腹主动脉瘤患者外周血CD4+CD28-调节性T细胞的变化及意义

目的：探讨CD4+ CD28-T细胞在不同阶段腹主动脉瘤患者血清中的数量变化及临床意义。
方法:腹主动脉瘤患者20例,其中小腹主动脉瘤10例,大腹主动脉瘤10例,正常健康对照组10例,采用流式细胞术测定3组受检者外周血CD4+,CD8+,CD4+ CD28-T亚群数量及占淋巴细胞的比例,运用酶联吸附实验（ELISA）方法检测上述各组人群血浆中Th1细胞因子（IFN-γ,TNF-α）表达水平。
结果:腹主动脉瘤组CD4+ CD28-T细胞表达水平显著升高,其中CD4+ CD28-T细胞在小腹主动脉瘤组升高最为显著（P＜0.05）。Th1细胞因子（IFN-γ,TNF-α）表达水平亦呈显著升高。
结论:不同发展阶段腹主动脉瘤患者血清中CD4+ CD28-T细胞表达升高,AAA患者外周血T淋巴细胞处于活化状态,在AAA发生发展的过程中,机体存在全身免疫反应的激活,外周血中T细胞亚组发生改变,导致炎症及自身免疫反应性扩展。

     

CD4+CD25+T细胞联合CD154单抗抑制大鼠肝移植急性排斥反应的研究

目的探讨CD4 CD25 T细胞联合应用CD154单抗在抑制大鼠肝移植急性排斥反应中的作用。方法分离Lewis大鼠脾脏CD4 _CD25 T细胞后与DA大鼠脾细胞单向混合淋巴细胞反应行体外激活。用"二袖套法"行DA到Lewis的原位肝移植48例。A组为对照组;B、C组单独术前回输体外激活的CD4 CD25 T细胞或术后腹腔注射抗CD154单抗;D组联合应用CD4 CD25 T细胞和CD154单抗。每组大鼠12对。术后7 d各组处死6只受体,检测移植肝内T细胞亚群和细胞因子水平。余大鼠观察生存情况,死亡大鼠观察移植肝病理变化。结果D组受体生存期(52.00±10.64)d明显长于B、C组(P<0.01);移植肝内CD4 CD25 T细胞比例(16.43±4.28)%明显高于B、C组(P<0.05、P<0.01),而淋巴细胞浸润数量[(3.47±1.21)%×106]和(CD8 T细胞百分比(14.19±3.02)%明显低于B、C组(P<0.05、P<0.01);移植肝内白细胞介素- 2(IL-2)水平(6.44±1.83)ng/L低于B、C组(P<0.05),IL-10(43.72±7.55)ng/L和转化生长因子-β1(TGF-β1)(270.06±46.91)ng/L明显高于B、C组(P<0.05、P<0.01)。结论联合应用CD154单抗能明显增强CD4 CD25 调节性T细胞对大鼠肝移植急性排斥反应的抑制作用。     

手术联合树突状细胞治疗肾透明细胞癌的效果

目的 观察手术联合树突状细胞治疗肾透明细胞癌的临床效果,为肾癌治疗提供依据.方法 入组患者分为A、B两组,A组(n=33)为术后接受细胞治疗组,B组(n=37)为单纯手术治疗组.所有入组患者分别于手术治疗前8周及治疗后8周采外周血行流式细胞术检测T淋巴细胞亚群(CD3+、CD4+、CD8+、CD4 +/CD8+比值),了解治疗前后机体免疫水平.结果 Ⅰ、Ⅱ期患者A、B组治疗前后T淋巴细胞亚群差异无统计学意义(P＞0.05);Ⅲ期患者A组治疗前后T淋巴细胞亚群差异有统计学意义(P＜0.05),B组治疗前后T淋巴细胞亚群差异无统计学意义(P＞0.05).A组治疗前后T淋巴细胞亚群差异有统计学意义(P＜0.05),B组差异无统计学意义.结论 手术联合树突状细胞治疗可明显提高Ⅲ期肾透明细胞癌患者术后免疫功能,对肾癌治疗具有积极意义.     

CD8+CD28-T细胞在大鼠肾脏缺血预处理保护效应中的作用

Burne-Taney等[1]研究表明造成肾脏缺血组织损伤的T淋巴细胞功能减退起到了类似缺血预处理样作用,提出T淋巴细胞中可能存在对肾脏缺血再灌注损伤(IRI)起保护作用的亚群,其中造成缺血组织损伤的免疫细胞主要是表达CD28+的CD4+T细胞、NKT细胞等,而起保护作用的T细胞亚群尚不清楚.我们在建立大鼠肾脏热IRI模型的基础上通过对外周血免疫抑制性细胞CD8+CD28-T的检测探讨其在肾脏缺血预处理(IP)中的作用.     

全身照射预处理供体对异基因大鼠肝移植的作用及对受体内CD4~+CD25~+调节性T细胞的影响

目的 探讨全身照射(TBI)预处理诱导大鼠肝移植术后急性排斥反应的发生机制,及CD4~+ CD25~+调节性T细胞的变化在诱导免疫耐受中的作用.方法 以雄性Lewis、DA大鼠为供、受体,随机分为正常对照组、同种肝移植组、自发免疫耐受组、急性排斥反应组.观察各组受体的生存时间及生存率,检测受体术后外周血中ALT、TB含量、Foxp3~+ CD4~+ CD25~+ 调节性T细胞和T细胞亚群上GITR的表达,检测受体术后第14天移植肝的病理变化和受体脾脏CTL杀伤活性.结果 自发免疫耐受组,术后经历短暂排斥反应最终获得免疫耐受并长期存活.急性排斥反应组,在术后第17～21天死亡,与其他组相比,外周血血清中ALT、TB含量明显升高,而Foxp3~+ CD4~+ CD25~+调节性T细胞比例明显降低.TBI预处理大鼠供肝致受体外周血中CD3~+ CD4~+ T细胞上GITR表达降低,CD3~+CD8~+T细胞上GITR表达增加,提高CTL的杀伤活性.结论 通过TBI清除供体大鼠肝移植物中携带的旁路淋巴细胞,致受体外周血中Foxp3~+ CD4~+ CD25~+调节性T细胞表达降低,而使CD3~+ CD8~+T细胞上GITR表达增加,共同诱导大鼠肝移植术后急性排斥反应发生和耐受障碍.     

大鼠肝癌组织中巨噬细胞与细胞毒性T细胞关系

目的 观察大鼠肝癌组织中巨噬细胞对细胞毒性T细胞浸润和凋亡的影响.方法 构建Wistar大鼠Walker-256肝癌模型,将大鼠分为3组:A组(腹腔注射氯磷酸盐脂质体)、B组(腹腔注射PBS脂质体)、C组(腹腔注射生理盐水).12 d后摘取肿瘤组织,测量肿瘤大小、免疫组织化学检测肿瘤组织中浸润的CD68、CD163、CD8及GranzymeB阳性细胞个数;CD8与TUNEL双标方法检测肿瘤组织中T细胞的凋亡.结果 (1)A、B、C3组肿瘤体积比较,差异均无统计学意义(P＞0.05).(2)A、B、C3组中,CD68阳性细胞个数在A、B组之间比较;CD163阳性细胞个数在A、B组之间比较;CD8阳性细胞个数在A、B组,A、C组之间比较;Granzyme阳性细胞个数在A、C组之间比较,差异有统计学意义(P＜0.05).其余各组间比较,差异无统计学意义(P＞0.05).其中,CD68、CD163阳性细胞个数在A组明显少于B、C组;而CD8、GranzymeB阳性细胞个数在A组明显多于B、C组.(3)相关分析显示,针对全部肿瘤组织,CD68与CD8阳性细胞之间,CD163与GranzymeB阳性细胞之间呈负相关,差异有统计学意义(P＜0.05).CD68与GranzymeB阳性细胞之间,CD8与CD163阳性细胞之间无明显相关.(4)A、B、C3组中T细胞(CD8)凋亡率在A、C组之间比较,差异有统计学意义(P＜0.05);其余各组间比较,差异无统计学意义(P＞0.05).A组T细胞凋亡率明显低于B、C组.结论 氯磷酸盐脂质体可有效清除大鼠肝癌组织中巨噬细胞.巨噬细胞清除后,细胞毒性T细胞的浸润增多、凋亡减少.肿瘤巨噬细胞通过促进细胞毒性T细胞凋亡而抑制其对肿瘤细胞的杀伤活性,在肿瘤发展中起重要作用.     

大鼠小肠移植排斥反应时外周血T淋巴细胞上CD2分子的表达

目的探讨大鼠小肠移植急性排斥反应时外周血T淋巴细胞上CD2分子的表达。方法实验分3组进行,A组为假手术对照组(n=18),给予普通饲料喂养;B组(n=18)行SD大鼠到SD大鼠的同系小肠移植,术后常规补液,给予抗生素;C组(n=18)行SD大鼠到Wistar大鼠的小肠移植,术后处理同B组。各组于术后3、5、7d取肝素抗凝血,行流式细胞术检测,同时取移植肠组织,进行病理学检查。结果术后C组动物的存活时间为(7.0±2.1)d,B组为(33.3±2.3)d,A组>90d,C组与A、B组相比,差异有统计学意义(P<0.05);术后3、5、7d的外周血CD2阳性T淋巴细胞,A组分别为70.2%、69.8%和70.3%;B组为71.3%、69.7%和70.2%;C组为95.6%、88.1%和81.2%,C组各时点的CD2阳性细胞均高于A、B组相应时点(P<0.05);C组移植肠可见排斥反应的病理改变,且随术后时间的延长逐渐加重。结论小肠移植术后发生急性排斥反应时外周血CD2阳性T淋巴细胞表达率升高;术后早期CD2表达率的突然增高,提示可能发生急性排斥反应。     

利用RNA干扰诱导产生的CD8+CD28-抑制性T淋巴细胞的免疫学特性

目的 探讨通过RNA干扰技术诱导产生的CD8+ CD28-抑制性T淋巴细胞(Ts细胞)的免疫学特性.方法 取SD大鼠骨髓,培养分离树突状细胞(DC),设计、合成主要组织相容性复合物(MHC)Ⅰ类小片段干扰RNA(siRNA),以MHC Ⅰ siRNA转染DC.先以Wistar大鼠肠系膜淋巴组织液刺激转染MHCI siRNA的DC,然后将DC与从SD大鼠脾脏分离得到的CD8+T淋巴细胞共同培养,通过磁珠法分离出Ts细胞.分别在由SD大鼠脾脏淋巴细胞(反应细胞)和Wistar大鼠肠系膜淋巴组织细胞(刺激细胞)组成的混合淋巴细胞培养体系中加入数量不等的Ts细胞,检测反应细胞增殖情况;分别以Wistar大鼠肠系膜淋巴组织细胞和卵白蛋白(OVA)刺激SD大鼠脾脏淋巴细胞,然后再按不同比例加入Ts细胞,检测各组脾脏淋巴细胞的增殖情况;在由SD大鼠脾脏淋巴细胞、Wistar大鼠肠系膜淋巴组织液和Ts细胞组成的混合淋巴细胞培养体系中加入可溶性重组白细胞介素2(rrIL-2),观察IL-2对Ts细胞功能的影响;采用实时定量聚合酶链反应(PCR)测定Ts细胞中转化生长因子β(TGF-β和γ干扰素(IFN-γ)mRNA的表达,流式细胞仪和实时PCR检测Ts细胞上CD25分子的表达.结果 Ts细胞对SD大鼠脾脏淋巴细胞和Wistar大鼠肠系膜淋巴组织细胞之问的混合淋巴细胞反应(MLR)具有抑制作用,但对于SD大鼠脾脏淋巴细胞和OVA之间的MLR则无抑制作用.在SD大鼠脾脏淋巴细胞、Wistar大鼠肠系膜淋巴组织液和Ts细胞组成的混合淋巴细胞培养体系中加入rrIL-2后,SD大鼠脾脏细胞的增殖并无明显增加(P＞0.05).与CD8+CD28+T淋巴细胞和CD8+ T淋巴细胞比较,Ts细胞的TGF-β和IFN-γ mRNA的表达量明显升高(P＜0.01,P＜0.05),而CD25的表达量明显降低(P＜0.05).结论 采用经MHC I siRNA干扰的DC能够诱导CD8+T淋巴细胞产生CD8+ CD28-Ts细胞;Ts细胞在体外具有免疫抑制特性,其免疫抑制作用不被外源性IL-2所逆转,且其免疫调节作用具有抗原特异性.     

T细胞共刺激分子及其亚群在胃癌和大肠癌组织中表达的意义

目的 探讨T细胞共刺激分子及其亚群在胃癌、大肠癌发生及预后中的作用.方法 应用流式细胞术检测38例胃癌、42例大肠癌患者和21例健康人(对照组)外周血T细胞亚群及其共刺激分子CD28的表达.结果 T细胞共刺激分子CD28(CD28 CD3 )表达 胃癌组为(25.80±10.56)%,大肠癌组为(28.95±9.29)%,均明显高于对照组的(0.82±0.98)%,P＜0.01; 总T细胞(CD3 )表达 胃癌组为(53.61±13.84)%,大肠癌组为(55.96±10.68)%,均明显低于对照组的(72.07±7.83)%,P＜0.01; CD4 T细胞(CD4 CD3 )表达 胃癌组为(29.84±9.71)%,大肠癌组为(33.75±9.04)%,也均明显低于对照组的(38.79±5.08)%,P＜0.01, P＜0.05; 细胞毒T细胞(CTL,CD8 CD28 CD3 )表达 胃癌组为(1.57±1.99)%,大肠癌组为(1.93±2.61)%,均明显高于对照组的(0.02±0.04)%,P＜0.01; 胃癌组CD8 抑制性T细胞(CD8 CD28-CD3 )和CD4/CD8比值明显低于对照组[(16.06±6.94)% vs (20.56±6.54)%,P＜0.05; (1.10±0.51)% vs (1.36±0.31)%,P＜0.05]; 大肠癌组调节性T细胞(CD4 CD25 CD3 )明显高于对照组[(19.74±6.89)% vs (13.72±3.08)%, P＜0.01].胃癌组和大肠癌组患者手术前和手术后1周外周血T细胞亚群(除外胃癌组的CD3 细胞和CD28 CD3-细胞)的差异无统计学意义(P＞0.05). 结论胃癌和大肠癌患者T细胞数量明显减少,T细胞共刺激分子CD28表达增高.胃癌患者CD4 T细胞显著减少; 大肠癌患者调节性T细胞显著增加.     

人CD8+CD28-抑制性T淋巴细胞的体外扩增及其抗原特异性调节作用

目的 探讨在体外大量扩增CD8+CD28-抑制性T淋巴细胞(Ts细胞)的方法,并检验其免疫调节作用.方法 分离健康志愿者全血中CD8+T淋巴细胞,在含不同细胞因子和异体抗原提呈细胞(APC)的培养条件下进行体外扩增.应用流式细胞术对扩增过程中CD28 -细胞亚群的比例进行监测.分为3组进行混合淋巴细胞培养,反应细胞均为CD4+T淋巴细胞:B-APc组以来源于原致敏供者的APC作为刺激细胞,I-APC组以HLA-A、B、DR全错配的无关供者的APC作为刺激细胞,Dynabeads组以包被有抗CD3和CD28单克隆抗体的免疫微球Dynabeads作为刺激细胞;扩增后的Ts细胞作为第三方调节细胞加入混合淋巴细胞培养中,测定其对CD4+T淋巴细胞增殖的抑制作用.结果 在含白细胞介素2(IL-2)+ IL-7+ IL-15的培养条件下,CD8+T淋巴细胞培养后CD8+CD28-T淋巴细胞亚群的比例最高(P＜0.05).B-APC组不加入Ts细胞时,增殖的CD4+T淋巴细胞比例为66.7％,当加入的Ts细胞与反应细胞的比例分别为0.5∶1、0.1∶1和0.02∶1时,增殖的CD4+T淋巴细胞比例分别为16.5％、34.1％和62.6％.Ts细胞对CD4+T淋巴细胞的增殖有明显抑制作用,而在I-APC组和Dynabeads组中,此抑制作用不明显.结论 应用含IL-2+ IL-7+ IL-15的培养条件联合异体APC刺激可以在体外大量扩增Ts细胞,扩增所得Ts细胞在体外对供者CD4+T淋巴细胞的增殖有明显抗原特异性抑制作用.     

Pathways of helper CD4 T cell allorecognition in generating alloantibody and CD8 T cell alloimmunity

BACKGROUND: The relative contributions of the "direct" and "indirect" pathways of CD4 T cell allorecognition in providing help for generating effective humoral and CD8 T cell alloimmunity remain unclear. Here, the generation of alloantibody and cytotoxic CD8 T cell responses to a vascularized allograft were examined in a murine adoptive-transfer model in which help could only be provided by transferred CD4 T cells recognizing alloantigen exclusively through the direct pathway. METHODS: Rejection kinetics and the development of alloantibody and cytotoxic CD8 T cell responses to MHC-mismatched H-2d heart grafts were compared when CD4 T cell help was present (wild-type H-2d recipients), or absent (CD4 T cell deficient, MHC class II-/- H-2b recipients [B6CII-/-]), or available only through the direct pathway (B6CII-/- mice reconstituted with wild-type CD4 T cells). RESULTS: BALB/c allografts were rejected by B6 mice rapidly (median survival time [MST] 7 days) with strong CD8 T cell effector and alloantibody responses, but were rejected by B6CII-/- mice more slowly (MST 23 days), with markedly reduced CD8 T cell responses and no detectable alloantibody. CD4 T cell reconstitution of B6CII-/- recipients accelerated heart graft rejection to near that of wild-type recipients (MST 13 days), with complete restoration of cytotoxic CD8 T cell responses but without detectable IgM or IgG alloantibody. CONCLUSIONS: Different pathways of helper T cell allorecognition are responsible for generating humoral and CD8 T cell alloimmunity. CD4 T cell help provided exclusively through the direct pathway generates strong cytotoxic CD8 T cell responses that effect rapid heart graft rejection.     

CD4+CD25+ cells regulate CD8 cell anergy in neonatal tolerant mice

BACKGROUND: Injection of neonatal BALB/c mice with semi-allogeneic splenocytes leads to antigen-specific tolerance lasting into adulthood. Tolerant mice accept A/J skin grafts and fail to generate CD8 cytotoxic T lymphocyte (CTL) activity against A/J targets. Anergic CD8 T cells are present in tolerant mice, and CD4 regulatory cells function to maintain CD8 cell anergy. METHODS: Neonatal BALB/c mice were injected with 108 live CAF, splenocytes, and mice were deemed tolerant by accepting A/J grafts over 40 days. CD8 cell proliferation was measured by in vitro incorporation of bromodeoxyuridine coupled with fluorescence-activated cell sorter analysis. Alloantigen-specific cytotoxicity was tested using 51Cr release assays of A/J or third-party targets. RESULTS: We demonstrate that A/J-specific anergic CD8 cells are present in neonatal primed mice that develop tolerance but not in neonatal primed mice that reject A/J skin grafts. Anergic CD8 cells show decreased proliferation and no CTL activity against A/J targets. Addition of interleukin-2 (IL-2) to unfractionated cultures fails to restore CTL activity against A/J targets. However, addition of IL-2 to CD4-depleted cultures restores A/J-specific CD8 CTL activity. Removal of CD4+/CD25+ cells, but not CD4+/CD25- cells, also restores CD8 CTL activity against A/J in the presence, but not the absence, of IL-2. Moreover, when added back into cultures, purified CD4+/CD25+ cells from tolerant mice inhibit the generation of CD8 CTL against A/J targets. CONCLUSION: These data indicate that CD8 anergy is associated with the state of tolerance, and that CD4+CD25+ cells from tolerant mice function to maintain A/J-specific CD8 cell anergy in vitro.     

Immunohistochemical analysis of CD83, CD8 and CD4 positive cells in renal cell carcinoma

OBJECTIVES: In order to evaluate the immunological milieu in renal cell carcinoma (RCC), we investigated infiltration by mature dendritic cells (CD83 positive cells), cytotoxic-T cells (CD8 positive cells) and helper-T cells (CD4 positive cells) in RCCs, as well as in surrounding normal tissues and correlations between the cell types. MATERIALS AND METHODS: Specimens from 33 surgically resected RCCs were embedded in paraffin and then stained for CD4, CD8, CD83. Each section contained three areas, tumor tissue, tumor margin and normal renal parenchyma. Cells positive for CD4, CD8 and CD83 were counted each area. RESULT: Cells positive for CD4, CD8 and CD83 were observed predominantly in the tumor margins, rather than tumor tissue and normal renal parenchyma. The differences were significant in the number of immune positive cells between tumor margin and tumor tissue, and between tumor margin and normal renal parenchyma. A significant correlations was found between CD4 and CD83 positive cells (r = 0.805, p < 0.0001), and also between CD8 and CD83 positive cells (r = 0.505, p < 0.0001). CONCLUSION: It has been reported that mature dendritic cells induces cytotoxic-T cell and helper-T cell responses. Infiltrating mature dendritic cells, cytotoxic-T cells and helper-T cells were present only in the tumor margin. This may reflect significant immune reaction around the tumor margin.     

血清微环境对小鼠T细胞衰老的调节作用

目的：探讨血清微环境对小鼠T细胞衰老的调节作用。方法：分别取年老（12～14月龄）及年轻（1.5～2月龄）小鼠各10只,提取其脾脏淋巴细胞及血清,实验分4组。组I为年老鼠T淋巴细胞＋10%年轻鼠血清;组II为年老鼠T淋巴细胞＋10%年老鼠血清;组III为年轻鼠T淋巴细胞＋10%年轻鼠血清;组IV为年轻鼠T淋巴细胞＋10%年老鼠血清。培养48h后,经流式细胞术研究CD8＋CD28＋共表达率差异。结果：组I和组II T细胞表面的CD8＋CD28＋共表达率分别是（10.84±0.6841）%和（3.18±0.1789）%,组III和组IV T细胞表面的CD8＋CD28＋共表达分别是（12.5±0.9445）%和（8.36±0.2074）%。各组间对比有统计学差异（P〈0.05）结论：血清微环境具有调节小鼠T细胞衰老的作用,年轻鼠血清能使年老鼠的T细胞表面的CD8＋CD28＋共表达率提高,年老鼠的血清能使年轻鼠的T细胞表面的CD8＋CD28＋共表达率降低。     

CD2 and CD3 receptor-mediated tolerance: constraints on T cell activation

BACKGROUND: Antigen specific allograft tolerance is induced in mice by anti-CD2 plus anti-CD3epsilon monoclonal antibody (mAb) treatment. Because anti-CD2 mAb inhibits several aspects of anti-CD3epsilon driven T cell activation, we investigated what components of T cell activation are required or may be dispensed with for tolerance induction. Anti-CD3epsilon-mediated T cell activation depends on FcgammaR interactions. METHODS: To assess the role of FcgammaR-mediated T cell activation in tolerance induction, FcgammaR binding IgG or non-binding IgG3 anti-CD3epsilon mAbs were examined. RESULTS: These mAbs, administered in conjunction with anti-CD2, were equally effective in inducing tolerance. Moreover, in vivo administration of a blocking mAb directed against the FcgammaR, or the use of allograft recipients deficient in FcgammaR, had no effect on tolerance induction. Blocking IL-2 using mAb directed against IL-2 or IL-2R also did not prevent the induction of tolerance. These results suggest that complete T cell activation was not required for tolerance induction. However, substitution of a partially activating mAb, directed against the T cell receptor (TCR) beta subunit for anti-CD3epsilon, failed to synergize with anti-CD2 mAb to induce tolerance. The anti-TCRbeta mAb and anti-CD3epsilon mAb were found to differentially down modulate expression of TCR/CD3 complex subunits. In particular, anti-CD3epsilon caused transient down modulation of the TCRbeta receptor subunit and the TCRzeta signaling module, and this pattern was enhanced and prolonged by anti-CD2. Anti-TCRbeta caused persistent TCRzeta modulation but no TCRbeta modulation, and anti-CD2 did not influence this pattern. CONCLUSIONS: These results suggest that, although full T cell activation is not required for the induction of tolerance by anti-CD2 plus anti-CD3epsilon mAb, a signal transduction pathway that is associated with TCRbeta and TCRzeta expression, and, specifically, is perturbed by mAb binding of the CD3epsilon epitope, is critical.     

CD40分子在肾癌组织中的表达研究

目的探讨肾癌组织中CD40分子的表达与癌发生浸润转移的关系。方法用免疫组织化学二步法（Envision法）对甲醛固定石蜡包埋的肾细胞癌组织（32例）和癌旁组织（10例）中CD40的表达情况进行研究，并分析CD40分子表达水平与肾癌临床分期、病理分级和发生淋巴结转移的相关性。结果CD40在肾细胞癌组织中表达的阳性率为87．5％，与肾癌癌旁组织组相比，具有显著性差异（P〈0．01）；CD40的阳性过度表达与肿瘤临床分期、病理组织学分级和淋巴结转移显著相关（P〈0．05）。结论CD40分子在肾癌中的异常表达可为肾癌的诊断、治疗及指导预后提供实验依据，并为进一步研究肾癌的生物学，尤其是为抑制Fas和TNFR介导的细胞凋亡与基因治疗肾细胞癌打下基础。     

短发夹RNA对SMMC-7721肝癌细胞株ICAM-1基因表达的影响

目的探讨细胞间黏附分子-1(ICAM-1,CD54)基因的短发夹结构RNA(shRNA)表达载体pGenesil-1/CD54对SMMC-7721肝癌细胞株CD54表达的抑制作用。方法设计CD54基因shRNA片段和阴性对照shRNA片段,构建pGenesil-1/CD54和阴性对照表达载体,载体在coli菌扩增、鉴定后,转入SMMC-7721细胞。应用免疫组织化学技术检测肝癌细胞中CD54基因的表达。结果阳性和阴性表达载体酶切,电泳有65 bp和4.9 kb条带,载体插入片段测序,与设计序列相同,表达载体构建正确。免疫组织化学检测表明,阴性对照表达载体转染SMMC-7721细胞,细胞膜和细胞质着黄色和棕色,pGenesil-1/CD54转染SMMC-7721细胞,不着色或着淡黄色。结论CD54的shRNA能特异性地抑制CD54在肝癌细胞SMMC-7721中的表达。     

Apoptosis and CD8 and CD54 cell expression in rat small bowel transplantation

The importance of activated CD8 cells expressing IL-2R in small bowel and other organ rejection has been reported. Some authors even consider that a positive correlation might be demonstrated between the number of apoptotic enterocytes and the degree of graft rejection. In addition, moderate to intense activation of endothelial molecules in small bowel allograft in rats has been reported in chronic rejection. The aim of the present paper is to ascertain, in a heterotopic small bowel transplantation (HSBT) in rats, whether CD3, CD4, CD8, and CD54 cell expression in the allograft infiltrates shows some relationship with allograft enterocyte apoptosis when rejection is present. Wistar Furth male rats were allotted to two groups: group A was the control group without transplantation; group B received a heterotopic small bowel allograft from Fisher rats and an im dose of FK506 (0.25 mg/kg/day). A significant increase of CD8, CD54 cell receptor expression, and apoptosis in the group undergoing HSBT showed rejection. No significant differences have been observed in the variables under study between the control and HSBT without rejection groups or in CD3 and CD4 among the three groups. We observed a significant correlation between apoptosis and rejection, between CD8 and CD54 with apoptosis and with rejection, and between CD8 and CD54. This indicates that the activation of endothelial molecules and cells may play an important role in established HSBT chronic rejection. We consider that this study may contribute to the knowledge of small bowel allograft chronic rejection and its immunomodulation.     

Quantitative analysis of EBV-specific CD4/CD8 T cell numbers, absolute CD4/CD8 T cell numbers and EBV load in solid organ transplant recipients with PLTD

Post transplant lymphoproliferative disease (PTLD) in solid organ transplant (SOT) recipients is assumed to be the result of impaired Epstein-Barr Virus (EBV)-specific cellular immunity. We analyzed the absolute CD4 and CD8 T cell counts as well as the EBV-specific CD4 and CD8 T cell responses in relation to EBV load in SOT recipients with PTLD. A prospective, single center study was initiated and 10 immunosuppressed patients with diagnosis of PTLD were analyzed and compared to 3 patients without PTLD (2 SOT recipients with EBV-reactivation, 1 patient with Infectious Mononucleosis) and 6 healthy EBV positive controls. EBV-specific CD8 T cells were enumerated using HLA class I tetramers and the IFN-gamma cytokine secretion assay. EBNA1-specific CD4 T cells were analyzed after protein stimulation and EBV load was quantified by real-time PCR. Absolute CD8 T cell counts were highly variable in all 19 cases analyzed. In contrast, the absolute EBV-specific CD8 T cell count was found to be low in 7/9 patients with PTLD (<5/microl whole blood). These frequencies were similar to absolute EBV-specific CD8 T cell numbers observed in healthy EBV positive donors, but much lower compared to patients with EBV reactivation but no PTLD. Absolute CD4 T cell counts were significantly lower in PTLD patients (mean: 336/microl+/-161 vs. controls 1008/microl+/-424, p=0.0001), with EBNA1-specific CD4 T cell responses being also low, but highly variable. Moreover, low absolute CD4 T cell counts (<230/microl) were associated with an elevated EBV load (>1000 copies/microg DNA). We conclude that SOT recipients with PTLD have an inadequate functional EBV-specific T cell response. Our data suggest that the frequency and function of circulating EBV-specific CD8 T cells are dependent on absolute CD4 T cell counts. Further studies are needed to verify if a low absolute CD4 T cell count presents a risk factor for the development of PTLD in SOT recipients.     

Effect of CD4(+) and CD8(+) cell depletion on wound healing

BACKGROUND: Depression of the immune system can result in poor or delayed wound healing. METHODS: Thymectomized rats were depleted of CD4(+) and CD8(+) lymphocytes by intraperitoneal injection of Medical Research Council Oxford (MRC OX)38 antibodies and MRC OX8. Significant depletion was demonstrated throughout the wound healing process by immunofluorescence studies of peripheral blood. Following depletion the rats underwent laparotomy incisions which were allowed to heal for 10 weeks. Differences in healing were demonstrated by analysing the wounds biomechanically by tensiometry to obtain values of ultimate strength, resilience, toughness, maximum extension and elastic constant. RESULTS: Wounds of animals depleted of CD4+ lymphocytes showed a significant decrease in ultimate strength, resilience and toughness. Wounds of animals depleted of CD8(+) lymphocytes showed a significant increase in ultimate strength, resilience and toughness. CONCLUSION: Wounds healed in the absence of T lymphocytes. However, the subsets have an opposing regulatory role, with CD4(+) lymphocytes upregulating and CD8(+) lymphocytes downregulating wound healing. Presented to the Surgical Research Society in Nottingham, UK, 11 July 1997 and published in abstract form as Br J Surg 1997; 84: 1618