Biological advances in acute graft-versus-host disease after allogeneic hematopoietic stem cell transplantation

Allogeneic hematopoietic stem cell transplantation is a potentially curative therapy for many malignant and non-malignant hematological diseases. Unfortunately, na?ve donor T cells that are critical for the success of this effective therapy also cause its most severe toxicity--graft-versus-host disease (GVHD). Recent experimental observations have brought into focus a critical role for additional cell populations in the pathogenesis and modulation of GVHD. A better understanding of the complex interactions involving these cellular subsets and the inflammatory cascades is likely to be critical in the pathophysiology of GVHD. This review will primarily focus on the immunobiology of experimental acute GVHD with an emphasis on the recent observations on the novel role of innate and adaptive cellular subsets in the activation, effector phases, and target organ specificity of acute GVHD.     

Pre-transplantation vitamin E levels and acute graft-versus-host disease after non-myeloablative allogeneic hematopoietic cell transplantation

BackgroundLow pre-transplantation plasma vitamin E levels have been associated with increased risk of acute graft-versus-host disease (GvHD) after myeloablative allogeneic hematopoietic cell transplantation (allo-HCT). We aimed to investigate the association between pre-transplantation plasma vitamin E levels and acute GvHD in patients undergoing allo-HCT with non-myeloablative conditioning.MethodsIn a cohort of 194 adults who underwent non-myeloablative allo-HCT at Rigshospitalet between July 2015 and December 2019, we measured pre-transplantation plasma vitamin E levels by high-performance liquid chromatography in stored plasma samples. Univariable ordinary least squares linear models were used to investigate associations between vitamin E levels and patient characteristics. A multivariable logistic regression model was used to estimate the association between vitamin E levels and grade II–IV acute GvHD, adjusted for recipient age, donor age, female-male donor-recipient pairing, and donor type.ResultsThe median (Q1, Q3) pre-transplantation plasma vitamin E level was 32.3 (26.4, 40.4) μmol/L. No patients had a vitamin E level below the normal reference range. Vitamin E levels were higher in females (mean difference: 8.0 μmol/L, 95% confidence interval [CI]: 4.9, 11.1 μmol/L) and in patients transplanted for acute leukemia (mean difference: 6.2 μmol/L, CI: 3.0, 9.4 μmol/L). Grade II–IV acute GvHD developed in 33 (17%) patients. Patients who developed acute GvHD had similar pre-transplantation vitamin E levels compared with patients who did not develop grade II–IV acute GvHD (mean difference: 0.7 μmol/L, bootstrap CI: −3.3, 4.7 μmol/L). In the adjusted logistic regression model, an increase in the pre-transplantation vitamin E level from 26.4 (Q1) to 40.4 (Q3) μmol/L was associated with an odds ratio of grade II–IV acute GvHD of 1.17 (CI: 0.64, 2.12).ConclusionsContrary to the previously reported association between pre-transplantation vitamin E levels and acute GvHD after myeloablative allo-HCT, we did not find support for an association in patients who received non-myeloablative conditioning. The potential protective effects of vitamin E may not be efficacious in the reduced inflammatory response following non-myeloablative conditioning.     

Both genetic and clinical factors predict the development of graft-versus-host disease after allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Graft-versus-host disease is the main complication of hematopoietic stem cell transplantation. Recently, pro- and anti-inflammatory cytokines and mismatches of minor histocompatibility antigens between HLA-identical sibling donor/recipient pairs have been implicated in the development of acute graft-versus-host disease. It is not known, however, whether these factors are independent of other clinically recognized risk factors such as age and disease stage. METHODS: In this study, we searched for risk factors of acute graft-versus-host disease using multivariate Cox regression analysis in 100 consecutive patients who underwent allogeneic stem cell transplantation from an HLA-identical sibling donor. Eight polymorphisms from five different cytokine genes were studied (tumor necrosis factor alpha, tumor necrosis factor beta, interleukin (IL) 6, IL-10, and interferon gamma). Mismatches for the minor histocompatibility antigen HA-1 were searched in HLA-A*0201 individuals. In addition to these new risk factors, patient, donor, disease, and transplant risk factors were analyzed by multivariate analysis using the Cox proportional hazards model. RESULTS: Acute graft-versus-host disease was independently associated with IL-10 gene polymorphisms both from the recipient (relative risk=7.9, P<0.0001) and the donor (relative risk=3.5, P=0.02), a donor's positive serology for cytomegalovirus, and HA-1 mismatches in HLA-A*0201 individuals (relative risk=2.8, P=0.05). Chronic graft-versus-host disease was independently associated with IL-6 gene polymorphism from the recipient (relative risk=4.2, P=0.02), older age (relative risk=2.5, P=0.0009), and previous acute graft-versus-host disease (relative risk=9.7, P=0.003). CONCLUSION: In addition to previously described clinical risk factors, genetic risk factors are independently associated with the risk of developing graft-versus-host disease and may, thus, be considered for the selection of the donor.     

环孢素A联合钴原卟啉预防小鼠HSCT后移植物抗宿主病

目的 探讨应用环孢素A(CsA)联合钴原卟啉(CoPP)预防小鼠异基因造血干细胞移植(allo-HSCT)后移植物抗宿主病(GVHD)的作用.方法 以C57BL/6小鼠为供鼠,Balb/c小鼠为受鼠,分为4组:清髓性全身照射(TBI)组,受鼠TBI后经尾静脉注射磷酸盐缓冲液;常规异基因移植组(BS组),受鼠TBI后经尾静脉注射供鼠骨髓细胞与脾细胞的混合悬液(含骨髓细胞5×106个和脾细胞5×105个);CsA组,受鼠HSCT后腹腔注射CsA;CsA联合CoPP干预组(联合组),受鼠HSCT后腹腔注射CsA和CoPP.观察受鼠存活情况,记录体重.获取受鼠肝脏、小肠和皮肤组织样本,观察GVHD的病理变化;行Masson染色,观察肾组织纤维化;检测受鼠肾组织血红素氧合酶1(HO-1)mRNA的表达.结果 与BS组和CsA组相比较,联合组GVHD程度较轻,体重下降较少且恢复较快.移植后30d时BS组累积存活率为36.8％,联合组为69.6％,CsA组为53.5％,差异有统计学意义(P＜0.05).联合组小鼠肝脏、小肠和皮肤组织可见细胞水肿、变性均较轻,坏死少见,炎症细胞浸润较少,其病理改变较BS组和CsA组轻.联合组几乎无肾间质小管纤维化,而CsA组则较明显.联合组HO-1 mRNA表达高于BS组,CsA组HO-1 mRNA表达低于前两组(P＜0.05).结论 CsA和CoPP联合应用可增强CsA抗GVHD效应,并可减轻CsA的不良反应,其机制可能与HO-1表达上调有关.     

利用脊柱骨来源骨髓细胞建立急性移植物抗宿主病模型

目的探讨利用脊柱骨来源骨髓细胞建立小鼠异基因造血干细胞移植(allogeneic hematopoietic stem cell transplantation,Allo-HSCT)急性移植物抗宿主病(aGVHD)模型的可行性。方法选择C57BL/6(H-2b)雄性小鼠为供体鼠,BALB/c(H-2d)雌性小鼠为受体鼠。制备供体鼠的脾细胞和脊柱骨来源骨髓细胞悬液。受体鼠采用药物加小剂量辐照的预处理方式,于移植前8d～移植前4d腹腔注射氟达拉滨(200mg/kg),接着移植前3d～移植前1d腹腔注射环磷酰胺(60mg/kg),最后在移植前进行全身照射(total-body irradiation,TBI),照射剂量为4Gy(戈瑞)。18只受体鼠经预处理后随机分为3组,每组6只:(1)骨髓移植组,只输入1×107个脊柱骨来源骨髓细胞;(2)aGVHD组,输注1×107个脊柱骨来源骨髓细胞和5×106个脾细胞,建立aGVHD模型;(3)空白对照组,不输入任何细胞。观察3组小鼠生存状态及存活率,取aGVHD组与骨髓移植组存活21d的受体鼠进行病理学检查,取aGVHD组移植后21～28d存活的小鼠的脾脏进行流式细胞术检测骨髓细胞嵌合度。结果骨髓移植组小鼠全部存活,可重建造血,单纯输注骨髓细胞不会诱发aGVHD。aGVHD组小鼠出现aGVHD表现,100%发生aGVHD相关死亡,中位生存期为18d;病理检查结果显示符合aGVHD病理表现,移植后21～28d存活的小鼠诊断为供受体混合嵌合状态,符合aGVHD诊断标准。结论用脊柱骨来源骨髓建立的aGVHD模型完全符合标准,且更加经济,适合大规模建模。     

异基因造血干细胞移植后并发中枢神经系统急性移植物抗宿主病1例并文献复习

目的观察急性白血病患者接受异基因造血干细胞移植（allogeneic hemotopoietic stem cell transplantation,allo-HSCT）后并发中枢神经系统急性移植物抗宿主病（aGVHD）的临床特点。方法回顾性分析1例急性髓细胞白血病（acute myeloid leukemia,AML）M5b型患者于allo-HSCT后并发中枢神经系统aGVHD的临床资料。结果患者接受allo-HSCT后40d突发癫,根据头颅磁共振成像（MRI）表现以及相关科室会诊意见,考虑中枢神经系统aGVHD,经大剂量甲泼尼龙冲击治疗后病情好转,1个月后复查头颅MRI示病灶基本消失。结论急性白血病接受allo-HSCT后发生中枢神经系统aGVHD较少见,临床表现缺乏特异性,加强监测排除其他疾病,确诊后及早使用肾上腺皮质激素（激素）,可使患者获得较好的转归。     

Impact of thrombophilic gene mutations and graft-versus-host disease on thromboembolic complications after allogeneic hematopoietic stem-cell transplantation

BACKGROUND: Hemostatic events in patients undergoing allogeneic hematopoietic stem-cell transplantation (HSCT) increase the morbidity and mortality in this cohort. Little is known about the impact of graft-versus-host disease (GvHD) or of thrombophilic gene mutations/polymorphisms on these complications. STUDY DESIGN: Eighty-nine allogeneic stem-cell recipients and their donors were evaluated prospectively for the presence of the factor V G1691A mutation, the prothrombin G20210A mutation, the 5,10-methylenetetrahydrofolate-reductase (MTHFR) C677T mutation, the glycoprotein IIIa PI(a1/a2) polymorphism, the fibrinogen-beta-chain 455G/A polymorphism, the plasminogen activator inhibitor-1 -675 4G/5G polymorphism, and the angiotensin-converting enzyme intron 16 I/D polymorphism. These mutations/polymorphisms and GvHD parameters were correlated to hemostatic and toxic complications after transplantation. The data were compared with those of 128 healthy controls. RESULTS: The PAI-1 4G/4G polymorphism increases the risk for catheter thrombosis after HSCT 5.7-fold (32.2% vs. 71.4%, P<0.05). In patients with hepatic veno-occlusive disease, the frequency of the PAI-1 4G allele is also increased (83.3% vs. 55.1%, NS). Thrombophilic mutations/polymorphisms in donors do not influence complications in the corresponding recipients. The MTHFR TT genotype does not modify severity and duration of mucositis and aplasia in patients receiving methotrexate prophylaxis. Patients with chronic GvHD have a higher risk of thromboembolism (12.9% vs. 1.7%, P<0.05). CONCLUSION: Thrombophilic gene mutations have only a moderate influence on hemostatic complications in patients undergoing HSCT. This may be because of the overwhelming immunologic impact of GvHD on hemostasis in the allogeneic transplantation setting.     

Factors influencing T-lymphopoiesis after allogeneic hematopoietic cell transplantation

In adult recipients of allogeneic hematopoietic cell transplants (HCT) studied at 1 year after grafting, there was a significant correlation between the counts of T cell receptor excision circle (TREC)-containing CD4 T cells (presumed recent thymic emigrants) and the counts of total T cells (r=0.65, P<0.001). Thus, the reconstitution of CD4 T cell pool depends on T cell generation from hematopoietic stem cells (T-lymphopoiesis). We evaluated factors that could affect T-lymphopoiesis. Low TREC-containing CD4 T cell counts were associated with older patient age (r=-0.41, P=0.01) but not with donor age, graft type (marrow vs. blood stem cells), CD34 cell dose, conditioning (with vs. without irradiation), acute graft-versus-host disease (aGVHD), or chronic graft-versus-host disease (cGVHD) in multivariate analysis. We conclude that patient age is the primary determinant of CD4 T-lymphopoiesis after allogeneic HCT.     

Activated circulating dendritic cells after hematopoietic stem cell transplantation predict acute graft-versus-host disease

BACKGROUND: Dendritic cells (DC) are central to the development of acute graft-versus-host disease (GVHD) following allogeneic hematopoietic stem cell transplantation (alloHSCT). We hypothesized that DC activation status determines the severity of GVHD and that activated DC may be detected in the circulation prior to clinical presentation of GVHD. METHODS: Following transplant, blood samples were obtained twice weekly from alloHSCT patients. Myeloid (CD11c+) and plasmacytoid (CD123hi) DC were enumerated by flow cytometry, and activated myeloid DC were identified using the CMRF-44 monoclonal antibody. RESULTS: Of 40 alloHSCT patients, 26 developed acute GVHD. Severity of GVHD was associated with low total blood DC counts (P=0.007) and with low myeloid and plasmacytoid DC numbers (P=0.015 and 0.003). The CMRF-44 antigen was expressed on blood CD11c+ DC in all cases prior to GVHD onset, whereas of the 14 patients without GVHD, seven had no CMRF-44+ CD11c DC. Patients with CMRF-44+ CD11c+ DC in more than 20% of samples were more likely to subsequently develop acute GVHD (P=0.001, odds ratio=37.1), while patients who developed grade 2-4 GVHD had prior higher percentages of CMRF-44+ CD11c+ DC compared to grade 0-1 GVHD patients (P=0.001). CMRF-44 expression on >7.9% CD11c+ DC predicted for subsequent development of GVHD with a sensitivity of 87.5% and specificity of 79.2%. CONCLUSIONS: Activation status, as assessed by CMRF-44 antigen expression, of blood CD11c+ DC is highly associated with acute GVHD and these cells may be targets for therapeutic intervention.     

High serum leptin in patients with chronic graft-versus-host disease after hematopoietic stem cell transplantation

BACKGROUND: Increased serum leptin has been described after various organ transplants, with a mechanism that is still unclear. METHODS: We measured serum leptin in 60 patients before and after allogeneic (allo) or autologous (auto) stem cell transplant (SCT) and in 60 healthy controls, matched for age and body mass index (BMI). RESULTS: Serum leptin was higher in patients after SCT than before and in controls. Leptin production was higher after allo- than after auto-SCT; the presence of chronic graft-versus-host disease (cGVHD) was associated with the highest values. The physiological correlation with BMI was lost in the allogeneic setting, indicating a strong influence of factors other than the nutritional status on circulating leptin. No relationship was found between serum leptin levels and time from transplant, age, cortisol, C-reactive protein, and T-lymphocyte CD4-to-CD8 ratio. Among the cytokines secreted by type-1/type-2 T-helper lymphocytes, only serum interferon-gamma significantly correlated with serum leptin levels. Anti-leptin blocking antibodies partially inhibited T-cell activation in mixed lymphocyte reaction, suggesting a link between leptin and T-lymphocyte activation in the allo-SCT setting. CONCLUSION: Taken together, these findings suggest that increased serum leptin concentrations may contribute to T-cell activation during development of cGVHD.     

T helper 17 and regulatory T-cell profile and graft-versus-host disease after allogeneic hematopoietic stem cell transplantation in pediatric patients with beta-thalassemia

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is an effective treatment option for hereditary hemoglobin disorders, such as beta-thalassemia; However, this procedure is not without constraints, mainly engendering complications such as acute graft-versus-host disease (aGvHD), chronic GvHD (cGvHD), and susceptibility to infections. The clinical outcomes of allo-HSCT are highly dependant on the quality and quantity of T-cell subsets reconstitution. Following the allo-HSCT of six pediatric patients afflicted with beta-thalassemia, their mononuclear cells were isolated, and then cultured with a combination of phorbol myristate acetate (PMA)/ionomycin and Brefeldin A. The content of CD4 T-cell subsets, including T helper 17 (Th17) cells and regulatory T cells (Tregs), were determined by specific conjugated-monoclonal antibodies three and six months post-HSCT. An increased frequency of total CD4 T-cells, Tregs and Th17 cells was observed at day 90 and 180 after allo-HSCT, albeit the numbers were still lower than that of our healthy controls. In patients who developed cGvHD, a lower Th17/Treg ratio was observed, owing it to a decreased proportion of Th17 cells. In conclusion, creating balance between Th17 and Treg subsets may prevent acute and chronic GvHD in patients after allo-HSCT.     

Antineutrophil cytoplasmic antibody-associated glomerulonephritis in chronic graft-versus-host disease after allogenic hematopoietic stem cell transplantation

Graft-versus-host disease (GVHD) is one of the most frequent complications that occur after hematopoietic stem cell transplantation (HSCT). Recently, renal involvement, including membranous nephropathy, focal segmental glomerulosclerosis, and minimal change disease, has been described as a manifestation of chronic GVHD. This case report describes a patient who developed antineutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis after HSCT. Following preparation with chemotherapy, a 29-year-old man with chronic myeloid leukemia underwent allogenic peripheral blood stem cell (PBSC) transplantation, after which first acute and then chronic GVHD developed. Treatment with prednisone resulted in improvement in the patient's GVHD. After the termination of steroid therapy and about 10 months after PBSC transplantation, nephritic syndrome appeared and the patient's serum creatinine value increased to 1.7 mg/dL. Laboratory evaluation revealed perinuclear antineutrophilic cytoplasmic antibody (p-ANCA) in the serum. Histological examination of renal biopsy tissue showed focal segmental proliferative glomerulonephritis with glomerulosclerosis in 20% of available glomeruli, large cellular crescents in 6% of glomeruli, and no staining of immunoglobulins or complement along the capillary walls. Electron microscopy revealed no immune deposits. After treatment with prednisone 60 mg/d, diltiazem 120 mg/d, and enalapril 10 mg/d, the proteinuria gradually decreased, and p-ANCA was undetectable. These findings suggest that in this patient the ANCA-associated glomerulonephritis was associated with renal involvement that occurred during the course of chronic GVHD.     

Acute graft-versus-host disease of the lung after liver transplantation

We describe the first case of a man who developed acute graft-versus-host disease (GVHD), isolated to the lung, after an orthotopic liver transplant from a female donor. Our patient experienced dyspnea, worsening hypoxemia, and a progressive obstructive ventilatory defect 12 days after liver transplantation. Open-lung biopsy revealed grade 2 lymphocytic bronchiolitis, the pathologic and immunologic correlate of acute pulmonary GVHD. Fluorescent in situ hybridization confirmed donor cells at sites of peribronchiolar inflammation. High-dose corticosteroids were given with a return to baseline pulmonary function. The current case should alert clinicians to investigate pulmonary GVHD as a potential cause of postoperative dyspnea in liver transplant recipients. (Liver Transpl 2002;8:968-971.)     

Mycophenolate mofetil and cyclosporine as graft-versus-host disease prophylaxis after allogeneic blood stem cell transplantation

BACKGROUND: Mycophenolate mofetil (MMF) is an inhibitor of purine nucleotide de novo synthesis leading to impaired proliferation of activated lymphocytes. Studies in animals show a synergistic effect of MMF and cyclosporine (CsA) in preventing acute graft-versus-host disease (GVHD) after allogeneic bone marrow transplantation. We performed a pilot study evaluating the feasibility of the combined application of MMF and CsA as GVHD prophylaxis after allogeneic blood stem cell transplantation. Toxicity and the bioavailability of MMF in this setting were investigated. METHODS: Fourteen patients who had received grafts from HLA-compatible siblings received 2 g of oral MMF from day 1 to 14 combined with intravenous CsA at 4 mg/kg starting at day-1. Plasma levels of mycophenolic acid (MPA) and its glucoronide were measured by high-performance liquid chromatography. Fifteen patients treated with a combination of CsA and methotrexate at the same institution were referred to as the control group. RESULTS: Trilineage engraftment was achieved in all study and control patients. Acute GVHD > or = grade II was observed in 46.5% and 60% of the study and control patients, respectively. No major differences in the rate of acute toxicities were detectable. The mean trough blood level of MPA in 10 patients was 0.28 microg/ml, and 5.7 microg/ml for MPA glucoronide. Reduced peak levels of MPA indicate a reduced absorption rate of MMF in the early posttransplant phase. CONCLUSIONS: The combined administration of MMF and CsA was shown to be feasible in patients after allogeneic blood stem cell transplantation. Because of the decreased bioavailability of MMF, dose-finding studies for an intravenous formulation are warranted.     

Regulatory T-cells in the graft and the risk of acute graft-versus-host disease after allogeneic stem cell transplantation

BACKGROUND: FOXP3+ regulatory T-cells (Treg) are important regulators of allo-reactivity and may therefore represent an important predictor for the risk of graft versus-host disease (GVHD) after allogeneic stem cell transplantation. METHODS: To determine the clinical significance of Treg-content in stem cell grafts, we analyzed 58 human leukocyte antigen (HLA)-identical sibling donors (34 patients received myeloablative and 24 patients reduced intense conditioning regimens) and correlated the Treg frequency with clinical outcome after stem cell transplantation (SCT). RESULTS: A mean value of 9.1 x 10(6) CD4+ FOXP3+ Treg per kg body weight (bw) of the recipient was transplanted (ranging from 0.7 to 33.7 x 10(6) Treg/kg bw). Graft content of Treg correlated with mononuclear cells and CD3+ T-cells. Patients receiving low numbers of Treg (Treg(low)) after myeloablative conditioning for SCT had a significantly increased cumulative incidence of 76% for acute GVHD when compared with 23% for individuals receiving high numbers of Treg (Treg(high)). This observation, however, was not made in patients after reduced intense conditioning-SCT. Notably, relapse rate was not significantly different between Treg(low) and Treg(high) patients in either patient group and overall survival was even increased in Treg(high) patients after myeloablative SCT. Finally, low Treg graft levels represent an independent prognostic factor in multivariate analysis for the appearance of acute GHVD. CONCLUSION: Donor-derived Treg might be of particular significance for the development of acute GVHD after myeloablative SCT using HLA-identical sibling donors.     

Hepatitic graft-versus-host disease after hematopoietic stem cell transplantation: clinicopathologic features and prognostic implication

BACKGROUND: Graft-versus-host disease (GVHD) of the liver after allogeneic hematopoietic stem cell transplantation classically presents with increased bilirubin and alkaline phosphatase (ALP) levels. A hepatitic variant was recently recognized, with more than a 10-fold increase in aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels. This study defines the clinicopathologic features and prognostic implications of hepatitic GVHD compared with classic liver GVHD. METHOD: A total of 38 cases of hepatitic GVHD, 68 cases of classic liver GVHD, and 13 cases of hepatitis B virus (HBV)-related hepatitis after hematopoietic stem cell transplantation were analyzed. RESULTS: Hepatitic GVHD cases showed significantly higher ALT, AST, and ALP levels compared with classic liver GVHD cases (at onset, mean ALT: 154 vs. 58 U/L, P <0.001; AST: 167 vs. 77 U/L, P <0.001; at peak, ALT: 435 vs. 112 U/L, P <0.001; AST: 587 vs. 150 U/L, P <0.001; ALP: 416 vs. 238 U/L, P =0.001), persisted longer (74 vs. 32 days, P =0.006), and showed more lobular pathologic changes in biopsy (lobular changes: 16/26 vs. 4/19, P =0.007; hepatocyte necrosis: 16/26 vs. 6/19, P =0.008; acidophil bodies: 15/26 vs. 4/19, P =0.014) but less cholestasis (4/26 vs. 8/19, P =0.045). However, cumulative doses of immunosuppressants prescribed, response, and outcome were similar. Compared with hepatitic GVHD, HBV-related hepatitis occurred later (95 vs. 184 days, P =0.049), but clinical and biochemical profiles were similar, requiring liver biopsies for their distinction. CONCLUSIONS: Hepatitic and classic liver GVHD differed biochemically and pathologically, but these differences showed no obvious impact on outcome. The distinction of hepatitic GVHD from other hepatitis is mandatory.