Isolated Donor Specific Alloantibody-Mediated Rejection after ABO Compatible Liver Transplantation

Antibody-mediated rejection (AMR) after liver transplantation is recognized in ABO incompatible and xeno-transplantation, but its role after ABO compatible liver transplantation is controversial. We report a case of ABO compatible liver transplantation that demonstrated clinical, serological and histological signs of AMR without evidence of concurrent acute cellular rejection. AMR with persistently high titers of circulating donor specific antibodies resulted in graft injury with initial centrilobular hepatocyte necrosis, fibroedematous portal expansion mimicking biliary tract outflow obstruction, ultimately resulting in extensive bridging fibrosis. Immunofluorescence microscopy demonstrated persistent, diffuse linear C4d deposits along sinusoids and central veins. Despite intense therapeutic intervention including plasmapheresis, IVIG and rituximab, AMR led to graft failure. We present evidence that an antibody-mediated alloresponse to an ABO compatible liver graft can cause significant graft injury independent of acute cellular rejection. AMR shows distinct histologic changes including a characteristic staining profile for C4d.     

The Significance of Donor‐Specific HLA Antibodies in Rejection and Ductopenia Development in ABO Compatible Liver Transplantation

The role of humoral alloreactivity in ABO‐compatible liver transplantation remains unclear. To understand the significance of donor‐specific HLA alloantibodies (DSA) in liver rejection, we applied the currently used strategy for detection of antibody‐mediated rejection of other solid allografts. For this purpose we reviewed the data on 43 recipients of ABO identical/compatible donor livers who had indication liver biopsy stained for complement element C4d and contemporaneous circulating DSA determination. Seventeen (40%) patients had significant circulating DSA in association with diffuse portal C4d deposition (DSA+/diffuse C4d+). These DSA+/diffuse C4d+ subjects had higher frequency of acute cellular rejection (ACR) 15/17 versus 13/26 (88% vs. 50%), p = 0.02, and steroid resistant rejection 7/17 versus 5/26 (41% vs. 19%), p = 0.03. Based on detection of the combination DSA+/diffuse C4d+, 53.6% of cases of ACR had evidence of concurrent humoral alloreactivity. Six of the 10 patients with ductopenic rejection had circulating DSA and diffuse portal C4d, three of whom (2 early and 1 late posttransplantation) developed unrelenting cholestasis, necessitating specific antibody‐depleting therapy to salvage the allografts. Thus, in ABO‐compatible liver transplantation humoral alloreactivity mediated by antibodies against donor HLA molecules appears to be frequently intertwined with cellular mechanisms of rejection, and to play a role in ductopenia development.     

Control of antidonor antibody production with tacrolimus and mycophenolate mofetil in renal allograft recipients with chronic rejection

BACKGROUND: In renal transplantation, chronic rejection is a major cause of late allograft loss. Recent studies indicate that a subset of chronic rejection is associated with anti-HLA donor specific antibodies (DSA) and complement C4d deposition in peritubular capillaries (PTC). Since rescue therapy with tacrolimus and mycophenolate mofetil has been found to limit antidonor B-cell responses in recipients with acute humoral rejection, we sought to determine whether a similar immunosuppressive regimen might be effective in patients with 'chronic humoral rejection'. METHODS: Four renal allograft recipients with 'chronic humoral rejection' were prospectively identified. The diagnosis was based on: (1) progressive rise in serum creatinine over 12 months; (2) typical pathologic features by light microscopy (transplant arteriopathy and glomerulopathy); (3) widespread C4d deposits in PTC by immunofluorescence; (4) detection of 'de novo' DSA at the time of biopsy. Maintenance immunosuppression was CsA, prednisone and azathioprine (n=3) or prednisone and azathioprine (n=1). Rescue therapy with tacrolimus and mycophenolate mofetil was initiated in all patients, 12 hr after cyclosporine and azathioprine discontinuation. RESULTS: At diagnosis, the mean serum creatinine was 3.9 mg/dl (range: 3.3 to 5.4 mg/dl). DSA was an IgG directed against HLA class II (n=3) or class I (n=2), that is one patient had both anti-HLA class I and class II antibodies. Pretreatment antibody titers varied between 1:8 and 1:128. Rescue therapy was associated with a rapid and sustained decrease in antibody titers. In two patients, DSA became undetectable after 9 months and a repeat biopsy performed after 12 months revealed a decrease in C4d deposition in PTC. CONCLUSION: These results suggest that a decrease in DSA production can be induced in renal allograft recipients with 'chronic humoral rejection' by using an immunosuppressive regimen that combines tacrolimus and mycophenolate mofetil. Limitation of antidonor antibody synthesis may be important for the treatment or the prevention of chronic rejection in organ transplantation.     

Endothelial C4d deposition is associated with inferior kidney allograft outcome independently of cellular rejection.

BACKGROUND: Capillary deposition of complement split product C4d has been suggested to be a valuable marker for humoral rejection. In this retrospective study we evaluated the clinical impact of C4d deposition in renal allografts with special emphasis on associations between C4d staining patterns and histological features of acute rejection. METHODS: One hundred and two allograft biopsies obtained from 61 kidney transplants (1-532 days after transplantation; median 14 days) were examined by immunohistochemistry on routine paraffin sections using a novel anti-C4d polyclonal antibody (C4dpAb). RESULTS: Fourty-two of 102 biopsies showed endothelial C4d deposits in peritubular capillaries (PTC). Histopathological analysis revealed a significantly lower frequency of positive C4d staining in biopsies with rather than in those without acute cellular rejection defined by the Banff grading schema (P<0.01). For clinical evaluation, patients were classified according to C4d staining in allografts (C4d(PTC) positive in at least one biopsy, n=31 vs C4d(PTC) negative in all biopsies, n=30). C4d(PTC) positive patients had significantly higher serum creatinine levels than C4d negative patients. Even in the absence of morphological evidence for rejection, differences in serum creatinine levels between C4d(PTC) positive and negative recipients were significant (6 months: 2.01+/-0.75 vs 1.41+/-0.27 mg/dl; 12 months: 1.95+/-0.60 vs 1.36+/- 0.34 mg/dl; 18 months: 1.98+/-0.50 vs 1.47+/-0.31 mg/dl; P<0.05). All patients with rejection resistant to conventional therapy (n=4) were in the C4d(PTC) positive subgroup. All recipients with panel reactive antibodies (PRA) >50% (n=8) were C4d(PTC) positive. CONCLUSIONS: Our data indicate that endothelial C4d deposition is associated with inferior graft outcome. We provide evidence that this immunohistochemical finding and its clinical impact are not associated with morphological signs of cellular rejection.     

Significance of C4d deposition in the diagnosis of rejection after liver transplantation

C4d immunohistochemical staining of liver allograft biopsies was performed to assess its relationship to other pathological changes in the liver. C4d deposition was detected in 69.2% of liver graft biopsies from patients under going rejection, 33.3% of liver graft biopsies from patients with hepatitis B relapse after transplantation, and 28.6% of liver biopsies from patients with hepatitis B. When rejection occurred C4d deposition was located in the vascular walls of portal areas and hepatic sinusoidal walls. Examination of biopsies from patients with hepatitis B relapse after transplantation or hepatitis B infection showed C4d deposition only in the vascular walls of the portal area. C4d deposition in both vascular walls of portal area and hepatic sinusoidal walls was detected in only one of 12 ischemia-reperfusion damage cases. Repeated biopsy of the same patient 1 month later revealed acute cellular rejection. No C4d deposition was found in biopsies from a patient with bile duct occlusion after liver transplantation. C4d might serve as a sensitive marker for the diagnosis of liver rejection.     

Role of CXCR3 in cellular but not humoral renal allograft rejection

CXCR3, a chemokine receptor mainly expressed by T cells, is involved in animal transplant models and in human allograft rejection. CXCR3 expression was localized in formalin-fixed, paraffin-embedded renal allograft biopsies without signs of rejection (C4d-negative, Banff 0, n = 16), with C4d deposits as a sign of humoral rejection (C4d-positive, Banff 0, n = 8), with cellular rejection (C4d-negative, Banff I, n = 7) and with signs of both cellular and humoral rejection (C4d-positive, Banff 1, n = 5). Small, round infiltrating cells were CXCR3-positive. A high number of these cells was present in biopsies with cellular rejection (independent of C4d deposition). CXCR3-positive cells diffusely infiltrated the interstitium, including the tubular epithelium (tubulitis). CXCR3 scores and the area of CXCR3 staining were significantly higher in cellular rejection, when compared to biopsies without rejection, and with deposition of C4d alone. CXCR3-positive cells infiltrate renal allografts during cellular rejection, whereas C4d deposition is not associated with the recruitment of these cells.     

Capillary deposition of complement split product C4d in renal allografts is associated with basement membrane injury in peritubular and glomerular capillaries: a contribution of humoral immunity to chronic allograft rejection

Endothelial deposition of the complement split product C4d is an established marker of antibody-mediated acute renal allograft rejection. A contribution of alloantibody-dependent immune reactions to chronic rejection is under discussion. In this study, the association of immunohistochemically detected endothelial C4d deposition in peritubular capillaries (PTC) with morphologic features of chronic renal allograft injury was investigated in a large study cohort. C4d deposits in PTC were detected in 73 (34%) of 213 late allograft biopsies performed in 213 patients more than 12 mo after transplantation (median, 4.9 yr) because of chronic allograft dysfunction. Endothelial C4d deposition was found to be associated with chronic transplant glomerulopathy (CG) (P < 0.0001), with basement membrane multilayering in PTC (P = 0.01), and with an accumulation of mononuclear inflammatory cells in PTC (P < 0,001). Furthermore, C4d deposits in PTC (in biopsies with normal glomerular morphology) were associated with development of CG in follow-up biopsies. Other morphologic features of chronic allograft nephropathy (with exception of tubular atrophy) were not associated with C4d deposits in PTC. Analyses of previous and follow-up biopsies revealed that C4d deposits may occur de novo and may also disappear at any time after transplantation. In conclusion, the data suggest that complement activation in renal microvasculature, indicating humoral alloreactivity, contributes to chronic rejection characterized by chronic transplant glomerulopathy and basement membrane multilayering in PTC.     

Implications of immunohistochemical detection of C4d along peritubular capillaries in late acute renal allograft rejection

BACKGROUND: Immunohistochemical detection of the C4d complement product along peritubular capillaries (PC) may indicate humoral rejection of renal allografts. We examined the frequency of PC C4d expression in renal-allograft biopsies with acute rejection (AR) arising more than 6 months after transplantation and the impact of this finding. METHODS: C4d was detected by immunoperoxidase in 2-micron paraffin sections of consecutive biopsies obtained over a 3-year period. The extent was classified as diffuse (> or =50% PC C4d+), focal (<50% C4d+), and negative (C4d-). Clinical data were obtained by retrospective chart review. Fifty-five AR episodes with Banff 97 types 1A (n = 13), 1B (n = 26), 2A (n = 11), 2B (n = 3), and 3 (n = 2) met inclusion criteria. RESULTS: PC C4d expression was diffuse in 23 (42%), focal in 9 (16%), and negative in 23 (42%) biopsies. AR episodes with focal and diffuse C4d expression had higher proportionate elevation of serum creatinine at biopsy and 4 weeks after diagnosis (P< or =0.05). Biopsies with diffuse PC C4d had interstitial hemorrhage (56.5%) and plasmacytic infiltrates (52%) more frequently than C4d- biopsies (22% and 16%), P = 0.02, but had no other distinctive histologic features. Graft loss was greater in diffuse (65%) compared with focal C4d+ (33%) and C4d- (33%) groups 1 year after diagnosis, P = 0.03. Other clinical and pathologic parameters did not differ significantly, including treatment received for AR. CONCLUSION: Evidence of acute cellular with occult humoral rejection is identified in more than 40% of late AR episodes. Late acute humoral rejection may be associated with interstitial hemorrhage and plasma cells and contributes significantly to graft loss.     

C4d in Acute Rejection After Liver Transplantation—A Valuable Tool in Differential Diagnosis to Hepatitis C Recurrence

Hepatitis C is the most common indication for liver transplantation. Recurrence of HCV is universal leading to graft failure in up to 40% of all patients. The differentiation between acute rejection and recurrent hepatitis C is crucial as rejection treatments are likely to aggravate HCV recurrence. Histological examination of liver biopsy remains the gold standard for diagnosis of acute rejection but has failed in the past to distinguish between acute rejection and recurrent hepatitis C. We have recently reported that C4d as a marker of the activated complement cascade is detectable in hepatic specimen in acute rejection after liver transplantation. In this study, we investigate whether C4d may serve as a specific marker for differential diagnosis in hepatitis C reinfection cases. Immunohistochemical analysis of 97 patients was performed. A total of 67.7% of patients with acute cellular rejection displayed C4d-positive staining in liver biopsy whereas 11.8% of patients with hepatitis C reinfection tested positive for C4d. In the control group, 6.9% showed C4d positivity. For the first time we were able to clearly demonstrate that humoral components, represented by C4d deposition, play a role in acute cellular rejection after LTX. Consequently C4d may be helpful to distinguish between acute rejection and reinfection after LTX for HCV.     

体液性排斥反应患者移植物组织C4d沉积和浆细胞聚集性浸润初探

目的检测移植肝及肾组织中浆细胞的浸润和补体C4裂解产物C4d的沉积情况,分析浆细胞浸润、C4d沉积与体液性排斥反应的相关性。方法25例肝移植术后出现不同程度肝脏损害的患者的34次经皮肝脏穿刺活体组织病理检查标本与43例因排斥反应而丧失功能的移植肾组织,进行HE染色和免疫组织化学染色,对排斥反应进行病理分型,检测移植物组织中C4d和CD138的表达,分析二者之间的相关性。同时以10例非排斥反应导致移植肾功能丧失者和供肝术前标本作为对照。结果34个移植肝组织标本病理检查诊断为急性排斥反应16个,慢性排斥反应9个,非排斥反应9个。移植前供肝病理标本中均未见C4d沉积,急性排斥反应和慢性排斥反应标本中分别有9／16（56．3％）和5／9（55．6％）见到C4d沉积现象,非排斥反应标本只有1例非吻合口胆管狭窄患者C4d染色也呈阳性（11．1％）。移植肝排斥反应标本中11／25（44．0％）CD138阳性,8／25（32．0％）标本C4d和CD138均阳性。C4d的沉积与CD138的表达存在相关性（r=0．346,P〈0．05）。43例移植肾排斥反应中,超急性排斥反应5例,急性排斥反应9例,慢性排斥反应29例;43例中,C4d阳性19例（44．2％）,CD138阳性24例（55．8％）;有10例（23．3％）C4d和CD138均阳性,C4d的沉积与CD138的表达亦存在相关性（r=0．5051,P〈0．01）。10个对照标本中,C4d阳性1个,无CD138阳性标本。结论CD138与C4d的沉积存在相关性,提示移植肝和肾组织中聚集性浸润的浆细胞可能通过局部分泌抗体的方式参与体液性排斥反应。     

The role of C4d immunostaining in the evaluation of the causes of renal allograft dysfunction.

BACKGROUND: Renal biopsy is the gold standard for diagnosis of acute rejection in renal transplant recipients. The Banff (1997) classification was revised in 2003 incorporating morphological criteria and C4d immunostaining for the diagnosis of acute antibody-mediated rejection. AIMS: The aim of this study was to evaluate the role of histomorphology and C4d immunostaining in indicated renal allograft biopsies with a clinical follow-up for a minimum duration of 1 year. MATERIAL AND METHODS: Histological analysis and C4d immunostaining were performed on 132 needle core biopsies and 2 nephrectomy specimens from 107 patients from July 2004 to June 2005. RESULTS: Histological analysis revealed 59 cases of acute rejection, 10 biopsies of acute tubular necrosis, 41 cases of chronic allograft nephropathy (CAN), either alone or in combination with other diseases, and 18 biopsies of normal morphology. There were four cases of BK nephropathy (BK N) and eight cases had miscellaneous diagnoses. C4d immunostaining was performed on 126 biopsies. Overall, the prevalence of C4d positivity was 45% (57 of 126). Fifty-five percent (28 of 51) of the cases of acute rejection showed C4d positivity including 81% of presumptive antibody-mediated rejection (P-AbAR), 20% acute cellular rejection and 58% acute cellular rejection + P-AbAR. Overall C4d positivity was 37% in chronic allograft nephropathy. Acute tubular necrosis and borderline rejection showed 25 and 50% C4d positivity, respectively. Amongst various histological features, capillary margination of polymorphs and dilatation of peritubular capillaries (PTC-D) showed significant association with C4d positivity (P < 0.005). In cases of CAN, transplant glomerulopathy had significant association with C4d positivity. C4d-positive cases had a higher mean value of serum creatinine at the time of biopsies. CONCLUSION: It is concluded that C4d staining is a useful adjunct marker of the humoral limb of rejection, both in early and late post-transplant periods.     

移植肝内C4d沉积在肝移植急性和慢性排斥反应诊断和治疗中的意义

目的 观察移植肝组织中补体C4的裂解片断CAd沉积情况,探讨其与排斥反应的关系,为临床诊断和治疗排斥反应提供思路和依据.方法 25例肝移植患者术后共行移植肝穿刺36次,取34次肝脏穿刺样本入组研究.穿刺时间为移植后7 d至25个月,所有供肝移植前均留取样本.穿刺所得移植肝组织进行HE染色和免疫组织化学染色,观察CAd的沉积情况.结果 34次病理检查中,诊断为急性排斥反应16次,慢性排斥反应9次,非排斥反应9次(包括胆道并发症3次,药物性肝损害5次,乙型肝炎复发1次).移植前供肝组织中均未见CAd沉积.发生急性排斥反应者中,9例(9/16)移植肝组织中可见到C4d沉积,发生慢性排斥反应者中,5例(5/9)移植肝组织中可见到CAd沉积,非排斥反应者中,仅1例(1/9)非吻合口胆管狭窄患者的肝组织中有CAd沉积.排斥者和非排斥者的CAd沉积部位无明显差异.急性排斥反应者和慢性排斥反应者的C4d阳性率均高于非排斥反应者(P<0.05,P<0.05);CAd阳性率与肝损伤程度无明显相关性.CAd阳性者激素冲击有效率为35.7%(5/14),远低于CAd阴性者的63.6%(7/11).结论 CAd沉积可提示体液性排斥反应,可作为肝移植后肝损害时病理检查的常规免疫组织化学项目,有利于监测和预警体液性排斥反应的发生.并为选择冲击治疗方案提供依据.     

Capillary deposition of complement C4d and C3d in pediatric renal allograft biopsies

BACKGROUND: Peritubular capillary deposition of C4d (C4d(PTC)) is a marker of antibody-mediated alloresponse and is associated with poor graft survival in adults. C3d(PTC) has received less attention; its significance is unclear. To date no information has been gained in children. METHODS: The prevalence of C4d(PTC) and C3d(PTC) in pediatric renal allograft biopsies (n=77, 31 cadaveric kidneys) was analyzed retrospectively. Associations with histology, donor-specific antibodies (DSAs), and outcome were investigated. RESULTS: The overall prevalence of C4d(PTC) and C3d(PTC) was 52% and 48%, respectively. C3d(PTC) was associated with C4d(PTC) (P<0.0001). Thirty-six percent of acute rejections were cellular, 28% were humoral, and 36% were combined cellular and humoral. C3d(PTC) was found in 57% of acute rejection biopsies. C4d(PTC), but not C3d(PTC), was associated with accumulation of polymorphonuclear cells in peritubular capillaries (P=0.02). Fifty-one percent of late biopsies (>6 months posttransplantation) had features of chronic allograft nephropathy: 50% were C4d(PTC_ positive, and 50% were C3d(PTC) positive. C4d(PTC) positive chronic allograft nephropathy biopsies had more transplant glomerulopathy (P=0.020) and mesangial matrix increase (P=0.026). C3d(PTC) tended to be associated with transplant glomerulopathy (P=0.06), but not with mesangial matrix increase. C4d(PTC) was correlated with DSA (P=0.011). Excluding early nonrejection graft losses, more grafts were lost in the C4d(PTC) positive group (P=0.019). C3d(PTC) was not associated with DSA or graft outcome. CONCLUSIONS: Our results support C4d(PTC) being a hallmark of humoral rejection in pediatric renal transplantation; its presence was associated with DSA and poorer immunologic graft outcome. In contrast, C3d(PTC), although highly associated with C4d(PTC), did not correlate with DSA or outcome.     

The impact of c4d staining as a humoral injury marker

Purpose

Acute and chronic humoral injuries in renal tranplant recipients are the main reasons for graft rejection and failure. Histological and clinical characteristics of humoral rejection and symptoms are variable and not always helpful for differential diagnosis. Clinical monitoring of the allograft, an elevated serum panel-reactive antibody (PRA), and the presence of donor-specific antibody (DSA) during immune monitoring as well as C4d staining of biopsy material can establish the differential diagnosis. Even without a cellular component, humoral rejection reaction is serious because the target tissue is the graft endothelium. Because the kidney graft has a rich vascular structure this attack causes permanent injury to the kidney in the long term. Graft dysfunction in this setting is usually more severe, requiring dialysis therapy, compared with acute cellular reactions. Positive C4d staining of peritubular capillaries in biopsy material represent a hallmark of complement-dependent cytotoxicity, supporting the diagnosis of humoral rejection. We analyzed C4d staining as a hallmark of humoral rejection.

Methods

From 2009 to 2011, we analyzed the relationship between pathological findings of C4d immunohistochemistry staining and the clinical outcomes of 45 kidney transplant recipients who underwent a kidney biopsy because of graft dysfunction due to possible humoral rejection.

Results

Biopsy specimens of 15 patients stained C4d positive; the remaining 30 showed negative results. Intravenous steroids, PP + IVIG with or without antithymocyte globulin (ATG), was administered for treatment. Sixty six percent (n = 10) of patients were C4d positive with 16% (n = 5) of those showing C4d-negative biopsy results, losing their grafts, and returning to hemodialysis.

Conclusions

C4d staining refractory humoral rejection injury was related to poor graft outcomes.     

肝移植术后发生急性排斥反应受者淋巴细胞亚群的变化及意义

目的  探讨肝移植受者术后发生急性排斥反应时淋巴细胞亚群的变化及意义。方法  选取接受肝移植且发生急性排斥反应的受者作为排斥组(17例),利用倾向评分匹配方法按1∶1比例选取肝功能稳定的肝移植受者作为对照组(17例)。分析肝移植术后急性排斥反应的发生情况,对比两组受者他克莫司浓度。比较两组受者外周血淋巴细胞亚群的绝对值和比例,采用受试者工作特征(ROC)曲线分析淋巴细胞亚群对肝移植术后急性排斥反应发生的诊断价值。比较排斥组治疗前后淋巴细胞亚群绝对值和比例的变化。结果  排斥组17例受者中,4例在术后28 d内发生急性排斥反应,13例在术后29~180 d发生急性排斥反应。两组他克莫司谷浓度差异无统计学意义(P=0.295)。与对照组比较,排斥组受者外周血T细胞、CD4⁺T细胞、B细胞和自然杀伤(NK)T细胞的比例均上升(均为P < 0.05)。肝移植术后早期NKT细胞比例升高是肝移植术后发生急性排斥反应的独立危险因素[比值比(OR)1.774,95%可信区间(CI)1.059~2.971,P=0.029]。ROC曲线分析结果提示,CD4⁺T细胞、B细胞和NKT细胞比例的曲线下面积(AUC)分别为0.76、0.73和0.77。联用CD4⁺T细胞、B细胞和NKT细胞比例的AUC为0.89,当临界值为0.69时,灵敏度为0.706,特异度为0.941。排斥组所有受者治疗后均逐渐恢复,最终肝功能正常,治疗后T细胞、CD4⁺T细胞、CD8⁺T细胞和NK细胞比例均降低(均为P < 0.05)。结论  NKT细胞比例升高提示肝移植术后急性排斥反应发生风险增加,联用CD4⁺T细胞、B细胞和NKT细胞比例可早期发现和诊断肝移植术后急性排斥反应。     

Acute humoral rejection in renal allograft recipients: I. Incidence, serology and clinical characteristics

BACKGROUND: Acute rejection (AR) associated with de novo production of donor-specific antibodies (DSA) is a clinicopathological entity that carries a poor prognosis (acute humoral rejection, AHR). The aim of this study was to determine the incidence and clinical characteristics of AHR in renal allograft recipients, and to further analyze the antibodies involved. METHODS: During a 4-year period, 232 renal transplants (Tx) were performed at our institution. Assays for DSA included T and B cell cytotoxic and/or flow cytometric cross-matches and cytotoxic antibody screens (PRA). C4d complement staining was performed on frozen biopsy tissue. RESULTS: A total of 81 patients (35%) suffered at least one episode of AR within the first 3 months: 51 had steroid-insensitive AR whereas the remaining 30 had steroid-sensitive AR. No DSA were found in patients with steroid-sensitive AR. In contrast, circulating DSA were found in 19/51 patients (37%) with steroid-insensitive AR, and widespread C4d deposits in peritubular capillaries were present in 18 of these 19 (95%). In at least three cases, antibodies were against donor HLA class II antigens. DSA were not found in the remaining 32 patients but C4d staining was positive in 2 of 32. The DSA/C4d positive (n=18) and DSA/C4d negative (n=30) groups differed in pre-Tx PRA levels, percentage of re-Tx patients, refractoriness to antilymphocyte therapy, and outcome. Plasmapheresis and tacrolimus-mycophenolate mofetil rescue reversed rejection in 9 of 10 recipients with refractory AHR. CONCLUSION: More than one-third of the patients with steroid-insensitive AR had evidence of AHR, often resistant to antilymphocyte therapy. Most cases (95%) with DSA at the time of rejection had widespread C4d deposits in peritubular capillaries, suggesting a pathogenic role of the circulating alloantibody. Combined DSA testing and C4d staining provides a useful approach for the early diagnosis of AHR, a condition that often necessitates a more intensive therapeutic rescue regimen.     

Immunosuppression withdrawal after auxiliary liver transplantation for acute liver failure

BACKGROUND: The potential for immunosuppression withdrawal is the rationale for auxiliary liver transplantation (AUX) in patients with acute liver failure (ALF). PATIENTS AND METHODS: Forty-four AUX were performed in 28 adults and 16 children with ALF secondary to seronegative hepatitis (n = 20; 45%), paracetamol hepatotoxicity (n = 14; 32%), acute viral hepatitis (hepatitis B virus [HBV] n = 3, Epstein-Barr virus n = 1; 9%), drug-induced hepatitis (n = 3; 7%), autoimmune hepatitis (n = 2; 5%), and mushroom poisoning (n = 1; 2%). All patients fulfilled the King's College Hospital transplant criteria for ALF. After partial hepatectomy, 38 patients received a segmental auxiliary graft and six, a whole auxiliary graft. Immunosuppression was based on calcineurin inhibitors and steroids. RESULTS: Thirty-four patients (77%) are alive after a median follow-up of 30 months (range 4 to 124). Eight adults and two children died of sepsis (n = 6; 14%) at a median interval of 30 days (range 2 to 66), intraoperative cardiac failure (n = 1), brain edema on postoperative day 8 (n = 1), sudden death on day 35 (n = 1), and multiple organ failure associated with HBV recurrence 4 years after transplantation (n = 1). Three patients underwent retransplantation for small-for-size graft syndrome with sepsis on postoperative day 15 (n = 1) and for ductopenic rejection 4 and 15 months after AUX (n = 2). In 10/31 (32%) survivors (6/18 adults and 4/13 children) immunosuppression was completely withdrawn after a median of 19 months. CONCLUSION: Complete immunosuppression withdrawal can be achieved in a significant proportion of patients after AUX for ALF.     

C4d-positive acute humoral renal allograft rejection: effective treatment by immunoadsorption.

There is increasing evidence for an important pathogenetic role of alloantibodies in acute renal allograft rejection. Acute humoral rejection (AHR) has been reported to be associated with a poor transplant survival. Although treatment modalities for cellular rejection are fairly well established, the optimal treatment for AHR remains undefined. Ten of 352 kidney allograft recipients transplanted at the authors' institution between November 1998 and September 2000 were diagnosed as having AHR, supported by severe graft dysfunction, C4d deposits in peritubular capillaries (PTC), and accumulation of granulocytes in PTC. AHR was diagnosed 18.9 +/- 17.5 d posttransplantation. All patients were subjected to immunoadsorption (IA) with protein A (median number of treatment sessions, 9; range, 3 to 17). Seven recipients with additional signs of cellular rejection (according to the Banff classification) received also antithymocyte globulin. In nine of ten patients, AHR was associated with an increase in panel reactive antibody reactivity. A pathogenetic role of alloantibodies was further supported by a positive posttransplant cytotoxic crossmatch in all tested recipients (n = 4). In nine of ten recipients, renal function recovered after initiation of anti-humoral therapy. One patient lost his graft shortly after initiation of specific therapy. Another recipient with partial reversal of AHR returned to dialysis 8 mo after transplantation. Mean serum creatinine in functioning grafts was 2.2 +/- 1.2 mg/dl after the last IA session (n = 9) and 1.5 +/- 0.5 mg/dl after a follow-up of 14.2 +/- 7.1 mo (n = 8). In conclusion, this study suggests that AHR, characterized by severe graft dysfunction, C4d staining, and peritubular granulocytes, can be effectively treated by timely IA. In the majority of patients, IA treatment can restore excellent graft function over a prolonged time period.     

C4d staining of perioperative renal transplant biopsies

BACKGROUND: Deposition of C4d in peritubular capillaries (PTCs) has been shown to be a sensitive marker for antibody-mediated (humoral) rejection in renal transplant biopsies. Some studies also suggest that C4d in PTCs is specific for humoral rejection or, at least, for the presence of donor-specific antibodies. However, in other studies, PTC C4d deposits were noted in more than 40% of renal transplant biopsies performed for graft dysfunction and capillary C4d deposition in heart transplants may result from ischemic injury. METHODS: To test the specificity of C4d staining as a marker for acute humoral rejection ACR in renal allografts, indirect immunofluorescence using a monoclonal anti-C4d antibody and a fluorescein-isothiocyanate-conjugated secondary antibody was performed on cryostat sections of 90 renal transplant biopsies, including 35 pairs of preimplantation and 1-hr postreperfusion biopsies of the same graft, postreperfusion biopsies of 12 additional grafts, and 8 positive controls (biopsies with known C4d-positive AHR). Eighteen grafts were cadaveric, 17 grafts were liviing-related, and 12 grafts were living-unrelated (excluding controls). Included in these grafts were 13 grafts that developed AHR 3 to 34 days posttransplantation. RESULTS: Only 2 of 82 perioperative biopsies showed C4d staining in PTCs. Both perioperative biopsies were postreperfusion biopsies of grafts diagnosed with AHR 5 and 34 days posttransplantation, respectively, and, in each case, the recipient had been treated with plasmapheresis before transplantation because of a positive crossmatch (cytotoxic and flow cytometric) and continued to have a weakly positive flow crossmatch at the time of transplantation. In one biopsy, C4d staining was focal, and in the other biopsy, it was diffuse; in both biopsies, C4d staining was relatively mild (1+ on a 0-4+ scale). No C4d staining was noted on preimplantation biopsies of each graft. All biopsies that contained glomeruli showed linear capillary loop or blotchy mesangial staining, or both, which was similar in prereperfusion and postreperfusion biopsies. All positive controls showed diffuse C4d staining in PTCs.CONCLUSIONS: C4d staining in PTCs may be seen as early as 1 hr posttransplantation in some recipients with low levels of antidonor antibodies. However, this was not observed as a feature of ischemic or ischemia-reperfusion injury in perioperative renal transplant biopsies, including those of cadaveric grafts with cold ischemia times of as long as 41 hr.