Clinicopathologic analysis of renal biopsies after haematopoietic stem cell transplantation

Aim: The ever‐growing number and increasing survival of haematopoietic stem cell transplantation (HSCT) allow better recognition of its associated renal injuries. We aimed to study the clinicopathologic features of renal biopsies after HSCT by reviewing 13 percutaneous renal biopsies in our institute (Queen Mary Hospital). Methods: A retrospective clinicopathologic study of all renal biopsies archived to the Department of Pathology, Queen Mary Hospital during the period January 1999 to December 2006 was performed. Biopsies from patients with HSCT were selected. Clinical data on presentation and follow up were retrieved from hospital records and physicians. Results: In the 8‐year period, a total of 2233 native renal biopsies were archived. Thirteen renal biopsies were selected from 12 patients with HSCT (11 allogeneic, one autologous). All but one patient were male. The age at renal biopsy ranged from 7 to 63 years (median: 32 years). The median interval of renal biopsy after HSCT was 24 months (range 1–134 months). Evidence of graft‐versus‐host disease was found in nine patients. The most common presentation was significant proteinuria (10 cases) and renal impairment (eight cases). The predominant histological changes were membranous glomerulonephritis (n = 4) and thrombotic microangiopathy (n = 4). One case of focal segmental glomerulosclerosis, IgA nephropathy, minimal change disease, acute tubular necrosis and hypertensive nephrosclerosis were also recorded. Four of our patients died at 0–11 months after renal biopsy. Of the remaining eight patients with a mean follow up of 43.6 months (range, 10–98 months), chronic renal impairment were found in three (37.5%) patients and significant proteinuria also persisted in three. One patient had cytogenetic evidence of relapse of underlying haematological malignancy after HSCT. Conclusion: Among the various renal lesions after HSCT, membranous glomerulonephritis and thrombotic microangiopathy were the most common. Mechanisms of renal injury varied from graft‐versus‐host disease‐associated immune complex deposition to non‐immune complex injury on endothelial cells, glomerular epithelial cells and tubular epithelium. Pathologists and clinicians should attend to the histological and temporal heterogeneity of renal injury when managing patients after HSCT.     

Pre-transplant co-infusion of donor-adipose tissue derived mesenchymal stem cells and hematopoietic stem cells may help in achieving tolerance in living donor renal transplantation

Transplantation tolerance is still a Utopian dream for many transplanters. Mesenchymal stem cells (MSC) have shown immuno-modulatory and tolerogenic effects in experimental models. We present a 29-year-old male with end stage renal disease (ESRD) who was transplanted with HLA 4/6 matched kidney from 51-year-old father in June 2010 preceded by co-infusion of donor-adipose tissue derived mesenchymal stem cells (AD-MSC) and bone marrow derived hematopoietic stem cells (BM-HSC) under non-myeloablative conditioning for deleting rejecting T and B-cells. He has maintained fairly stable graft function with serum creatinine (SCr) between 1.5 and 1.8?mg/dL at 3 years post-transplant with absence of donor specific antibodies (DSA), normal protocol graft biopsy, and peripheral T-regulatory cell levels (pTregs) (CD127^low/?CD25^highCD4⁺) of 4.57% on zero immunosuppression since 6 months.     

Diagnostic value of donor-derived cell-free DNA to predict antibody-mediated rejection in donor-specific antibody-positive renal allograft recipients

Circulating donor-specific antibodies (DSA) do not necessarily indicate antibody-mediated rejection (ABMR). Here, we evaluated the diagnostic value of donor-derived cell-free DNA (dd-cfDNA) as an add-on to DSA detection. The study included two independent cohorts of DSA⁺ kidney allograft recipients, 45 subclinical cases identified by cross-sectional antibody screening (cohort 1), and 30 recipients subjected to indication biopsies (cohort 2). About 50% of the DSA⁺ recipients had ABMR and displayed higher dd-cfDNA levels than DSA⁺ABMR⁻ recipients (cohort 1: 1.90% [median; IQR: 0.78–3.90%] vs. 0.52% [0.35–0.72%]; P < 0.001); (cohort 2: 1.20% [0.82–2.50%] vs. 0.59% [0.28–2.05%]; P = 0.086). Receiver operating characteristic (ROC) analysis revealed an area under the curve (AUC) of 0.89 and 0.69 for dd-cfDNA, and 0.88 and 0.77 for DSA mean fluorescence intensity (MFI), respectively. In combined models, adding dd-cfDNA to DSA-MFI or vice versa significantly improved the diagnostic accuracy. Limited diagnostic performance of dd-cfDNA in cohort 2 was related to the frequent finding of other types of graft injury among ABMR⁻ recipients, like T cell-mediated rejection or glomerulonephritis. For dd-cfDNA in relation to injury of any cause an AUC of 0.97 was calculated. Monitoring of dd-cfDNA in DSA⁺ patients may be a useful tool to detect ABMR and other types of injury.     

Quality of life and emotional responses in cadaver and living related renal transplant recipients.

BACKGROUND: The specific impact of transplantation on living related donor (LRD) and cadaver (CAD) kidney transplant recipients and their health-related quality of life (HQoL) has received little attention. This study examined the role of sociodemographic, medical and psychological factors in these two groups. METHODS: A total of 347 transplant recipients (76 LRD and 271 CAD patients) completed the Short Form 36 Health Survey and Transplant Effects Questionnaire. RESULTS: Overall, transplant patients showed satisfactory HQoL particularly with respect to emotional well being. HQoL levels were found to be equivalent in both transplant groups. ANCOVAs showed that LRD recipients expressed more guilt in relation to the donor (P<0.001). Multivariate analysis revealed that worry about the viability and functioning of the transplant alone predicted 15.1% of the variance in the SF-36 mental composite score (MCS) whereas age, income, comorbidities and time on dialysis explained 37.8% of the variance in the SF-36 physical composite score (PCS). Multiple regression analyses performed separately for LRD and CAD patients showed that predictors of MCS and PCS between the two groups were similar. CONCLUSIONS: Our results indicate that different forms of transplantation (LRD vs CAD) may lead to different emotional responses albeit with no apparent quality of life differences. In particular, feelings of guilt appear to be prominent in LRD transplantation.     

Clinical significance of in vitro donor-specific hyporesponsiveness in renal allograft recipients as demonstrated by the MLR

A longitudinal study was carried out on 19 recipients of cadaveric renal allografts, monitoring their anti-donor and anti-third party responses in the mixed lymphocyte reaction (MLR) at the time of transplantation and at 3, 6, and 12 months post-transplant. Two patterns of responses were identified: in the first (n=11), patients showed, or later developed, donor-specific hyporesponsivenes, and in the second (n=8), patients had persistent antidonor and anti-third party responses. After 1 year, the serum creatinine, number of episodes of acute rejection and biopsy findings were compared in both groups. In the first group, the mean serum creatinine was 136.4 mmol/l, the total number of acute rejection episodes was three and in nine of the ten available biopsies, there were minimal cellular infiltrates and normal appearance of the glomeruli, tubules and blood vessels. In the second group, the mean serum creatinine was 163 mmol/l, the total number of acute rejection episodes was 12 and in five of the seven biopsies available, evidence of ongoing rejection was obtained. The difference in mean serum creatinine was not statistically significant (P>0.05), but the difference in the numbers of acute rejection episodes was (P>0.05). It is concluded that in some renal allograft recipients, a state of donor-specific hyporesponsiveness develops, and this state may be associated with better graft out-come at 1 year. These data may be useful in selecting patients for reduced immunosuppressive therapy.     

输注供鼠骨髓间充质干细胞延长肾移植大鼠的存活时间

目的 探讨在同种大鼠肾移植中输注供者骨髓间充质干细胞(MSC)对急性排斥反应的影响以及延长受鼠存活时间的作用.方法 取Wistar大鼠骨髓,分离和培养其MSC.以Wistar 大鼠为供者,Lewis大鼠为受者,建立同种大鼠肾移植模型.根据受鼠处理方式的不同,分为低剂量MSC组、高剂量MSC组、CsA组及对照组,低剂量MSC组和高剂量MSC组于移植前后分别多次输注1×106个和t×107个供者MSC,CsA组于术后2d开始腹腔内注入CsA 0.5 mg·kg-1 ·d-1,以腹腔内注射PBS作为对照.移植后,比较各组受鼠的存活时间,观察各组移植肾功能及移植肾组织病理学改变.结果 低剂量MSC组、高剂量MSC组、CsA组及对照组受鼠的存活时间分别为(21.7±7.2)d、(31.2±14.3)d、(34.9±15.7)d及(9.0±2.3)d;低剂量MSC组、高剂量MSC组和CsA组存活时间均明显长于对照组(P＜0.01),而低剂量MSC组存活时间明显短于高剂量MSC组和CsA组(P＜0.05).术后第4天,高剂量MSC组和CsA组移植肾组织形态和结构基本正常;对照组移植肾组织则表现出典型的急性排斥反应,出现广泛间质性浸润,肾小管炎症和片状坏死、出血,肾小球炎症浸润严重;而与对照组相比,低剂量MSC组急性排斥反应的病理表现则要明显减轻.结论 同种肾移植大鼠输注供者MSC后,可以达到有效的免疫调节作用,并且可明显延长大鼠的存活时间,呈MSC剂量依赖性.     

Renal transplantation after myeloablative and non-myeloablative hematopoietic cell transplantation from the same donor

Hematopoietic cell transplantation is a key treatment to prolong patient survival for many hematological disorders. Renal impairment is well recognized as a significant complication of hematopoietic cell transplantation, which can progress to end-stage renal disease. Herein, we report our experience of two patients who underwent renal transplantation from the same donor who provided cells for the preceding hematopoietic cell transplantation. One patient had undergone peripheral blood stem cell transplantation with a non-myeloablative conditioning regimen, whereas the other had received bone marrow transplantation with a myeloablative regimen. Chronic immunosuppressive therapy was not needed in either one to maintain the kidney graft function. Not only bone marrow transplantation with a myeloablative conditioning regimen, but also peripheral blood stem cell transplantation with a non-myeloablative regimen can confer immunological tolerance.     

输注供者自然杀伤细胞对小鼠单倍型相合造血干细胞移植的影响

目的 探讨输注供者自然杀伤(NK)细胞对小鼠单倍型相合造血干细胞移植的影响.方法 选取C57BL/6(H-2b)雄性小鼠为供者、CB6F1(H-2d/b)雌性小鼠为受者.移植前制备供者的骨髓细胞(BMC)、脾细胞(SC)及脾NK细胞,NK细胞经体外培养扩增和激活;所有受者均接受直线加速器X线全身照射(TBI)预处理.TBI后将受者分为4组(每组10只),分别进行单倍型相合造血干细胞移植.单纯TBI组:TBI后不输注细胞,仅作为对照;单纯BMC输注组:输注5×106个BMC;诱发GVHD组:输注5×106个BMC+1.5×107个SC;NK细胞输注组:输注5 x 106个BMC+1.5×107个SC+1×107个NK细胞,并腹腔注射100 ng重组人白细胞介素2(rhIL-2)和1μg rhIL-15,持续7 d.移植后观察各组受者GVHD的发生情况,并对各组受者进行组织病理学、供者细胞嵌合度和免疫功能重建等检测.另取TBI后受者20只,设白血病复发组和白血病治疗组,每组10只.白血病复发组:输注5×106个BMC+1×107个SC+2×106个白血病细胞株EL9611;白血病治疗组:在白血病复发组的基础上再输注1 x 107个NK细胞,并腹腔注射100 ng rhIL-2和1μg rhIL-15,持续7 d.观察两组受者白血病复发情况和移植后100 d的存活率.结果 单纯BMC输注组受者无GVHD发生,NK细胞输注组受者GVHD的评分和组织病理学改变均较诱发GVHD组轻(P<0.05)f诱发GVHD组的免疫功能重建较NK细胞输注组延迟.白血病复发组和白血病治疗组移植后100 d的存活率分别为20%和90%,两组比较,差异有统计学意义(P<0.01).结论 输注激活的供者NK细胞可以减轻小鼠单倍型相合造血干细胞移植后的GVHD,减少白血病复发,促进免疫功能重建.     

活体肾移植供体应用乌司他丁对受体肾功能影响的研究

研究乌司他丁应用于活体肾移植供体对受体术后移植肾功能的影响.方法 选择行活体肾移植的供体和受体40对,按其供体是否应用乌司他丁分为乌司他丁组和对照组,每组20对.患者均签署知情同意书,符合医学伦理学规定.乌司他丁组供体在麻醉前即以乌司他丁5 kU/kg静脉泵入.比较两组受者的一般资料;比较两组受者麻醉前、麻醉后、受体血管开放时和手术结束时的平均动脉压和心率;采用酶联免疫吸附试验(ELISA)法检测两组受者麻醉前、手术结束时和手术结束后24 h血浆胱抑素C和α1-微球蛋白(α1-microglobulin,α1-MG)水平.结果 两组受体的年龄、性别、体重指数、手术时间、术中输液量、术后24 h尿量、冷缺血时间和热缺血时间比较差异无统计学意义(均为P >0.05).两组受体的平均动脉压和心率在各个时间点比较差异无统计学意义(均为P >0.05).受体手术结束时、手术结束后24 h两个时间点的血浆胱抑素C和α1-MG水平较麻醉前时间点明显下降,两组在同时间点比较差异亦无统计学意义(均为P >0.05).结论 本研究供体所应用乌司他丁的剂量和方法对移植后肾功能的改善效果不显著.     

供受者围手术期应用乌司他丁对移植肾功能保护机制的探讨

目的 研究活体肾移植供受者围手术期应用乌司他丁(ulinastatin,UTI)对受者肾功能的影响,探讨其肾功能保护机制. 方法 将40对全身麻醉下活体肾移植供、受者按随机数字表法分为两组(每组20对):UTI组(U组)和对照组(C组).U组供者于阻断肾动脉前1h静脉泵注UTI5 000 U/kg(稀释至50 ml,输注时间20 min),受者于吻合肾血管开放后即刻泵注UTI 5 000 U/kg(稀释至50 ml,输注时间20min);C组供、受者以同U组相同方案泵注等量生理盐水.两组受者在麻醉诱导前(T1)、吻合血管开放前15 min(T2)、吻合血管开放即刻(T3)、手术结束时(T4)、术后6 h(T5)、术后24 h(T6)时的血浆血栓素A2(thromboxane A2,TXA2)、前列环素I2(prostacyclin I2,PGI2)浓度[采用ELISA法检测T1～T6时待测血样中血栓素B2(thromboxane B2,TXB2)、6-酮-前列腺素F1α(6-ketone-prostaglandin F1α,6-keto-PGF1α)浓度(TXA2和PGI2在血浆中极不稳定,但通过测量两者的稳定代谢产物TXB2、6-keto-PGF1α可间接了解TXA2和PGI2的生成情况)及TXA2/PGI2比值和T11T6、术后48 h(T7)时的血肌酐(serum creatinine,Scr)、半胱氨酸蛋白酶抑制剂C(cystatin C,CysC)浓度及尿量. 结果 与T11比较,两组受者T2～T6时血浆TXB2浓度均明显升高(P＜0.05),T4时达到峰值,之后降低,T6时仍高于术前水平;T2～T6时血浆6-keto-PGF1α浓度均升高(P＜0.05),T3时达到峰值,之后降低,T6时升高虽有统计学意义(P＜0.05),但基本恢复至术前水平;T2～T6两组TXB2/6-keto-PGF1α均明显升高(P＜0.05);T6～T2时Scr及CysC水平明显降低(P＜0.05),尿量明显增多(P＜0.05).与C组比较,U组T2～T6时血浆TXB22度明显降低(P＜0.05);T4～T5时血浆6-keto-PGF1α浓度降低幅度明显下降(P＜0.05);T2～T6时TXB2/6-keto-PGF1α明显降低(P＜0.05);T6～～7时Scr、CysC水平明显降低(P＜0.05);T6～～7时虽尿量增多,但差异无统计学意义(P＞0.05). 结论 UTI对TXA2/PGI2比例失衡有一定的调节作用,可通过此机制改善移植肾的血流灌注,对缺血//灌注损伤(ischemia/reperfusion injury,I/RI)导致的移植肾功能受损有一定保护作用.     

Association of donor hypertension and recipient renal function in living donor kidney transplantation: A single‐center retrospective study

Transplant centers now accept living donors with well‐controlled hypertension. Little is known whether hypertension in living donors affects recipient's kidney function. We aimed to examine potential differences in kidneys from hypertensive donors compared to normotensive donors with respect to renal function over 36 months and histologic findings at transplantation (T0) and 12 months after transplantation (T1). Retrospective single‐center analysis of 174 living donor‐recipient pairs (age > 18; transplantation date 1/2008‐3/2016). Hypertension in donors was defined as being on antihypertensive medication. All biopsies were assessed by the same blinded, experienced renal pathologist. Biopsies were scored for glomerulosclerosis, IFTA, and arteriosclerosis. Regression models were used to examine the relationship of donor hypertension with renal function and histologic changes. Hypertensive donors were significantly older than normotensive donors. Chronic changes such as tubular atrophy and atherosclerosis were more evident in kidneys from hypertensive donors at T0 as well as T1. Donor hypertension was independently associated with histologic changes at T0 and T1 but not with renal function over the follow period. Despite more pronounced histologic changes in kidneys from hypertensive living donors, these grafts exhibited a similar functional outcome. However, they subsequently might be at a greater risk and warrant thorough follow‐up care.     

Effect of co-transplantation of mesenchymal stem cells and hematopoietic stem cells as compared to hematopoietic stem cell transplantation alone in renal transplantation to achieve donor hypo-responsiveness

Introduction  

We evaluated donor hypo-responsiveness in renal allograft recipients to donor adipose tissue-derived mesenchymal stem cell (h-AD-MSC) +hematopoietic stem cell transplantation (HSCT) vs. HSCT alone.     

骨髓间充质干细胞对大鼠急性缺血再灌注肾损伤的修复作用

目的 观察骨髓间充质干细胞(BMSCs)移植对大鼠缺血再灌注(IR)急性肾损伤(AKI)的修复作用.方法 将50只SD大鼠随机分为3组,干细胞移植组(IR+ BMSCs组)18只,生理盐水注射组(IR组)18只,正常组(Sham组)14只.分离及培养大鼠BMSCs到第3代,建立大鼠缺血再灌注AKI模型,24h后移植干细胞;检测血清及尿肌酐值,并以细胞核增殖抗原(Ki-67)进行肾脏免疫组织化学观察.结果 BMSCs培养后,得到纯度高、生长状态良好的第3代(P3) BMSCs.造模后第4天,细胞治疗组血清肌酐浓度为(37.34±4.22) μmol/L,尿肌酐水平为(22.75±6.23) μmol,而生理盐水注射组血清肌酐浓度为(238.34±32.42) μmol/L,尿肌酐水平为(7.68±1.03) μmol,两组比较差异有统计学意义(P＜0.01);组织切片学观察,与对照组比较,IR+ BMSCs组肾小管结构改善及上皮细胞数量增多、刷状缘部分恢复,Millar法评分IR+ BMSCs组为0.799±0.023,IR组为2.798±0.055,两组比较差异有统计学意义(P＜0.01).以Ki-67进行免疫组织化学IR+ BMSCs组有大量增殖抗原阳性表达.结论 BMSCs对缺血再灌注性AKI有良好的修复作用.     

血管内皮生长因子基因转染骨髓间充质干细胞移植于梗塞心肌的实验研究

目的 探讨骨髓间充质干细胞（MSCs）移植联合血管内皮生长因子（VEGF）基因转染对大鼠梗塞心肌组织的修复重建、血管再生及梗塞后心功能的影响。方法 体外分离、培养、纯化SD大鼠的MSCs，以BrdU标记MSCs，腺病毒介导VEGF基因转染MSCs。建立大鼠急性心肌梗死模型4周后，随机分为4组（每组10只），分别行梗塞心肌内注射：转染VEGF基因的MSCs移植组（组Ⅰ）、单纯MSCs移植组（组Ⅱ）、单纯VEGF基因治疗组（组Ⅲ）和以注射无血清IMDM培养液为对照组（组Ⅳ）。移植4周后观察移植细胞的分化和新生血管的形成，并通过超声多普勒检测心功能变化。结果 组Ⅰ和组Ⅱ中，梗塞心肌处可见BrdU标记的移植细胞，cTnT染色阳性。超声心动图检查发现，组Ⅰ和组Ⅱ的左室射血分数（LVEF）的改善显著高于对照组（P均〈0．01），而组Ⅰ的LVEF改善程度要明显高于组Ⅱ；部分BrdU染色阳性的细胞可以分化成为内皮细胞，参与构成了梗塞区域的新生毛细血管。相对于对照组，组Ⅰ和组Ⅲ都有明显的血管新生（P均〈0．01）。结论 MSCs移植联合VEGF基因转染可以通过促进心肌再生和新生血管的形成来重建缺血心肌，显著改善心功能。     

供体骨髓干细胞输注对肝移植术后早期恢复的疗效观察

目的探讨同期供体骨髓干细胞输注对肝脏移植受者术后早期康复的影响。方法以2008年3月至12月本中心的30例肝移植患者为研究对象,并进行随机分组。实验组8例,行同期供体骨髓干细胞联合肝脏移植;对照组22例,仅行同种异体肝移植,不行骨髓干细胞输注。观察两组患者术后肝功能恢复情况、急性排斥发生情况、感染并发症发生情况以及术后住院时间和费用。结果实验组术后甲基强地松龙用量和出院时每日FK506用量低于对照组[甲基强地松龙用量：（1884±256）mgys（1314±105）mg,P〈0．01;FK506用量：（4．93±0．62）mg,dvs（3．73±0．35）mg/d,P〈0．01];实验组术后3个月内急性排斥发生率显著低于对照组（0／8vs4／22,P〈0．05）,术后感染并发症发生率两组间无显著性差异;术后第1、3天实验组血清谷丙转氨酶、谷草转氨酶均显著低于对照组（P〈0．05）,术后第7天两组间无显著差异,而血清总胆红素、谷氨酰转肽酶、白蛋白等指标于术后第1、3、7天两组间对比均无显著差异;两组患者平均术后住院时间、住院费用无显著性差异。结论同期供体骨髓干细胞联合肝脏移植可以降低术后免疫抑制剂用量,减少急性排斥反应发生,一定程度上可减低早期肝脏再灌注损伤,不会增加患者的治疗风险、术后住院时问和费用,是一种安全有效的治疗方法。     

An investigation to assess the potential of CD25highCD4+ T cells to regulate responses to donor alloantigens in clinically stable renal transplant recipients.

Regulatory T cells are enriched within CD25(high)CD4(+) leukocytes, however their role in renal transplant recipients with stable function vs. recipients with biopsy-proven chronic allograft dysfunction remains unclear. We therefore studied the number, phenotype, and function of CD25(high)CD4(+) cells in the peripheral blood of 30 renal transplant recipients of living-related grafts, comprising 15 rejection-free recipients with stable graft function (Group A) and 15 with biopsy-proven chronic graft dysfunction (Group B). A higher absolute number of CD25(high)CD4(+) cells were present in the peripheral blood of rejection-free recipients (Group A) vs. those recipients with chronic graft dysfunction (Group B) (P = 0.019); but there was no significant difference with healthy volunteers (P = 0.084). In carboxyfluorescein diacetate succinimidyl ester-mixed leukocyte culture assays, depletion of CD25(high)CD4(+) revealed active regulation in 11 (74%) of 15 rejection-free recipient samples (Group A) in response to donor- but not third party-leukocytes, whereas no regulatory activity was observed in any samples from recipients with chronic graft dysfunction (Group B). In conclusion, these data provide evidence for the presence of an increased number of CD25(high)CD4(+) T cells with donor-specific regulatory activity in the peripheral blood of renal transplant recipients with stable graft function compared with recipients with chronic graft dysfunction.