肝移植术后潜在免疫耐受者外周血自然杀伤细胞、调节性T细胞和记忆性T细胞的表达及其临床意义

目的 探讨肝移植术后潜在免疫耐受者免疫状态,为临床指导抗排斥治疗提供依据.方法 纳入1999年1月至2007年12月在本中心行肝移植存活至今潜在免疫耐受者29例,分为3～5年组(3Y组)10例,5～11年组(5Y组)19例,另纳入肝移植术后反复排斥反应者(RJ)10例,年龄匹配健康对照组(HC)17例;流式检测外周血单个核细胞中自然杀伤(NK)细胞、记忆性T细胞、调节性T细胞(Treg)3个亚群的表达.结果 3Y、5Y组CD4+ CD25+T细胞(3Y:27.56±4.63;5Y:30.07±3.91)、静息性Treg占CD4+T细胞比例(3Y:0.22±0.05;5Y:0.17±0.03)比R J组明显升高(CD4+ CD25+ T:11.78±4.28;静息性Treg:0.05±0.02)(P＜0.05);5Y组NK细胞占淋巴细胞比例(29.11±3.31)比RJ组(13.92±3.11)明显升高(P＜0.05);效应性记忆性CD8+T细胞比例3Y组(12.92±2.05)比RJ组(8.74±1.69)明显升高(P＜0.05).结论 肝移植术后潜在免疫耐受者外周血NK细胞、静息性调节性T细胞和效应性记忆性CD8+T细胞比例相对排斥反应者明显升高,可作为潜在的耐受标志物.     

Reduced size liver transplantation,split liver transplantation,and living related liver transplantation in relation to the donor organ shortage

Because of the shortage of cadaveric donors, three techniques of partial liver grarting have been developed. These techniques are placed in perspective in relation to the organ shortage. Reduced size liver transplantation (RSLTx) is widely used and has results comparable to those from whole liver grafting. However, this technique, while benefitting pediatric patients, reduces the adult donor liver pool. It also makes inefficient use of an available adult donor liver. In split liver transplantation (SPLTx), the whole liver is used after bipartition for two recipients. The results are comparable to those of RSLTx. The problem with SPLTx is that it is a very demanding technique applied only in centers with extensive experience with liver resection and reduction. Living related liver transplantation (LRLTx) yields excellent results; however, it places an otherwise healthy person at risk. It is argued that instead of performing risky operations on healthy persons, the health authorities should take specific measures to alleviate the organ shortage. In the meantime, SPLTx should be developed further because of its optimal use of donor tissue. As for LRLTx, its excellent results and the present shortage of size-matched pediatric liver donors justify its use, at least for now.     

大鼠肝移植急性排斥反应时T淋巴细胞免疫相关基因表达的基因芯片研究

目的 从基因水平了解T淋巴细胞在急性排斥反应中作用的分子机制。方法 建立大鼠肝移植急性排斥反应模型，并设对照组，利用4096条大鼠cDNA克隆的基因芯片从来自排斥组和对照组的T淋巴细胞中抽提、纯化mRNA，逆转录成cDNA，荧光标记后与芯片杂交，扫描后筛选出差异表达的基因。结果 在4096个基因中共发现差异表达基因190条，其中10条差异表达明显、已知功能的免疫相关基因为：主要组织相容性复合物(3条)和CD3抗原(1条)相关基因；干扰素调节因子1、上皮生长因子受体、转化生长因子相关基因各1条；白细胞蛋白酶抑制因子、急性期蛋白抑制因子3相关基因各1条；补体因子1基因1条。结论 T淋巴细胞在肝移植术后急性排斥反应中的作用机制涉及多个基因；基因芯片技术是一种高效、稳定的方法。     

Infections in human liver recipients: different patterns early and late after transplantation

The first 49 consecutive patients who underwent orthotopic liver transplantation between 1984 and 1989 in our department were studied with regard to symptomatic and asymptomatic post-transplantation infections. The major infections carrying a risk of fatal outcome are presented. During the first 4 weeks, fungal and bacterial infections predominated, the percentages of patients affected being 27% and 35%, respectively. Eight patients (17%) suffered from bacterial septicemia, which in six cases was due to gram-negative micro-organisms. The bacterial septicemia was often associated with severe ischemic damage to the graft, rejection, or cholangitis. In addition, a concomitant invasive fungal infection supervened in seven out of eight septic patients, further aggravating the patients' condition. Seventeen of the 49 patients (35%) died after transplantation within 3.3 years. Infection was the cause of death in nine patients (18%), with bacterial septicemia and/or fungemia in eight of these. Cytomegalovirus (CMV) disease was the dominant cause of illness after the 1st month. While only 5 of the 49 patients developed CMV disease during the 1st month (10%), as many as 16 of the 40 recipients who survived beyond that time suffered from symptomatic CMV viremia (40%). CMV mismatching, i.e., the donation of a CMV-positive organ to a CMV-seronegative recipient, entailed the highest risk for CMV disease. Pneumocystis carinii pneumonia occurred within 4 months in 10% of the patients. The four liver recipients affected were among the 20 patients not receiving trimethoprim-sulfamethoxazole prophylaxis. None of the 28 patients who received this prophylaxis over a 12-month period developed this complication (P<0.05). The time-related panorama of infectious complications observed in this study has immediate clinical implications for the screening, prophylaxis, and therapy of infections following liver transplantation.     

大鼠小肠移植后外周血T淋巴细胞亚群的变化

利用免疫荧光染色技术及流式细胞仪对大鼠小肠移植后外周血Ｔ淋巴细胞亚群变化进行连续监测，以探讨细胞免疫功能变化在排斥反应中的意义。结果表明，排斥反应时首先出现ＣＤ４阳性细胞显著增高，随后出现ＣＤ８阳性细胞明显下降及ＣＤ４阳性细胞／ＣＤ８阳性细胞比值增高，在排斥反应后期其比值下降；使用环孢素Ａ作免疫抑制的大鼠Ｔ淋巴细胞各亚群均无显著性变化。本实验证明ＣＤ４阳性细胞及ＣＤ８阳性细胞共同参与了排斥反应，根     

A prospective randomised trial of bile duct reconstruction at liver transplantation: T tube or no T tube?

A prospective randomised study of end-to-end bile duct reconstruction with or without T-tube drainage during orthotopic liver transplantation (OLT) was undertaken in 60 patients well matched for age, sex, aetiology of liver disease, operative blood loss, cold ischaemic time, preoperative serum bilirubin level and Child-Pugh score. Significant biliary complications in the T tube group occurred in five patients and included bile duct stricture (n=2), bile leak/peritonitis (n=1) and cholangitis (n=2). Bile duct strictures occurred in six patients in the no T tube group (P>0.05, NS). Hepatic artery stenosis was identified in one patient from each group in association with a biliary stricture. Biliary complications in both groups were associated with a prolonged graft cold ischaemic time (P<0.01). As no significant difference was noted in the number of early and late biliary complications between the two groups, the routine use of a T tube has been discontinued.     

大黄素对大鼠肝移植后CD4~+CD25~+调节性T细胞的影响

目的研究大黄素对大鼠肝移植后CD4+CD25+调节性T细胞的比例以及对其免疫抑制功能的影响。方法双袖套法建立大鼠原位肝移植模型,以大黄素和环孢素A(CsA)分别在术后腹腔给药,并以给予PBS作为模型对照组,观察不同药物移植后大鼠存活时间的影响,以及对移植术后大鼠外周血中CD4+CD25+调节性T细胞(Tregulatory cells,Treg)亚群的变化以及免疫功能的影响。结果大黄素能显著延长大鼠原位肝移植的术后成活时间,大黄素组大鼠的平均存活时间(17.4±2.5)d与肝移植模型对照组(8.8±1.9)d相比差异具有统计学意义;大鼠肝移植后大黄素给药可显著上调受体大鼠外周血以及肝内CD4+CD25+Treg的比例,并显著增强CD4+CD25+Treg抑制效应性T细胞的增殖能力(P0.05)。结论大黄素能够延长大鼠原位肝移植的术后成活时间,并可能通过上调外周血以及肝内CD4+CD25+Treg的数量及免疫抑制功能来实现移植后免疫排斥反应的抑制。     

脂肪沉着供肝在肝脏移植中的应用

目的探讨供肝脂肪浸润程度与肝脏移植病人预后的关系。方法天津市第一中心医院2002年1～12月间供体采用UW液灌注的首次肝脏移植病人71例，根据供肝脂肪浸润程度分为四组，比较各组问术后谷丙转氨酶（ALT）、谷草转氨酶（AST）水平、ICU时间及1年移植物存活率等各项指标。结果轻度脂肪肝组与无脂肪肝组的术后ALT、AST、ICU时间、1年移植物存活率均无显著性差异，中度脂肪肝组的术后ALT、AST、ICU时间均高于轻度及无脂肪肝组，但1年移植物存活率一致，三组均无移植物原发无功（PNF）发生。重度脂肪肝组只有2例，故未作统计学分析，其中1例发生PNF，于术后第2天行再次移植手术。结论轻、中度脂肪肝均可应用于l临床肝移植，对病人预后无影响；重度脂肪肝PNF发生率较高，不宜应用。     

稳定存活肝移植受者血清细胞因子水平的研究

目的 研究稳定存活的肝移植受者血清细胞因子的表达状况及意义。方法ELISA检测22例原位肝移植患者、13例原发性肝癌患者及12例正常人血清细胞因子的表达情况。流式细胞术分析外周血T细胞表型。结果外周血CD3^ 、CD8^ T细胞百分比以及CD4^ ／CD8^ 比值,各组间比较无统计学差异。肝移植组CD3^ CD25^ T细胞百分比高于正常人组(P=0．022)。血清IL-2、IFN-γ、IL-10、IL-4及INF-α水平,各组间比较差异无显著性。IL-6、ICAM-1和P-seleetin水平肝移植组显著高于正常人组(P值分别为0．048、0．000和0．025)。结论稳定存活的肝移植患者体内效应T细胞仍处于低活化状态,炎症性细胞因子(TNF-α、IL-6)和黏附分子(ICAM-1、P-Seleetin)可能在移植物的慢性损伤过程中发挥了作用。     

肝癌肝移植术前循环肿瘤细胞检测的预后价值

目的研究分析肝移植术前循环肿瘤细胞(CTC)检测评估预测受者肝癌复发与生存的应用价值。方法收集中山大学附属中山医院于2015年10月至2019年10月期间62例肝癌患者肝移植术前通过Cyttel法检测分析CTC,应用X-tile软件通过Kaplan-Meier法确定术前CTC最佳的截止值,并分析CTC与临床因素的关系;单因素及多因素COX回归分析影响其预后的独立危险因素,采用Kaplan-Meier法描绘肝移植术后无瘤生存期和总生存期的生存曲线。结果确定术前CTC最佳临界值为3个/3.2 ml,将CTC≥3/3.2 mL设置为CTC阳性组,CTC<3个/3.2 ml设置为CTC阴性组;肝移植术前CTC阳性/阴性与术前甲胎蛋白(AFP)水平、最大肿瘤直径、淋巴转移、肝移植标准、分化程度呈显著相关(P<0.05);单因素及多因素COX回归模型法系发现术前CTC个数(HR:1.262,95%CI:1.069~1.489,P=0.006)、微血管侵犯(HR:2.657,95%CI:1.120~6.305,P=0.027)是肝癌肝移植术后无瘤生存期的独立危险因素,而微血管侵犯(HR:3.738,95%CI:1.219-11.459,P=0.027)是影响肝癌肝移植术后总生存期唯一的独立危险因素;术前CTC阳性/阴性与肿瘤复发或转移(未复发、肝内复发与远处转移)差异有统计学意义(χ²=7.790,P=0.020);术前CTC阴性、阳性患者在1年、2年、3年无瘤生存率分别为82.90%、68.70%、58.90%和49.00%、29.40%、22.10%;术前CTC阴性、阳性患者在1年、2年、3年总生存率为85.50%、77.10%、69.79%和64.90%、47.20%、40.50%。术前CTC阴性的无瘤生存率曲线高于CTC阳性,差异有统计学意义(P=0.001);术前CTC阴性的总生存率曲线高于CTC阳性,差异有统计学意义(P=0.005)。结论术前CTC检测对评估肝癌肝移植术后预后具有重要的临床意义及应用前景。     

CD4+T细胞ATP含量与肝移植后急性排斥反应的关系

目的 探讨CD4+T细胞ATP含量与肝移植术后围手术期急性排斥反应(acute rejection,AR)的关系.方法 以2009年2月至2009年10月在本院行肝移植术的77例病人为研究对象.肝移植病人术前以及术后1、2、4周各采集1份全血标本,发生AR当日和激素冲击治疗后1周各采集1份全血标本.每次均送检1～2份健康志愿者全血标本作为对照.用ImmuKnowTM免疫细胞功能测定试剂盒检测样本中CD4+T细胞ATP值.结果 AR组病人ATP值在术后第1周达到高峰,显著高于同期非排斥组.术后第1周的高ATP值对诊断肝移植术后围手术期AR具有较好的敏感度和特异度.AR组病人排斥当日的ATP值与排斥活动指数(RAI)呈显著正相关,经激素冲击治疗1周后,AR均达到逆转,ATP值明显降低.结论 肝移植术后围手术期,CD4+T细胞ATP值动态变化与移植肝急性排斥反应密切相关,有望成为诊断和预防围手术期AR发生以及抗排斥治疗效果的无创监测指标.     

The characterization of reconstituted passenger leukocytes on the induction of tolerance in rat liver transplantation

The tolerance induced by orthotopic liver transplantation [DA (RT1^a) rats to PVG (RT1^c) rats] can be prevented by total body irradiation of the donor rat. Reconstitution of the irradiated donor with DA splenic leukocytes reintroduces this tolerance. To investigate the major histocompatibility complex (MHC) specificity of passenger leukocytes, irradiated DA donors were reconstituted by third-party BN (RT1ⁿ) splenic leukocytes. The reconstitution with BN splenocytes re-established DA-specific tolerance in PVG recipients, as confirmed by subsequent DA cardiac allografting, while BN hearts were rejected with second-set tempo. To determine which cell components play an important role in re-establishing liver graft tolerance, DA splenic leukocytes were further purified into three types: T, B, and adherent cells. Only “T-cell-enriched” preparations restored liver graft tolerance in three out of five PVG recipients. These results suggest that passenger leukocytes of differing MHC types can help to induce liver-specific tolerance and that T cells in the liver graft may be essential to regulate tolerance induction. Received: 31 December 1996 Received after revision: 14 April 1997 Accepted: 15 April 1997     

大鼠肝移植后树突状细胞的迁移变化

目的：研究大鼠肝移植后树突状细胞（DC）的迁移情况。方法：制作Wistar大鼠到SD大鼠的肝移植模型（实验组），并设Wistar大鼠间肝移植作为对照（对照组）。分别于术后第3、5、7天处死受者，切取移植肝组织及腹腔淋巴结，进行组织学观察，并以免疫组织化学染色观察移植肝组织和淋巴结中S-100^＋DC的动态变化，以及CD25^＋T淋巴细胞在淋巴结的活化增殖。结果：肝组织病理学检查显示，实验组移植后第5天即出现轻至中度急性排斥反应，至第7天发展成中至重度排斥反应，受者存活时间（9．7±1．4）d。免疫组织化学染色显示，术后第3天实验组移植肝组织中DC数量明显增多，于第5天达到高峰，第7天则出现下降，明显高于对照组，差异有统计学意义（P〈0．05）。从第3天开始，淋巴结中DC数量明显增多，第5、7天持续上升，与对照组相比，差异有统计学意义（P〈0．05）。移植术后第3天，实验组淋巴结中CD25^＋T淋巴细胞明显增多，呈现明显的T淋巴细胞活化增殖反应。结论：同种异体肝移植后，DC对抗原的摄取和递呈加速，产生对T淋巴细胞强烈而持久的刺激，最终激活T淋巴细胞，导致排斥反应的发生。     

Increase in natural killer cell activity following living-related liver transplantation

Abstract We monitored the serial changes of natural killer cell (NK) activity in eight recipients of living-related liver transplantation. The HLA types of all eight patients were haplotypically identical with those of their donors. Tacrolimus and methylprednisolone were used for immunosuppression. The NK activity before transplantation was 24.1 ± 20.2 % which is surprisingly low when compared with the value for normal individuals (67.7 ± 13.2%, P < 0.01) or a liver dysfunction group (49.4 ± 21.9%, P < 0.05). Serial changes in NK activity revealed a minimum of 6.1 ± 3.6% 1 week after transplantation, gradually increasing to 49.2 ± 12.5 % at 2 months after transplantation. These results suggest that the diseased liver might play an important role in the suppression of NK activity.     

肝移植患者外周血辅助性T 17细胞的变化及意义

目的探讨肝移植术后患者外周血辅助性T(Th)17细胞(CD4+IL-17+T淋巴细胞)与急性排斥反应的关系。方法本文研究对象为2008年6月至2012年12月在首都医科大学附属北京朝阳医院肝胆胰脾外科因良性终末期肝病行原位肝移植术的76例患者。根据患者术后有否发生急性排斥反应,分为排斥组(17例)和非排斥组(59例)。所有患者均按常规定期随访。记录患者排斥反应发生情况及治疗经过。排斥组患者发生急性排斥反应时给予肝穿刺活组织检查确定排斥反应严重程度。所有患者分别于肝移植术前、出院后1年内定期(间隔3~6个月)、或急性排斥反应治疗前和缓解后(3~6个月),检测外周血CD4+IL-17+T淋巴细胞占CD4+T淋巴细胞百分比(CD4+IL-17+T%)。比较两组患者各时间点的CD4+IL-17+T%。分析CD4+IL-17+T%与排斥活动指数(RAI)、免疫抑制剂血药浓度的相关性。结果急性排斥反应发生在术后0.7~12.0个月,中位数2.5个月。肝移植术后,与非排斥组比较,排斥组患者CD4+IL-17+T%明显升高[(1.79±0.44)%比(2.56±0.43)%,P0.001]。排斥组患者发生急性排斥反应时,CD4+IL-17+T%均较未发生急性排斥反应时明显升高[(2.56±0.43)%比(1.50±0.25)%,P0.001)]。非排斥组患者不同时间点的CD4+IL-17+T%变化不明显(P0.05)。排斥组患者发生急性排斥反应时的CD4+IL-17+T%与RAI呈正相关(r=0.72,P=0.001)。排斥组和非排斥组患者他克莫司、环孢素血药浓度与CD4+IL-17+T%无明显相关性(r=0.21,-0.13;均为P0.05)。结论外周血CD4+IL-17+T%可作为诊断和评估肝移植术后急性排斥反应严重程度的监测指标,外周血CD4+IL-17+T%升高提示急性排斥反应严重。     

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目的 总结人工肝脏支持系统（ALSS）应用于人体原位肝脏移植围手术期的经验体会。方法 回顾性分析1993年4月至2001年5月连续实施的55例肝脏移植中9例病人在移植前后进行ALSS治疗的临床资料。结果 55例肝脏移植中有9例病人在移植前和（或）后进行了36例次的ALSS治疗。其中移植前有8例病人进行共24例次的ALSS治疗。原发疾病为慢性重型肝炎4例，终末期肝硬化4例。治疗后病人病情好转，肝功能等各项指标显著改善，顺利实施肝脏移植。移植术后3例病人出现排斥反应，在调整免疫抑制剂方案同时辅以共12例次的ALSS治疗，此后肝功能趋于好转，排斥反应得以扭转。结论 ALSS能纠正机体术前的内环境失衡，为供肝等待和增加手术耐受创造条件，应作为肝脏移植前积极准备的重要部分；ALSS为术后移植物功能发挥欠佳者提供了有力支持和恢复功能的机会。     

Combined liver and kidney transplantation

Patients with end-stage renal and hepatic failure may be treated with combined liver and kidney transplantation (CLKTx). We reviewed the indications and outcomes of 16 CLKTx performed at the University of Minnesota between 1980 and 1994. The majority of the recipients (87.5%) were young patients affected by congenital hepatic anomalies and concomitant end-stage renal failure. Fourteen were treated with cyclosporin-based immunosuppression and had an excellent outcome: with an average of 6 years of follow-up, patient survival was 85.7%, liver graft survival 85.7%, and kidney graft survival 72%. The incidence of rejection episodes was similar to the rate of rejection in our solitary kidney and liver transplants. In conclusion, our experience supports the value of CLKTx in treating patients with simultaneous failure of both organs or with congenital enzymatic hepatic deficits leading to renal failure.     

慢性乙型肝炎病毒感染者外周血T细胞亚群及NK细胞的特点及意义

目的探讨慢性乙型肝炎病毒（hepatitis B virus,HBV）感染者外周血T细胞亚群及NK细胞的特点及临床意义。方法采用流式细胞术检测各研究组,包括慢性乙型肝炎（chronic hepatitis B,CHB）组33例、乙型肝炎失代偿期肝硬化（1iver cirrhosis,LC）组20例、慢性乙型重型肝炎（chronic severe hepatitis B,CSH）组17例及健康对照组（Control）20例的外周血T细胞亚群及NK细胞相对计数,并检测肝功能、HBVDNA含量及HBV血清标志物。结果按Control、CHB、LC、CSH顺序,CD3^＋T细胞、CD8^＋T细胞百分比依次升高,而CIM^＋T细胞、CD4^＋／CD8^＋比值及NK细胞百分比依次降低,且CHB、LC、CSH组与Control组及CHB组与CSH组相比,差异均有统计学意义（P〈0．05或P〈0．008）。CHB患者的CD3^＋T细胞与血清总胆红素（total bilirubin,TB）、HBVDNA含量（log_10）呈正相关（P〈0．001;P〈0．001）;CD8^＋T细胞与HBVDNA含量（log_10）呈正相关（P=0．007）,NK细胞与HBVDNA含量（log_10）（P=0．001）呈负相关。CHB组乙型肝炎e抗原（HBeAg）阳性者的CD4^＋T细胞及CD4^＋／CD8^＋比值低于HBeAg阴性者（P=0．018;P〈0．001）,而HBVDNA含量（log_10）和CD8^＋T细胞高于HBeAg阴性者（P=0．012;P=0．019）。结论慢性HBV感染者外周血T细胞亚群及NK细胞相对值紊乱,且与临床类型、病情、血清HBVDNA水平及HBeAg相关。     

Migration of host and donor T cells in small bowel transplantation

Abstract In this study migration of host and donor CD4⁺ andCD8⁺ T cells in a fully allogeneic model was described and compared with the migration pattern in a graft-versus-host reaction (GVHR) model, where the T-cell traffic in the graft served as a physiological control. Heterotopic small bowel transplantations were performed in a rat model, with animals being sacrificed on postoperative days (POD) 2, 3, 4, 5, and 7. Graft and host mesenteric lymph nodes were harvested, homogenized, and stained with monoclonal antibodies against MHC class I, CD4 ⁺, and CD8 ⁺ antigens. The host and donor T cell migration patterns were studied using a double-staining flow cytometric technique. We found that during the development of rejection, the normal physiological circulation of graft and host T cells was disrupted. In the graft of the allogeneic model, a shift from host cell to graft cell dominance occurred on POD 3–4. This change in migration pattern coincided in the host with a 6 % peak in graft cell infiltration, which disappeared on POD 7. These patterns of T-cell migration may be further explored for diagnostic purposes.