Antigen recognition by MHC-incompatible cells of a human mismatched chimera

Tetanus toxin (TT)-specific T cell clones of donor origin were obtained from a patient with severe combined immunodeficiency (SCID) successfully reconstituted by transplantation of allogeneic fetal liver and thymus cells from two different donors performed 10 yr ago. A series of these clones recognized TT in the context of "allo" class II HLA determinants expressed by recipient APC. The restriction element of two T cell clones with the HLA phenotype of the first donor (HLA-DR1,8) and one T cell clone with the HLA phenotype of the second transplant (HLA-DR3,9) was HLA-DR4 of the recipient, whereas other T cell clones derived from the second transplant recognized TT in the context of HLA-DR5 of the recipient's APC. These latter T cell clones were not able to proliferate in response to TT when autologous APC were used. These data demonstrate that recipient and donor cells having different HLA phenotypes could cooperate across the allogeneic barrier and that MHC restriction of antigen (Ag) recognition is independent from the MHC genotype of the T cells but is influenced by the environment in which the T cells mature. We also isolated T cell clones that were able to recognize processed TT presented by all allogeneic EBV cell lines tested, indicating that the Ag specificity of these clones was not restricted by a particular class II MHC molecule. The Ag-specific proliferative response of one of these clones could be blocked by anti-class II MHC mAbs. These results demonstrate that in addition to Ag recognition in the context of specific class II MHC Ags, other types of Ag-specific responses may occur in this human chimera. It is not clear whether this "allo" plus Ag recognition is the result of education of transplanted fetal cells in the host thymus. Taking into consideration our previous findings indicating that alloreactive T cell clones specific for the recipient cells could be isolated in vitro from the PBL of the same patient, our data suggest that the mechanism for deletion of self-reactive clones and the generation of MHC-restricted responses are different.     

Autoreactive T cell clones specific for class I and class II HLA antigens isolated from a human chimera

T cell clones of donor origin that specifically react with recipient cells were obtained from a SCID patient successfully reconstituted by allogeneic fetal liver and thymus transplantation performed 10 yr ago. The majority of these clones displayed both cytotoxic and proliferative responses towards PBL and an EBV-transformed B cell line derived from the patient. In addition, these T cell clones had proliferative and cytotoxic responses towards the parental PBL, EBV cell lines, and PHA blasts. Blocking studies with anti-class I and anti-class II HLA mAbs indicated that the activity of the CD4+ T cell clones was specifically directed against class II HLA antigens of the recipient. On the other hand, the cytotoxic and proliferative responses of the CD8+ T cell clones were specific for class I HLA antigens which are ubiquitously expressed on the recipient cells. Thus, the establishment of transplantation tolerance observed in this stable human chimera is not due to the elimination of host-reactive T cells from the repertoire and suggests the presence of a peripheral autoregulatory suppressor mechanism.     

Major histocompatibility restriction of antigen recognition by T cells in a recipient of haplotype mismatched human bone marrow transplantation.

Immune T cells proliferate in response to antigen that is recognized in association with self-Ia determinants. T cells from a patient with severe combined immunodeficiency that has been successfully reconstituted with haplotype-mismatched, maternal bone marrow were studied in an attempt to understand the development of Ia restriction of antigen recognition in man. All the patient's T cells were of maternal origin as determined by HLA typing. The patient received a series of three immunizations with tetanus toxoid (TT) antigen between the 6th and 14th week posttransplant. TT-specific T cell lines were established from the patient's peripheral blood at 6 and 8 mo posttransplantation and were maintained in culture in the presence of irradiated monocytes from the patient, TT antigen, and interleukin-2. HLA typing of the two T cell lines revealed them to be exclusively of donor origin. Both T cell lines could proliferate to TT in the presence of monocytes derived from either the patient's mother or father. In contrast, a TT-specific T cell line obtained from the patient's mother proliferated to TT in the presence of autologous monocytes, but not in the presence of monocytes derived from the patient's father. Studies using monocytes from a panel of HLA-typed donors indicated that the patient's T cell lines proliferated to TT in the presence of monocytes that expressed the paternal DR antigen (HLA-DR4) inherited by the patient but not in the presence of monocytes that expressed the paternal DR antigen (HLA-DR1) not inherited by the patient or in the presence of monocytes bearing irrelevant DR antigens. Monocytes that expressed either one of the two maternal DR antigens (HLA-DR3 and DR5) could support the proliferation of the patient's T cell lines in response to TT antigen. HLA typing of the patient's monocytes at 6 mo post-transplant revealed only recipient HLA-DR antigens (HLA-DR3 and DR4). At 12 mo posttransplant, the patient's monocytes expressed recipient HLA-DR antigens as well as the non-shared HLA-DR5 antigen of donor origin. The results of the present study indicate that T cells of human bone marrow chimera recognized antigen in the context of Ia determinants of recipient origin. The apparent recognition of antigen by the chimera's T cells in the context of donor Ia determinants that were not shared with the recipient is discussed.     

Human hematopoietic cells and thymic epithelial cells induce tolerance via different mechanisms in the SCID-hu mouse thymus

To study the role of thymic education on the development of the human T cell repertoire, SCID-hu mice were constructed with fetal liver and fetal thymus obtained from the same or two different donors. These animals were studied between 7 and 12 mo after transplantation, at which times all thymocytes and peripheral T cells were derived from stem cells of the fetal liver graft. Immunohistology of the thymus grafts demonstrated that thymic epithelial cells were of fetal thymus donor (FTD) origin. Dendritic cells and macrophages of fetal liver donor (FLD) origin were abundantly present in the medullary and cortico-medullary areas. Thymocytes of SCID-hu mice transplanted with liver and thymus of two different donors (FLDA/FTDB animals) were nonresponsive to Epstein-Barr virus-transformed B cell lines (B-LCL) established from both the FLDA and FTDB, but proliferated vigorously when stimulated with third-party allogeneic B-LCL. Mixing experiments showed that the nonresponsiveness to FTDB was not due to suppression. Limiting dilution analysis revealed that T cells reacting with the human histocompatibility leukocyte antigens (HLA) of the FLD were undetectable in the CD8+ T cell population and barely measurable in the CD4+ subset. On the other hand, CD4+ and CD8+ T cells reactive to the HLA antigens of the FTD were readily detectable. These results indicate that FLD-reactive cells were clonally deleted, whereas FTD-reactive cells were not. However, the frequencies of FTD-reactive T cells were consistently twofold lower than those of T cells specific for any third-party B-LCL. In addition, the cytotoxic activity and interleukin 2 production by FTD-specific T cells were lower compared with that of third-party-reactive T cell clones, suggesting that FTD-specific cells are anergic. These data demonstrate that T cells become tolerant to autologous and allogeneic HLA antigens expressed in the thymus via two different mechanisms: hematopoietic cells present in the thymus induce tolerance to "self"-antigens by clonal deletion, whereas thymic epithelial cells induce tolerance by clonal energy and possibly deletion of high affinity clones.     

Influence of obesity on the metabolism of apolipoprotein B in humans.

After successful bone marrow transplantation, patient hematopoietic and lymphoid cells are replaced by cells derived from the donor marrow. To document and characterize successful engraftment, host and donor cells must be distinguished from each other. We have used DNA sequence polymorphism analysis to determine reliably the host or donor origin of posttransplant cell populations. Using a selected panel of six cloned DNA probes and associated sequence polymorphisms, at least one marker capable of distinguishing between a patient and his sibling donor can be detected in over 95% of cases. Posttransplant patient peripheral leukocytes were examined by DNA restriction enzyme digestion and blot hybridization analysis. We have studied 18 patients at times varying from 13 to 1,365 d after marrow transplantation. Mixed lymphohematopoietic chimerism was detected in 3 patients, with full engraftment documented in 15. One patient with severe combined immunodeficiency syndrome was demonstrated to have T cells of purely donor origin, with granulocytes and B cells remaining of host origin. Posttransplant leukemic relapse was studied in one patient and shown to be of host origin. DNA analysis was of particular clinical value in three cases where failure of engraftment or graft loss was suspected. In two of the three cases, full engraftment was demonstrated and in the third mixed lymphohematopoietic chimerism was detected. DNA sequence polymorphism analysis provides a powerful tool for the documentation of engraftment after bone marrow transplantation, for the evaluation of posttransplant lymphoma or leukemic relapse, and for the comprehensive study of mixed hematopoietic and lymphoid chimeric states.     

Circulating red cells usually remain of host origin after bone marrow transplantation for severe combined immunodeficiency

Background: Patients with severe combined immunodeficiency (SCID) treated with allogeneic bone marrow transplantation often receive a milder conditioning regimen than patients who undergo transplantation for hematologic malignancy, and they regularly retain circulating white cells of host origin. The origin of circulating red cells following successful bone marrow transplantation to treat SCID is not known. Study Design and Methods: Review of the medical records identified all patients with SCID who underwent ABO-mismatched bone marrow transplantation at the University of California, San Francisco, between 1982 and 1994. The ABO and Rh phenotype at>6 months after transplantation was determined for all successful transplants by review of the medical record or the taking of a fresh blood sample for analysis. Patient-conditioning and donor bone marrow-preparative regimens were reviewed to assess their possible influence on the red cell phenotype after successful bone marrow transplantation. Results: Nine of 35 SCID patients who underwent successful transplantation received marrow from ABO-mismatched donors. Eight of the nine patients had only host red cells circulating at 6 to 84 months after transplantation, while one patient had only donor red cells circulating at 48 months after transplantation. None of the patients had circulating red cells of both host and donor origin. Conditioning regimens included cyclophosphamide and antithymocyte globulin for all nine patients; only three patients also received total body irradiation. Seven of the nine patients received related-donor, HLA- mismatched bone marrow, and two patients received HLA-identical bone marrow; eight patients received T-cell-depleted bone marrow. The one patient whose red cell phenotype converted to that of the donor received T-cell-depleted, haploidentical marrow, and the preparative regimen included chemotherapy and total body irradiation. Conclusion: SCID patients successfully treated with allogeneic bone marrow transplantation typically fail to show circulating red cells of donor phenotype; this finding is in contrast to the universal presence of circulating donor red cells following successful bone marrow transplantation to treat hematologic malignancies and other diseases. The milder conditioning regimens typically given to patients with SCID, along with T-cell depletion and HLA mismatching, may play a role in this different outcome. It is not known whether the inability to find circulating red cells of donor origin is due to a failure to engraft donor pluripotent stem cells or a failure of engrafted donor stem cells to differentiate along the erythroid lineage.     

非血缘异基因脐血造血干细胞移植现状、问题与对策

非血缘异基因脐血移植（UCBT）自1988年第一例发展至今已经经历了20年。全世界报道脐血移植的病例已经累积10000例以上。大量的临床研究报告和统计学处理结果已有力地证实非血缘异基因脐血移植是有显效的,HLA 0—1位点不合的UCBT的临床TRM、复发率和无复发生存率等数据和HLA全相合骨髓移植的相仿,但UCBT的重度GVHD发生率明显减少。已经公认：异基因脐血无疑是一个造血干细胞随时可取到的新来源,尤其当病人不能及时找到合适的HLA全相合非血缘骨髓供者,也没有单倍体相合的外周血供者时。近年来,异基因脐血移植的两大新发展,一是脐血双份移植,一是脐血移植用于非恶性的难治性疾病治疗（如重症免疫缺损、地中海贫血、骨髓衰竭综合症和代谢障碍病等等）。脐血双份移植用于大体重儿童或成人病例并获得成功报告越来越多,其发展势头超过了儿童单份UCBT病例。双份异基因脐血移植的临床成功实践给有关基础研究提出许多令人困惑而又饶有兴趣的问题,例如HLA互不相合的双份脐血在体内的斗争究竟是什么,为什么不破坏整个移植,反而提高了植入率？双份脐血有核细胞剂量之和并不高于单份脐血移植成功的剂量,而每单份的细胞剂量都低于单份脐血移植成功需要的剂量,这样反而提高了植入率,为什么？双份脐血之间,受者之间的HLA都互不相舍,为什么受者可以在第一阶段容纳双份脐血同时植入,以后双份脐血中只有一份可长期植入,又为什么？那一份脐血细胞能优胜而长期植入的因素是什么？本文参考国际UCBT文献,就如何评价非血缘性异基因UCBT存在的问题,采取对策及其发展前景,加以讨论。     

异基因造血干细胞移植后T细胞再生及其临床意义

异基因造血干细胞移植包括已广泛开展的异基因骨髓移植（Ａｌｌｏ－ＢＭＴ）异基因外周血干细胞移植（Ａｌｌｏ－ＰＢＳＣＴ）以及脐带血血干细胞移植（ＣＢＴ）。它是治愈血液系统恶性肿瘤和其它多种疾病的有效手段。移植后在造血系统恢复的同时,免疫系统也在经历一个重建过程,Ｔ细胞再生通过胸腺依赖途径和胸腺非依赖途径。成人由于胸腺功能退化,Ｔ细胞再生更多是靠成熟Ｔ细胞的外周扩增和胸腺外分化,造成Ｔ细胞库多样性受限,     

Immunomonitoring of graft‐versus‐host minor histocompatibility antigen correlates with graft‐versus‐host disease and absence of relapse after graft

BACKGROUND: After HLA‐identical hematopoietic stem cell transplantation, minor histocompatibility (mH) antigen alloreactivity plays a dominant role in the development of graft‐versus‐host disease (GVHD) and graft versus leukemia (GVL). STUDY DESIGN AND METHODS: We have analyzed the mH alloreactivity (enzyme‐linked immunospot [ELISpot] for interferon‐γ[IFN‐γ] assay) from 24 donor/recipient pairs over a period of 2 years of follow‐up and correlated such alloreactivity with the development of GVHD or absence of relapse. Circulating specific T cells anti‐mH with multimer HLA‐peptides were also studied. RESULTS: We show by ELISpot IFN‐γ assay that alloreactivity during the first 3 months from donor versus recipient or donor versus mismatched identified mH antigens is associated with acute GVHD and GVL effect. In addition, we demonstrate that the donor‐versus‐recipient reactivity observed after the third month is highly associated with chronic GVHD and GVL (p = 0.0007). Finally, we show by multimer HLA‐peptide assay that mH epitope‐specific T cells present after 3 months are statistically related to the GVL effect. CONCLUSIONS: Our results provide a robust method to monitor mH antigen graft‐versus‐host reaction and suggest that current identified mH have predictive value on GVHD and GVL.     

供者活化型KIR2DS5在单倍体相合造血干细胞移植预后中的意义

本研究探讨供者杀伤细胞免疫球蛋白样受体（KIR）和受者人类白细胞抗原（HLA）基因型在单倍体相合造血干细胞移植（HSCT）预后中的意义。26例患者接受单倍体相合HSCT,均未进行体外T细胞去除（TCD）。采用序列特异性引物聚合酶链式反应（PCR-SSP）分型技术对供、受者HLA和供者KIR基因型进行检测。根据供者KIR和受者HLA-Bw4组、-C1组、-C2组等位基因分析结果进行供受者KIR/HLA分组。对KIR/HLA不合与相合组、Bw4不合与相合组、C2不合与相合组、Bw4和C2均不合组与至少其一相合组在造血重建、移植物抗宿主病（GVHD）、无病生存（DFS）、移植后感染及移植相关死亡（TRM）等方面的差异进行比较;并就供者活化型KIR对单倍体相合HSCT预后的影响进行评价。结果表明：各组在造血重建、急性或慢性GVHD发生率、DFS、移植后感染及TRM等方面均无显著性差异（p均〉0.05）,但C2不合组发生严重GVHD4例。供者活化型KIR2DS5阳性组2年DFS明显优于阴性组,分别为（85.7±13.2）%、（31.2±12.8）%,p〈0.05〕。结论：供者KIR及受者HLA基因型对单倍体相合HSCT预后有影响,供者携带活化型KIR2DS5可能有助于提高患者的DFS。     

Transfusion‐associated graft‐versus‐host disease in a liver transplant recipient: an unusual presentation and review of the literature

BACKGROUND: Transfusion‐associated graft‐versus‐host disease (TA‐GVHD) is a rare, nearly universally fatal complication from transfusion of nonirradiated cellular blood components, occurring when a recipient's immune system is unable to recognize and destroy transfused T lymphocytes. Irradiation of cellular components eliminates this risk. We present an unusual case of a liver transplant recipient developing TA‐GVHD 13 weeks after transfusion of a random unit of nonirradiated red blood cells (RBCs) that happened to be from a donor homozygous for an HLA haplotype shared by the patient. STUDY DESIGN AND METHODS: This study was a single case review of a liver transplant recipient who developed skin GVHD and marrow aplasia. Clinical course and the chimerism studies involving the patient, the liver donor, and the blood donor are detailed. RESULTS: The patient presented 3 months posttransplant with GVHD of his skin and marrow aplasia. In addition to standard antigraft immunosuppression, this patient had started the interleukin‐1 receptor antagonist anakinra on Posttransplant Day 13 for an acute gout flare. Chimerism studies on the patient's peripheral blood identified a population of CD3 cells that did not originate with either the patient or his liver donor. HLA studies and microsatellite profiling of the unknown CD3 population identified the source of the patient's TA‐GVHD, a unit of nonirradiated, nonleukoreduced apheresis RBCs. CONCLUSION: Use of an immunomodulating agent may have contributed to the development of TA‐GVHD in a liver transplant patient who received a random unit of nonirradiated RBCs by chance from an unrelated haploidentical donor.     

免疫活性细胞表达杀伤细胞抑制性受体与造血干细胞移植后移植物抗宿主病的关系

本研究的目的是探讨造血干细胞移植后杀伤细胞抑制性受体(KIR)CD158和CD94与GVHD发生的关系。用流式细胞术检测分析T淋巴细胞和NK细胞表达CD158a，CD158b和CD94表达状况，同时用PCR方法分析供受HLA—Cw配型，比较异基因外周血造血干细胞移植(allb-PBSCT)和脐血移植(UCBT)后该KIR分子变化和HLA—Cw相合或错配与GVHD发生的关系。结果表明：无论是PBSCT或是UCBT，移植后CD3^ CD158a^ 和CD3^ CD158b^ T细胞均增高并以CD8^ CD158b^ 细胞升高为主。发生急性与慢性GVHD组CD3^ CD158b^ 细胞均呈不同程度升高，在急性GVHD病例中升高最明显。急性GVHD期(即移植早期)KIR表达增高，慢性GVHD阶段(即移植后期)KIR呈减低趋势。CD94主要表达于CD3^ CD8^ T细胞，在UCBT或PBSCT后CD94在CD4^ T细胞和CD8^ T细胞均明显增高。5例HLA—Cw相合无1例发生重症GVHD；2例HLA—Cw不相合病例，有1例发生急重症GVHD，并死于间质性肺炎(IP)，另1例为AML(M5)，完全植入，无重度GVHD发生，但于53天后复发。4例相关相合移植中2例无急性GVHD，而无关相合4例均发生不同程度GVHD。结论：GVHD的发生与KIR表达有一定关系。CD158b分子可能是移植后早期T淋巴细胞活化的负调控分子。从T细胞表达KIR来理解GVHD发生机理，特别是与HLA-Cw特异性识别结合将有利于寻找一条新的特异性建立免疫耐受和减低GVHD的新策略。     

混合脐血移植中HLA差异对植入的影响

目的 探讨HLA不同差异混合脐血移植的可行性和植入特性。方法 分别将2份人HLA半相合或不相合混合脐血输入经亚致死量照射后的严重联合免疫缺陷（SCID）小鼠，观察两组脐血在SCID小鼠体内的植入状况及造血和免疫功能重建特性。结果 两组混合脐血均可在受鼠体内植入，形成供－受混合嵌合体，并能重建多系造血，植入率差异无显著性（P＞0．05），且未见明显不良反应。用多聚酶链反应－序列特异性寡核苷酸（PCR－SSO）探针检测人HLA－DQB1基因发现，在多数受鼠体内只用多聚酶链反应－序列特异性寡核苷酸（PCR－SSO）探针检测人HLA－DQB1基因发现，在多数受鼠体内只有1份脐血植入，该份脐血为体外集落形成能力较高者，HLA半组合组有3例出现来自供－受三者的多嵌合体。结论 HLA相合及不合混合脐血移植可重建SCID小鼠造血和免疫功能，HLA相合程度不同的混合脐血移植与植入率无明显相关性，但相合程序较高者，似易于2份脐血同时植入。     

Acute graft versus host disease due to T lymphocytes recognizing a single HLA-DPB1*0501 mismatch.

Analysis of a large number of unrelated bone marrow transplantations (BMT) has shown that HLA-DP incompatibility did not detectably influence the risk for acute graft-versus-host disease (aGVHD). Accordingly, it was proposed that HLA-DP determinants did not function as transplantation antigens in the same way as HLA-A, -B, or -DR. We have previously shown that HLA-DP (as well as HLA-A, -B, -DQ, or -DR)-specific T cells could be isolated from skin biopsies of patients who developed an aGVHD after semiallogeneic BMT. Nevertheless, whether a single HLA-DP mismatched allele could induce a detectable allo-specific reaction in vivo after BMT remained to be established. To directly address this issue we studied one patient who presented aGVHD after receiving purified CD34+ bone marrow (BM) cells from an unrelated donor with a single HLA-DP mismatch in the GVHD direction. To characterize the immunological events associated with GVHD, we analyzed the peripheral T cell repertoire, the T cell receptor Vbeta diversity, and the specificity of T cells invading a skin biopsy at the onset of GVHD. Our results demonstrated that a large fraction of skin-infiltrating lymphocytes, which expressed diverse T cell receptors, were reactive against this single HLA-DPB1 *0501 mismatch and consequently that a single HLA-DP mismatch between BM donor and recipient can activate a strong T cell response in vivo.     

异基因骨髓移植小鼠供体骨髓细胞在受体内的分布及淋巴细胞供受体来源比例分析

目的观察小鼠异基因骨髓移植(alloBMT)后供体骨髓细胞在受体内的分布以及嵌合体内T、B淋巴细胞增殖分化规律。方法以绿色荧光蛋白(GFP)转基因C57BL/6J(H2b)小鼠为供体,以137Cs5.5Gy×2照射的BALB/c(H2d)小鼠为受体,分别于移植后3天(+3天)、+7天、+21天、+35天和+70天取外周血、骨髓、胸腺、脾脏、肠集合淋巴结及肝脏,用流式细胞仪和荧光显微镜观察供体来源的骨髓细胞分布。用藻红蛋白标记的抗CD4、抗CD8、抗B220抗体检测+21天小鼠体内骨髓、外周血及其他免疫器官中T、B淋巴细胞的构成比例。结果①+3天、+7天供体骨髓细胞在受体脾脏中分布最多,分别为(1.06±0.02)%、(76.60±1.80)%;骨髓中分别为(0.37±0.06)%、(39.70±5.38)%。②+21天受体骨髓、脾脏、淋巴结、胸腺及外周血中供体细胞达60%～90%。③+3天受体肠集合淋巴结中供体骨髓细胞达(0.36±0.04)%,与受体骨髓中供体骨髓细胞的比例相当。结论①alloBMT后早期供体骨髓细胞可分布到受体的骨髓、脾脏、肠集合淋巴结中,脾脏中最多。②+21天受体内骨髓及其他免疫器官以供体细胞为主。③受体肠道集合淋巴结也是移植后供体骨髓细胞的早期寄居点和B淋巴细胞增殖后的迁移部位。     

Long-term human hematopoiesis in the SCID-hu mouse

Coimplantation of small fragments of human fetal thymus and fetal liver into immunodeficient SCID mice resulted in the formation of a unique structure (Thy/Liv). Thereafter, the SCID-hu mice showed reproducible and long-term reconstitution of human hematopoietic activity. For periods lasting 5-11 mo after transplantation, active T lymphopoiesis was observed inside the grafts and cells that were negative for T cell markers were found to have colony-forming units for granulocyte/macrophage (CFU-GM) and erythroid burst-forming unit (BFU-E) activity in the methylcellulose colony assay. In addition, structures similar to normal human bone marrow were observed inside the Thy/Liv grafts, consisting of blast cells, mature and immature forms of myelomonocytic cells, and megakaryocytes. These data indicate long-term maintenance, in vivo, of human progenitor cells for the T lymphoid, myelomonocytic, erythroid, and megakaryocytic lineages. The role of the implanted fetal liver fragments was analyzed using HLA-mismatched Thy/Liv implants. The HLA type of the liver donor was found on T cells and macrophages in the graft. In addition, cells grown in the methylcellulose colony assay and cells in a bone marrow-like structure, the "thymic isle," expressed the HLA type of the liver donor. Thus, the Thy/Liv implants provided a microenvironment in which to follow human hematopoietic progenitor cells for multiple lineages. The formation of the Thy/Liv structures also results in a continuous source of human T cells in the peripheral circulation of the SCID-hu mouse. Though present for 5-11 mo, these cells did not engage in a xenograft (graft-versus-host) reaction. This animal model, the first in which multilineage human hematopoietic activity is maintained for long periods of time, should be useful for the analysis of human hematopoiesis in vivo.     

Recent advances in the treatment of graft-versus-host disease

Graft-versus-host disease (GVHD) is a lethal complication of allogeneic hematopoietic stem cell transplantation (HSCT) where immunocompetent donor T cells attack the genetically disparate host cells. The predominant symptoms of acute GVHD occur in the skin, liver, and intestine. Induction of acute GVHD can be divided into three phases: recipient conditioning, donor T cell activation, and effector cell-mediated GVHD. Chronic GVHD usually appears up to 100 days after HSCT and is characterized by symptoms similar to those observed for autoimmune disease. It is possible that chronic GVHD is the result of autoreactive T cells that escaped negative selection due to damage to the thymus from conditioning regimens, acute GVHD, and/or age related atrophy. Recent advances in the understanding of the basic mechanisms involved in GVHD pathophysiology have led to new strategies designed to block GVHD. This review focuses on recent developments in the treatment of GVHD, including insights gained from our own experimental studies.     

Designed transfer of specific immune responses with bone marrow transplantation.

Bone marrow transplant donors were immunized with tetanus/diphtheria toxoids 6-7 d before bone marrow donation to investigate the role of B cell subpopulations in reconstitution of humoral immunity. Lymphoblastoid B cells spontaneously producing IgG antitetanus and/or antidiphtheria toxoid were detected in the donor marrows at the time of transplantation. Recipients rapidly demonstrated 3-90-fold increases in serum IgG antitetanus and antidiphtheria toxoid levels. Antidiphtheria fragment A antibody in three donor/recipient pairs demonstrated spectrotypic identity indicating transfer of the donors' response. Reimmunization of three recipients 64-154 d after transplant revealed an IgG antibody response associated with reappearance of spontaneous antibody-producing B cells and an antidiphtheria fragment A response characteristics of the donor's immune response. These observations extend the understanding of the role of B cell subpopulations and provide a basis for specific modulation of immunity in the setting of bone marrow transplantation.     

Treatment of aplastic anemia by marrow transplantation from HLA identical siblings. Prognostic factors associated with graft versus host disease and survival.

73 consecutive patients with severe aplastic anemia were treated by marrow transplantation from hematologically normal HLA identical siblings. 68 patients lived long enough to document marrow engraftment. 21 rejected the graft and 19 of these died. 47 sustained engraftment and 18 of these died. In 16 patients, death was associated with graft versus host disease. 29 patients with sustained engraftment are alive with complete hematologic restoration between 8 mo and 5 yr. This analysis, by using a proportional hazards regression model, was directed at identifying factors that predicted survival (and absence of graft versus host disease). Of the 24 factors entered into the analysis only two strongly correlated with survival: (a) sex match of donor and recipient (P less than 0.01), and (b) absence of refractoriness to random donor platelets at the time of transplantation (P less than 0.05). Refractoriness adversely influenced the survival of the sex mismatched patients, These data suggest that X and Y-associated transplantation antigen systems are important determinants of the outcome of marrow grafts between HLA identical siblings for the treatment of aplastic anemia. The machanism by which refractoriness to random donor platelets influences survival is currently unclear.     

HLA gene amplification and hybridization analysis of polymorphism. HLA matching for bone marrow transplantation of a patient with HLA-deficient severe combined immunodeficiency syndrome

The treatment of choice for certain immunodeficiency syndromes and hematological disorders is bone marrow transplantation (BMT). The success of BMT is influenced by the degree of HLA compatibility between recipient and donor. However, aberrant expression of HLA sometimes makes it difficult, if not impossible, to determine the patient's HLA type by standard serological and cellular techniques. We describe here the application of new molecular biological techniques to perform high resolution HLA typing independent of HLA expression. A patient with HLA-deficient severe combined deficiency was HLA typed using in vitro amplification of the HLA genes and sequence-specific oligonucleotide probe hybridization (SSOPH). Two major advances provided by this technology are:detection of HLA polymorphism at the level of single amino acid differences; and elimination of a requirement for HLA expression. Although the patient's lymphocytes lacked class II HLA proteins, polymorphism associated with DR7,w53;DQw2;DRw11a (a split of DR5), w52b (a split of DRw52);DQw7 were identified. The patient's class I expression was partially defective, and typing was accomplished by a combination of serological (HLA-A and -C) and SSOPH analysis (HLA-B). Complete patient haplotypes were predicted after typing of family members [A2;B35(w6); Cw4; DRw11a(w52b);DQw7 and A2;B13(w4); Cw6;DR7(w53); DQw2]. Potential unrelated donors were typed and a donor was selected for BMT.