三种细胞因子与白血病异基因造血干细胞移植后移植物抗宿主病的关系(英文)

背景：目前转化生长因子α与移植物抗宿主病关系的报道较少，急、慢性移植物抗宿主病患者转化生长因子α与多种因子联合的检测值得研究。目的：分析血清肿瘤坏死因子α、白细胞介素4和转化生长因子α在白血病患者异基因造血干细胞移植后的变化与发生移植物抗宿主病轻重程度的关系。设计：病例对照观察。单位：解放军总医院第二附属医院器官移植中心血液科及解放军总医院第二附属医院核医学科。对象：选择2005—06／2007—06在解放军总医院第二附属医院器官移植中心血液科及解放军307医院移植科住院首次接受异基因造血干细胞移植的42例白血病患者，男23例，女19例，年龄16～68岁，平均35岁。其中急性粒细胞性白血病12例，急性淋巴细胞性白血病15例，慢性粒细胞性白血病15例，移植患者中行外周血移植37例、骨髓移植5例。异基因造血干细胞移植后发生急性移植物抗宿主病20例（Ⅰ～Ⅱ度18例、Ⅲ～Ⅳ度3例），未发生急性移植物抗宿主病21例，发生慢性移植物抗宿主病14例（局限型5例、广泛型9例），未发生慢性移植物抗宿主病28例。选择30名同期到院健康体检者为正常对照组，男18名，女12名，年龄20～70岁，平均44岁。所有受试对象均对实验项目知情同意，实验经过医院伦理委员会批准。方法：用放射免疫分析法检测并比较白血病异基因造血干细胞移植患者及正常对照者血清肿瘤坏死因子α、白细胞介素4和转化生长因子α水平，同时比较移植后发生或未发生急性移植物抗宿主病、发生或未发生慢性移植物抗宿主病及发生轻重程度的白血病异基因造血干细胞移植患者3种细胞因子水平。主要观察指标：血清肿瘤坏死因子α、白细胞介素4、转化生长因子α水平比较。结果：白血病患者42例及健康对照者30名均进入结果分析。①急、慢性移植物抗宿主病患者血清肿瘤坏死因子α、白细胞介素4、转化生长因子α水平均高于正常对照组，差异有显著性意义（P〈0．05～0．01），未发生急性移植物抗宿主病患者血清肿瘤坏死因子α、白细胞介素-4水平高于正常对照组，差异有显著性意义（P〈0．01，0．05）。急性移植物抗宿主病患者血清肿瘤坏死因子α、白细胞介素4、转化生长因子α水平高于未发生者，差异有显著性意义（P〈0．05）。慢性移植物抗宿主病患者血清肿瘤坏死因子α、白细胞介素4、转化生长因子α水平高于未发生者，差异有显著性意义（P〈0.05）。②急性移植物抗宿主病Ⅲ-Ⅳ度或慢性移植物抗宿主病广泛型患者血清肿瘤坏死因子α及转化生长因子α水平均高于急性移植物抗宿主病Ⅰ—Ⅱ0或慢性移植物抗宿主病局限型，差异有显著性意义（P〈0．05～0．01）。结论：异基因造血干细胞移植术后动态监测肿瘤坏死因子α、白细胞介素4和转化生长因子α水平变化有助于对发生急性或慢性移植物抗宿主病预测，对判断移植物抗宿主病的轻重程度有一定意义。

Setting up the Model of Xeno-graft Verse Host Disease

Objective: To observe human to mouse one way mixed lymphocyte(MLC); And to set up the xeno-grats verse host disease Xeno-graft host disease(XGVHD) model,probing its immunologic mechamism.Methods: Mouse splenic lymphocyte were collected in asepsis and treated by mitomycin as activating cell. Human Peripheral blood lymphocytes (hPBL)were separated and gathered as reacting cell; Mouse splenic lymphocyte and hPBL were mixed to incubate for a week. Destroying recipient (mouse) immune system by total body irradiation (TBI) and intraperitoneal injecting CTX、MTX; Separating and collecting hPBL; Injecting hPBL to mouse by caudal vein. Results; ①HPBL in the experiment groups(mixed mouse lymphocyte) proliferated obviously, the amount of 3H-TdR in corporation increased evidently(P<0.05); The mean percentage of CD 4、CD 8、IgG 、IgM positive cells rose markedly. ②Experiment groups,the hPBL were found in the spleen and kidney tissue, fas protein expressing, we occasionally discovered and apoptosis cells.Conclusion: The human to mouse one-way MLC has obvious lymphocyte proliferation. By these means,we succeed in inducing XGVHD and setting up a XGVHD model.

Graft-versus-host disease after liver transplantation:A comprehensive literature review

AIM： To determine the factors affecting mortality in pa- tients who developed graft-versus-host disease （GvHD） after liver transplantation （LT）. METHODS： We performed a review of studies of GvHD following LT published in the English literature and ac- cessed the PubMed, Medline, EBSCO, EMBASE, and Google Scholar databases. Using relevant search phras- es, 88 articles were identified. Of these, 62 articles con- raining most of the study parameters were considered eligible for the study. Risk factors were first examined using a univariate Kaplan-Meier model, and variables with a significant association （P 〈 0.05） were then sub- jected to multivariate analyses using a Cox proportional- hazards model. RESULTS： The 61 articles reported 87 patients, 58 male and 29 female, mean age, 40.4 ± 15.5 years （range： 8 mo to 74 years）, who met the inclusion criteria for the present study. Deaths occurred in 59 （67.8%） patients, whereas 28 （32.2%） survived after a mean follow-up period of 280.8 ± 316.2 d （range： 27-2285 d）. Among the most frequent symptoms were rash （94.2%）, fever （66.6%）, diarrhea （54%）, and pancytopenia （54%）. The average time period between LT and first symptom on- set was 60.6 ± 190.1 d （range： 2-1865 d）. The Kaplan- Meier analysis revealed that pancytopenia （42.8% vs 59.3%, P = 0.03）, diarrhea （39.2% vs 61.0%, P = 0.04）, age difference between the recipient and the donor （14.6 ± 3.1 years vs 22.6 ± 2.7 years, P 〈 0.0001）, and time From first symptom occurrence to diagnosis or treatment （13.3 ± 2.6 mo vs 15.0 ± 2.3 mo, P 〈 0.0001） were significant factors affecting mortality, whereas age, sex, presence of rash and fever, use of immunosuppressive agents, acute rejection before GvHD, etiological causes, time of onset, and donor type were not associated with mortality risk. The Cox proportional-hazards model, de- termined that an age difference between the recipient and donor was an independent risk Factor （P = 0.03; hazard ratio, 7.395, 95% confidence interval, 1.2-46.7）. CONCLUSION： This study showed that an age differ- ence between the recipient and donor is an independent risk factor for mortality in patients who develop GvHD after LT.

Gastrointestinal and hepatic complications of hematopoietic stem cell transplantation

Recognition and management of gastrointestinal and hepatic complications of hematopoietic stem cell transplantation has gained increasing importance as indications and techniques of transplantation have expanded in the last few years.The transplant recipient is at risk for several complications including conditioning chemotherapy related toxicities,infections,bleeding,sinusoidal obstruction syndrome,acute and chronic graftversus-host disease(GVHD) as well as other long-term problems.The severity and the incidence of many complications have improved in the past several years as the intensity of conditioning regimens has diminished and better supportive care and GVHD prevention strategies have been implemented.Transplant clinicians,however,continue to be challenged with problems arising from human leukocyte antigen-mismatched and unrelated donor transplants,expanding transplant indications and age-limit.This review describes the most commonly seen transplant related complications,focusing on their pathogenesis,differential diagnosis and management.

Alpha-GalCer Administration after Allogeneic Bone Marrow Transplantation Improves Immune Reconstitution in Mice

Objective To explore the effect of α-galactosyleramide （α-GalCer） on immune reconstitution under acute graft-versus-host disease （aGVHD）. Methods BALB/c mice were transplanted with allogeneic C57BL/6 bone marrow cells and splenocytes （both 1 × 10^7） after receiving lethal total-body irradiation, α-GalCer （100 ug/kg） or vehicle （dimethyl- sulfoxide） was administered intraperitoneally immediately after transplantation. The effects of α-GalCer on immune reconstitufion, proliferation of T cells and B cells, hematopoiesis, and thymic microenvironment were assessed. Results The α-GalCer group exhibited higher percentages of CD3^＋, CD4^＋, CD8^＋, B220^＋, CD40＋, and CD86＋cells compared with the vehicle group. The number of colony forming unit per 1000 CD34^＋ cells in the et-GalCer group was higher than in the vehicle group （P=0.0012）. In vitro proliferation assays showed that the α-GalCer group had higher percentages of CD3^＋, CD4^＋, CD8^＋, and B220^＋ cells compared with the vehicle group. As for the results of in vivo proliferation assays, the numbers of CD3^＋, CD4^＋, CD8^＋, and B220^＋ cells were higher in the α-GalCer group than in the normal group, especially the number of B220^＋ cells （P=0.007）. Significant difference was not found in thymocyte count between the α-GalCer group and the vehicle group, nor in the percentages of CD3^＋, CD4^＋, and CD8^＋ cells. Conclusion Administration of tl-GalCer after allogeneic bone marrow transplantation may promote immune reconstitution in the presence of aGVHD.

Induction of graft-versus-leukemia effect using a mixture of syngeneic plus G-CSF primed haploidentical bone marrow grafts in mice

Objective： To explore whether the graft-versus-leukemia （GVL） effects could be enhanced and acute graft-versus-host disease （aGVHD） could be relieved by syngeneic bone marrow mixed with G-CSF-primed H-2 haploidentical marrow grafting. Methods： Female L615 （H-2^k） mice were recipient mice and male （615×BALB/c） F1 （6BF1） （H-2^k×H-2^d） mice were donors respectively. Donor mice were injected subcutaneously with G-CSF daily at 0.01 μg/g for 6 days, and splenocytes were harvested on day 7. A total of 615 mice were loaded with L615 tumor cells and received 8.5 Gy （^60Co γ-ray） irradiation three days later, followed by a mixed bone marrow transplantation （MBMT）. The allo-grafts consisted of a mixture of syngenetic plus G-CSF-mobilized （control diluents） H-2 haploidetical marrow cells. GVL effects were monitored by survival time and survival rate of recipient mice. GVHD was assessed by clinical signs of weight loss, ruffled fur, diarrhea and histological changes of skin, liver and small intestines. AIIogeneic chimerism was detected using cytogenetic analysis. Enzyme-linked immunosorbent assay （ELISA） method was used to detect cytokines （IL-2, IL-4 and IFN-γ）. Fluorescence-activated cell sorting （FACS） analysis was used to detect T-cell phenotype. Results： （1） The mice merely received L615 leukemia cells were all died of leukemia. The L6t5 mice of 3：1 and 4：1 MBMT groups survived longer than those post syngeneic BMT （P 〈 0.01）. （2） The survival time of mice in the G-CSF-treated MBMT groups was longer than that of non-primed MBMT groups （P 〈 0.05）. Administration of G-CSF-treated 6BF1 mice could markedly increase the survival rate of 3：1 and 4：1 MBMT mice （P 〈 0.01） with little or a little GVHD. （3） As the post-transplanted time prolonged, the rates of allogeneic chimerism were decreased. The chimerism rates decreased to zero when the mice died of leukemia relapse. （4） L3T4^＋ cells and relative ratio in both subsets were significantly reduced in G-CSF-treated donor mice. After G-CSF-treated donors, splenocytes from recipients at day 14 post-MBMT showed an increased production of IL-4 and a decreased production of IL-2 and IFN-γ. Conclusion： Syngeneic bone marrow mixed with H-2 haploidentical marrow grafts had a potential way to increase GVL effects, and this GVL effects could be enhanced with little or a little GVHD by G-CSF pretreatment of donors. Improved survival in recipients of G-CSF-mobilized donors is associated with increased IL-4 production and decreased IL-2 and IFN-γ production.

Haploidentical stem cell transplantation used in treating primary plasma cell leukemia: a case report

Here we report a successful protocol in treatment of a patient with primary plasma cell leukemia (PPCL) using haploidentical stem cell transplantation (hi-HSCT). During first complete remission after routine chemotherapy, the patient received autologous blood stem cell transplantation, but he had relapse later. He gained a second CR after chemotherapy and underwent hi-HSCT from his daughter, who had HLA mismatched at three loci. Recovery of hemopoiesis was found at day 14 and complete donor chimerism was confirmed by PCR-STR on day 34, 95 and 238. The patients have survived disease-free for 56 months since hi-HSCT, without serious graft-versus-host-disease.

Might liver transplantation recipients with primary hepatocellular carcinoma benefit from GVT effect of aGVHD?

We aimed to access if acute graft-versus-host disease （aGVHD） in liver transplantation recipients of hepatocellular carcinoma （HCC） might develop a graft-versus-tumor effect （GVT） other than immunological damage which would benefit prophylaxis of tumor recurrence. Methods： Dynamic observation of 3 cases of liver transplantation recipients of HCC and cirrhosis, which developed manifestations of fever, skin rash, watery diarrhea, pancytopenia and were finally diagnosed as aGVHD. Two of which got recovered from intravenously pulse methylprednisolone, high-dose intravenous immunoglobulin, antibiotics administration simultaneously and promptly withdrawal of oral immunosuppressants. Two survivors were follow-up regularly with biological monitoring and imaging surveillance for tumor recurrence thereafter. Results： Two recipients survived healthily with stable liver graft function and normal serum AFP level and blood routine test. No sign of tumor recurrence was found in repeat imaging examinations for liver graft, lung, brain and other tissue or organs within a period of 96 months and 17 months to date, respectively. Conclusien： Despite of the fatal damage to according organs and tissue, it suggest that aGVHD in liver recipients of HCC may also develop a GVT effect and benefit prophylaxis of tumor recurrence and result in a long-term healthy recipients survival.

Protective effect of human CD40-Ig fusion protein in a murine model of acute graft-versus-host disease

Abstract:Objective To investigate the protective effects of blocking CD40/CD40L interactions with human CD40-Ig fusion protein in a murine graft-versus-host disease model.Methods Human CD40 gene extracellular region was inserted into plasmid pIG1, which contains genomic human IgG1 Fc gene. A transient vector containing CD40-Fc fusion gene was transfected into COS-7 cells. The CD40-Ig fusion protein was detected through enzyme-linked immunosorbent assay (ELISA). A constitutive vector was also generated by ligating the CD40-Fc fusion gene into pcDNA3.1 and transfecting it into CHO cells. CD40-Ig was purified by protein A affinity chromatography. SDS-PAGE, Western blot and ligand binding assay were used to identify the qualities of CD40-Ig. Murine acute graft-versus-host disease (GVHD) was induced by intravenous injection of C57BL/6J (H-2b) spleen cells into sub-lethally irradiated BALB/c (H-2d) mice. Protective effects against murine graft-versus-host disease by in vivo administration of CD40-Ig were evaluated.Results Mammalian expression vectors pIG/40Ig and p3.1/40Ig were constructed as described above. Chimeric proteins were expressed in COS-7 and CHO cell culture supernatant and confirmed by ELISA and Western blot. SDS-PAGE showed that fusion proteins had a disulfide-bonded dimeric structure and existed as homodimer. Purified CD40-Ig could bind to CD40L. In vivo administration of CD40-Ig could prevent the development of GVHD and significantly prolong the mean survival time of mice with graft-versus-host disease.Conclusions These results demonstrate that CD40/CD40L interactions play an important role in the pathogenesis of graft-versus-host disease and suggest clinical potential for CD40-Ig in the prevention and treatment of human graft-versus-host disease.

Stable clonal expansion of the T-cell receptor Vβ6, Vβ17 and Vβ19 T cells in a cGVHD case using genescan analysis

Objective To investigate the distribution and clonality of the T-cell receptor (TCR) Vβ repertoire in chronic graft versus host disease (cGVHD).Methods The complementarity determining region 3 (CDR3) of the TCRβ gene with 24 variable regions was amplified in peripheral blood mononuclear cells drawn from one cGVHD patient after allogenic bone marrow transplantation (allo-BMT) 35, 39, 43 or 45 months respectively, using RT-PCR, to observe the expression of TCR Vβ repertoire T cells. The PCR products were further analyzed by genescan to evaluate clonality of T cells. Ressults Fourteen or 16 TCR Vβ subfamily T ceils were detected in each sample of cGVHD case. Oligoclonal T cells were identified in TCR Vβ 6, 16, 17, 19 and 21 subfamilies. The stable clonal T cells in all samples were identified in Vβ6, Vβ17 and Vβ21 subfamilies.Conclusion Skewing distribution and stable clonal expansion of T cells can be found in cGVHD cases and it may be related to the initiation of cGVHD.

Allogeneic peripheral blood stem cell transplantation for leukemia

Objective To observe engraftment kinetics, the incidence and severity of graft-versus-host disease (GVHD), and clinical outcome on 40 recipients undergoing allogeneic peripheral blood stem cell transplantation (allo-PBSCT).Methods From June 1997 to May 1999, forty leukemia patients with a median age of 35 years underwent allo-PBSCT. PBSC were mobilized with G-CSF at a dose of 5 μg/kg s.c. every 12 hours for 5 days. A median of 7.7 (2.0 - 16.8) × 106 CD34+ cells/kg was infused into the recipients. Busulfancyclophosphamide (BU-CY) was used as the conditioning regimen. All patients received cyclosporine A and either methotrexate ( n = 34) or methylprednisolone ( n = 6) for GVHD prophylaxis.Results Engraftment of neutrophils and platelets was achieved at a median of 13 days (9- 28 days) and 12 days (7- 60 days) respectively. Patients receiving ≥4×106 CD34+ cells/kg or given G-CSF post transplant had significantly accelerated neutrophil and platelet engraftment. Acute GVHD occurred in 17 of 40 patients (42.5％), with grade Ⅱ-Ⅳ acute GVHD in 10 patients (25％). Chronic GVHD developed in 21 (9 extensive, 12 limited) out of 30 evaluable patients (21/30, 70％) with a median follow up of 380 days (180-900 days). Transplant related mortality was 17.5％ end the relapse rate was 10％. The probability of leukemia free survival at 3 years was 72.5％.Conclusion Allo-PBSCT can provide rapid hematopoietic reconstitution without an increased incidence of acute GVHD, but may be associated with a high risk of chronic GVHD.

Long-term follow-up of haploidentical transplantation in relapsed/refractory severe aplastic anemia:a multicenter prospective study

In recent decades,haploidentical stem cell transplantation(haplo-SCT)to treat severe aplastic anemia(SAA)has achieved remarkable progress.However,long-term results are still lacking.We conducted a multicenter prospective study involving SAA patients who underwent haplo-SCT as salvage therapy.Long-term outcomes were assessed,mainly focusing on survival and quality of life(QoL).Longitudinal QoL was prospectively evaluated during pretransplantation and at 3 and 5 years posttransplantation using the SF-36 scale in adults and the PedsQL 4.0 scale in children.A total of 287 SAA patients were enrolled,and the median follow-up was 4.56 years(range,3.01–9.05 years)among surviving patients.During the long-term follow-up,268 of 275 evaluable patients(97.5%)obtained sustained full donor chimerism,and 93.4%had complete hematopoietic recovery.The estimated overall survival and failure-free survival for the whole cohort at 9 years were 85.4%±2.1%and 84.0%±2.2%,respectively.Age(≥18 years)and a poorer performance status(ECOG>1)were identified as risk factors for survival outcomes.For Qo L recovery after haplo-SCT,we found that QoL progressively improved from pretransplantation to the 3-year and 5-year time points with statistical significance.The occurrence of chronic graft versus host disease was a risk factor predicting poorer QoL scores in both the child and adult cohorts.At the last followup,74.0%of children and 72.9%of adults returned to normal school or work.These inspiring long-term outcomes suggest that salvage transplantation with haploidentical donors can be routine practice for SAA patients without human leukocyte antigen(HLA)-matched donors.

Virus genomes and virus-host interactions in aquaculture animals

Over the last 30 years,aquaculture has become the fastest growing form of agriculture production in the world,but its development has been hampered by a diverse range of pathogenic viruses.During the last decade,a large number of viruses from aquatic animals have been identified,and more than 100 viral genomes have been sequenced and genetically characterized.These advances are leading to better understanding about antiviral mechanisms and the types of interaction occurring between aquatic viruses and their hosts.Here,based on our research experience of more than 20 years,we review the wealth of genetic and genomic information from studies on a diverse range of aquatic viruses,including iridoviruses,herpesviruses,reoviruses,and rhabdoviruses,and outline some major advances in our understanding of virus–host interactions in animals used in aquaculture.

Potential value of autoantibodies as biomarkers of chronic graft-versus-host disease after allogeneic stem cell transplantation

We investigated the value of autoantibodies as biomarkers of chronic graft-versus-host disease(cGVHD)by analyzing the autoantibody profiles of 65 patients(34 cGVHD and 31 non-cGVHD)surviving longer than three months after allogeneic hematopoietic stem cell transplantation(allo-HSCT).Autoantibodies to at least one antigen were detected in 45 patients(70.8%),with multiple autoantibodies detected in 30 patients(46.2%).Antinuclear antibodies(ANAs)were the most frequently detected autoantibodies,with a significantly higher prevalence in non-cGVHD patients and c GVHD patients than that in healthy controls(HCs).ANA-nucleolar(ANA-N)was the main immunofluorescence pattern of ANA-positivity in both the non-cGVHD and c GVHD groups.There was a higher prevalence of anti-Ro52-positivity in non-cGVHD and cGVHD patients than in HC.Liver cGVHD was significantly associated with anti-Ro52-positivity.However,cGVHD activity and severity were not associated with the presence of autoantibodies.Similarly,there were no significant differences in overall survival or relapse among the four groups of patients expressing autoantibodies.Our results suggest that autoantibodies have limited value in predicting cGVHD.