联体共生诱导大鼠心脏移植免疫耐受的实验研究

目的　通过联体共生模型 ,建立供、受者嵌合体 ,探讨嵌合体与免疫耐受的关系。方法　纯系雄性DA(RT1a)大鼠为供者 ,Lewis(RT11)大鼠为受者 ,随机分成 3组 ,每组供、受者各 15只。Ⅰ组 (未处理组 ) :仅行DA到Lewis大鼠的腹部心脏移植 ,手术前后不作任何处理。Ⅱ组 (环磷酰胺组 ) :DA到Lewis大鼠的心脏移植前后分别经腹腔注射环磷酰胺 80mg/kg。Ⅲ组 (联体组 ) :0d :供、受者大鼠腹腔注射环磷酰胺 80mg/kg ;第6d :供、受者联体 ;第 16d :联体大鼠腹腔注射环磷酰胺80mg/kg ;第2 1d :分开联体 ,行DA到Lewis大鼠的心脏移植。观察各组移植心存活时间 ,供心病理学改变 ,供、受者间的混合淋巴细胞反应 (MLR)。结果　Ⅲ组形成了稳定的供、受者嵌合体 ,供心平均存活时间为 :(76 .33± 10 .71)d ,较Ⅰ组 (7.17± 1.17)d、Ⅱ组 (8.5 0± 1.87)d显著延长 ,差异有显著性 (P <0 .0 1) ;Ⅲ组的供心仅见少量炎性细胞浸润 ;供、受者间MLR较正常对照组显著降低 ,差异有显著性 (P <0 .0 1)。结论　联体共生可形成稳定的外周和中枢嵌合体 ,嵌合体在同种心脏移植的免疫耐受中起重要作用。     

阻断B7诱导大鼠心脏移植免疫耐受的实验研究

目的　研究阻断B7方法诱导大鼠心脏异位移植的免疫耐受。　方法　采用Ono法建立大鼠异位心脏移植模型 ,实验组腹腔注射B7阻断剂CTLA 4Ig,观察大鼠移植的心脏存活天数 ,病理改变和术后IL 2 ,IgG以及IgM含量的变化。　结果　实验组移植的心脏存活时间为 :(31 6 0± 1 82 )d ,对照组 (7 2 5± 0 71)d ,差异有显著性意义 (P <0 0 1)。组织学改变 :实验组见局灶性血管周围或间质内淋巴细胞浸润和局灶性心肌损伤 ,病理分级Ⅰ～Ⅱ级。对照组可见大量淋巴细胞浸润 ,心肌细胞损伤和坏死 ,间质内出血水肿 ,病理分级Ⅳ级。术后IL 2含量实验组比对照组明显下降 ,有显著性差异 ,P <0 0 1。IgG和IgM含量差异无显著性意义 (P >0 0 5 )。　结论　阻断B7能诱导大鼠心脏移植免疫耐受 ,为移植排斥反应提供了新的治疗方法     

多次输注供体脾细胞诱导心脏移植免疫耐受的实验研究

目的　探讨供体脾细胞诱导心脏移植免疫耐受的作用。方法　将 5 0只行腹部心脏移植的纯系雄性Lewis大鼠 ,随机分为未处理组、一次脾细胞组、环磷酰胺组、一次脾细胞 环磷酰胺组、多次脾细胞 环磷酰胺组 ,每组 10只大鼠 ,以 5 0只纯系雄性DA大鼠为供体。观察移植心脏平均存活时间 (MST) ,移植后第 6天观察供体心脏病理学改变 ,供受体间的混合淋巴细胞反应 (MLR)、外源性白细胞介素 2 (IL 2 )对MLR的影响及体外过继转移实验。结果　多次脾细胞 环磷酰胺组供体心脏MST为 (85 3± 7 5 )d ,较未处理组 (7 3± 1 0 )d、一次脾细胞组 (7 9± 0 9)d、环磷酰胺组(8 1± 1 2 )d、一次脾细胞 环磷酰胺组 (2 5 8± 3 5 )d显著延长 (t=0 ,P <0 0 1) ;供体心脏仅见少量炎性细胞浸润 ;供受体间MLR较DA Lewis对照组显著降低 ,差异有显著意义 (P <0 0 1) ;外源性IL 2可以部分逆转DA Lewis耐受组MLR的低反应性 ;其免疫耐受状态可过继转移给正常的同系大鼠。结论　多次输注供体脾细胞联合应用环磷酰胺 ,可成功诱导同种大鼠心脏移植的免疫耐受。     

腹腔注射豚鼠血管内皮细胞诱导豚鼠-大鼠心脏移植免疫耐受的研究

目的探讨腹腔注射豚鼠血管内皮细胞诱导豚鼠-大鼠心脏移植免疫耐受的可行性.方法心脏移植前14 d注射豚鼠血管内皮细胞至大鼠腹腔,观察豚鼠心脏存活时间.供心标本行苏木素-伊红(HE)染色和IgM、C3免疫组织化学检查.A组为实验对照组.B组给予2×106个豚鼠血管内皮细胞.C组予以肌注环磷酰胺(CY)40 mg/kg体重.D组为腹腔注射内皮细胞2.5×106个联合肌注CY 40 mg/kg体重组.结果腹腔注射豚鼠血管内皮细胞明显延长供心存活时间[B组为(35.12±5.24)min,与A组(14.75±7.22)min比较,差异有非常显著性(P＜0.001),与C组(42.34±5.43)min比较,P＜0.05].联合使用CY与内皮细胞能促进诱导耐受[(50.33±6.21)min],C组与D组相比P＜0.05.各组的病理表现为典型的超急性排斥反应改变.结论腹腔注射豚鼠血管内皮细胞具有延长供心存活时间,诱导免疫耐受的作用.联合使用环磷酰胺与内皮细胞能促进诱导免疫耐受.     

胸腺注射供体脾细胞诱导移植心脏免疫耐受的实验研究

目的探讨诱导心脏移植免疫耐受的方法及其产生的可能机制. 方法采用大鼠腹部心脏移植模型,随机分成未处理(Ⅰ)组,胸腺注射供体脾细胞(Ⅱ)组,腹腔注射兔抗鼠淋巴细胞血清(Ⅲ)组,胸腺注射供体脾细胞联合应用兔抗鼠淋巴细胞血清(Ⅳ)组,每组6只大鼠.Ⅱ、Ⅳ组在移植前2 1 d将供体脾细胞2.5×107个注射到受体胸腺,Ⅲ、Ⅳ组受体腹腔注射兔抗鼠淋巴细胞血清(ALS)1 ml,然后行异位心脏移植.观察移植心脏存活时间,供心病理学改变及供、受体间的混合淋巴细胞反应(MLR). 结果Ⅳ组供心平均存活时间(MST)为(81.8±7.6)d,较Ⅰ组(7.3±1.0)d、Ⅱ组( 7.8±1.0)d、Ⅲ组(8.2±1.2)d显著延长,差异有显著性意义(P＜ 0.01 );供心仅见少量炎性细胞浸润;供、受体间MLR较正常对照组显著降低,差异有显著性意义(P＜0.01). 结论胸腺注射供体脾细胞联合应用ALS能成功诱导心脏移植的免疫耐受;胸腺内特异性T细胞克隆消除可能与免疫耐受的形成有关.     

胸腺内注射异基因抗原诱导鼠神经移植免疫耐受的实验研究

目的探讨小鼠胸腺内注射异基因抗原在同种异体异基因坐骨神经移植免疫耐受中的作用。方法自供体小鼠C57BL／6的脾细胞中提取MHC抗原注人受体鼠Balb／c小鼠胸腺内，于2周后移植供体鼠坐骨神经。48只Balb／c小鼠随机分为4组，A组（胸腺内注射组）、B组（自体神经移植组）、C组（冷冻异体神经移植组）、D组（异体神经移植加用免疫抑制剂组）。于3周后进行电生理学、组织学、免疫学检测。结果A组运动神经传导速度（38．23m／s）与D组（36．39m／s）相比无显著性差异（P〉0．05），组织学、电镜、免疫学（混合淋巴细胞培养及迟发性超敏反应）检测结果均证实B组分别优于A组、D组和C组。结论胸腺内注射异基因MHC抗原可诱导大鼠对异体坐骨神经移植的特异性免疫耐受。     

重组pEGFP-N1-H2Bl质粒载体诱导心脏移植免疫耐受

目的 探讨H2-Bl基因诱导小鼠心脏移植免疫耐受的机制和效果.方法 建立小鼠颈部心脏异位移植模型,供体心脏经主动脉根部灌注H2-Bl质粒真核表达载体后进行心脏移植.实验分4组:对照组、环孢素A(CsA)组、H2-Bl质粒转染组、H2-Bl质粒转染+CsA组.各组于术后1、3和7天各动态枪测供心病理改变,免疫组织化学方法测供心CD40表达情况,流式细胞仪检测供心血清中Th1/Th2细胞因子变化,记录移植心脏存活时间.结果 对照组移植后排斥反应最重,其余组与之比较均有所减轻,以H2-Bl质粒转染+CsA组排斥反应最轻.免疫组化显示术后7天时H2-Bl质粒转染组CD40与对照组相比差异有统计学意义(P＜0.05),CsA组、H2-Bl+CsA组CD40与对照组相比差异有统计学意义(P＜0.01).H2-Bl+CsA组Th2细胞因子表达较其余组增加而Th1细胞因子则减少,到术后7天时各组Th细胞因子与对照组相比差异均有统计学意义(P＜0.05).与对照组相比,其余各组小鼠供心存活天数均延长(P＜0.05).结论 H2-Bl基因干预可一定程度诱导移植心脏免疫耐受,延长同种异体小鼠颈部移植心脏存活时间.     

嵌合体在同种心脏移植免疫耐受中的作用

目的探讨嵌合体在同种心脏移植免疫耐受中的作用.方法采用大鼠腹部心脏移植模型,将30只Lewis大鼠随机分成正常对照组(Ⅰ组)、排斥反应组(Ⅱ组)、免疫耐受组(Ⅲ组),每组10只.观察移植心存活时间,供心病理学改变,供、受者间的混合淋巴细胞反应(MLR)和脾、胸腺嵌合体.结果Ⅲ组供心平均存活时间(85.28±7.48天)较Ⅱ组(7.33±1.03天)显著长(P＜0.01);Ⅱ组供心见大量炎性细胞浸润,Ⅲ组供心仅见少量炎性细胞浸润;Ⅲ组供、受者间MLR较Ⅰ组显著低(P＜0.01);Ⅲ组受者的脾、胸腺形成了稳定的供者细胞嵌合体.结论移植免疫耐受的受者形成了稳定的中枢和外周嵌合体,嵌合体的形成对移植耐受起重要的作用.     

联合阻断CD28/B7与CD40/CD40L共刺激通路诱导抗原特异性免疫耐受

目的探讨阻断CD28／B7与CD40／CD40L共刺激通路对同种异体小鼠移植心脏存活时间的影响及其机理。方法实验分4组进行，均以C57BL／6小鼠为受者，BALB／c小鼠为供者，施行腹部异位心脏移植，根据分组要求，MR1组于移植当天静脉内注射抗CD40L单克隆抗体（MR1抗体）0．25mg／d，移植后第2、4天改为腹腔注射0．25mg／d；抗B7组于移植当天至术后第4天腹腔内注射抗B7—1和抗B7—2抗体各0．1nag／d；联合处理组术后联合使用MR1抗体和抗B7抗体，二者的用法同MR1组和抗B7组；对照组术后不使用任何抗体。记录各组移植心的存活时间；移植后60d时对移植心脏组织行病理学检查。联合处理组的受者于心脏移植后150d分别接受供者来源（BALWc小鼠）及无关供者来源（C3H小鼠）的皮肤移植，对照组的受者也同时接受两种皮肤移植，术后不进行处理，术后观察移植皮片的存活时间。结果对照组移植心脏存活时间为（7．86±1．57）d，与对照组比较，MR1组、抗B7组和联合处理组的移植心脏存活时间均得到显著延长，但联合处理组延长最为明显，均超过150d；MR1组和抗B7组移植心脏组织病理学检查均呈慢性排斥反应改变，而联合处理组未见明显慢性排斥反应征象。联合处理组移植的供者来源皮肤存活时间均超过50d，而无关供者来源的皮肤则被很快排斥；对照组两种来源的皮肤移植后存活时间均较短。结论联合阻断CD28／B7与CD40／CD40L共刺激通路可延长移植心脏存活时间，诱导出抗原特异性免疫耐受。     

维生素D衍生物联合供者骨髓细胞输注诱导异基因大鼠心脏移植免疫耐受

目的 探讨同种异基因心脏移植后免疫耐受的诱导。方法 建立大鼠颈部异位心脏移植模型，按分组分别给予门静脉输注供者骨髓细胞(DBMC)(B组)、骨化三醇灌胃(C组)、输注DBMC及骨化三醇灌胃(D组)以及环孢素A(CsA)灌胃(E组)。观察移植心脏的存活时间及心肌组织病理改变，测定心肌组织中肿瘤坏死因子及细胞间粘附分子-1 mRNA的表达以及血清钙、磷浓度，进行受者与供者及无关第三品系大鼠脾细胞混合淋巴细胞培养(MLR)。结果 D组移植心脏的存活时间较其它各组显著延长(P＜0．05)；术后7d，C组、D组、E组受者脾细胞均能显著抑制供者及第三方无关供者脾细胞作为刺激细胞引起的MLR；D组手术前后血磷、血钙浓度的差异无显著性(P＞0．05)；各组急性排斥反应的程度，D组最轻；D组肿瘤坏死因子及细胞间粘附分子-1 mRNA的表达受到显著抑制，与对照组(A组)、B、C组比较，差异有显著性(P＜0．05)。结论 骨化三醇灌胃联合DBMC输注可显著延长移植心脏的存活时间，二者具有协同作用。     

胸腺修饰诱导同种异体静脉移植的免疫耐受研究

目的 观察大鼠胸腺内注射异基因抗原在同种异体异基因股静脉移植免疫耐受中的作用.方法 将48只SD大鼠随机分为4组:自体股静脉移植组(A组)、异体股静脉移植组(B组)、异体股静脉移植免疫抑制剂组(C组)、胸腺内注射供体组织相容性(MHC)抗原后移植组(D组).于2周后进行影像学、组织学、免疫学检测.结果 组织学检测结果显示:D组、C组急性排斥反应损伤较轻,B组血管壁的各层结构破坏最重,可见大量炎性细胞浸润.B组受体大鼠血清干扰素(IFN)-γ浓度为(86.707±10.928)ng/L,显著高于A、C、D组[(29.328±4.170)、(69.076±8.059)、(63.355±4.895)ng/L,P<0.05];B组受体大鼠血清白细胞介素(IL)-4浓度为(23.656±3.369)ng/L,显著低于C、D组[(29.425±4.174)、(31.000±4.659)ng/L,P<0.05].结论 胸腺内注射异基因MHC抗原可诱导大鼠对同种异体血管移植的特异性免疫耐受.     

IFN-γ对小鼠心脏移植免疫耐受的影响

目的探讨γ干扰素(IFN-γ)对阻断共刺激信号CD40-CD40配体所诱导的小鼠心脏移植免疫耐受的作用。方法移植心脏和脾脏取自行心脏移植后的同种同系受体和同种异系受体(包括接受和未接受抗CD40配体抗体治疗),使用实时定量RT-PCR方法测定IFN-γ的表达。比较野生型小鼠和IFN-γ基因去除小鼠受体移植物的存活时间。同时比较接受MR-1治疗的野生型小鼠受体和IFN-γ基因去除小鼠受体的CD4+T细胞混合淋巴细胞反应和CD8+T细胞的细胞毒性。结果发生排斥反应的移植物较免疫耐受移植物表达更高的IFN-γ。IFN-γ基因去除小鼠受体如未接受免疫抑制治疗,移植物存活时间无明显延长,较之野生型小鼠受体反而有所缩短。使用MR-1在野生型小鼠受体中诱导出移植物的长期存活,但在IFN-γ基因去除小鼠受体中却无类似效果。IFN-γ缺失使T细胞的增殖活性及细胞毒性增强。结论在阻断共刺激信号CD40-CD40配体所诱导的移植免疫耐受中,IFN-γ能够促进免疫耐受状态的形成。     

胸腺修饰诱导同种异体静脉移植的免疫耐受研究

Objective To investigate the possibility of inducing immune tolerance to vein trans-plantation in rats by intrathymic injection of allogene. Methods MHC antigen extracted from splenic cells of donor Wistar rats was intrathymically injected to recipients SD rats, and donor Wistar femur vein was transplanted after 2 weeks. Forty-eight SD rats were randomly divided into 4 groups: group A ( femur vein autograft) ,group B ( femur vein allograft) ,group C ( femur vein allograft with use of immunosuppressant), group D (intrathymus injection). Imaging, histology and immunological assays of these groups were carried out 2 weeks after the transplantation. Results Histologic parameters that were tested were better in group D than those in group B. The concentration of serum IFN-γ in group B was significantly higher than that in groups A, C and D [ ( 86. 707±10.928 ) ng/L vs (29.328±4. 170), (69.076±8.059) and (63.355± 4.895) ng/L respectively, P < 0.05 ]. The concentration of serum IL-4 in group B was obviously lower than in groups C and D [ (23.656±3.369) ng/L vs (29.425±4.174) and ( 31.000±4.659) ng/L re-spectively,P < 0.05 ]. Conclusion Intrathymus injection of allogenic antigen might induce specific im-mune tolerance to femur vein transplantation in rats.     

胸腺修饰诱导同种异体静脉移植的免疫耐受研究

Objective To investigate the possibility of inducing immune tolerance to vein trans-plantation in rats by intrathymic injection of allogene. Methods MHC antigen extracted from splenic cells of donor Wistar rats was intrathymically injected to recipients SD rats, and donor Wistar femur vein was transplanted after 2 weeks. Forty-eight SD rats were randomly divided into 4 groups: group A ( femur vein autograft) ,group B ( femur vein allograft) ,group C ( femur vein allograft with use of immunosuppressant), group D (intrathymus injection). Imaging, histology and immunological assays of these groups were carried out 2 weeks after the transplantation. Results Histologic parameters that were tested were better in group D than those in group B. The concentration of serum IFN-γ in group B was significantly higher than that in groups A, C and D [ ( 86. 707±10.928 ) ng/L vs (29.328±4. 170), (69.076±8.059) and (63.355± 4.895) ng/L respectively, P < 0.05 ]. The concentration of serum IL-4 in group B was obviously lower than in groups C and D [ (23.656±3.369) ng/L vs (29.425±4.174) and ( 31.000±4.659) ng/L re-spectively,P < 0.05 ]. Conclusion Intrathymus injection of allogenic antigen might induce specific im-mune tolerance to femur vein transplantation in rats.     

胸腺修饰诱导同种异体静脉移植的免疫耐受研究

Objective To investigate the possibility of inducing immune tolerance to vein trans-plantation in rats by intrathymic injection of allogene. Methods MHC antigen extracted from splenic cells of donor Wistar rats was intrathymically injected to recipients SD rats, and donor Wistar femur vein was transplanted after 2 weeks. Forty-eight SD rats were randomly divided into 4 groups: group A ( femur vein autograft) ,group B ( femur vein allograft) ,group C ( femur vein allograft with use of immunosuppressant), group D (intrathymus injection). Imaging, histology and immunological assays of these groups were carried out 2 weeks after the transplantation. Results Histologic parameters that were tested were better in group D than those in group B. The concentration of serum IFN-γ in group B was significantly higher than that in groups A, C and D [ ( 86. 707±10.928 ) ng/L vs (29.328±4. 170), (69.076±8.059) and (63.355± 4.895) ng/L respectively, P < 0.05 ]. The concentration of serum IL-4 in group B was obviously lower than in groups C and D [ (23.656±3.369) ng/L vs (29.425±4.174) and ( 31.000±4.659) ng/L re-spectively,P < 0.05 ]. Conclusion Intrathymus injection of allogenic antigen might induce specific im-mune tolerance to femur vein transplantation in rats.     

胸腺修饰诱导同种异体静脉移植的免疫耐受研究

Objective To investigate the possibility of inducing immune tolerance to vein trans-plantation in rats by intrathymic injection of allogene. Methods MHC antigen extracted from splenic cells of donor Wistar rats was intrathymically injected to recipients SD rats, and donor Wistar femur vein was transplanted after 2 weeks. Forty-eight SD rats were randomly divided into 4 groups: group A ( femur vein autograft) ,group B ( femur vein allograft) ,group C ( femur vein allograft with use of immunosuppressant), group D (intrathymus injection). Imaging, histology and immunological assays of these groups were carried out 2 weeks after the transplantation. Results Histologic parameters that were tested were better in group D than those in group B. The concentration of serum IFN-γ in group B was significantly higher than that in groups A, C and D [ ( 86. 707±10.928 ) ng/L vs (29.328±4. 170), (69.076±8.059) and (63.355± 4.895) ng/L respectively, P < 0.05 ]. The concentration of serum IL-4 in group B was obviously lower than in groups C and D [ (23.656±3.369) ng/L vs (29.425±4.174) and ( 31.000±4.659) ng/L re-spectively,P < 0.05 ]. Conclusion Intrathymus injection of allogenic antigen might induce specific im-mune tolerance to femur vein transplantation in rats.     

大鼠辅助性肝移植诱导免疫耐受模型的建立

目的利用PVG（TR-1^c）和DA（RT-1^a）大鼠分别为供受体，联合进行辅助性肝移植和异位心脏移植，建立大鼠辅助性肝移植诱导免疫耐受模型。方法辅助性肝移植采用重建门静脉法，异位心脏移植采用腹腔吻合法。结果同品系对照组和实验组动物及其移植心脏存活超过100d，而异品系对照组移植心脏只存活7d。结论在受体肝脏存在的情况下，辅助性供肝依然发挥其诱导耐受的效应。经长期观察，供肝无萎缩，耐受诱导效应持续存在，指示心脏搏动良好。因此，该模型在肝移植基础研究中具有实用价值。     

胸腺修饰诱导同种异体静脉移植的免疫耐受研究

Objective To investigate the possibility of inducing immune tolerance to vein trans-plantation in rats by intrathymic injection of allogene. Methods MHC antigen extracted from splenic cells of donor Wistar rats was intrathymically injected to recipients SD rats, and donor Wistar femur vein was transplanted after 2 weeks. Forty-eight SD rats were randomly divided into 4 groups: group A ( femur vein autograft) ,group B ( femur vein allograft) ,group C ( femur vein allograft with use of immunosuppressant), group D (intrathymus injection). Imaging, histology and immunological assays of these groups were carried out 2 weeks after the transplantation. Results Histologic parameters that were tested were better in group D than those in group B. The concentration of serum IFN-γ in group B was significantly higher than that in groups A, C and D [ ( 86. 707±10.928 ) ng/L vs (29.328±4. 170), (69.076±8.059) and (63.355± 4.895) ng/L respectively, P < 0.05 ]. The concentration of serum IL-4 in group B was obviously lower than in groups C and D [ (23.656±3.369) ng/L vs (29.425±4.174) and ( 31.000±4.659) ng/L re-spectively,P < 0.05 ]. Conclusion Intrathymus injection of allogenic antigen might induce specific im-mune tolerance to femur vein transplantation in rats.     

胸腺修饰诱导同种异体静脉移植的免疫耐受研究

Objective To investigate the possibility of inducing immune tolerance to vein trans-plantation in rats by intrathymic injection of allogene. Methods MHC antigen extracted from splenic cells of donor Wistar rats was intrathymically injected to recipients SD rats, and donor Wistar femur vein was transplanted after 2 weeks. Forty-eight SD rats were randomly divided into 4 groups: group A ( femur vein autograft) ,group B ( femur vein allograft) ,group C ( femur vein allograft with use of immunosuppressant), group D (intrathymus injection). Imaging, histology and immunological assays of these groups were carried out 2 weeks after the transplantation. Results Histologic parameters that were tested were better in group D than those in group B. The concentration of serum IFN-γ in group B was significantly higher than that in groups A, C and D [ ( 86. 707±10.928 ) ng/L vs (29.328±4. 170), (69.076±8.059) and (63.355± 4.895) ng/L respectively, P < 0.05 ]. The concentration of serum IL-4 in group B was obviously lower than in groups C and D [ (23.656±3.369) ng/L vs (29.425±4.174) and ( 31.000±4.659) ng/L re-spectively,P < 0.05 ]. Conclusion Intrathymus injection of allogenic antigen might induce specific im-mune tolerance to femur vein transplantation in rats.