大鼠心脏移植与心肌细胞坏死

目的 探讨心肌细胞坏死在心脏移植中的发生和发展情况，为心脏移植的抗排斥反应治疗和预防提供依据。方法 用供体脾细胞（ＳＰＣ）和环磷酰胺（ＣＰ）预处理移植受体，ＨＥ染色对移植心脏的炎细胞浸润和心肌细胞坏死进行分析。结果 经ＳＰＣ和ＣＰ预处理后，移植心脏的存活时间明显延长，炎细胞浸润和心肌细胞变性坏死明显减轻。结论（１）急性排斥反应时，移植心脏内大量的炎细胞浸润和心肌细胞坏死。（２）大鼠心脏移植的急性排     

移植心脏动脉硬化和心肌细胞坏死的研究

目的 探讨心肌细胞坏死、纤维化和动脉硬化在移植心脏中的发生和发展情况,为心脏移植的抗排斥反应治疗和预防提供依据。方法 用供体脾细胞（ＳＰＣ）和环磷酰胺（ＣＰ）预处理移植受体,然后行异位心脏移植术,ＨＥ、Ｍａｓｓｏｎ、Ｖａｎ、Ｇｉｅｓｏｎ染色对移植以及瓣炎细胞浸润、心肌细胞坏死、心肌纤维化的冠状动脉硬化进行分析。结果 ＳＰＣ和ＣＰ预处理后,移植心脏的存活时间明显延长,炎细胞浸、心肌细胞坏死、心肌纤维     

中华眼镜蛇蛇毒对大鼠异种心脏移植超急性排斥反应的影响

试用中华眼镜蛇蛇毒消耗补体，观察其对豚鼠到ＳＤ大鼠异种心脏移植超急性排斥反应的影响。一次性给予小剂量ＣＣＶ腹腔注射后可明显降低大鼠血清补体溶血活性，４－２４小时后渐降至给药前１／２以下水平。ＣＣＶ可明显延长异种移植心脏４ 存活时间达２．５－３６小时，对照组仅为２５分钟；ＣＣＶ与脾切除，前列腺素Ｅ１联用可进一步延长移植心的存活时间，最长１例达４０小时。     

国产雷帕霉素对小鼠淋巴细胞增殖和DTH,GVHR的免疫抑制 …

目的 开发研制国产雷帕霉素（Ｒａｐａｍｙｃｉｎ,ＲＰＭ）,探讨其免疫抑制作用机理。方法 观察：⑴ＲＰＭ对刀豆素Ａ（ＣｏｎＡ）诱导的小鼠脾细胞（Ｔ淋巴细胞）增殖的抑制作用;⑵ＲＰＭ对脂多糖（Ｌｐｓ）诱导的小鼠脾细胞（Ｂ淋巴细胞）增殖的抑制作用;⑶ＲＰＭ对二硝基氯苯（ＤＮＣＢ）诱导的小鼠迟发性过敏反应（ＤＴＨ）的抑制作用;⑷ＲＰＭ对大鼠宿主抗移植物反应（ＧＶＨＲ）的抑制作用。结果 ⑴ＲＰＭ对Ｔ淋巴细胞     

大鼠移植心脏心肌纤维化的实验研究

目的 分析研究大鼠移植心脏急、慢性排斥反应的心肌纤维化变化。方法 采用供体（Wistar大鼠）脾细胞（CP）预处理移植受体（SD大鼠）,然后行同种异体心脏移植术。对移植心脏的心肌纤维化进行Masson染色分析。结果 急性排斥反应组和环孢素（CsA）组中,移植心脏呈现出严重的心肌纤维化;SPC和CP处理后,移植心脏的存活时间明显延长,其心肌纤维化的程序明显减轻。结论 经供体SPC和CP预处理,可以预防和减缓受体移植心脏心肌纤维化的发生和发展。     

胸腺内注射可溶性异基因主要组织相容性复合物抗原诱导皮肤移植特异性耐受

用３ｍｏｌ／ＬＫＣｌ从Ｃ５７ＢＬ／６小鼠脾细胞提取可溶性主要组织相容性复合物（ＭＨＣ）抗原，注射到ＢＡＬＢ／Ｃ受体鼠胸腺内，诱导了成年小鼠对该异基因小鼠皮肤移植物的耐受。除在胸腺注射当天及第３天给予抗Ｔ细胞单克隆抗体外，不使用免疫抑制剂。实验组移植皮肤平均存活时间（ＭＳＴ）为８３天，对照组ＭＳＴ为１１天。诱导耐受的小鼠对第３供体的移植皮肤仍正常排斥（ＭＳＴ为１２天）。单向混合淋巴细胞反应，耐受小鼠脾脏淋巴细胞对特异供体的脾细胞无反应，对第３供体的脾细胞反应正常，对丝裂原刺激的增殖反应正常。显示诱导的耐受是供体特异性的，无非特异性免疫抑制。     

新生大鼠胸腺内脾细胞注射对同种移植心存活的影响

目的探讨诱导新生大鼠免疫耐受的方法.方法分别给新生SD大鼠胸腺或腹腔内注射2.5×107个Wistar大鼠脾淋巴细胞,8～10周龄时行Wistar及第三品系大鼠供心移植.结果胸腺或腹腔注射Wistar大鼠脾淋巴细胞者移植Wistar大鼠心脏后移植物的存活时间显著延长,胸腺注射者平均存活时间超过60.4*!d(P＜0.001),腹腔注射者平均存活时间为15.8*!d(P＜0.02),而第三品系大鼠的心脏移植后平均存活9.2*!d(P＞0.05).结论单一胸腺或腹腔内注射同种脾细胞可以诱导新生大鼠的特异性免疫耐受.     

脾切除对心脏移植大鼠外周血淋巴细胞凋亡及调节性T淋巴细胞的影响

目的 探讨脾切除对同种异体心脏移植大鼠外周血淋巴细胞凋亡及调节性T淋巴细胞的影响.方法 以Wistar大鼠为供者、SD大鼠为受者,进行腹部异位心脏移植,同时切除受者的脾脏(心脏移植切脾组),并以不切脾者为对照(心脏移植对照组),另设不行任何处理的对照组和单纯切脾的单纯切脾组.术后第1、3、5、7天.取各组受者的移植心脏和外周血,观察移植心脏的组织学变化和细胞超微结构改变情况,以流式细胞仪检测外周血淋巴细胞的凋亡率及CD4+ CD25+ T淋巴细胞的变化,逆转录聚合酶链反应检测CD4+ CD25+ T淋巴细胞上Foxp3 mRNA的表达情况,记录移植心脏的存活时间.结果 心脏移植对照组移植心脏存活时间为(7.47±2.24)d,心脏移植切脾组移植心脏存活时间为(17.63±4.54)d,二者间的差异有统计学意义(P<0.05).心脏移植对照组的移植心脏肿胀,质硬,色暗,间质水肿、出血,弥漫性炎症细胞浸润,大量心肌细胞坏死、溶解,横纹不清;心脏移植切脾组的移植心脏质软,色红,局部灰白,外膜下以及细胞间局灶性水肿,炎症细胞浸润,心肌细胞结构完整,横纹清晰;心脏移植切脾组的细胞超微结构改变轻于心脏移植对照组.心脏移植切脾组术后第5天和第7天的淋巴细胞凋亡率分别为(7.62±2.15)%和(9.41±3.82)%,明显高于心脏移植对照组(P<0.05,P<0.05).心脏移植切脾组术后第3、5、7天时的CD4+ CD25+ T淋巴细胞明显多于心脏移植对照组(P<0.01,P<0.01,P<0.01),其Foxp3 mRNA的表达也较心脏移植对照组明显上调.结论 脾切除使心脏移植大鼠外周血淋巴细胞凋亡率增加,调节性T淋巴细胞增多,其Foxp3 mRNA表达上调,这些变化与移植心脏病理改变呈负相关.     

落新妇甙对肺移植术后急性排斥反应的作用

目的探讨落新妇甙对大鼠肺移植后机体急性排斥反应的影响和机制，以明确落新妇甙对大鼠肺移植急性排斥反应的作用。方法建立大鼠原位肺移植模型，术后将60只受体大鼠随机分为两组，对照组：术后用生理盐水1ml／d灌胃，实验组：术后用落新妇甙1ml／kg·d灌胃。观察肺移植后大鼠的存活时间、大鼠脾细胞T淋巴细胞转化率、脾淋巴细胞白细胞介素2（IL-2）的活性以及外周血中活化T淋巴细胞凋亡情况。在电子显微镜下观察肺血管超微结构变化。结果实验组大鼠肺移植后存活时间较对照组明显延长（25．4±2．1d vs．13．4±1．2d；t=2．042，P〈0．05）。实验组脾细胞T淋巴细胞转化率较对照组明显降低（23465．8±8783．4 cpm vs．74567．3±12874．6cpm；t=2．284，P〈0．05）；实验组移植大鼠脾淋巴细胞IL-2活性较对照组明显降低（4．25±2．65U／ml vs．23．46±1．82 U／ml；t=3．165，P〈0．01）。实验组能有效地诱导急性排斥反应中活化T淋巴细胞凋亡。实验组肺组织超微结构损伤较对照组减轻。结论落新妇甙通过下调IL-2产生，诱导活化T淋巴细胞凋亡，抑制T淋巴细胞增殖分化，广泛抑制了以T淋巴细胞为主的肺移植术后急性排斥反廊，从而延长肺移槽大鼠的存活时间．     

雷公藤多甙在大鼠肾移植模型中的实验研究

用显微外科技术建立大鼠肾移植模型（Ｗ→ＳＤ）。将大鼠分为四组：对照组仅给生理盐水，雷公藤多甙（Ｔ_Ⅱ）组给Ｔ_Ⅱ３０ｍｇ·ｋｇ~（１）·ｄ~（－１），ＣｓＡ组给ＣｓＡ１５ｍｇ·ｋｇ~（－１）·ｄ~（－１），ＣｓＡ＋Ｔ_Ⅱ组同时给ＣｓＡ１５ｍｇ·ｋｇ~（－１）·ｄ~（－１）＋Ｔ_Ⅱ３０ｍｇ·ｋｇ~（－１）·ｄ~（－１）。对照组大鼠肾移植后平均存活时间为７．３±２．２天，Ｔ_Ⅱ组平均存活时间为１８．１±３．７天，两组存活时间有显著性差异（Ｐ＜０．０５）。Ｔ_Ⅱ＋ＣｓＡ组平均存活时间为３４．６±５．２天，ＣｓＡ组为２６．１±４．６天。单纯用药的两组分别与联合用药组比较均有显著性差异（Ｐ＜０．０５）。Ｔ_Ⅱ３０ｍｇ·ｋｇ~（－１）·ｄ~（－１）对大鼠肝、肾、心脏及白细胞总数无影响，但显著抑制大鼠脾淋巴细胞的转化。Ｔ_Ⅱ也显著抑制大鼠外周血白细胞介素－２受体水平，但抑制能力不及ＣｓＡ。     

T细胞疫苗诱导免疫耐受与外周血T细胞凋亡的关系探讨

目的　探讨T细胞疫苗诱导特异性免疫耐受与外周血T细胞凋亡的关系。方法　制备针对Wistar大鼠的SD大鼠T细胞疫苗 ,然后用该疫苗免疫健康SD大鼠 ,每周 1次 ,连续 3周 ,作为实验组 (n =6)。对照组 (n =6)用RPMI 164 0培养液替代T细胞疫苗。以被免疫的SD大鼠的脾细胞作为反应细胞 ,以Wistar大鼠的脾细胞作为刺激细胞 (经丝裂霉素处理 ) ,于接种前和每次接种后第 5天进行单向混合淋巴细胞反应 (3 H TDR掺入法 ) ,测定其cpm值 ;用流式细胞仪对外周血T细胞凋亡情况进行检测。结果　混合淋巴细胞反应结果显示 ,实验组SD大鼠脾细胞免疫应答能力于接种后受到显著抑制 ,其cpm值较接种前显著降低 (P<0 .0 1) ;对照组接种前后各时点比较 ,其差异无显著性(P>0 .0 5 ) ;接种后相同时点的cpm值组间比较 ,实验组显著低于对照组 (P <0 .0 1) ;与接种前比较 ,接种后实验组外周血T细胞凋亡率显著增高 (P<0 .0 1) ,且随着疫苗接种次数的增加而升高 ,而对照组接种后各时点T细胞凋亡率与接种前比较差异无显著性 (P>0 .0 5 )。结论　T细胞疫苗可以诱导同种特异性免疫耐受 ,其机理可能是通过诱导外周血T细胞凋亡 ,清除抗原特异性反应性T细胞克隆来实现的。     

Role of CD4+ T cells in the rat to mouse cardiac xenotransplantation

Abstract T cell subsets involved in rejection of xenografts were analyzed using a rat to mouse cardiac xenotransplant model. Proliferating response and interleulin-2 (IL-2) production in recipients' spleen cells were almost completely abrogated by elimination of L3T4⁺ T cells, but not by elimination of Lyt2.1⁺ T cells. Cytotoxic T lymphocyte (CTL) activities were mediated by both L3T4⁺ and Lyt2.1⁺ T cells with the help of IL-2-producing L3T4⁺ T cells. Administration of anti-L3T4 monoclonal antibody (mAb) into recipient mice resulted in a significant prolongation of graft survival (mean graft survival was 29.2 days). Moreover, anti-L3T4 mAb treatment plus thymectomy led to indefinite graft survival. Anti-rat endothelial cell (EC) antibody production in the grafted mice was remarkably suppressed by anti-L3T4 mAb treatment. In contrast, Lyt2.1 mAb treatment did not prolong the graft survival and did not suppress anti-EC antibody production. These results indicated the absolute requirement of L3T4⁺ T cells in the rejection of rat to mouse cardiac xenografts.     

供体抗原特异性T细胞疫苗诱导大鼠移植心存活延长

采用大鼠异位(腹腔)心脏移植模型,使T细胞疫苗的提供者和接受者成为同一个体,探讨供体抗原致敏的T细胞疫苗接种对移植物存活期的影响,为其应用于临床防治排斥反应提供可行性依据。结果显示,T细胞疫苗可明显延长同种移植物的存活。注射T细胞疫苗后,虽然也使第三供体的移植心存活期明显延长(21.57±8.02天),但与特异供体移植心的存活期(40.75±25.15天)相比,有明显差异,证明其作用具有供体抗原特异性。本实验结果也表明使疫苗的提供者和接受者成为同一个体是可行的。     

Prolongation of allograft survival by Nippostrongylus brasiliensis is associated with decreased allospecific cytotoxic T lymphocyte activity and development of T cytotoxic cell type 2 cells

BACKGROUND: We have demonstrated that infection with Nippostrongylus brasiliensis (Nb), which induces strong type 2 responses, prolongs kidney allograft survival in rats. Here, we confirm that this effect is not species-specific and address immune modulation in allospecific T-cell responses mediated by nematode infection. METHODS: C57BL/6 mice were injected with Nb or phosphate-buffered saline. Four days later, mice were transplanted with BALB/c hearts and graft survival was assessed. In other experiments, Nb-infected mice were immunized with BALB/c spleen cells and allospecific T-cell responses were determined in vitro. RESULTS: In this study, we show that Nb prolongs cardiac allograft survival in mice. Further, spleen T cells from Nb-infected, allo-immunized mice exhibit reduced allospecific cytotoxic T-lymphocyte activity. In contrast, allospecific proliferation of T cells in the mixed lymphocyte reaction was not reduced by Nb, ruling out immunosuppression as the mechanism of Nb-induced allograft survival. Nb infection induced IL-4 and IL-6 and inhibited IFN-gamma production by T cells in response to allo-antigen. Furthermore, anti-IL-4 treatment reduced allospecific T-cell proliferation from Nb-infected but not control mice, indicating that type 2 allospecific T cells develop in the presence of Nb. We also double-stained T cells for CD8 and IL-4 and showed that Nb induces an 8-fold increase in Tc2 cell numbers. CONCLUSIONS: These results are consistent with a hypothesis that Nb mediates prolongation of allograft survival through induction of type 2 immunity, including the development of regulatory Tc2 cells, and subsequent inhibition of allospecific cytotoxic T-lymphocyte activity.     

协同信号途径阻断和供体脾细胞输注诱导预存同种记忆性T淋巴细胞大鼠心脏移植物的耐受

目的 联合阻断OX40/OX40L和CD28/B7双协同信号途径和供体特异性脾细胞输注,诱导预存同种反应性记忆T淋巴细胞大鼠对同种心脏移植物的耐受.方法 建立大鼠预存同种反应性记忆T淋巴细胞的移植模型,采用免疫磁珠法分选CD8+CD44-记忆性T细胞;对过继转移供体特异性CD8+记忆性T细胞3 d的Lewis大鼠分别/合并输注AdCTLA4Ig、AdOX40Ig、供体脾细胞(DST),同时移植来至DA大鼠的心脏;48 h后取出心脏进行组织学分析、细胞因子表达分析,同时观察不同处理移植心脏存活时间.结果 AdCTLA4Ig+AdOX40Ig+DST组分别与AdCT-LA4Ig,AdOX40Ig和DST比较,心脏组织病理级别较低,白细胞介素(IL)-2及干扰素(IFN)-γ的表达水平也明显比其他组低很多,而心脏存活时间显著延长.结论 联合阻断OX40/OX40L和CD28/B7和供体特异性脾细胞输注能较好地诱导大鼠心脏移植物耐受.     

CTLA4-Ig abrogates the anti-globulin response and prolongs cardiac allograft survival after anti-CD2 treatment

CD2 is expressed on T cells and NK cells and is important in T cell activation, making it a potential target for immune intervention. Here, we report a series of experiments aimed at defining the ability of mAbs directed against the CD2 molecule to prevent cardiac allograft rejection in low and high responder rat strain combinations. Administration of the mouse anti-rat CD2 mAbs OX34 or OX55 around the time of transplantation prolonged survival of fully allogeneic Lewis (RT1l) cardiac allografts in low responder DA (RT1a) recipients (MST 14 days for OX55 and >100 days for OX34). Treatment with OX34 prolonged graft survival in the reciprocal high responder DA to Lewis rat strain combination (MST 19 days) and when combined with CTLA4-Ig resulted in long-term graft survival (MST>100 days). Despite these in vivo effects, OX34 had little effect on in vitro assays of lymphocyte activation. Instead, the ability of OX34 to extend allograft survival correlated with T cell depletion. Administration of OX34 induced a similar degree of CD4 T cell depletion in DA and Lewis recipients, but the CD4 depletion observed was more transient in Lewis recipients. Lewis, but not DA strain rats, developed an anti-murine Ig response. Combined treatment with CTLA4-Ig abolished the anti-globulin response to OX34 in Lewis recipients, prolonged circulation of OX34 and increased the extent and duration of CD4 depletion. We conclude that anti-CD2 treatment effectively prolongs cardiac allograft survival and addition of CTLA4-Ig increases its efficacy by abrogating the production of neutralising antibodies.     

Prolongation of corneal xenotransplant survival by T-cell vaccination-induced T-regulatory cells

Abstract: Background: Corneal xenotransplantation is an alternative approach for overcoming shortage of allograft in clinics. However, the mechanism of acute corneal xenograft rejection and the method of prolonging xenograft survival have not been well defined. Methods: In this study, we used an orthotopic corneal guinea pig‐to‐rat xenotransplantation model to study the effects of CD4 and CD8 T cells, T‐cell vaccination (TCV) and TCV‐induced T‐regulatory (Treg) cells on xenograft survival. Results: The acute rejection of xenografts occurred in untreated rats as early as 6 days post‐transplantation, while TCV significantly prolonged xenograft survival from 6–12 to 21–27 days. The lymph node cells of the TCV‐treated rats exhibited significant response to the anti‐guinea pig T cells and the responding cell populations contained two Treg cell subsets, CD4⁺ CD25^? and CD8⁺ CD28^? T cells, both of which lack expression of Foxp3. Adoptive transfer of CD8⁺ CD28^? T cells resulted in profound inhibition of corneal xenograft rejection, while transfer of CD4⁺ CD25^? T cells alone exhibited no significant inhibition. However, transfer of the CD4⁺ CD25^? and CD8⁺ CD28^? T‐cell mixture remarkably enhanced the in vivo protective activity against xenograft rejection. Conclusions: These data suggest that TCV induces the activation of specific Treg cell subsets, CD4⁺ CD25^? and CD8⁺ CD28^? T cells, which may act cooperatively to mediate prolongation of corneal xenograft survival. Therefore, TCV can be used as immunotherapy for suppression of acute xenograft rejection.     

Immunomodulatory effects of mesenchymal stem cells in a rat organ transplant model

BACKGROUND: Recent reports suggest that mesenchymal stem cells (MSCs) have immunomodulatory properties. Mesenchymal stem cells can suppress the immune response toward alloantigen in vitro by inhibiting T cell proliferation in mixed-lymphocyte reactions (MLRs). However, relatively little has been reported regarding the immunomodulative potential of MSCs in vivo. Herein the authors confirm the immunomodulatory effects of rat MSCs in vitro and tested for tolerogenic features in a model of allogeneic heart transplantation. METHODS: Mesenchymal stem cells were obtained from bone marrow aspirates of male Lewis rats (major histocompatibility complex [MHC] haplotype RT1) and ACI rats (RT1). Lewis MSCs were cocultured with ACI spleen cells to reveal direct effects of MSCs on lymphocytes. In addition, MSCs were added to MLRs between ACI T cells as responders and irradiated Lewis spleen cells as stimulators. Finally, MSCs were administered in an allogeneic heart transplantation model at different doses (with and without cyclosporin A [CsA]). RESULTS: Mesenchymal stem cells appeared with typical spindle-shaped morphology in culture and readily differentiated into adipocytes when exposed to differentiation media. Mesenchymal stem cells expressed MHC class I, but not class II or costimulatory molecules. In vitro, MSCs phagocytosed ACI spleen cells. When introduced into an MLR, donor MSCs suppressed the proliferation of stimulated T cells. However, in vivo, MSC injection did not prolong allograft survival. In addition, concurrent treatment with low-dose CsA and MSCs accelerated allograft rejection. CONCLUSIONS: The present data confirm previous reports suggesting that MSCs have immunomodulatory properties in vitro. However, their tolerogenic properties in vivo must be questioned, as MSC injections were not only ineffective at prolonging allograft survival, but tended to promote rejection.     

AdCTLA-4Ig combined with donor splenocytes,bone marrow cells and anti-ICOS antibody treatment induce tolerance in a rat heart transplantation model

It is difficult to induce rat cardiac allograft tolerance by co-stimulator blockade of the B7-CD28 pathway with CTLA-4Ig monotherapy alone. However, combined therapies of AdCTLA-4Ig plus donor-specific spleen-cell infusion, bone marrow cell infusion, and anti–ICOS antibody have been demonstrated to effectively induce indefinite acceptance of rat cardiac allografts. In this report, we compared the tolerance of cardiac allograft tolerant recipients induced by the above three strategies. The results show that treating Lewis recipients of a DA cardiac allograft with a combination of AdCTLA4-Ig and anti-ICOS antibody, donor splenocytes or bone marrow cells produced indefinite graft survival. Second transplantation of donor type skin or heart grafts could not affect the survival of primary heart graft in anti-ICOS treated groups, but results in rejection of primary heart grafts in other two groups, and that co-stimulator blockade, CD28 and ICOS simultaneously with CTLA-4Ig and anti-ICOS antibody, facilitates the development of CD25+ CD4+ regulatory T cells and induces stable transplantation tolerance in the rat cardiac allograft model. This also provides an effective therapy in clinical transplantation for promoting permanent graft survival by generating regulatory T cells.Abbreviations BMC bone marrow cells - ICOS inducible co-stimulator - pfu plaque-forming units - SPC spleen cell