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。     

ImmuknowTM法检测细胞免疫功能在老年肾移植中的应用52例

目的 探讨CD4+T淋巴细胞内腺苷三磷酸(ATP)含量(ImmmuknowTM法)检测细胞免疫功能在老年肾移植中的应用价值.方法 采用ImmuknowTM免疫细胞功能监测仪通过生物发光法测定52例老年肾移植受者移植前后全血CD4+T淋巴细胞ATP含量的变化.结果 52例中,33例术后移植肾功能稳定(稳定组),未发生感染或排斥反应,其ATP值在相对稳定的水平,与普通尿毒症患者(对照组)比较,差异无统计学意义(P＞0.05);11例出现肺部感染和皮肤疱疹病毒感染(感染组),跟踪监测其ATP值进行性降低,其中4例ATP值降至100 μg/L以下,出现重症肺部感染,与稳定组比较,差异有统计学意义(P＜0.01);8例出现急性细胞性排斥反应(AR组),跟踪监测其ATP值进行性升高,与稳定组和感染组比较,差异有统计学意义(P＜0.01).结论 术后动态监测老年肾移植受者ImmuknowTM ATP水平的变化,有助于了解术后细胞免疫状态,以利于预防排斥反应和感染等.     

肾移植术后并发巨细胞病毒肺炎患者的细胞免疫状态

目的 探讨CD4+T淋巴细胞ATP含量检测在肾移植术后并发巨细胞病毒(CMV)肺炎治疗中应用价值.方法 以187例首次肾移植受者作为研究对象,分别于术前,术后30、60、90和180d,发生CMV肺炎时,以及治疗4周后采集受者外周血,应用ImmuKnowTM免疫细胞功能测定试剂盒检测CD4+T淋巴细胞内ATP含量.采用方差分析对不同检测时间点及术后有无并发CMV肺炎者的外周血CD4+T淋巴细胞ATP含量进行比较,采用Pearson-Spearman秩和检测对ATP含量与感染的相关性进行分析.结果 187例受者中发生CMV肺炎17例,发生率为9.1 ％(17/187),发生时间为术后(2.8±1.2)个月.术后所有时间点CD4+T淋巴细胞ATP含量均明显低于术前(P＜0.01),ATP含量在术后90d时达最低点,与术后其他时间点比较,差异有统计学意义(P＜0.05).发生CMV肺炎者术前外周血CD4+T淋巴细胞ATP含量为(376±182) μmol/L,术后30和90 d分别为(283±146) μmol/L和(196±112) μmol/L,发生CMV肺炎时和治疗4周后分别为(145士102)μmol/L和(236±117) μmol/l,发生CMV肺炎时ATP含量与其他各个时间点比较,差异均有统计学意义(P＜0.05).相关分析表明,CD4+T淋巴细胞内ATP含量降低与CMV肺炎的发生具有显著相关性(相关系数=0.5106,P＜0.01).结论 肾移植后测定受者外周血CD4+T淋巴细胞ATP含量,可反映受者的细胞免疫状态及判断CMV肺炎的严重程度和临床预后,并可指导CMV肺炎的治疗.     

Assessing immunologic function through CD4 T-lymphocyte ahenosine triphosphate levels by ImmuKnow assay in Chinese patients following renal transplantation

Objective

Balancing immunosuppression to prevent rejection while minimizing infection/drug toxicity risk is a challenge in organ transplantation. Drug monitoring alone or with functional monitoring is inadequate to measure the immune response after transplantation. The Food and Drug Administration (FDA)-approved immune monitoring assay, ImmuKnow, offers an noninvasive method to assess the immune status of transplanted patients by measuring adenosine triphosphate (ATP) released from CD4 T cells. Herein, we have evaluated ATP levels reflecting the immune responses of Chinese kidney transplant recipients as a monitoring parameter to guide treatment after transplantation.

Methods

From October 2008 to March 2010, we recruited 259 kidney transplant patients who were divided into four groups: stable (n = 174), postoperative infection (n = 32), postoperative rejection (n = 16), and high-dose corticosteroid treatment (n = 33). The ImmuKnow assay was performed to measure CD4 T-cell ATP levels. No prisoners or organs from prisoners were used in the study.

Results

Receiver operating characteristics measurements indicated an ATP predictive range of 238 to 497 ng/mL to monitor immune responses after transplantation and immunosuppressive therapy. To identify patients with infection, we used a cutoff ATP value of 238 ng/mL with 100% specificity and positive predictive value and 92.9% sensitivity. To identify patients with rejection, we used a value of 497 ng/mL with 91.5% sensitivity. Compared with the 225 to 525 ng/mL ATP levels recommended by the FDA, our target values showed similar or better diagnostic accuracy.

Conclusion

We provide additional data to monitor immunosupressant treatment of Chinese kidney transplant patients.     

Gonadal function and immunosuppressive therapy after renal transplantation

End-stage renal disease is associated with disorders in hypothalamic-pituitary-gonadal function. Immunosuppressive therapies may influence the restoration of normal levels of gonadal hormones after renal transplantation. The aim of the present study was to evaluate the hormonal status of successful renal transplant recipients who were treated with different immunosuppressive agents.

Methods

Testosterone, luteinizing hormone (LH), and follicle stimulating hormone (FSH) were measured in 59 male renal transplant recipients with stable graft function with serum creatinine <2.5 mg/dL. Patients were treated with three different immunosuppressive regimens: group I, calcineurin inhibitors (CI; n = 15), group II, sirolimus without calcineurin inhibitors (SRL; n = 15), group III, sirolimus in combination with calcineurin inhibitors (SRL * CI; n = 29).

Results

Testosterone was significantly lower in group II versus group I (3.12 ± 1.23 versus 4.39 ± 1.53 ng/mL; P < .0197). Group III had higher testosterone values than group II, but lower than group I. FSH and LH were also higher in the SRL group, but the differences were not statistically significant, perhaps because of the small number of patients. No relationship was found between testosterone blood levels and age, posttransplant follow-up, renal function, time on dialysis, body mass index, steroid use, or posttransplant diabetes.

Conclusion

Sirolimus seems to impair the improvement of gonadal function after renal transplantation. Further prospective studies are needed to confirm these data before patients are advised of this potential side effect.     

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.     

Drawbacks of the use of indirect estimates of renal function to evaluate the effect of risk factors on renal function

Many epidemiologic studies presently aim to evaluate the effect of risk factors on renal function. As direct measurement of renal function is cumbersome to perform, epidemiologic studies generally use an indirect estimate of renal function. The consequences of using different methods of renal function measurement in studies that evaluate the effect of cardiovascular risk factors on renal function were questioned. Data of the 8592 Prevention of Renal and Vascular End-stage Disease study participants, in whom the association was plotted between various cardiovascular risk factors and renal function measured either by creatinine clearance based on two 24-h urine collections or by the Cockcroft-Gault or Modification of Diet in Renal Disease formula were used. A repeated measurement analysis was used to compare the slopes of the linear regression lines of the risk factors and the different methods of renal function measurements. The relation between cardiovascular risk factors and renal function seems to be different when different methods for renal function are used. This was most pronounced for age, weight, and body mass index and less pronounced (but still statistical significant) for BP, cholesterol, and glucose. The relation between weight or body mass index and renal function showed completely different directions, depending on the renal function method used. In conclusion, the interpretation of the relation of cardiovascular risk factors and renal function is affected by the method selected to estimate renal function. For studying the relation of risk factors and renal function in large population studies, indirect estimates of renal function should be used with caution.     

肾移植术后肺孢子菌肺炎感染与CD4细胞计数的相关性

目的 分析肺孢子菌肺炎感染与CD4细胞计数的相关性。方法 对武汉大学人民医院器官移植科2019年4月至2020年3月收治的134例肾移植术后1年内患者,其中54例患者确诊感染肺孢子菌肺炎,检测CD4细胞数量。分析肺孢子菌肺炎发病率与CD4细胞计数是否具有相关性,再根据CD4细胞计数分成3组,以t检验分别分析两组患者的住院天数、吸氧天数、肺部炎症吸收天数等相关指标,探究CD4细胞计数与肺孢子菌肺炎转归与预后之间的关系。结果 134例病例中有53例患者感染肺孢子菌肺炎,3组患病率分别0.237、0.720、0.813。根据分组分别用住院时间、吸氧时间、病灶吸收时间两两进行t检验,P值分别为0.667、0.517、0.779、0.335、0.863、0.150、0.404、0.139、0.405。结果显示均无统计学差异。结论 在肾移植患者术后,肺孢子菌肺炎的发病率随着CD4细胞计数的减少而上升,但CD4细胞计数与疾病预后没有明显关联。     

心脏死亡器官捐献供肾热缺血因素对移植肾功能的影响

目的结合心脏死亡器官捐献(DCD)肾移植受者术后随访情况,探讨供肾热缺血因素对移植肾功能的影响。 方法回顾性分析2011年5月至2015年6月浙江大学医学院附属第一医院肾脏病中心施行的肾移植术供、受者临床资料。移植术后1年根据受者估算肾小球滤过率(eGFR)≥60 mL·min^－1·(1.73 m²)^－1和<60 mL·min^－1·(1.73 m²)^－1将受者分为高肾功能组与低肾功能组,最终纳入340例受者,其中高肾功能组259例,低肾功能组81例。根据DCD供者手术记录表,整理分析两组供者不同收缩压(SBP)及血氧饱和度(SpO₂)热缺血时间段。符合正态分布计量资料以均数±标准差( ±s)表示,采用t检验比较高、低肾功能组供、受者一般资料、供者不同SBP和SpO₂热缺血时间段差异;采用Wilcoxon符号秩和检验比较两组受者供肾获取时肾小球和肾小管病理评分。计数资料以百分数表示,采用卡方检验比较两组供、受者性别和移植肾功能延迟恢复(DGF)发生率。P<0.05为差异有统计学意义。 结果截至2016年6月,所有受者随访(28.4±2.8)个月(13.1～62.5个月)。术后1年内高、低肾功能组DGF发生率分别为14.7%(38/259)、22.2%(18/81),差异无统计学意义(χ²＝2.557,P>0.05)。高肾功能组平均年龄和BMI均低于低肾功能组,男性比例和捐献时eGFR高于低肾功能组,差异均有统计学意义(t＝－6.363、－2.049、4.190, χ²＝4.863,P均<0.05);高肾功能组供肾获取时肾小球病理评分低于低肾功能组,差异有统计学意义(Z＝－2.606,P<0.05)。高肾功能组受者年龄小于低肾功能组,而男性比例高于低肾功能组,差异均有统计学意义(t＝－2.790, χ²＝9.658,P均<0.05)。高、低肾功能组初始SpO₂降低40%、撤除生命支持至SpO₂测不出以及90%、80%、70%、60%初始SpO₂至SpO₂测不出的平均时间分别为(5.9±4.3)和(4.8±3.3)、(8.0±5.2)和(6.1±4.4)、(4.5±3.6)和(3.5±2.8)、(4.0±3.7)和(2.9±2.4)、(4.0±3.6)和(2.8±2.7)、(3.6±3.5)和(2.4±2.5) min,差异均有统计学意义(t＝2.088、2.983、2.328、2.622、2.557、2.759,P均<0.05)。高、低肾功能组初始SpO₂降低10%、40%、撤除生命支持至SpO₂测不出、60%初始SpO₂至SpO₂测不出平均变化速率分别为(2.40±1.78)和(2.90±1.70)、(8.71±6.96)和(15.01±12.97)、(19.60±17.49)和(25.80±22.85)、(22.41±15.94)和(29.93±19.36) %/min,差异均有统计学意义(t＝－2.230、－5.647、－2.577、－3.514,P均<0.05)。 结论DCD肾移植预后受供、受者年龄、BMI等一般因素影响。供者高、低SpO₂时间段长短及变化速率与移植肾功能相关,DCD供肾移植过程中可通过优化手术流程等方法缩短低SpO₂时期时间,以减少供肾热缺血损伤,改善受者预后。     

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.     

抗体诱导免疫抑制对同种异体肾移植术后肾功能恢复的影响

为探讨同种异体肾移植术使用抗体诱导免疫抑制方案对移植肾术后早期功能恢复的影响，我们对相关病例进行了研究，现报告如下。     

前列腺素E1对大鼠肝移植肾功能的保护作用

目的探讨术中应用前列腺素E1（prostaglandin E1，PGE1）对大鼠肝移植肾功能的保护作用。方法大鼠原位肝移植术中经颈内静脉灌注PGE1为治疗组，生理盐水和空白为对照组，观察术后1周存活率、1h的尿量，测定血浆肌酐、尿素氮和肾组织中丙二醛（malondjaldehyde，MDA）、谷胱甘肽（glutathione，GSH）含量，肾组织病理检查。结果PGE1治疗组术后1h尿量较对照组明显增加，肌酐和尿素氮水平均较对照组降低，PGE1治疗组肾组织中GSH含量显著高于两对照组，MDA含量低于两对照组。病理检查PGE1治疗组肾脏组织形态学损伤明显减轻。结论术中应用PGE1能显著改善大鼠肝移植后的肾功能，其机制可能与对抗氧自由基损伤作用有关。     

个体化免疫诱导方案在肾移植中的应用进展

肾移植术后早期急性排斥反应发生的风险较高,严重影响受者的生存质量。2009年,改善全球肾脏病预后组织（KDIGO）建议将免疫诱导药物纳入肾移植术前免疫诱导方案中,其目的就是针对这一关键时期提供一定强度的免疫抑制,从而有效减少术后急性排斥反应的发生。目前全球各移植中心对于免疫诱导药物的选择及其有效性、安全性仍不确定。本文通过汲取国内外学者的研究成果,对比分析单克隆抗体包括白细胞介素-2受体拮抗剂、阿伦单抗、利妥昔单抗及多克隆抗体抗胸腺细胞球蛋白在肾移植术前免疫诱导中的应用效果,旨在为推动肾移植免疫诱导药物的个体化选择,提高受者的生存质量提供参考。     

Nonsustained effect of short-term bisphosphonate therapy on bone turnover three years after renal transplantation

BACKGROUND: We recently showed that two doses of 4 mg of zoledronic acid (ZOL) ameliorated the bone loss and improved bone histology within the first six months after kidney transplantation. The aim of the present study was to evaluate whether this early short-term intervention exhibited a sustained bone-sparing effect. METHODS: A homogenous group of 20 de novo renal transplant recipients were equally randomized to two infusions of 4 mg of ZOL or placebo at two weeks and three months after engraftment. Patients were followed up for three years by sequential determination of bone densitometry and specific biochemical markers. RESULTS: From month six to three years after transplantation, both treatment groups exhibited an improvement of bone mineralization. Femoral neck bone mineral density z-scores increased statistically significantly from -1.3 (2.6) to -0.2 (3.6) in the placebo group and from -1.6 (2.9) to -1.2 (1.9) in the ZOL group (median, range). Biochemical parameters of osteoblast activity such as osteocalcin and bone-specific alkaline phosphatase did not increase significantly in both groups. Osteoprotegerin, a marker of osteoclast inhibition, was significantly elevated over the first six months in the ZOL group, but decreased to similar levels, as in the placebo group, over the next two and a half years. Other markers of osteoclast activity such as c-telopeptide of type 1 collagen, calcitonin, and intact parathyroid hormone were not different between six months and three years in either group. CONCLUSION: The early bone-sparing effect of short-term ZOL therapy confers no sustained benefit versus placebo at three year post-transplantation.     

The effect of renal transplantation on pulmonary function.

In patients with chronic renal failure, mechanical and hemodynamic changes could occur in the lungs without obvious pulmonary symptoms and findings and their effects could pave the way to pulmonary functional disorders. In this study, pulmonary functional disorders and especially alveolocapillary defects, which are frequently seen in uremia, were determined in renal transplanted patients. Pulmonary functions and diffusion capacity were assessed in uremic patients (n = 20) and in successfully transplanted patients (n = 20) without any lung disease or pulmonary edema symptoms and findings. Patients were selected randomly among outpatients who were followed up in a Nephrology and Transplantation Unit. Forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1), and peak expiratory flow (PEF25-75) were measured. Single breath carbon monoxide diffusion test and diffusion lung capacity adjusted for hemoglobin concentration (DLAdj) were done. The means of the spirometric values such as FVC, FEV1 and FEV1/FVC were normal in the nondialyzed uremic group, but the PEF25-75 value (68.7%) and diffusion capacity (DLAdj 72.7%) were found to be slightly low. There were 2 patients with normal values and 18 patients with some functional abnormalities in this nondialyzed uremic group. The means of all spirometric parameters and diffusion capacities were found to be normal in the transplanted group. There were 7 patients with normal function and 13 patients with some functional abnormalities in this transplanted group. When the nondialyzed uremic group and the transplanted group were compared statistically, significant differences were found between their spirometric values (except for FVC) and their diffusion capacities. Even though the uremic patients did not show any symptoms, their pulmonary function tests, especially diffusion capacity, were found to be disturbed. Although the transplanted patients as a group had normal mean spirometric values and diffusion capacity there were nevertheless many individual transplanted patients with defective diffusion capacity and abnormal spirometric values.     

Obesity in renal transplantation: the role of induction therapy on long-term outcomes

Obesity is a burgeoning problem among renal transplant recipients given its association with increased morbidity, graft loss, and mortality. The long-term influence of different induction therapies in obese compared to nonobese patients is uncertain. We examined the long-term effect of low-dose rabbit antithymocyte globulin (rATG; 3-5 mg/kg) induction therapy compared to two doses of 20 mg basiliximab (BSX) in nonobese and obese renal transplant patients. The medical records of all adult (>18 years) recipients of kidney transplants between June 2001 and June 2009 in our center were reviewed. Patients whose body mass index (BMI) was greater than 30 were considered to be obese. The average dose of rATG was 3.2 ± 1.6 mg/kg. A total of 475 patients were included. In the nonobese group with a BMI less than 30, 68 received BSX and 247, rATG. In the obese group, 27 patients were given BSX and 133 were given rATG. Mean follow-up was 1523 days. These four groups were similar in baseline characteristics including: donor and recipient age, percent diabetes, living donors, panel-reactive antibodies > 35, HLA mismatch, race, gender, and maintenance immunosuppression. Serum creatinine levels at 3 months and 1, 5, and 7 years were not statistically different between groups. Compared to BSX induction therapy, rATG was associated with better graft survival at 47.4 ± 10 months in obese (63.6% vs 90.3%, P < .05, respectively) as well as nonobese patients (68.2% vs 88.7%, P < .05, respectively). Rejections were numerically lower in rATG-treated obese patients, which reached statistical significance in nonobese patients. Wound and viral infections were not statistically different between rATG and BSX groups. Therefore, low-dose rATG is associated with a better long-term graft survival rate in obese patients without incurring an increased risk of infectious complications. When rATG was used in obese and nonobese patients, there was no difference in graft and patient survival.     

Studies on the multiple-drug induction immunosuppressive therapies with cyclosporine after renal transplantation

Our induction immunosuppressive therapies were carried out on patients split into three groups. The first group of 25 recipients were treated with regimen I [cyclosporin (CsA); 12 mg/kg/day and prednisolone (Pred)]. The second group of 16 recipients were treated with regimen II [CsA; 6 mg/kg/day, Pred and mizoribine (MIZ) or azathioprine (AZA)]. The third group of 14 recipients were treated with regimen III [CsA; 10 mg/kg/day, Pred and MIZ or AZA]. There was no significant difference among the three groups in renal function three months after renal transplantation. The frequency and grade of rejection were significantly higher in Group II than in the other groups. One of group I had CsA nephrotoxicity and none of group III had liver dysfunction three months after renal transplantation. Group I had a higher incidence of posttransplant hypertension. Hypertension of group I was very severe. We concluded that the triple-drug therapy on group III was the best induction immunosuppressive therapy after renal transplantation of the above three.