供体脾灌注对高度致敏肾移植受者嵌合体形成的影响

目的探讨供体脾灌注对高度致敏肾移植受者稳定期嵌合体形成及移植肾功能的影响。方法对16例高度致敏患者进行配对分组,实验组肾移植术中先予供体脾灌注40min,前瞻性观察脾灌注对患者术后6个月内嵌合体形成、移植肾排斥反应发生及移植肾功能的变化。结果脾灌注后受者外周血中供者来源的有核细胞数量显著增加,受者形成稳定嵌合体的时间较早,例数比对照组更多;肾移植术后脾灌注组排斥反应发生时间较对照组延迟,排斥反应严重程度明显较对照组轻微;术后6个月时,脾灌注组患者血肌酐值低于对照组。结论供体脾灌注可以显著提高高敏肾移植受者外周血中供者来源的有核细胞数量,减轻排斥反应强度,从而促进受者嵌合体的形成,有利于改善稳定期移植肾功能。     

犬肾移植致敏模型的建立

目的建立犬致敏后的肾移植动物模型。方法雄性家犬各6条配对作为淋巴细胞供受体,采用小剂量（0.4×10^7-1.2×10^7个/kg）淋巴细胞多部位多次输注的方法诱导致敏。用补体依赖的淋巴细胞毒（complement deendent cytotoxicity,CDC）和混合淋巴细胞培养（mixed lymphocyte culture,MLC）试验进行检测。当CDC转为阳性和MLC显示反应淋巴细胞增殖明显活跃后,淋巴细胞供受体犬间进行交叉肾移植,术后定期ECT动态显像观察移植肾功能变化;分批摘取移植肾行病理检查,观察排斥反应发生情况。结果实验犬均在输注淋巴细胞3-4次后CDC转为阳性,MLC显示反应淋巴细胞增殖明显活跃。肾移植术后4天ECT检查有4条致敏犬移植肾血流灌注下降,肾功能出现损害,摘取的3只移植肾病理检查表现为抗体介导的排斥（延缓）或急性排斥反应;对照组未见异常。术后7天,致敏犬移植肾均因急性排斥失功;对照组3条犬有2条出现肾功能损害,病理检查示急性排斥反应。结论小剂量淋巴细胞多部位多次输注可以诱导出家犬的免疫致敏状态。致敏犬肾移植术后移植肾排斥反应出现较早,肾功能损害更严重。     

肾移植群体反应性抗体检测及其临床意义

目的 探讨群体反应性抗体(PRA)检测在肾移植中的意义。方法 采用酶联免疫吸附法(ELISA)对360例肾移植受者移植前、移植后1周和2周的血清PRA进行检测,并分析其与术后急性排斥反应（AR）的关系。 结果 肾移植术前PRA阳性者59例,术后20例发生AR（33.9%）;术前PRA阴性者301例,术后58例发生AR（19.3%）,两组比较有显著性差异(P<0.05)。移植术前PRA阴性,移植后转为阳性者66例,术后19例发生AR（28.8%）;手术前后PRA持续为阴性者235例,术后39例发生AR（16.6%）,两组比较有显著性差异(P<0.05)。结论 PRA的检测是肾脏移植术前筛选致敏受者的重要指标,手术前后检测PRA对预测移植肾排斥反应有重要意义。     

肾移植受者血清群体反应性抗体水平与术后早期排斥反应间的关系

目的 探讨肾移植受者血清群体反应性抗体(PRA)水平与术后早期排斥反应间的关系.方法 对256例肾移植受者术前测定其血清PRA水平,并对PRA与术后早期排斥反应(超急性、加速性、急性排斥反应)之间的关系进行分析.结果 PRA阴性组、轻度致敏组、高致敏组3组间术后早期排斥反应发生率差异有统计学意义(χ2=78.23,P＜0.01),组间两两比较,差异有统计学意义(q=-8.32～-2.87, P＜0.05、0.01).男性组PRA阳性率低于女性组(χ2=14.18,P＜0.01);无输血史组PRA阳性率低于有输血史组(χ2=5.39,P＜0.05);有妊娠史组与无妊娠史组PRA阳性率比较差异无显著性(χ2=0.66,P＞0.05);二次移植组PRA阳性率明显高于初次移植组(χ2=20.10,P＜0.01).结论 输血、再次移植是PRA阳性的危险因素.随着PRA水平的升高,术后早期排斥反应的发生率升高.肾移植受者血清低水平的PRA可降低术后早期排斥反应的发生,提高人、肾生存期.     

致敏受者肾移植急性排斥反应的影响因素

目的：探讨致敏受者肾移植急性排斥反应的影响因素。方法：对102例术前致敏患者临床资料进行回顾性分析．探讨群体反应抗体(PRA)水平、氨基酸残基配型、术后PRA水平升高及细胞因子基因型对急性排斥反应发生率的影响。结果和结论：102例致敏肾移植受者术后随访期间发生急性排斥反应33例次，其中PRA水平、氨基酸残基相配程度、术后PRA水平升高、TNF-α高产量基因型和IL-10高产量基因型对移植肾的急性排斥发生率均有显著性影响。术前综合评估这些因素，有利于制订合理的免疫抑制方案。     

肾移植术后受者高PRA介导超急性排斥反应

为探讨群体反应性抗体（Penalreactiveantibody，PRA）对肾移植的独立影响，对9例移植受者术后PRA及其与超急排斥性反应和预后进行关联调查，发现术后PRA明显升高，接受供肾HLA（Humanleucocyteantigen）配合≤3个抗原的受者升值均＞34％，8／9例发生超急性排斥反应和移植肾无功能，2／9例死于排斥并发症；接受供肾HLA配合5个的1例受者PRA升值15％，无排斥反应，说明HLA配合差的移植和输血均可导致高PRA，但是前者PRA高幅＞34％。研究揭示，PRA作为HLA体液免疫性抗体诱导移植物超急性排斥反应，术后PRA检测有助早期识别严重排斥反应。     

输供体骨髓对可溶性人类白细胞抗原-I的影响

目的：通过测定肾移植病人各处理组的可溶性人类白细胞抗原-1(sHLA-Ⅰ)含量,探讨sHLA-Ⅰ在排斥及预后中的意义。方法：80例肾移植病人分为四组,第一组术前用免疫抑制剂骁悉(MMF) 输供体骨髓;第二组术前用MMF;第三组术前输供体骨髓;第四组术前不用MMF也不输供体骨髓。术前、术后6月和12月采血测sHLA-Ⅰ含量。结果：第一组与第二组、第四组相比有极显著差异,第三组与第二、第四组有明显差异;第一、第三组术前术后相比有明显差异,且术后12月表现更为显著;第二、四组之间无差异。结论：输供体骨髓后,血清中sHLA-Ⅰ含量增加,sHLA-Ⅰ以阻断受者识别供体抗原,形成免疫耐受,这可能是移植后排异反应减少的原因之一;免疫抑制剂MMF对sHLA-Ⅰ影响不大,说明MMY、是通过非sHLA-Ⅰ途径起作用的。     

致敏受者肾移植急性排斥反应的影响因素

目的 探讨致敏受者肾移植急性排斥反应的影响因素。方法 对102例术前致敏患者临床资料进行回顾性分析,探讨群体反应抗体(PRA)水平、氨基酸残基配型、术后PRA水平升高及细胞因子基因型对急性排斥反应发生率的影响。结果和结论 102例致敏肾移植受者术后随访期间发生急性排斥反应33例次,其中PRA水平、氨基酸残基相配程度、术后PRA水平升高、TNF-α高产量基因型和IL-10高产量基因型对移植肾的急性排斥发生率均有显著性影响。术前综合评估这些因素,有利于制订合理的免疫抑制方案。     

无功能移植肾切除对群体反应抗体的影响

目的 观察无功能移植肾切除对肾移植受者群体反应抗体(panel reactive antibody,PRA)及再次肾移植的影响.方法 收集我院2004-2007年收治的15例移植肾切除患者,分别于肾移植前,移植肾切除前,移植肾切除后1个月、半年、1年检测其PRA水平,同时行移植肾病理检查.部分患者经HLA配型检查后进行再次肾移植.结果 肾移植后受者PRA高于移植前,移植肾切除后1个月PRA水平进一步升高,在移植后半年及1年后逐渐降低.采用LAT板检查可发现新的HLA致敏位点,肾脏病理检查肾间质有多量C4d沉积.通过HLA配型避开致敏位点选择供肾,7例患者行再次肾移植均获成功.结论 移植肾切除可导致血清中PRA水平显著升高,PRA检查可发现新的HLA致敏位点,经良好HLA配型不影响再次肾移植.     

PRA配型技术在致敏受者肾移植术中的应用研究

目的 探讨群体反应性抗体(PRA)配型技术在致敏受者肾移植中的临床效果.方法 应用抗原板(LAT),采用酶联免疫吸附法(ELISA)检测肾移植受者术前的PRA;采用PRA配型技术进行术前配型.结果 12例致敏受者组采用PRA配型技术,肾移植术后肾功能恢复正常,无1例发生超急性排斥反应,术后1个月内急性排斥反应的发生率为25%;同期43例非致敏受者组,术后1个月内急性排斥反应的发生率为18.6%,虽较致敏受者组低,但两组之间差异无统计学意义.结论 PRA配型技术对减少致敏受者肾移植排斥反应,提高移植物存活率具有重要意义.     

群体反应性抗体对肾移植后人/肾存活率影响的临床分析

目的回顾性调查群体反应性抗体(PRA)对肾移植术后1、3、5年人/肾存活率的影响。方法纳入北京军区第二八一医院2000年1月~2012年3月肾移植受者469例,采用酶联免疫吸附法(ELISA)检测受者移植前PRA水平,以PRA≤10%为阴性结果,10%40%为高度致敏,并对1、3、5年人/肾存活率进行统计。结果 469例肾移植受者中,PRA阴性349例(阴性组),PRA阳性120例(阳性组),其中Ⅰ类抗体阳性66例,Ⅱ类抗体阳性31例,23例同时存在Ⅰ类和Ⅱ类抗体。术前PRA阴性组与阳性组1、3、5年人存活率分别为98.3%、90.9%、80.7%和93.3%、82.9%、62.9%;1、3、5年肾存活率分别为96.8%、87.9%、75.9%和86.7%、76.1%、50.5%,二组相比1、3、5年人/肾存活率均差异有统计学意义(P<0.05),轻度致敏组的1、3、5年人存活率、肾存活率与PRA阴性组相比,差异无统计学意义(P>0.05);中度致敏组的1、3、5年人/肾存活率与阴性组比较,差异有统计学意义(P<0.05);高度致敏组的1、3、5年人/肾存活率与阴性组比较,差异有统计学意义(P<0.05)。结论 PRA是肾脏移植术前筛选致敏受者的重要指标,PRA阳性会影响移植物的长期存活,PRA阳性患者在肾移植手术前必须进行严格的组织配型,采取措施降低抗体水平,选择合适的供体与手术时机,以提高人/肾存活率。     

Impact of human leukocyte antigen matching and recipients′ panel reactive antibodies on two-year outcome in presensitized renal allograft recipients

Background Renal transplantation in sensitized candidates remains a highly significant challenge worldwide. The production of panel reactive antibody （PRA） against human leukocyte antigen （HLA） is a major risk factor in presensitized recipients. The aim of this study was to evaluate the impact of HLA matching and recipients＇ PRA on two-year outcome in presensitized renal allograft recipients.
Methods We determined the percentage of panel reactivity and specificity of anti-HLA immunoglobulin （Ig） G antibodies in 73 presensitized renal allograft recipients compared with 81 unsensitized recipients （control group）. HLA genotyping of both recipients and corresponding donors was performed by PCR with sequence-specific primers （PCR-SSP）. We analyzed the factors influencing the early graft outcome （two-year rejection rates and survival rates of the grafts）, including HLA mismatching, class and degree of panel reactivity, and target antigen of donors.
Results Presensitized recipients had a worse two-year outcome than unsensitized recipients （P=0.019 for rejection rate, P=0.01 for survival rate）. The difference in number of HLA-mismatched alleles with either 6-antigen matching （Ag M） standard or amino acid residue matching （Res M） standard was not significant between the rejection and non-rejection groups of presensitized recipients or between the graft survival group and graft loss group. Compared with the control group, recipients with both PRA-I and PRA-II antibodies had a significantly worse two-year outcome （P=0.001 for rejection rate, P=0.002 for survival rate）. The two-year outcomes of the peak PRA 〉50% group and its subgroup, at-transplant PRA 〉50% group, were significantly worse compared with the control group （P=0.025 and P=0.001 for rejection rate, P=0.043 and P=0.024 for survival rate）. The rejection rates of the at-transplant target antigen positive group and its subgroup, HLA-I target antigen positive group, were significantly higher than the control group （P=0.001 and P=-0.001）, target antigen negative group （P=0.003 and P=0.001）, and peak target antigen positive with negative at-transplant target antigen group （P=0.024 and ,0=-0.002）. Two-year graft survival rates of the target antigen positive group and HLA-I target antigen positive group were significantly lower than the control group （P=0.012 and ,P=0.001）. The two-year outcome of target antigen unknown group was similar to that of the target antigen positive group. Presensitized recipients with pre-transplant plasmapheresis or immunoadsorption （PRA prepared group） had a better but non-significant two-year outcome than the control group. However, the PRA unprepared presensitized recipients were different to the control group （P=-0.004 for rejection rate and P=-0.005 for survival rate）. Hyperacute rejection （HR） occurred in three recipients with positive HLA-I target antigen and without mismatch according to Res M and in one case with positive PRA-II （for an unknown target antigen）. No HR occurred in eight cases with positive HLA-II target antigens.
Conclusions Pre-transplant PRA preparations might improve the access of presensitized patients to renal donors. Avoiding antigen-positive donors remains a fundamental measure in preventing HR and early rejections.     

Establishment of a sensitized canine model for kidney transplantation

Objective:To establish a sensitized canine model for kidney transplantation. Methods: 12 male dogs were averagely grouped as donors and recipients. A small number of donor canine lymphocytes was infused into different anatomic locations of a paired canine recipient for each time and which was repeated weekly. Specific immune sensitization was monitored by means of Complement Dependent Cytotoxicity (CDC) and Mixed Lymphocyte Culture (MLC) test. When CDC test conversed to be positive and MLC test showed a significant proliferation of reactive lymphocytes of canine recipients, the right kidneys of the paired dogs were excised and transplanted to each other concurrently. Injury of renal allograft function was scheduled determined by ECT dynamic kidney photography and pathologic investigation. Results:CDC test usually conversed to be positive and reactive lymphocytes of canine recipients were also observed to be proliferated significantly in MLC test after 3 to 4 times of canine donor lymphocyte infusions. Renal allograft function deterioration occurred 4 d post-operatively in 4 of 6 canine recipients, in contrast to none in control dogs. Pathologic changes suggested antibody-mediated rejection (delayed) or acute rejection in 3 excised renal allograft of sensitized dogs. Seven days after operation, all sensitized dogs had lost graft function, pathologic changes of which showed that the renal allografts were seriously rejected. 2 of 3 dogs in control group were also acutely rejected. Conclusion：A convenient method by means of repeated stimulation of canine lymphocyte may induce specific immune sensitization in canine recipients. Renal allografts in sensitized dogs will be earlier rejected and result in a more deteriorated graft function.     

腺病毒介导人白介素-10转基因对大鼠同种异体移植血管IL-2、TNF-α的影响

目的:观察腺病毒介导人白介素-10(hIL-10)转基因对大鼠同种异体移植血管IL-2、TNF-α的影响及其可能的作用机制。方法:以Wistar大鼠为供体,SD为受体,采用颈动脉显微外科原位吻合技术,将经直接浸泡法转染腺病毒介导hIL-10的离体供血管植入受体血管。随机分为3组:I组(空白对照组,n=9),II组(空载体组,n=10),III组(基因转染组,n=10)。移植后5d,观察移植物病理组织学变化及免疫排斥级别;RT-PCR、ELISA、免疫组化染色检测移植血管hIL-10基因产物;免疫组化染色检测IL-2、TNF-α。结果:基因转染组移植血管有效地表达特异性hIL-10基因产物。与对照组比较,急性排斥级别降低(P<0.01);IL-2、TNF-α的表达量明显下降(P<0.01)。结论:腺病毒介导人白介素-10转基因能有效地下调IL-2、TNF-α表达,诱导移植血管局部免疫抑制状态,抑制同种异体移植血管的急性排斥反应。     

地塞米松诱导骨髓来源抑制细胞对同种异系小鼠皮肤移植排斥反应的作用

目的 探讨地塞米松诱导骨髓来源抑制细胞（MDSCs）在同种异系小鼠皮肤移植模型中的表达及免疫负调节作用。方法 以雌性BALB/c小鼠皮肤为供体,雄性C57BL/6小鼠为受体,建立同种异系小鼠尾—背皮肤移植模型（n=40）。受体小鼠随机分为实验组和对照组（n=20）,实验组每日地塞米松磷酸盐溶液5 mg/kg腹腔注射,对照组等体积生理盐水腹腔注射,观察小鼠移植皮肤排斥反应情况,确定移植皮肤排斥时间;术后第7日切取小鼠移植皮肤行病理学检查;术后第9日处死部分小鼠,获取脾脏及移植皮肤引流淋巴结,行流式细胞术检测。结果 实验组小鼠移植皮肤的中位生存期为（24±3.062）d,显著长于对照组小鼠的（9±0.816）d （P〈0.05）。石蜡切片苏木精-伊红（HE）染色可见,与对照组比较,实验组移植皮肤的排斥反应显著减轻。流式细胞术检测结果显示：在脾脏组织,与对照组比较,实验组CD11b＋GR1＋ 细胞比例明显增高（P〈0.05）,肿瘤坏死因子-α （TNF-α）表达下调（P〈0.05）、白介素-10 （IL-10）表达上调（P〈0.05）,并伴有趋化因子受体CXCR2和黏附分子CD44、CD62L表达增加（P〈0.05）;在引流淋巴结,与对照组比较,实验组CD11b＋GR1＋ 细胞比例显著增高（P〈0.05）,而CD4＋ T细胞比例及γ干扰素（IFN-γ）表达显著减少（P〈0.05）。结论 地塞米松可能通过诱导产生MDSCs,使其向移植物局部迁移并作用于CD4＋ T细胞,抑制其功能,从而达到减轻移植物排斥反应及延长移植物生存的效应。     

不同miR-146a表达条件下Treg细胞体内回输对小鼠心脏移植免疫排斥反应的影响

目的:本研究探讨调控miR-146a Treg细胞体内回输对小鼠心脏移植免疫排斥反应的影响。方法:流式分选Treg细胞并进行体外扩增。转染试剂分别上调和下调Treg miR-146a表达。建立小鼠心脏移植模型,设立对照组,空白Treg组,miR-146a上调组,miR-146a下调组。移植后回输Treg,观察生存曲线,病理分级,测定受体脾T细胞亚群,RT-PCR检测供心IFN-γ、IL-4、IL-17表达。结果:细胞扩增10倍后miR-146a表达无明显变化,并能有效调控miR-146a表达。回输后空白Treg组（中位生存期11 d）及上调组（中位生存期13 d）生存时间延长,病理分级降低（P<0.05）;上调组更为明显（P<0.05）;下调组（中位生存期7 d）生存时间缩短,病理分级升高（P<0.05）,Th1细胞数量（28.6%±2.7%）及供心IFN-γ表达（1.12±0.11）升高（P<0.05）。结论:体外能有效地分选和扩增Treg细胞,并保证miR-146a的表达。回输Treg明显抑制小鼠心脏移植急性排斥反应,上调组抑制功能增强,下调组抑制功能减弱。     

Protein A immunoadsorption combined with rituximab in highly sensitized kidney transplant recipients

Background The number of highly sensitized patients is rising, and sensitization can lead to renal transplant failure. The present study aimed to investigate the safety and efficacy of protein A immunoadsorption combined with rituximab （RTX） in highly sensitized recipients of kidney transplants.
Methods Seven highly sensitized recipients of living-related renal transplants （4 men and 3 women, mean aged 42.5 years old （range 33-51）） were pretreated with this combination. Human leukocyte antigen （HLA） mismatch number was 2-5. Panel reactive antibody （PRA） of class Ⅰwas high in 2 cases and that of class Ⅱwas high in 1 case. All patients were pretreated with immunoadsorption 2-10 times. Immunoglobulin and PRA changes were monitored before and after absorption. The operation was conducted when PRA or immunoglobulin levels were at or below normal levels. Immunosuppressive drugs were provided 3-5 days before the operation, and one dose of RTX （375 mg/m^2） was infused with polyclonal antibody on the day of operation. Postoperative creatinine （Cr）, creatinine clearance rate （Ccr）, PRA ratio, and immunoglobulin changes were monitored.
Results All 7 patients had good recovery without delayed graft function. Acute rejection occurred in 3 cases at postoperative days 8, 10, and 14, respectively. The Banff 07 biopsy grades were la in 1 case and lla C4d0 in 2 cases. Successful reversion was achieved after giving methylprednisolone or antithymocyte immunoglobulin ＋ cyclophosphamide. All patients were discharged with normal renal function, mean class Ⅰ PRA was 14% and mean class ⅡPRA was 35%. PRA was completely negative in 3 cases.
Conclusion Protein A immunoadsorption combined with RTX can safely reduce the occurrence of humoral rejection in highly sensitized renal transplant recipients.