一氧化氮(NO)合酶和NO在延迟性异种移植排斥反应中的作用

目的利用小鼠至大鼠异位心脏移植模型,研究诱导性一氧化氮合酶(iNOS)和受体血清一氧化氮(NO)在延迟性异种移植排斥反应(DXR)中的作用.方法将大鼠随机分为4组A组(6只),空白对照;B组(5只),来氟米物(Lef)+环孢素A(CsA);C组(6只),氨基胍;D组(6只),氨基胍+Lef+CsA.利用免疫组织化学染色检测CD68和NOS2,原位杂交技术检测iNOS mRNA表达.于移植前3 d和移植心脏排斥时分别采集血清检测NO含量.结果所有被排斥心脏中均见巨噬细胞(MФ)浸润,Lef+CsA显著延长移植心脏存活(与A和C组相比,P＜0.05),单用氨基胍使移植心脏存活(3 83±1.47)d(与A组比较,P＜0.05),氨基胍联用Lef和CsA使移植心脏存活(8.67±1.76)d(与A、B和C组比较,P＜0.05).发生DXR时浸润的MФ均有NOS2蛋白和mRNA阳性表达,且不受氨基胍影响.发生DXR时大鼠血清NO水平较移植前显著升高(P＜0.01),氨基胍可显著降低排斥时NO水平.结论小鼠至大鼠心脏移植发生DXR时浸润的MФ表达iNOS增多,且血清NO升高.抑制iNOS活性,降低NO水平可显著延长移植物存活时间,提示iNOS和NO是DXR发生的可能机制之一.     

青藤碱和环孢素A联合应用延长大鼠移植心的存活时间

目的探讨青藤碱在大鼠单个核细胞（PBMC）增殖中的作用；观察青藤碱和环孢素A（CsA）联合应用对大鼠心脏移植术后移植心存活时间的影响。方法分离出大鼠外周血单个核细胞，用刀豆蛋白A（ConA）和混合淋巴细胞培养（MLC）激活单个核细胞增殖，采用四甲基偶氮唑盐（MTT）比色法及磷脂酰丝氨酸外翻分析（Annexin V）法检测单纯应用青藤碱或青藤碱＋CsA联合应用后对单个核细胞增殖作用的影响；以Wistar大鼠为供者，SD大鼠为受者，建立改良的颈部异位心脏移植模型，根据术后用药不同将其分为5组。（1）对照组：肌肉注射生理盐水1ml；（2）青藤碱组：肌肉注射青藤碱20mg·kg^-1·d^-1；（3）CsA组：肌肉注射CsA2mg·kg^-1·d^-1;（4）青藤碱＋CsA组：肌肉注射青藤碱20mg·kg^-1·d^-1＋CsA2mg·kg^-1·d^-1；（5）大剂量青藤碱组：肌肉注射青藤碱40mg·kg^-1·d^-1。观察各组移植心存活时间及病理学改变。结果（1）青藤碱能够显著的抑制ConA和MLC激活的单个核细胞增殖，并与CsA有协同作用。（2）青藤碱组及大剂量青藤碱组与对照组比较，移植心存活时间虽延长，但差异无统计学意义（P〉0．05）；青藤碱＋CsA组移植心存活时间明显延长，与其它4组比较，差异均有统计学意义（P〈0．01）。结论青藤碱能够显著的抑制单个核细胞的增殖；青藤碱联合小剂量环孢素A能明显延长移植心存活时间。     

白细胞介素10基因转染抑制小鼠心脏移植排斥反应的实验研究

目的　探讨白细胞介素 1 0 (IL 1 0 )基因转染对小鼠心脏移植排斥反应的抑制作用。方法　采用小鼠颈部心脏移植模型。随机将心脏移植后的小鼠分为 4组 :(1 )对照组 :心脏移植后每天用生理盐水 1ml灌胃 ;(2 )环孢素A(CsA)组 :心脏移植后每天用CsA 5mg/kg 灌胃 ;(3)IL 1 0组 :心脏移植时用IL 1 0重组腺病毒 30 0 μl(腺病毒滴度为 5× 1 0 1 1 pfu/ml)经主动脉根部灌注小鼠供心。(4)IL 1 0 CsA半剂量组 :在IL 1 0组的基础上 ,每天用CsA 2 .5mg/kg灌胃。观察移植心脏的存活时间及心脏跳动情况。结果　IL 1 0组、IL 1 0 CsA半剂量组和CsA组移植心脏存活时间均较对照组显著延长 (P <0 .0 1 ) ;IL 1 0组移植心脏存活时间较CsA组明显延长 (P <0 .0 5 ) ;IL 1 0 CsA半剂量组移植心脏存活时间最长 ,优于IL 1 0组 (P <0 .0 5 )和CsA组 (P <0 .0 1 )。结论　IL 1 0基因转染对心脏移植排斥反应有较强的免疫抑制作用 ,可明显延长移植心脏的存活时间 ,并且与CsA有协同作用 ,共同应用时 ,可减少CsA的用量。     

氨基胍对大鼠胰腺移植保护作用的实验研究

目的:探讨诱导型一氧化氮合酶抑制剂氨基胍对大鼠移植胰腺的保护作用。方法:糖尿病大鼠模型30只随机分成3组：（1）空白对照组（n=6），仅开腹手术，不作移植;（2）移植对照组（n=6），仅作胰腺移植;（3）氨基胍处理组（n=18），移植胰腺恢复血运前经阴茎背静脉注入盐酸氨基胍（AG）溶液，剂量分别为60，80，100 mg/kg。再灌注4h后检测血清一氧化氮（NO）水平，血糖以及淀粉酶活性，定量分析胰腺组织中的结构型一氧化氮合酶（cNOS）和诱导型一氧化氮合酶（iNOS）活性，并对胰腺进行组织形态学和组织化学检查。结果:与移植对照组比较，氨基胍处理组血NO水平及淀粉酶活性明显降低，胰腺病理损害较轻，其中以AG 80 mg/kg亚组效果更显著（P＜0.01），且该亚组血糖及iNOS活性与表达也明显低于移植对照组（P＜0.01）。结论:诱导型一氧化氮合酶选择性抑制剂氨基胍在大鼠胰腺移植中起到保护作用。其作用机制可能与抑制NO的过量产生，减轻其作为自由基的细胞毒性有关。     

他克莫司联合青藤碱抑制大鼠心脏移植急性排斥反应

目的 探讨青藤碱（SIN）对大鼠心脏移植急性排斥反应抑制作用的机制，并评价青藤碱与他克莫司（FK506）联合作用的效果。方法 以PVG大鼠为供者，DA大鼠为受者，建立同种异体心脏移植模型。将受者随机分为4组，每组20只。对照组：采用生理盐水3ml·kg^-1·d^-1灌胃；SIN组：腹腔注射SIN 30mg·kg^-1·d^-1；FK506组：采用FK506 0．25mg·kg^-1·d^-1灌胃；联合组：腹腔注射SIN 30mg·kg^-1·d^-1并联合应用FK506 0．25mg·kg^-1·d^-1灌胃。各组均在术后24h内用药，用药时间共7d。观察各组移植心存活时间，术后第7d取部分受者的移植心做病理检查，检测外周血中肿瘤坏死因子-α（TNF-α）、干扰素-γ（IFN-γ）、T细胞亚群、一氧化氮合酶（iNOS）等的浓度。结果 对照组移植心平均存活时间为（6．0±1．054）d，FK506组为（10．2±2．2）d，SIN组为（9．5±1．84）d，联合组为（16．2±2．1）d，联合组与其他3组比较，差异有统计学意义（P〈0．05）。SIN组、FK506组及联合组与对照组比较，外周血中TNF-α、IFN-γ、iNOS的表达明显减少（P〈0．05），CD4^＋T细胞亚群增殖率也明显下降（P〈0．05）。其中联合组的效果更优于SIN组和FK506组（P〈0．05）。结论 SIN能明显地抑制大鼠心脏移植急性排斥反应的发生，与FK506联合应用有协同作用。     

大鼠心脏移植后动态监测血清RANTES和单核细胞趋化蛋白-1的意义

目的 探讨测定血清RANTES（regulated on activation normal T cell expressed and secreted)和单核细胞趋化蛋白－1（MCP－1）浓度在判断大鼠心脏移植急性排斥反应中的意义及环孢素A（CsA）对它们的影响。方法 施行大鼠异位心脏移植术,按术后免疫抑制方案的不同将动物分为未干预组（不给免疫抑制剂）、低剂量CsA组、高剂量CsA组,并设同系移植对照组;采用酶联免疫吸附法检测受者术前及术后不同时段血清RANTES和MCP－1的浓度。结果 血清RANTES浓度的动态变化与急性排斥反应的进程相关,峰值出现于排斥反应的早期;应用CsA者,其RANTES峰值出现延缓,但明显低于未干预组（P＜0．05）;血清MCP－1的浓度在术后6h及严重急性排斥反应时2次升高,应用CsA后第一峰出现延迟,但不影响其峰值浓度（P＞0．05）,高CsA组未出现第二个高峰。结论 术后动态观察受者血清RANTES和MCP－1的浓度,可能对诊断急性排斥反应及判断其严重程度有帮助。     

环孢素A与盐酸小檗碱联用预防大鼠心脏移植排斥反应

目的探讨环孢素A(CsA)与盐酸小檗碱联用对同种异基因大鼠心脏移植排斥反应的抑制效果。方法建立Wistar大鼠心脏到SD大鼠的异位(腹腔内)心脏移植模型,术后单独使用CsA、盐酸小檗碱以及联合使用CsA和盐酸小檗碱进行免疫抑制治疗,观察各组移植心脏的存活时间。结果单独使用不同剂量CsA者移植心脏的存活时间较安慰剂组明显延长(P<0.01);CsA和盐酸小檗碱联用组移植心脏的存活时间较单用小剂量CsA组明显延长(P<0.01);单用盐酸小檗碱组与安慰剂组相比,移植心脏存活时间的差异无统计学意义(P>0.05)。结论CsA与盐酸小檗碱联用预防大鼠心脏移植后的排斥反应具有协同作用。     

环孢素A联合供者骨髓细胞输注延长大鼠移植心脏存活时间

目的：探讨环孢素A（CsA）联合供者骨髓细胞（DBMC）输注对同种大鼠移植心脏存活时间的影响。方法：制作Lewis大鼠到BN大鼠的异位（腹部）心脏移植模型，并按对受者处理的不同分为四组，对照组不进行特别处理，CsA组术后连续7d给予CsA5mg·kg^-1·d^-1，联合处理组分别于术中及术后第6天输注1×10^8个DBMC，术后连续7d给予CsA5nag·kg^-1·d^-1，DBMC组分别于术中及术后第6天输注1×10^8个DBMC；另设BN大鼠间的心脏移植作为对照（BN对照组）。观察移植心脏的存活时间，观测术后第6天血清白细胞介素2（IL-2）、移植心脏组织中肿瘤坏死因子（TNF-α）mRNA表达以及组织学改变，流式细胞仪检测术后第6、12、18天时受者外周血有核细胞中的供者来源细胞、CD3^＋ CD25^＋细胞、CD4^＋ CD25^＋细胞的百分比以及共刺激分子CD86表达、CD4^＋ CD45RC^＋／CD4^＋ CD45RC^-等。结果：联合处理组移植心脏存活时间为（21．6±3．2）d，明显长于对照组和DBMC组（P〈0．05），其血清IL广2为（313±95）pg／ml，心肌组织中TNF-α mRNA的表达量为0．12±0．10，均明显低于对照组和DBMC组（P〈0．05），排斥反应程度轻于其它3个移植组。联合处理组大鼠外周血有核细胞上CD86表达受到明显抑制；术后6、12d，联合处理组的CD4^＋ CD45RC^＋／CD4^＋ CD45RC^-比值低于对照组和DBMC组，CD3^＋ CD25^＋细胞百分比也低于对照组和DBMC组；接受DBMC输注者外周血中供者来源的有核细胞明显多于未接受DBMC者。结论：短疗程CsA联合DBMC输注能减轻大鼠心脏移植急性排斥反应程度，延长移植心脏存活时间。     

AG490延长移植心存活时间的探讨

目的 研究AG490在大鼠心脏移植中免疫抑制及延长移植心存活时间的作用,探讨AG490的作用机制.方法 供者为SD大鼠,受者为Wistar大鼠,建立大鼠心脏移植模型.将受者分为4组.对照组(14只):术后1～7 d经尾静脉注射生理盐水0.2 ml·kg-1·d-1;AG490组(14只):术后1～7 d经尾静脉注射AG490 20 mg·kg-1·d-1;CsA组(10只):术后1～7 d经尾静脉注射CsA20 mg·kg-1·d-1;AG490+CsA组(10只):术后1～7 d经尾静脉注射AG490和CsA各20mg·kg-1·d-1.术后各组分别取10只受者,观察移植心存活时间,并在术后1、4和7 d时,检测受者外周血中白细胞介素(IL)-2和IL-6的水平.术后第7天,处死对照组和AG490组受者(各4只)后,分别取移植心组织进行病理学检测.结果 AG490组受者术后移植心存活时间为(26.6±3.81)d,CsA组为(28.4±4.25)d,AG490+CsA组为(31.8±4.39)d,均较对照组的(8.4±0.84)d显著延长(P＜0.05).与对照组相比,AG490组术后外周血IL-2水平显著降低(P＜0.05),IL-6水平虽有所降低,但差异无统计学意义(P＞0.05),而AG490+CsA组IL-2和IL-6水平下降更为显著,与对照组相比,差异均有统计学意义(P＜0.05).AG490组移植心组织中淋巴细胞浸润程度较对照组明显减轻.结论 AG490具有免疫抑制作用,能延长移植物的存活时间,与CsA联合应用时效果更加明显.AG490的主要作用机制包括降低IL-2表达的水平,抑制移植心组织中淋巴细胞的浸润.     

环孢菌素A抑制大鼠同种心脏移植急性排斥反应中趋化因子受体CCR5表达的研究

目的观察大鼠同种心脏移植急性排斥反应中趋化因子受体CCID的表达及环孢菌素A（CsA）的抑制作用。方法分两组建立同种大鼠颈部心脏移植模型：对照组（n=25）和新山地明组（CsA组，n=25），各5例观察移植心脏存活时间。于移植术后第1、5、7、14天分别取移植心组织各5例，逆转录-聚合酶链反应（RT—PCR）检测移植心组织内趋化因子受体CCR5mRNA不同时间点的表达水平，免疫组织化学方法检测移植心组织内趋化因子受体CCR5分子的表达差异。结果对照组移植心存活时间为（11．6±1．5）d，CsA组移植心存活时间为（22．4±5．1）d，两组问移植心存活时间差异有统计学意义（P〈0．01）。急性排斥组和CsA处理组大鼠移植心脏在术后第5、7、14天都可检测到趋化因子受体CCR5mRNA阳性表达，但CsA处理组趋化因子受体CCRSmRNA的表达明显弱于急性排斥组。同样急性排斥组和CsA处理组大鼠移植心脏在术后第5、7、14天都可检测到趋化因子受体CCR5分子的表达，CsA处理组趋化因子受体CCR5分子的表达相对较弱。结论趋化因子受体CCR5在早期移植免疫事件中具有重要的作用，CsA能部分抑制趋化因子受体CCR5的表达，并与移植物存活时间延长有一定的关系。     

不同免疫治疗方案诱导大鼠原位肝移植免疫耐受的实验研究

目的 观察抗CD-40L单抗加小剂量CsA联合免疫治疗对肝移植大鼠受体免疫耐受诱导的作用.方法 在建立稳定大鼠肝移植模型的基础上,将肝移植模型分为5组.A组为SD→SD对照组;B组为SD→Wistar对照组,A,B组术后不用任何治疗措施;C组为SD→Wistar,术后用CsA1～5 d;D组为SD→Wistar,术后用CsA 1～5 d加抗CD-40L(CD-154)单抗0～2d;E组为D组+术前供体特异性输血(DSBT).观察受体存活时间、移植肝病理改变以及术后外周血中细胞因子的变化.结果 A,D,E组受体大鼠存活时点(均>60 d)均明显长于B组和C组.D,E组移植肝急性排斥反应明显减轻.B组IL-2和IFN-γ的血清水平显著高于其余各组(P＜0.05).B,C,D,E 4组IL-4和IL-10较A组均有明显增加,尤其D,E组的IL-10表达较B组显著增高(P＜0.05). 结论 抗CD-40L单抗加小剂量CsA(伴或不伴DSBT)联合免疫治疗,可有效延长肝移植大鼠受体生存时间、减轻急性排斥反应并诱导Th2类细胞因子的高水平表达,有助于受体和移植肝的长期存活.     

The relation between apoptosis of acinar cells and nitric oxide during acute rejection of pancreas transplantation in rats

Apoptosis is an important mechanism of immune-mediated graft damage. Nitric oxide (NO) generated by inducible NO synthase (iNOS) has been demonstrated to induce apoptosis. This study investigated whether apoptosis occurs during pancreas allograft rejection and examined the relationship of apoptosis of acinar cells and NO. The rats were divided into three groups: untreated isograft group, untreated allograft group and aminoguanidine (AG)-treated group. The pancreatic grafts were harvested on the post-transplantation day 3, 5 and 7 and were used to detect the histopathological rejection grade, the expression of iNOS and the apoptotic index (AI) of the graft. iNOS presented faint positive in the acinar cells of untreated isografts and did not change greatly after transplantation (P>0.05), the level of iNOS in the untreated allografts increased progressively (P<0.01) and at the same time point was significantly higher than that of untreated isograft group and AG-treated group (P<0.01). The transferase-mediated dUTP nick end labeling showed that the apoptotic cells were mainly acinar cells. A significant correlation between AI and iNOS was noted (P<0.01, r=0.611). Therefore, NO-mediated apoptosis of acinar cells plays an important role in acute rejection of pancreas transplantation, AG can mitigate the damage of pancreas allografts.     

低分子肝素减轻大鼠移植心脏排斥反应的作用及其机制

目的 探讨低分子肝素对大鼠移植心脏急性和慢性排斥反应的影响及其机制.方法 以SD大鼠为供者,Wistar大鼠为受者,进行异位(腹部)心脏移植.将受者随机分为急性排斥反应组(简称"急排组")和慢性排斥反应组(简称"慢排组").急排组中,15只于移植当天开始皮下注射低分子肝素200 μg穔g-1穌-1(小剂量者),直至移植心脏停搏;15只皮下注射低分子肝素2000 μg穔g-1穌-1(大剂量者),用药时间同前;以不给低分子肝素者作对照.慢排组所有受者均于移植当天至移植术后第9天腹腔注射环孢素A 5 mg·kg-1·d-1,其中10只还于移植当天至移植后第90天皮下注射低分子肝素2000 μg穔g-1穌-1,10只注射低分子肝素的同时再皮下注射左旋精氨酸甲酯(L-NAME)10mg·kg-1·d-1,以不给低分子肝素和L-NAME者作对照.移植后第5天,处死急排组部分受者,切取移植心脏,根据Stanford标准进行移植物排斥反应的病理学诊断和分级;剩余受者观察移植心脏存活时间.移植后第90天,测定慢排组受者的血NO浓度和移植心脏组织中诱生型NO合酶(iNOS)mRNA表达强度,并行心脏移植物血管病(CAV)评分.结果 急排组中,对照者、小剂量者和大剂量者的排斥反应等级评分分别为(4.20±0.45)分、(3.60±0.55)分和(2.40±0.55)分,移植心脏存活时问分别为(7.30±1.49)d、(8.20±1.47)d和(9.20±1.23)d,大剂量者的排斥反应等级评分明显低于对照者和小剂量者,移植心脏存活时间明显长于对照者和小剂量者(P＜0.05).慢排组中,仅给予低分子肝素者的血NO浓度和iNOS mRNA表达强度明显高于对照者和加用L-NAME者,而其CAV评分明显低于对照者和加用L-NAME者,差异均有统计学意义(P＜0.01).结论 低分子肝素可减轻急、慢性排斥反应程度,延长移植心脏存活时间;其抑制CAV的作用可能是通过上调iNOS mRNA表达水平,进而增加NO的释放来实现的.     

骁悉、环孢素A和泼尼松在肾移植受者联合应用中最适剂量探讨

目的探讨国人肾移植后骁悉(MMF)、环孢素A(CsA)和泼尼松(Pred)联合应用的最适剂量。方法比较常规剂量组(104例)和低剂量组(96例)患者肾移植术后免疫抑制剂用量和急性排斥反应、肺部感染发生率和人/移植肾生存率。结果MMF和CsA用量在术后3个月、Pred用量在术后6个月内常规剂量组明显高于低剂量组。常规剂量组在6个月内包括活检证实、临床推断和亚临床排斥,其总的急性排斥发生率为17.3%,而低剂量组为19.8%,两组间差异无显著性意义(P=0.55)。两组患者在6个月内肺部感染发生率分别为40.4%和11.5%(P<0.0001),尤其是重症肺部感染发生率常规剂量组明显要高(26.9%比5.2%,P<0.0001)。两组患者人/肾1年生存率分别为87.4%/85.5%和97.9%/96.9%(P均<0.01)。排除感染所致的带功死亡后两组患者人/肾1年生存率差异无显著性意义。结论低剂量MMF、CsA和Pred联合应用并不增加急性排斥率、亚临床排斥发生率、排斥治疗逆转率和排斥反应的严重程度,但明显降低肺部感染发生率和病死率。     

Improved renal function in sirolimus-treated renal transplant patients after early cyclosporine elimination

BACKGROUND: Sirolimus (Rapamune; SRL) in combination with cyclosporine (CsA) reduces the incidence of acute rejection episodes in renal allograft recipients. This study evaluated whether renal function could be improved by elimination of CsA from an SRL-based regimen. METHODS: This phase 2, open-label, controlled, randomized study was conducted at 17 centers in the United States and Europe. Two hundred forty-six first cadaveric renal allograft recipients were enrolled, and 197 were randomized to full-dose CsA (microemulsion) plus fixed-dose SRL (2 mg/day; group A, n=97) or reduced-dose CsA plus concentration-controlled SRL (troughs 10-20 ng/mL; group B, n=100). Most patients with acute tubular necrosis-delayed graft function that resolved later than posttransplantation day 7 were not randomized but were assigned to a third group (nonrandomized, n=49) and received up to 5 mg per day of SRL as part of their individualized treatment regimen. All patients received standard doses of corticosteroids. At the end of posttransplantation month 2, eligible patients (those not treated for rejection within 3 weeks) in group B had CsA tapered and eliminated over the subsequent 4 to 6 weeks. RESULTS: At 12 months after transplantation, renal function was significantly better in the CsA-elimination arm. In patients who were on therapy and who had not experienced an acute rejection episode before month 6, serum creatinine level was significantly lower (1.38 mg/dL vs. 1.82 mg/dL, P < 0.001) and calculated glomerular filtration rate (Nankivell method) was significantly higher (73.5 mL/min vs. 57.1 mL/min, P < 0.001) in group B than in group A. In the intention-to-treat population, rates of biopsy-confirmed acute rejection at 12 months were similar between groups A and B (18.6% vs. 22.0%, respectively; P = 0.598). In addition, graft survival (92.8% and 95.0%) and patient survival (96.9% and 96.0%) rates at 12 months were not significantly different between groups A and B, respectively. Furthermore, there were no significant differences between black and nonblack recipients within treatment groups in terms of rejection rates and graft survival at 12 months. Black recipients in group B had better serum creatinine levels at 12 months compared with black recipients in group A (1.55 mg/dL vs. 2.69 mg/dL, respectively, P = 0.011), as did nonblack recipients in group B compared with nonblack recipients in group A (1.53 mg/dL vs. 1.75 mg/dL, respectively, P = 0.055). Black patients in group A had higher mean serum creatinine levels (2.69 mg/dL) than nonblack patients in group A (1.75 mg/dL, P = 0.028). Hypertension, edema, hypomagnesemia, and dyspnea were reported significantly less frequently in patients randomly assigned to undergo CsA elimination compared with patients in group A (P < 0.05); group B patients had a significantly greater (P < 0.05) incidence of abnormal liver function tests, diarrhea, hypokalemia, and thrombocytopenia. CONCLUSION: Concentration-controlled SRL with early elimination of CsA is safe and results in improved renal function. Reduced exposure to CsA does not result in a clinically significant increase in the incidence of acute rejection episodes. This is true for both black and nonblack recipients. SRL may be used to reduce the exposure of renal allograft recipients to the nephrotoxic effects of CsA.     

Trafficking of donor-derived bone marrow correlates with chimerism and extension of composite allograft survival across MHC barrier

We proposed to evaluate differences between recipient's immune response to vascularized skin and combined vascularized skin/bone allografts, under a 7-day alphabeta-TCR plus cyclosporine (CsA) treatment protocol. Thirty-six transplantations were performed in six groups: group I (isograft control-vascularized skin graft; n=6); group II (isograft control-combined vascularized skin/bone graft; n=6); group III (allograft rejection control group-vascularized skin graft; n=6); group IV (allograft rejection control-combined vascularized skin/bone graft; n=6); group V (allograft treatment-vascularized skin graft; n=6); and group VI (allograft treatment-combined vascularized skin/bone graft; n=6). Isograft transplantations were performed between Lewis rats and allografts were transplanted across the MHC barrier from Brown Norway to Lewis rats. In the allograft treatment group, a combined alphabeta-TCR+CsA protocol was applied for 7 days. All groups were compared clinically, immunologically and histologically. Statistical significance was determined with two-tailed Student's t test. Indefinite graft survival was achieved in the isograft control group (>300 days). Allograft rejection controls rejected within 5 to 9 days posttransplant; chimerism levels were undetectable (<.5%). Allografts under the alphabeta-TCR+CsA protocol had significantly extended survival when skin was combined with bone (61-125 days) compared to vascularized skin allografts (43-61 days). Lymphoid macrochimerism was significantly higher in group VI than group V. Histology confirmed skin and bone viability. Combined vascularized skin/bone allografts had higher and sustained levels of donor-specific chimerism and extended allograft survival.     

Beneficial effect of concomitant induction with antilymphoblast globulin, cyclosporine, and steroids on long-term renal allograft outcome

BACKGROUND: The addition of induction therapy with antilymphocytic antibodies to cyclosporine (CsA) based immunosuppression, has reduced acute rejection incidence and improved short-term survivals, but has not had well-established effects on long-term renal transplant survival. PATIENTS: We analyzed the long-term allograft outcome of patients included in a prospective randomized clinical study conducted in our center 15 years ago by comparing two strategies: (A) horse antilymphoblast globulin (ALG) given at 10 mg/kg on alternate days to a maximum of 6 doses with low-dose CsA started at 8 mg/kg per day and prednisone at 0.25 mg/kg per day, versus (B) CsA started at 15 mg/kg per day and prednisone at 0.5 mg/kg per day. Diabetic and highly sensitized patients (PRA > 70%) were excluded from the study. RESULTS: The characteristics of the 50 patients enrolled in each group were not different. Although patient survival was not different (88% in group A vs 77% in group B), recipients treated with ALG showed a lower incidence of acute rejection episodes (20% vs 44%, P = .01) and better death-censored renal allograft survival (57% vs 41%, P = .03). Among rejection-free patients, graft survival was 15% higher in group A (60% vs 45%, P = .12). Multivariate Cox regression analysis showed that an acute rejection episode (relative risk [RR]: 2.44, 95% confidence interval [CI] 1.36-4.39; P = .0029) rather than ALG immunosuppression (RR 0.74, 95% CI 0.41-1.33; P = NS) was an independent predictor of death-censored graft survival. CONCLUSIONS: In summary, we confirmed that concomitant induction therapy with ALG, CsA, and steroids improves long-term renal allograft survival.     

Effect of elimination of OX-19+ and OX-8+ T-cell subsets upon pancreatic allograft survival

Depletion of T cell subsets with monoclonal antibody (mAb) permits analysis of cellular events mediating allograft destruction. Mab OX-19 and mAb OX-8 were used singly and in combination together with a short pretransplant course of cyclosporine A (CsA) to deplete OX-19+ cells (all T cells) and OX-8+ cells (cytotoxic/suppressor and NK cells), respectively, in diabetic Lewis (Lew) recipients of a Wistar Furth (WF) pancreatic allograft. Depletion of lymph node T cell subsets was assessed at rejection (blood sugar greater than 250 mg/dl) by flow cytometry. Untreated Lew recipients (Group 1) rapidly rejected their allograft (11.5 +/- 2.5 days). MAb OX-19 administration on the day prior to surgery (Day -1), on the day of surgery (Day 0), and alternate days thereafter until rejection (Group 2) prolonged graft survival (15.0 +/- 1.6 days, P less than 0.05). MAb OX-19 administration on alternate days beginning 14 days prior to transplantation (Day -14) until rejection (Group 3) further prolonged graft survival (22.6 +/- 3.4 days, P less than 0.01). At rejection large numbers of OX-19+ cells were present in both groups. Administration of mAb OX-8 alone (Group 4) failed to prolong graft survival despite marked depletion of OX-8+ cells at rejection. Administration of mAb OX-19 from Day -14 together with CsA (15 mg/kg) from Days -14 to -8 inclusive (Group 5) resulted in a marked and sustained depletion of OX-19+ cells at rejection but only a modest prolongation of graft survival (27.6 +/- 6.0 days, P = 0.11). CsA alone from Days -14 to -8 failed to prolong graft survival.(ABSTRACT TRUNCATED AT 250 WORDS)     

The activity of inducible nitric oxide synthase in rejected skin xenografts is selectively inhibited by a factor produced by grafted cells

BACKGROUND: Production of nitric oxide (NO) by graft infiltrating macrophages has been suggested as an important effector mechanism of allograft rejection. Expression of the gene for the inducible NO synthase (iNOS) and the production of NO in rejected graft has been demonstrated in various models of allotransplantation. However, whether NO plays a role in rejection of skin xenografts has not been documented. METHODS: Explants of rejected skin allografts or xenografts (rat to mouse) were cultivated in vitro and the production of NO, interleukin (IL)-2, IL-4, IL-10 and interferon-gamma (IFN-gamma) by graft infiltrating cells was determined by the Griess reaction or ELISA. Effects of supernatants from cultures of xenograft explants on the expression of gene for iNOS, accumulation of iNOS protein and NO production were determined by RT-PCR or Western blots. Molecular mass of the factor with the suppressive activity was characterized by filtration on chromatography Sephacryl S-200 Superfine column. In addition, the effects of 2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine (AMT), a selective iNOS inhibitor, on survival of skin xenografts were tested. RESULTS: While explants of rejected mouse skin allografts produced substantial amounts of NO, undetectable or only very low levels of NO were found in supernatants from cultured rat skin xenografts. Cocultivation of bacterial lipopolysaccharide (LPS)-stimulated mouse macrophages which produce high quantities of NO, with pieces of rejected xenografts, but not of syngeneic grafts, allografts or normal rat skin, completely inhibited production of NO. Production of IL-6 and IL-10 by LPS-stimulated macrophages was not inhibited under the same conditions. The inhibition of NO production was mediated by a factor which was produced by rejected rat xenograft and which was eluted from chromatography Sephacryl S-200 Superfine column in a fraction representing a molecular mass of 67 kDa. The factor did not inhibit the expression of the gene for iNOS, reduce the level of iNOS protein in stimulated macrophages, or function as a scavenger of NO. Rather, the factor inhibited the function of iNOS. The finding that NO does not play an important role during rejection of skin xenografts is supported by the observation that treatment of graft recipients with AMT, a specific iNOS inhibitor, did not enhance xenograft survival, while the same treatment resulted in prolongation of survival of skin allografts. CONCLUSION: The results thus demonstrate that a 67-kDa molecule produced by rejected rat skin xenografts selectively inhibits iNOS activity in graft infiltrating macrophages. We suggest that NO does not play a significant role in rejection of skin xenografts as it does in the case of allograft rejection.