ImmuKnow检测免疫细胞功能在肾移植术后免疫功能监测中的应用

目的  探讨ImmuKnow检测免疫细胞功能在监测肾移植术后患者免疫功能变化的应用价值。方法  2013年1月至2014年12月在广州医科大学附属第二医院器官移植科实施肾移植手术的106例尿毒症患者, 分别于术前、术后12个月内发生感染或急性排斥反应时抽取血液标本。采用ImmuKnow测定CD4⁺ T细胞内的三磷腺苷(ATP)含量。观察与比较不同临床状态肾移植患者的ATP含量, 包括术前组、稳定组、急性排斥反应组和感染组(含重症肺炎)。检测外周血T细胞亚群CD4⁺T细胞、CD8⁺T细胞及自然杀伤(NK)细胞比例。采用Pearson相关分析法了解ATP值与他克莫司(FK506)和环孢素(CsA)血药谷浓度的关系。结果  感染组患者ATP含量低于术后稳定组患者(P < 0.001), 其中发生重症肺炎患者ATP含量低于发生其他感染的患者(P < 0.05)。感染组患者的CD4⁺T细胞百分比低于稳定组患者(P < 0.05)。ATP含量与移植患者术后FK506和CsA血药谷浓度无相关性。结论  ImmuKnow检测可用于监测肾移植患者术后免疫功能状态。CD4⁺T细胞内ATP含量检测对术后感染, 特别是对重症肺炎有提示和预警作用。     

检测肾移植受者CD4~+T淋巴细胞内三磷腺苷的意义

目的探讨检测CD4+T淋巴细胞内三磷腺苷(ATP)水平用于评价肾移植受者细胞免疫功能的价值。方法采用植物血凝素刺激、单克隆抗体包被免疫磁珠分离CD4+T淋巴细胞,以荧光试剂标记,测定27例肾移植受者(研究组)的术前,术后3d、7d、14d、30d以及发生感染时的CD4+T淋巴细胞内ATP水平,并以14名健康人作对照(对照组)。研究组患者术后均采用抗人T细胞CD3鼠单抗(OKT3)诱导治疗,并予免疫抑制剂治疗,其中21例采用以环孢素(CsA)为基础的三联用药方案,6例采用以他克莫司(FK506)为基础的三联用药方案,比较采用不同用药方案患者的ATP水平,采用Pearson相关分析评估肾移植患者ATP水平与环孢素或他克莫司血药浓度的关系。结果肾移植受者术前CD4+T淋巴细胞内ATP水平为(382±187)ng/ml,明显低于正常成人的(496±116)ng/ml(P0.05),术后7d降至最低,14d开始回升,30d时仍低于术前水平(P0.01)。3例术后并发肺部感染者,其CD4+T淋巴细胞内ATP水平下降,病情加重时下降更加明显,感染控制后有所回升。采用环孢素三联方案治疗的患者各时点的ATP水平与采用他克莫司三联方案治疗的患者比较差异无统计学意义。肾移植受者的ATP水平与上述两药血药浓度变化无关。结论测定肾移植受者CD4+T淋巴细胞内ATP水平可反映患者的整体细胞免疫状态,指导临床制定个体化治疗方案。     

细胞免疫能量测定在肾移植中的应用

目的 探讨适用于评价肾移植受者免疫状态的新方法.防止移植后的排斥反应和感染,提高人/肾生存质量和存活率.方法 应用ImmuKnow~(TM)-Cylex检测技术测定肾移植受者细胞免疫能量(三磷酸腺苷,ATP).收集62例肾移植受者术前(术前组)、术后稳定期(术后稳定组)、发生感染(术后感染组)和排斥反应(排斥反应组)等不同时期的肝素钠抗凝样本共150份.通过测量CD4+T淋巴细胞受刺激后释放的ATP浓度来判定细胞免疫力.并用秩和检验和两两比较统计方法,对结果进行分析和比较.结果 各组肾移植受者CD4+T淋巴细胞ATP浓度分别为:术前组(281.33±146.46)/μg/L、术后稳定组(310.19±147.12)/μg/L、术后感染组(142.41±118.26)μg/L、排斥反应组(332.77±154.44)μg/L;术后感染组ATP浓度显著低于其他各组.差异均有统计学意义(P<0.05).结论 LmmuKnow~(TM)-Cylex细胞免疫能量测定具有灵敏度高、特异性强、简便易操作等优点,适合于肾移植受者免疫状态的临床监测,特别是对术后感染有很好的预警作用,对指导感染后免疫抑制剂的个体化用药具有一定的参考价值.     

阿来佐单抗行肾移植免疫诱导治疗的有效性和安全性

目的 评价阿来佐单抗行肾移植免疫诱导治疗的有效性和安全性.方法 将89例肾移植受者随机分为试验组(43例)和对照组(46例).试验组于肾移植术前和术后24 h内分别静脉滴注阿来佐单抗15 mg,对照组不接受免疫诱导治疗.受者术后常规应用环孢素A(或他克莫司)+吗替麦考酚酯+泼尼松预防排斥反应.统计两组术后12月内的移植肾功能、急性排斥反应发生率、感染发生率、移植肾功能延迟恢复发生率、移植肾存活率及淋巴细胞计数,并用ImmuKnowTM免疫细胞功能测定法检测受者CD4+T淋巴细胞的三磷酸腺苷(ATP)值.结果 术后12个月内试验组7.0%(3/43)的受者发生病理证实的急性排斥反应,明显低于对照组的23.9%(11/46,P＜0.05).试验组和对照组总体的感染发生率为别为39.5%(17/43)和30.4%(14/46,P＞0.05),两组机会性感染的发生率分另为23.2%(10/43)和17.4%(8/46,P＞0.05).术后3个月内,试验组淋巴细胞计数低于对照组;术后6个月内,试验组CD4+T淋巴细胞ATP值低于对照组.结论 阿来佐单抗行肾移植免疫诱导治疗可维持受者的免疫抑制状态,未见严重不良反应.     

Cylex ImmuKnow测定稳定肝移植患者CD4+T淋巴细胞活性临床分析

目的 探讨中国人肝移植术后肝功能稳定患者ImmuKnow免疫细胞功能测定与临床免疫抑制状况的相关性.方法 收集46例肝移植术后肝功能稳定患者的临床资料,选取性别、年龄、病因、血常规、肝功能、肾功能、乙肝五项和丙肝病毒载量、CD淋巴细胞亚群、他克莫司血药谷浓度、ImmuKnow值作为观察指标,对术后肝功能稳定患者ImmuKnow值与其他各相关临床指标进行多因素回归分析.结果 肝移植术后(22±15)个月肝功能稳定患者ImmuKnow值为(203±114)ATPng/ml(38.47～524.06 ATP ng/ml).回归分析显示ImmuKnow值与白细胞计数、CD3+淋巴细胞绝对计数以及CD3+CD4+T淋巴细胞绝对计数有相关性(P<0.05).ImmuKnow值与他克莫司谷值浓度、他克莫司日用量以及患者性别、年龄及其他肝功能指标没有相关性(P>0.05).5例合并活动性丙型肝炎病毒感染者有较低的ImmuKnow值(<61 ATP ng/ml).结论 Cylex ImmuKnow检测可以用于稳定肝移植患者免疫抑制的监恻,具有潜在的临床应用价值.综合评价肝移植患者免疫状态需要联合ImmuKnow、T淋巴细胞亚群、血药浓度、肝功能指标检测等相关检查,才能有效指导肝移植受体免疫抑制剂的个体化使用.

Abstract：

Objective To identify the level of functional immunity as measured by the ImmuKnow assay in Chinese stable liver transplant recipients and to correlate these values with the dose and the trough levels of immunosuppressant, and with other clinical parameters of these patients. Methods Functional immune response was assessed by the ImmuKnow assay in 46 blood samples taken from 46 stable liver transplant recipients from Beijing Youan Hospital, Capital Medical University Liver Transplantation Center. Results The average ATP value in these stable liver transplant recipients was 203±114 ng/ml (range: 38.47 ATP ng/ml to 524.06 ATP ng/ml) at 22± 15 month post liver transplantation. There was no correlation either between ImmuKnow ATP values and the tacrolimus trough levels, or between ImmuKnow ATP values and the liver function (P<0. 05). Stepwise multiple regression analysis identified WBC and CD3+, CD3+ CD4+ as independent predictors of ImmuKnow assay levels when age, gender and underlying diagnosis were taken into account (P<0. 05). Five patients who were detected to have active HCV infection had lower ImmuKnow ATP values (<61 ng/ml). Conclusions The Cylex ImmuKnow assay ATP values were lower in Chinese stable liver transplant recipients compared with American patients. Further investigation is required to determine the role of the ImmuKnow assay in tailoring immunosuppressant therapy in liver transplant recipients.     

肝移植术后ImmuKnow细胞免疫功能与白细胞及T淋巴细胞计数的相关性研究

目的 探讨肝移植术后ImmuKnow细胞免疫功能测定值与白细胞分类计数和T淋巴细胞亚群计数的相关性,为临床提供一种价格低廉快速判断肝移植受者细胞免疫功能的方法.方法 选择49例行经典原位肝移植术受者术后2周至2个月内在无糖皮质激素应用情况下的外周血样本.分析ImmuKnow测定值与白细胞分类计数和T淋巴细胞亚群计数的相关性.并随机选择5例无激素免疫抑制的移植受者于术后2、3、4、6、8周分别重复检测上述指标,进一步验证其相关性.结果 白细胞总数与ImmuKnow ATP值相关性最高,相关系数为0.821;中性粒细胞计数与ImmuKnow ATP值相关性次之,相关系数为0.787;单核细胞计数相关系数虽然有统计学意义,但相关系数低于0.5.淋巴细胞计数和淋巴细胞亚群计数与ImmuKnow ATP值的相关性无统计学意义.5例无激素免疫抑制受者术后重复检测ImmuKnow ATP值的变化与细胞总数的变化呈正相关,相关系数均>0.5.结论 肝移植术后早期白细胞计数与CD4+T细胞ImmuKnow ATP值具有一定的正相关性,白细胞计数的变化,可以在一定程度上反映ImmuKnow ATP值的变化.

Abstract：

Objective To explore the relationship between peripheral differential blood count and ATP value in Cell CD4 + T tested by ImmuKnow method in liver transplants. Methods In this study 49recipients after classic orthotopic liver transplantation (OLT) were enrolled. In a period from two weeks to two months after transplantation when all were free of glucocorticoid. Blood were sent for WBC differential samples count and ATP value in Cell-CD4 + T tested by ImmuKnow method via SPSS17. 0 software. Five more samples were selected randomly for duplicated testing of the indices in Week2, 3, 4,6 and 8 after the transplanting operation to further verify the relativity. Results White blood cell count has the highest relativity with ImmuKnow ATP value at 0. 821. The 5 recipients were repeatedly tested for ImmuKnow ATP values that were found positively correlated to cell count with a coefficient of over 0. 5. Conclusions The peripheral leukocyte count in early stage after liver transplantation is in positive correlation with ATP value in Cell CD4 + T, and the changes of numeration of leukocyte reflect changes of ATP value.     

Prolonged survival of donor-specific rat intestinal allograft by administration of bone-marrow-derived immature dendritic cells

It has been reported that intraportal administration of donor antigens induced donor-specific hyporesponsiveness. We studied here the effects of transplantation of BM-derived immature dendritic cells (imDCs) and mature DCs (mDCs) via portal vein on rat small intestinal allograft survival. This study comprised four treatment groups: 1) untreated controls; 2) FK506 alone; 3) intraportal donor-specific BM-derived imDCs transplantation+FK506; 4) mDCs/Tx+FK506. Allograft survival was minimal in control group (5.2+/-0.8 days) and maximal in imDC+FK506 group (28.4+/-3.0 days). The rats in mDC+FK506 group showed systemic inflammatory reaction due to GVHR, and died within 10 days after transplantation. The in vitro MLR reaction using imDCs was also strongly inhibited both in direct and indirect recognition pathways. The impact of imDCs for the specific induction of transplant tolerance may suggest that immunization with donor-specific imDCs has therapeutic potential in organ transplantation.     

Role of CD4+ and CD8+ T cells in early and late acute rejection of small bowel allograft

BACKGROUND/PURPOSE: Results of small bowel transplantation remain unsatisfactory because of severe immune rejection. The current study aims to elucidate the role of activation of CD4+ and CD8+ T cells in early and late acute rejection of small bowel allograft and, hence, provide the immunologic basis for developing new therapeutic strategies. METHODS: We used an MHC fully mismatched (DA to Lewis) heterotopic rat small bowel transplant model and a unique FK506-based immunosuppressive regimen, which suppresses early acute rejection but does not prevent late acute rejection. Flow cytometric analysis was used to quantitate the number of activated CD4+ and CD8+ T cells in graft and host mesenteric lymph nodes. RESULTS: The survival (mean +/- SD) of intestinal allograft was significantly prolonged, from 6.6 +/- 0.84 days for the untreated group to 40.7 +/- 14.1 days for the FK506-treated group. Activation of CD4+ cells was suppressed significantly in the FK506-treated group on postoperative day 7 compared with the untreated group (29.4% +/- 3.55% v 52.83% +/- 11.9%; P <.01). Activation of CD8+ cells was similarly suppressed (31.5 +/- 10.34% v 48.53 +/- 14.34%; P <.05). Interestingly, at late acute rejection, activated CD4+ and CD8+ T cells remained at almost the same low levels as those on postoperative day 7 in the FK506-treated group. The spleen to body weight ratio was significantly increased in the untreated group (0.53 +/- 0.07), and slightly increased in the FK treated group (0.27 +/- 0.07, on postoperative day 7; 0.24 +/- 0.07 at late acute rejection) compared with the syngeneic group (0.18 +/- 0.02). CONCLUSION: The activation of CD4+ and CD8+ T cells was suppressed effectively by early potent immunosuppressive treatment resulting in prolonged survival of intestinal allograft. At late acute rejection, the CD4+ and CD8+ T cells remained at low-level activation status, in contrast to the surge of CD4+ and CD8+ activation during early acute rejection. This suggests that persistent T cell activation even at low level is sufficient to cause the late acute rejection eventually. A therapeutic strategy targeting these cells is needed for long-term engraftment.     

Steroid withdrawal 3 months after liver transplantation - does FK 506 confer any advantage over cyclosporin?

Abstract Eighty-one liver recipients were randomised to FK 506 Or cyclosporin (CyA) and azathioprine (AzA), both in combination with steroids. Twenty-even FK 506 and 29 CyA/AzA patients continued in the trial 3 months after transplantation. Steroids were ceased in 23 (85%) FK 506 patients and in 27 (93%) CyA patients. After steroid withdrawal, 2 FK 506 and 4 CyA patients were excluded from the study, all for reasons other than rejection. The median follow-up was 16 months for the FK 506, and 19 months for CyA group. There were no acute rejection episodes or graft losses in the FK 506 group. None of the CyA patients lost their graft but three (13%) had episodes of acute rejection requiring steroids to be recommenced in two cases. There was no evidence of chronic rejection in any of the annual review biopsies in either group. Our results suggested no advantage of FK 506 over CyA in its steroid-paring effect.     

A 39-month follow-up study to evaluate the safety and efficacy in kidney transplant recipients treated with modified-release tacrolimus (FK506E)-based immunosuppression regimen

Background

We initially performed a study to evaluate the safety and efficacy of modified-release tacrolimus (FK506E) in a phase 3, 2-arm, 6-month, randomized, open-label, multicenter trial in Korean living donor de novo kidney transplant recipients. We then performed an extended study to evaluate the long-term safety and efficacy of a FK506E-based regimen up to 45 months posttransplantation in recipients already treated with FK506E.

Methods

Initial study was designed as a randomized, open-label, comparative, multicenter study in de novo living donor kidney transplant recipients. The patients were randomized to an FK506E versus a control (FK506) group (1:1). Recipients who completed a 6-month FK506E treatment study were enrolled in the 39-month follow-up study. Primary end-points were patient and graft survivals at posttransplantation 45 months. Secondary end-point was the incidence of a clinical or biopsy-proven acute rejection episode between 6 and 45 months posttransplantation.

Results

In the initial 6-month de novo study 124 enrolled patients were randomized into either the FK506E (n = 62) or the control group (n = 62). The incidence of an acute rejection episode was 19.4% (n = 12) in the FK506E versus 16.1% (n = 10) in the control group (P = .638). There was no mortality or graft failure among the 44 recipients enrolled in this additional 39-month follow-up study. There was 1 patient with biopsy-proven acute cellular rejection episode (2.3%) who underwent steroid pulse therapy with renal function recovery. At the time of study completion 40/44 recipients (90.9%) maintained FK506E treatment.

Conclusion

This 39-month study following the initial 6-month FK506E study period showed an FK506E-based immunosuppressive regimen in living donor kidney transplantation recipients to be safe and effective.     

A prospective randomized trial comparing the efficacy of tacrolimus versus cyclosporine in black recipients of primary cadaveric renal transplants.

BACKGROUND: We performed a prospective randomized trial to compare the efficacy and safety of tacrolimus (FK506) versus cyclosporine (CSA) in black primary cadaveric renal transplant (CRT) recipients. METHODS: Between December 1994 and February 1997, 35 black primary CRT recipients were enrolled in this trial. All patients received 7 days of induction therapy with OKT3. Fourteen patients received FK506 and prednisone only. Twenty-one patients received CSA, azathioprine, and prednisone. The two groups were comparable in terms of age, gender, plasma renin activity, human leukocyte antigen mismatches, and cause of renal failure. RESULTS: Patient and graft survival were 12 of 14 (86%) for the FK506 group and 20 of 21 (95%) for the CSA group (P = 0.71). Three patients died owing to cardiac events with functioning grafts. Acute rejection was 2 of 14 (14%) for the FK506 and 8 of 21 (38%) for the CSA group (P = 0.25). Two other patients on CSA were converted to FK506 as rescue for OKT3-resistant rejection. Mean serum cholesterol at 1 year was 198 +/- 45 mg/dL for the FK506 group and 244 +/- 49 mg/dL for the CSA group (P = 0.03). Mean serum creatinine at 1 year was 1.39 +/- 0.38 mg/dL for the FK506 group and 1.94 +/- 0.64 mg/dL for the CSA group (P = 0.02). CONCLUSION: Patient and graft survival were similar in both groups at 1 year posttransplant. Although statistically not significant, the incidence of acute rejection was lower in the FK506 group. Furthermore, FK506-treated patients had significantly lower serum creatinine and cholesterol levels at 1 year posttransplant.     

Assessing relative risks of infection and rejection: a meta-analysis using an immune function assay

BACKGROUND: Long-term use of immunosuppressants is associated with significant morbidity and mortality in transplant recipients. A simple whole blood assay that has U.S. Food and Drug Administration clearance directly assesses the net state of immune function of allograft recipients for better individualization of therapy. A meta-analysis of 504 solid organ transplant recipients (heart, kidney, kidney-pancreas, liver and small bowel) from 10 U.S. centers was performed using the Cylex ImmuKnow assay. METHODS: Blood samples were taken from recipients at various times posttransplant and compared with clinical course (stable, rejection, infection). In this analysis, 39 biopsy-proven cellular rejections and 66 diagnosed infections occurred. Odds ratios of infection or rejection were calculated based on measured immune response values. RESULTS: A recipient with an immune response value of 25 ng/ml adenosine triphosphate (ATP) was 12 times (95% confidence of 4 to 36) more likely to develop an infection than a recipient with a stronger immune response. Similarly, a recipient with an immune response of 700 ng/ml ATP was 30 times (95% confidence of 8 to 112) more likely to develop a cellular rejection than a recipient with a lower immune response value. Of note is the intersection of odds ratio curves for infection and rejection in the moderate immune response zone (280 ng/ml ATP). This intersection of risk curves provides an immunological target of immune function for solid organ recipients. CONCLUSION: These data show that the Cylex ImmuKnow assay has a high negative predictive value and provides a target immunological response zone for minimizing risk and managing patients to stability.     

Measurement of CD4+ T‐cell function in predicting allograft rejection and recurrent hepatitis C after liver transplantation

Hashimoto K, Miller C, Hirose K, Diago T, Aucejo F, Quintini C, Eghtesad B, Corey R, Yerian L, Lopez R, Zein N, Fung J. Measurement of CD4+ T‐cell function in predicting allograft rejection and recurrent hepatitis C after liver transplantation.
Clin Transplant 2009 DOI: 10.1111/j.1399‐0012.2009.01169.x
© 2009 John Wiley & Sons A/S. Abstract: Recurrence of hepatitis C virus (HCV) can be difficult to distinguish from acute cellular rejection (ACR) following liver transplantation. The Cylex Immune Function Assay (ImmuKnow) provides objective measure of recipient’s immune function. The goal is to assess the ability of this assay to distinguish these similar conditions. A retrospective review was performed in 54 recipients with HCV. ImmuKnow assays were measured with allograft biopsies. Levels of adenosine triphosphate (ATP) release from CD4+ T cells (ng/mL) were compared with the following biopsy result classifications: 365 ± 130 with ACR (n = 11), 152 ± 100 with recurrent HCV (n = 26), 240 ± 71 with normal biopsies (n = 12), and 157 ± 130 with overlapping features of ACR and recurrent HCV (n = 5). Recipients with recurrent HCV had lower immune response than those with ACR (p < 0.0001).Using a cutoff level of 220, the sensitivity and specificity for distinguishing two conditions were 88.5% and 90.9%, respectively. When recipients with overlapping features had low immune response, three of four recipients’ subsequent biopsies showed recurrent HCV. In conclusion, the ImmuKnow assay can be a sensitive and specific additional test for distinguishing recurrent HCV from ACR and may be useful for predicting which recipients may be most vulnerable to recurrent HCV.     

肾移植受者应用他克莫司治疗窗浓度的探讨

目的　寻求适合国人肾移植受者他克莫司 (FK5 0 6 )理想治疗窗浓度范围。方法　应用微粒子酶免疫分析法测定 5 8例肾移植患者口服FK5 0 6后 12h的血药谷浓度 ,并观察排斥反应的发生及药物的肾毒性。结果　FK5 0 6的血药浓度 ,术后 1个月为 (13.0± 2 .1) μg/L ,2～ 3个月为 (9.4±1.6 ) μg/L ,3个月以后为 (6 .5± 1.3) μg/L ,比较各时期全血FK5 0 6谷浓度 ,差异均有极显著性 (P <0 .0 1) ;术后发生急性排斥反应 3例次 ,肾毒性 4例次。结论　FK5 0 6具有良好的免疫抑制效果 ,其治疗窗浓度范围 ,术后第 1个月为 11～ 15 μg/L ,第 2～ 3个月为 8～ 11μg/L ;第 3个月后为 5～ 8μg/L ,此浓度范围既能达到满意的免疫抑制效果 ,又能减少FK5 0 6的肾毒性     

两种包含他克莫司的免疫抑制方案在肾移植中的应用体会

目的　探讨适合中国人的他克莫司 (FK50 6)剂量。方法　对 8例使用高剂量FK50 6(0 .2mg·kg- 1 ·d- 1 )方案肾移植受者 (A组 )及 34例使用低剂量FK50 6(0 .1mg·kg- 1 ·d- 1 )受者 (B组 )术后早期血肌酐、排斥反应发生率及免疫抑制剂的不良反应进行比较。结果　A、B两组术后 2个月血肌酐水平的差异无显著性 (P >0 .0 5) ;A组半年内未见急性排斥反应发生 ;B组有 1例因血中FK50 6的浓度偏低而发生急性排斥反应 ,2例肾组织活检提示肾小管炎 ;两个组在随访期内均未见严重感染发生 ;药物的不良反应B组较少、较轻。结论　应用低剂量FK50 6组成的免疫抑制方案 ,在降低药物毒副作用的同时 ,并不影响其疗效     

他克莫司与来氟米特对异种胰岛移植术后T细胞亚群的影响

目的 研究他克莫司(FK506)和来氟米特(Lef)对异种胰岛移植后急性排斥反应的抑制作用。方法 将大鼠胰岛移植于糖尿病小鼠肾被膜下。观察移植后不用药对照组、单独应用FK506组、单独应用Lef组以及联合应用FK506和Lef组的胰岛存活情况；并于移植后第5d检测外周血T细胞亚群分布和血清IL-2含量。结果 单独用药组与不用药对照组比较，胰岛存活时间明显延长；联合用药组又明显长于单独用药组。各用药组外周血CD4 T淋巴细胞明显低于未用药组。结论 FK506与Lef通过抑制CD4 T淋巴细胞的增殖而阻止或延缓异种胰岛移植的急性排斥反应。     

采用程序化肾活检评估肾移植后低水平和标准水平他克莫司的疗效

目的以程序化肾活检评估低水平与标准水平他克莫司(FK506)的免疫抑制方案的疗效及其安全性。方法采用前瞻性、开放、随机对照研究,将48例首次接受尸体肾移植受者按随机数字分为两组:低水平FK506组(低FK506组,24例)和标准水平FK506组(标准FK506组,24例)。两组患者均采用麦考酚吗乙酯(MMF)+FK506+肾上腺皮质激素(激素)的三联免疫抑制方案,两组的MMF与激素用法相同。标准FK506组的FK506血药谷浓度:在入组后前3个月维持在10～12 ng/ml,3个月后维持在8～10 ng/ml。低FK506组的FK506血药谷浓度:入组后头2个月为8～10 ng/ml,第3个月为3～7 ng/ml,3个月后为3～5 ng/ml。术后定期随访1年,内容包括测定FK506血药谷浓度,检测肾功能[血清肌酐(Scr)、内生肌酐清除率(endogenous creatinine clearancerate,Ccr)]、空腹血糖、糖化血红蛋白、血清白蛋白、血脂。同时于术后3个月和12个月予以程序化移植肾活组织检查(活检),了解急性排斥反应(AR)发生情况、病理损伤指标评分及慢性移植物损伤指数(chronic allograft damage index,CADI)变化。观察术后1年的人、肾存活率及不良事件发生情况。结果移植术后1年,与标准FK506组比较,低FK506组:(1)FK506血药谷浓度较低(P<0.01),且达到目标水平的患者所占比例较高;(2)Ccr水平较高,(83±14)ml/min比(62±16)ml/min,P<0.05;(3)血糖水平较低(P<0.05);(4)根据程序化肾活检的结果,间质纤维化、肾小管萎缩和肾小球硬化评分较低(均为P<0.05);(5)AR发生率及其严重程度相似,术后1年人、肾存活率相近,均为100%(均为P>0.05)(6)术后1年的肺部感染与新发糖尿病的发生率较低(分别为P<0.05和P<0.01)。结论采用程序化肾活检评估疗效结果可靠,并有助于发现移植肾的慢性化改变,移植术后早期适度降低FK506血药谷浓度对于移植肾和患者均有较大益处,且不增加发生排斥反应的风险。     

FK506和RS-61443对大鼠异体肢体移植的联合免疫抑制作用

目的　通过大鼠异体肢体移植模型 ,旨在分析 FK5 0 6和 RS- 6 14 4 3对大鼠异体肢体移植中急性排斥反应的免疫抑制作用。　方法　选择雄性 Wistar和 SD大鼠为供、受体 ,以 FK5 0 6和 RS- 6 14 4 3为免疫抑制剂 ,对照组为术后不用药组 ,实验组根据用药剂量和药物不同分为 6组 ,各组用药时间均为 5周 (每日 1次共 2周 ,然后每周 2次共 3周 ) ,进行了 10 1例异体肢体移植动物实验。观察大鼠一般情况、移植肢体排斥反应及存活时间。　结果　对照组肢体平均存活时间为 (7.0 0± 0 .78)天 ;实验组 1～ 6组移植肢体平均存活时间分别为 (17.0 8± 4 .5 0、2 3.2 0± 5 .0 5、11.19±2 .2 8、16 .33± 1.83、13.33± 3.2 2和 5 8.76± 6 .81)天。　结论　 FK5 0 6和 RS- 6 14 4 3能抑制大鼠同种异体肢体移植术后急性移植排斥反应的发生 ,并能延长移植肢体的存活时间。     

Development of donor-specific immunoregulatory T-cells after local CTLA4Ig gene transfer to pancreatic allograft

BACKGROUND: CTLA4Ig gene transfer directly to graft tissue might have the potential to avoid the need for systemic immunosuppression. In our previous studies of bio-breeding (BB) rats, local adenovirus-mediated CTLA4Ig gene transfer protected the pancreas from autoimmune and alloimmune responses. This study investigated the potency of local CD28/B7 costimulatory blockade for induction of donor-specific tolerance and further examined the existing mechanisms. METHODS: Brown Norway (BN; RT1)-pancreaticoduodenal grafts transfected with Ad.CTLA4Ig via intraarterial ex vivo perfusion were transplanted into streptozotocin-induced diabetic Lewis (LEW; RT1) rats. RESULTS: Ad.CTLA4Ig transduced grafts combined with a short course of FK506 resulted in indefinitely prolonged survival (>156 days vs. 19.5 days with FK506 alone). CTLA4Ig was predominantly expressed in grafts on day 4. The expression was gradually diminished and was only slightly detectable at day >100. The proliferative responses against BN antigen were remarkably enhanced among recipients with rejected grafts, but the T-cells from tolerant recipients (>100 days) showed poor cytotoxic responses. On adoptive transfer assay, the splenic T-cells of tolerant recipients were able to suppress the rejection of BN, but not third-party Wistar Furth (WF; RT1) hearts in irradiated (480 cGy) LEW recipients. The percentage of CD4CD25 splenic T-cells was significantly increased in tolerant recipients (13.53 +/- 4.06% vs. 6.06 +/- 0.56% in naive rats). CONCLUSION: CTLA4Ig gene transfer to the pancreaticoduodenal allograft combined with a short course of FK506 induces donor-specific tolerance. The mechanism of maintaining tolerance could be explained by development of splenic T suppressor cells.