzed regulation of IL-27 and TGF-β1 expression in allograft response and their role in tolerance by using blocking anti-TGF-β antibody and by generating an adeno-associated virus encoding IL-27. RESULTS: Here, we confirmed the overexpression of IL-27 and TGF-β1 in tolerated cardiac allografts in two different rodent models. We observed that their expression correlates with inhibition of T helper 17 differentiation and with expansion of regulatory CD4CD25 T cells. We showed in a rat model that anti-TGF-β treatment abrogates infectious tolerance mediated by the transfer of regulatory CD4CD25 T cells. Moreover, overexpression of IL-27 by adeno-associated virus administration in combination with a short-term immunosuppression allows prolongation of cardiac allograft survival and one tolerant recipient. We found that IL-27 overexpression did not induce Foxp3CD4CD25 T-cell expansion but rather IL-10-expressing CD4 T cells in the tolerant recipient. CONCLUSIONS: Taken together, these data suggest that both TGF-β1 and IL-27 play a role in the mechanisms of tolerance. However, in contrast to TGF-β1, IL-27 seems not to be involved in regulatory CD4CD25 T-cell expansion but rather in their mode of action.     

他克莫司对小鼠T_H17型细胞分化增殖的影响及其机制

目的 观察在T_H17型细胞极化环境下,他克莫司(Tac)对小鼠T_H17型细胞分化增殖的影响及机制.方法 将小鼠脾脏初始CD4+CD25+T淋巴细胞使用抗CD3抗体及抗CD28抗体进行活化,使用转化生长因子β_1,(TGF-β_1,)和白细胞介素6(IL-6)等细胞因子进行诱导,促使其向T_H17型细胞方向分化.将体外培养的T_H 17型细胞分为4组,分别用不同剂量的Tac进行干预,即:Tac0 ng/ml组(对照组)、Tac 0.1 ng/ml组、Tac 1 ng/ml组和Tae 10 ng/ml组.应用流式细胞术检测各组T_H 17型细胞亚群纯度,使用逆转录聚合酶链反应(RT.PCR)检测各组IL-17 mRNA相对表达量.结果 Tac可抑制小鼠T_H17型细胞亚群的分化增殖,降低IL-17 mRNA的表达量,且呈剂量依赖性,各组间比较,差异均有统计学意义(P<0.05).结论 Tac通过抑制T_H17型细胞胞浆内的钙调磷酸酶,抑制T_H17型细胞亚群的分化增殖,抑制IL-17 mRNA产生并减轻炎症进程,从而抑制排斥反应.     

阿尔茨海默病患者外周血调节性T细胞稳定性的分析

目的:探讨阿尔茨海默病(AD)患者外周血调节性T细胞(Tregs)稳定性的特点.方法:分选AD患者外周血Tregs,通过流式细胞仪检测特征性标志物叉头转录因子(Foxp)-3和表面分子细胞毒性T淋巴细胞相关抗原(CTLA)-4及膜相关转化性生长因子(TGF)-β的表达水平,通过ELISA检测特征性细胞因子TGF-β和白细胞介素(IL)-10分泌水平,并进一步通过Transwell培养板培养方法,分析AD患者Tregs发挥免疫抑制功能的特点.结果:与对照组比较,AD组患者细微精神状态语言检查(MMSE)评分明显降低(P＜0.01);AD组外周血Tregs数目、特征性标志物Foxp-3及细胞表面标志物CTLA-4和膜相关TGF-β表达水平均均低于对照组,差异有统计学意义(P＜ 0.05或P＜0.01);特征性细胞因子TGF-β和IL-10分泌水平均无明显变化(P＞0.05);经Transwell培养板培养24h,能够降低AD组Tregs对CD4+ CD25-T细胞增殖功能的抑制作用,差异有统计学意义(P＜0.05).结论:AD能够降低患者外周血Tregs稳定性,其机制主要是降低其以细胞接触途径介导的免疫调节作用.     

NOD小鼠未成熟树突状细胞体外诱导淋巴细胞生成并扩增调节性T细胞的研究

目的 探索非肥胖糖尿病(non-obese diabetic,NOD)小鼠未成熟树突状细胞(immature dendritic cell,imDC)诱导并扩增调节性T细胞(regulatory T cells,Treg)的优化方案.方法 分离、培养NOD小鼠骨髓来源的imDC和淋巴细胞,并进行混合淋巴细胞培养,分别加...     

Küpffer cells promote acute rejection via induction of Th17 differentiation in rat liver allografts

Background

Th17, a newly identified CD4+ T-cell subset, has been implicated in transplant rejection. Differentiation of Th17 cells is associated with transforming growth factor-β (TGF-β) and interleukin-6 (IL-6), which are the main products of Küpffer cells.

Objective

To determine whether Küpffer cells promote acute liver allograft rejection by inducing Th17 cell differentiation.

Methods

A rat model of allogeneic liver transplantation using Dark Agouti (DA) to Brown Norway (BN) rats was established with or without gadolinium chloride (GdCl₃) pretreatment. Isogeneic liver transplantation (BN to BN) was performed as a control. Concentrations of cytokines secreted by Küpffer cells or Th17-related cytokines detected in the liver and peripheral blood were analyzed using immunohistochemistry assays, flow cytometry, and enzyme-linked immunosorbent assays. Survival differences were compared between treatment groups. In vitro, Küpffer cells from liver grafts were isolated and co-cultured with naïve CD4 T cells.

Results

Both Küpffer cells and Th17 cells infiltrated liver allografts, accompanied by an increase in concentrations of IL-6 and TGF-β. Pretreatment with GdCl₃ attenuated intragraft infiltration of Küpffer cells and Th17 cells, and decreased IL-6 and TGF-β concentrations. Liver function improved after pretreatment, and mean (SD) survival time was prolonged, compared with the control group (16.33 [0.96] days vs 11.50 [0.99] days, respectively; P < .01). In vitro, Küpffer cells from livers with allografts secreted significantly higher concentrations of IL-6 and TGF-β and induced Th17 differentiation more effectively compared with livers with isografts (30.8% vs 8.1%, respectively).

Conclusion

Küpffer cells have the potential to induce Th17 cells by secreting IL-6 and TGF-β, and as a result, promote acute liver allograft rejection.     

Evidence that genistein, a protein-tyrosine kinase inhibitor, inhibits CD28 monoclonal-antibody-stimulated human T cell proliferation.

In the present investigation, we compared the immunosuppressive effects of genistein and CsA on anti-CD28 stimulated human T cell proliferation, IL-2 production, and IL-2R expression. Genistein, an isoflavanoid compound, is a specific protein tyrosine kinase inhibitor and inhibited the PMA plus anti-CD28 stimulated T cell proliferation. In contrast, proliferation of T cells stimulated with PMA plus anti-CD28 is resistant to the inhibitory effects of CsA. Similar results were obtained with IL-2 synthesis and IL-2R expression. PHA plus anti-CD28 or PMA plus anti-CD28-induced IL-2 synthesis was inhibited by genistein, and CsA, though it inhibited the PHA plus PMA-stimulated IL-2 synthesis, failed to have any effect on PMA plus anti-CD28-induced IL-2 synthesis. Genistein at the concentration that inhibited T cell proliferation and IL-2 synthesis also showed significant inhibitory effects on PMA plus anti-CD28 stimulated IL-2R expression while CsA had no effect on IL-2R from these cultures. Our data suggest that genistein is a powerful immunosuppressive agent, with no toxic effects on T cells, and has the potential for use in the prophylaxis and treatment of allograft rejection. Since genistein blocks the CsA-resistant pathway of T cell proliferation, the combined usage of these two agents may provide better immunosuppressive effect and a lesser degree of CsA-induced nephrotoxicity.     

LAP+CD4+T细胞的研究进展

【摘要】 调节性T细胞是机体维持自身稳定的重要组成部分,对免疫反应具有抑制效应,在阻止自身免疫反应和维持机体免疫平衡等方面均具有重要作用。现在已发现不同的具有调节功能的T细胞亚群,如CD4+CD25+T细胞、Th3细胞和Tr1细胞等,这些T细胞多通过产生具有抑制功能的细胞因子,如IL-10和TGF-β等发挥免疫抑制效应。LAP+CD4+T细胞是近年发现的一种新的调节性T细胞亚群,在体内外实验中被证实具有免疫抑制作用,它通常以依赖TGF-β和IL-10等细胞因子的方式来发挥作用,且在一些疾病的动物模型中它影响着疾病的发生、发展,如自身免疫性疾病,炎症型疾病等。较少的研究提示LAP+CD4+T细胞在癌症病人中具有促进肿瘤进展的作用。     

Soluble CD83 inhibits acute rejection by up regulating TGF-β and IDO secretion in rat liver transplantation

BackgroundAllogeneic transplantation immune tolerance is currently a hot research issue and soluble CD83(sCD83) is a novel immunomodulator with great potential in inducing transplantation tolerance.ObjectiveTo investigate the mechanism of the immune tolerance effect of sCD83 on rat liver transplantation.MethodA rat liver transplantation model was established to study the effects of sCD83 on the expression levels of IL-2, IL-10, and TGF-β in peripheral blood and the mRNA expressions of foxp3, TGF-β, and Indoleamine 2,3-dioxygenase (IDO) in liver. The expression changes of costimulatory molecules CD80, CD86, and MHC-II on the surface of DC cells and the expressions of IDO ⁺ DC cell, TGF-β ⁺ CD4 ⁺ T cell, and CD4 ⁺ CD25 ⁺ Foxp3 ⁺ T cell were analyzed and compared.ResultssCD83 alleviated the rejection activity index (RAI) of rat liver transplantation in the early stage, increased the expressions of TGF-β, IL-10 in peripheral blood and the mRNAs of IDO, TGF-β and foxp3 in the transplanted liver, and down-regulated the expressions of MHC-II, CD86, and CD80 in DC cells, resulting in significant increased numbers of tolerogenic TGF-β ⁺ CD4 ⁺ T cells, Treg cells, and IDO ⁺ DC cells with low expression.ConclusionsCD83 inhibited acute rejection after liver transplantation in an allogeneic rat, and the mechanism was associated with the effect that sCD83 increased the expression of TGF-β, activated IDO immunosuppressive pathway, and increased tolerogenic DC cells and Treg cells.     

The effect of the CD28 activation pathway on the immunosuppressive action of cyclosporine.

The effect of the CD28 activation pathway on the immunosuppressive action of CsA was assessed. Human peripheral blood lymphocytes were stimulated with anti-CD3, bryostatin (Bryo) a novel activator of protein kinase C (PKC) and anti-CD28 singly or in combination, to which graded doses of CsA were added to determine relative sensitivity. Proliferation, IL-2 production, and IL-2 receptor expression were assessed and the IC50 determined. Lymphocytes stimulated with Bryo exhibited a marginal proliferative response but expressed the IL-2 receptor despite the presence of CsA. Addition of anti-CD3 or anti-CD28 to Bryo-stimulated lymphocytes promoted a vigorous proliferative response. CsA effectively inhibited the proliferative response and IL-2 production induced with anti-CD3 and Bryo but did not inhibit the response of cells stimulated with anti-CD28 and Bryo. However, II-2 receptor expression in both sets of cultures were comparable due to the induction of IL-2 receptor by Bryo and was not inhibited by CsA. Costimulation of lymphocytes with anti-CD3 plus anti-CD28 resulted in a 2-3-fold enhancement of proliferation compared with lymphocytes stimulated with anti-CD3 alone. Addition of CsA to lymphocytes stimulated with anti-CD3 resulted in the dose-dependent suppression of the proliferative response and IL-2 production (IC50 = 10-25 nM) but less so for IL-2 receptor expression (IC50 = 100-150 nM). In comparison, the proliferative response and IL-2 production elicited by anti-CD3 + anti-CD28 was more resistant to the effects of CsA (IC50 = 100-200 nM). However, IL-2 receptor expression exhibited comparable sensitivity to CsA (IC50 = 100-200 nM) in the presence of anti-CD28. Combination drug:drug studies revealed that CsA and the protein kinase C inhibitor H-7 were additive for both anti-CD3 and anti-CD3 plus anti-CD28 response. On the other hand, the cGMP-dependent protein kinase inhibitor H-8 was synergistic with CsA in inhibiting the response of lymphocytes to anti-CD3 plus anti-CD28 but only additive for responses to anti-CD3. Taken together, these data suggest that CsA inhibits T cell activation at two distinct levels, leading to inhibition of IL-2 production and inhibition of IL-2 receptor expression. Activation of the CD28 pathway partially overcomes the inhibitory activity of CsA on IL-2 production and may be mediated by indirect activation of a cGMP-dependent protein kinase.(ABSTRACT TRUNCATED AT 400 WORDS)     

Arsenic trioxide is a novel agent for combination therapy to prolong heart allograft survival in allo-primed T cells transferred mice

Alloreactive memory T cells are major barriers to transplantation acceptance due to their capacity to accelerate rejection. Here, we investigated the effects of combined treatment with arsenic trioxide (As(2)O(3)) and blocking monoclonal antibodies (mAb) against CD154 and LFA-1 (anti-CD154/LFA-1) on graft survival as well as changes in pathology and immunological responses in mice with adoptively transferred allo-primed T cells. The mean survival time (MST) for the cardiac allografts in recipient mice receiving the combination of As(2)O(3) and anti-CD154/LFA-1 was significantly longer (>113.7days) compared to those receiving anti-CD154/LFA-1 (23.2days), As(2)O(3) (12.5days) alone or no treatment (5.5days). This combined strategy distinctly inhibited lymphocyte infiltration in grafts, proliferation of splenic T cells and the generation of memory T cells in spleens. Moreover, the combined treatment caused the significant down-regulation of IL-2 and IFN-γ accompanied by increased expression of TGF-β and regulatory T cells (Tregs) in spleens, which led to long-term cardiac allograft survival in recipient mice. These results highlight the potential application of As(2)O(3) and its contribution in combination therapy with antibody blockade to delay rejection by memory T cells.