Allergy development after bone marrow transplantation from a non-atopic donor

BACKGROUND: Transfer of allergy from atopic bone marrow donors to recipients is known to occur. Development of allergy in a non-atopic patient transplanted from a non-atopic donor is an unfamiliar phenomenon in clinical practice. OBJECTIVES: To clarify the course of events causing a bone marrow recipient to acquire an allergic disease in such non-conducive circumstances. METHODS: Full medical history, prick and intradermal skin tests, and serum IgE levels were obtained from both donor and recipient patients. DNA and red blood cell phenotype analyses were used to detect the degree of chimerism. RESULTS: Only the recipient patient showed positive specific IgE antibodies and skin tests to house dust mite. The recipient patient displayed 100% donor chimera, based on all engraftment markers sought. CONCLUSION: Full engraftment after allogeneic bone marrow transplantation may be associated with modulation of T and B cell function, which in turn could cause the onset of allergic disease after bone marrow transplantation.     

骨髓干细胞移植后mdx鼠腓肠肌病理变化

目的 研究骨髓干细胞移植后mdx鼠腓肠肌组织病理变化. 方法 7～9周龄mdx鼠20只平均分为4组,放射处理后移植1.2×107细胞/只同种异基因全骨髓干细胞,于移植后4、8、12及16周用HE染色观察腓肠肌组织细胞形态及核中心移位纤维(CNF).C57鼠和未治疗mdx鼠各5只作阳性和阴性对照. 结果 CS7鼠腓肠肌横切面可见肌细胞大小形态基本一致,无核中心移位现象.各细胞移植治疗组和阴性对照组mdx鼠均有大量的炎细胞浸润,核中心移位明显.未治疗mdx鼠CNF最高,约达70%;移植后4、12和16周,CNF分别为55%、50%和44%. 结论 骨髓干细胞移植后mdx鼠腓肠肌CNF随移植时间延长逐渐减少,提示骨髓干细胞移植后长久持续参与受损骨骼肌的修复与再生.     

T-cell large granular lymphocyte leukemia of donor origin after allogeneic bone marrow transplantation

A 39-year-old man with chronic myeloid leukemia in accelerated phase underwent allogeneic bone marrow transplantation (BMT). At 6 months after BMT, lymphocytosis (WBC count, 23,100/microL [23.1 x 10(9)/L]; 80% (0.80) large granular lymphocytes [LGLs]) occurred. The LGLs were CD3+CD4-CD8+, with clonally rearranged T-cell receptor gamma gene, and of donor origin, as shown by analysis of polymorphic microsatellite markers. Epstein-Barr virus was not present. The diagnosis, therefore, was consistent with T-cell large granular lymphocytic (T-LGL) leukemia. Corticosteroids controlled the LGL count, but progressive pancytopenia led to death 4 months later. Retrospective analysis showed that the T-LGL leukemia apparently had arisen as early as 3 months after BMT. The distinguishing features of this case included donor origin, neoplastic nature, and the aggressive fatal outcome.     

Separating antiviral and GVHD activities of donor T cells prior to bone marrow transplantation

Approaches to speed immune reconstitution following bone marrow transplantation (BMT) or peripheral-blood hematopoietic stem-cell transplantation (HSCT) could markedly reduce morbidity and mortality, particularly following partially major histocompatibility complex (MHC)-matched related donor (PMRD) transplants. However, it is critical to simultaneously eliminate the subpopulation of donor T cells that are alloreactive with the recipient and may produce graft-vs-host disease (GVHD). In this article, we discuss a number of promising cellular engineering approaches that could be applied to this problem, including the use of veto cells, regulatory T-cell subsets, and psoralen-treated donor lymphocytes. Emphasis is placed on whether these approaches can simultaneously transfer broad-spectrum immunity to the recipient without producing GVHD.     

B7.2-/- mature dendritic cells generate T-helper 2 and regulatory T donor cells in fetal mice after in utero allogeneic bone marrow transplantation.

In utero hematopoietic stem cell transplantation (IUT) results in limited chimerism and tolerance to alloantigens. We studied the relative role of B7.1 and B7.2 expression by dendritic cells (DCs) in engraftment and in generating donor-specific tolerance in fetal mice. Mature dendritic cells (mDCs) from B7.1(-/-) or B7.2(-/-) donors and wild-type (WT) lineage-depleted (lin(-)) C57BL/6 (B6) bone marrow (BM) were injected into BALB/c fetuses. Six weeks after IUT, B7.1(-/-) recipients had multilineage engraftment (4.7% +/- 0.8% T cells and 5.7% +/- 1.1% granulocytes) associated with graft-versus-host disease (GVHD) and decreased survival, but by 12 weeks only donor CD3(+) cells (2.1% +/- 1.3%) were present. Recipients of B7.2(-/-) mDCs and lin(-) WT B6 BM had exclusively CD3(+)CD4(+) T cells (11.8% +/- 8.5% at 6 weeks and 6.5% +/- 2.5% at 12 weeks). Most of the cells were T-helper 2, although 10.4% +/- 1.4% were of the T-regulatory (T(reg)) phenotype, ie, CD4(+)CD25(+). Donor T(reg) cells were detected both in the thymus and spleen, thus suggesting an effect on both central and peripheral immunity. The animals with T(reg) cells had better survival (82.3% versus 47.4%; P < .01) and no GVHD (0% versus 65%; P < .001). This group alone demonstrated multilineage engraftment of donor hematopoietic cells after postnatal transplantation with megadoses of donor lin(-) BM. Both the engrafted donor CD4(+)CD25(-) and CD4(+)CD25(+) cells induced comparable in vitro suppression of T-cell proliferation, thus suggesting their role in the persistence of the donor T cells in vivo. The CD4(+)CD25(-) cells produced interleukin 10 or interleukin 4 and were inhibited by anti-T-helper 2 cytokine-neutralizing antibodies, whereas the CD4(+)CD25(+) cells showed no evidence of any involvement of a cytokine-like soluble mediator and expressed cytotoxic T-lymphocyte antigen 4 (CTLA-4) and foxp3 constitutively. Donor mDCs and donor CD4 T cells were detected among the thymocytes of the recipients of B7.2(-/-) mDCs and lin(-) WT B6 BM. Thus, it seems that costimulatory molecule expression of donor DCs can play a significant immunomodulatory role in survival, GVHD, engraftment, and homing of allogeneic BM cells after IUT through the generation of T(reg) cells.     

Persistence of recipient lymphocytes in NOD mice after irradiation and bone marrow transplantation

The non-obese diabetic (NOD) mouse is a unique and invaluable model of autoimmune disease, in particular type 1 diabetes. Bone marrow transplantation as a therapy for type 1 diabetes has been explored in NOD mice. NOD mice require higher doses of conditioning irradiation for successful allogeneic bone marrow transplantation, suggesting that NOD hematopoietic cells are radioresistant compared to those of other mouse strains. However, studies of hematopoietic reconstitution in NOD mice are hampered by the lack of mice bearing a suitable cell-surface marker that would allow transferred cells or their progeny to be distinguished. In order to monitor hematopoietic reconstitution in NOD mice we generated congenic NOD mice that carry the alternative allelic form of the pan-leukocyte alloantigen CD45. Following irradiation and congenic bone marrow transplantation, we found that the myeloid lineage was rapidly reconstituted by cells of donor origin but substantial numbers of recipient T lymphocytes persisted even after supra-lethal irradiation. This indicates that radiation resistance in the NOD hematopoietic compartment is a property primarily of mature T lymphocytes.     

Cytokines transduced bone marrow stromal cell lines promote immunohematopoietic reconstitution in mice after allogeneic bone marrow transplantation

Impaired immune reconstitution following allogeneic T-cell depleted bone marrow transplantation (allo-TCD-BMT) is a major obstacle to its clinical application. Stromal cell line QXMSC1, established from bone marrow cells of BALB/c(H-2d), was transfected with murine IL-3 and/ or IL-2 gene, and injected into lethally irradiated C57BL/6(H2b) mice. We evaluated its effects on immunologic and hematopoietic reconstitution after allo-TCD-BMT. The results showed that QXMSC1-IL-3 + IL-2 could significantly increase the numbers of hematopoietic primitive progenitors (CFU-S), committed progenitors (CFU-GM, and BFU-E), and lymphocytes (CD8+ cells, CD4+ cells, and B cells). Similarly, immune functions of recipient mice were significantly enhanced in the QXMSC1-IL-3 + IL-2 group. In addition, QXMSC1-IL-3 or QXMSC1-IL-2 also exerted apparent effects on accelerating immune reconstitution, but these effects were far less than that of QXMSC1-IL-3 + IL-2. Our results demonstrated that stromal cell-mediated IL-3 and IL-2 gene therapy may be a potent approach in promoting immunologic and hematopoietic reconstitution after allo-TCD-BMT.     

Histopathology of bone marrow reconstitution after allogeneic bone marrow transplantation

In order to study haematopoietic reconstitution in allogeneic bone marrow transplantation we investigated bone marrow histology in 61 biopsies of 37 patients, treated with HLA-compatible bone marrow grafts for leukaemia or severe aplastic anaemia. The biopsies were taken from the day of transplantation until 100 d after transplantation. Stromal changes, in particular oedema, fibrosis and granulomas, were found during the whole period of observation. These changes were more prominent in biopsies from leukaemia patients than from patients with aplastic anaemia. The cellularity in the biopsies increased until 28 d after bone marrow transplantation and was stable thereafter. Initially, only clusters of cells belonging to a single cell lineage were seen, suggesting that the first outgrowth of haematopoietic cells is by proliferation of committed precursor cells. Long-lasting abnormalities in localization of haematopoietic cells in the bone marrow space and of the myeloid: erythroid ratio were seen; dyserythropoiesis was common.     

Intra-bone marrow injection of donor bone marrow cells suspended in collagen gel retains injected cells in bone marrow, resulting in rapid hemopoietic recovery in mice

We have recently developed an innovative bone marrow transplantation (BMT) method, intra-bone marrow (IBM)-BMT, in which donor bone marrow cells (BMCs) are injected directly into the recipient bone marrow (BM), resulting in the rapid recovery of donor hemopoiesis and permitting a reduction in radiation doses as a pretreatment for BMT. However, even with this IBM injection, some of the injected BMCs were found to enter into circulation. Therefore, we attempted to modify the method to allow the efficient retention of injected BMCs in the donor BM. The BMCs of enhanced green fluorescent protein transgenic mice (C57BL/6 background) were suspended in collagen gel (CG) or phosphate-buffered saline (PBS), and these cells were then injected into the BM of irradiated C57BL/6 mice. The numbers of retained donor cells in the injected BM, the day 12 colony-forming units of spleen (CFU-S) counts, and the reconstitution of donor cells after IBM-BMT were compared between the CG and PBS groups. The number of transplanted cells detected in the injected BM in the CG group was significantly higher than that in the PBS group. We next carried out CFU-S assays. The spleens of mice in the CG group showed heavier spleen weight and considerably higher CFU-S counts than in the PBS group. Excellent reconstitution of donor hemopoietic cells in the CG group was observed in the long term (>100 days). These results suggest that the IBM injection of BMCs suspended in CG is superior to the injection of BMCs suspended in PBS.     

Optimal timing to repopulation of resident alveolar macrophages with donor cells following total body irradiation and bone marrow transplantation in mice

To determine the time required to repopulate mouse lungs with donor alveolar macrophages following total body irradiation (TBI) and bone marrow transplantation (BMT), C57Bl/6 mice were subjected to TBI with 900 cGy, followed by transplantation of bone marrow cells from mice expressing green fluorescent protein (GFP) in their somatic cells. The mice were euthanized at either 30 (n=5), 60 (n=5) or 90 (n=5) days following BMT. Thirty days following transplantation, 87.8 +/- 3.9% (mean +/- S.E.M.) circulating leukocytes in recipient mice were derived from the donor, as determined by fluorescence activated cell sorting (FACS) analysis for GFP. However, only 46.9 +/- 7.4% of the resident alveolar cells expressed GFP, indicating incomplete repopulation. By day 60 post-transplantation, the percentage of bronchoalveolar lavage fluid (BALF) cells expressing GFP reached 74.5 +/- 2.4%, remaining stable 90 days after transplantation (80.4 +/- 1.9%). We conclude that 60 days after TBI with 900 cGy and bone marrow transplantation, the majority of the lung resident alveolar macrophages is of donor origin. This study provides useful information regarding the time of reconstitution with donor alveolar macrophages in the pulmonary airspaces of recipient mice following marrow transplantation.     

Donor-derived human bone marrow cells contribute to solid organ cancers developing after bone marrow transplantation

Bone marrow-derived stem cells have been shown to participate in solid organ repair after tissue injury. Animal models suggest that epithelial malignancies may arise as aberrant stem cell differentiation during tissue repair. We hypothesized that if bone marrow stem cells participate in human neoplasia, then solid organ cancers developing after allogeneic bone marrow transplantation (ABMT) might include malignant cells of donor origin. We identified four male patients who developed solid organ cancers (lung adenocarcinoma, laryngeal squamous cell carcinoma, glioblastoma, and Kaposi sarcoma) after myeloablation, total body irradiation, and ABMT from female donors. Donor-derived malignant cells comprised 2.5%-6% of the tumor cellularity The presence of donor-derived malignant cells in solid organ cancers suggests that human bone marrow-derived stem cells have a role in solid organ cancer's carcinogenesis. However, the nature of this role is yet to be defined.     

Distribution of colony-forming cells in the bone marrow and spleen of irradiated mice

Two hours after transplantation of 2×10⁵ bone marrow cells, 1.7–2.8% and 14–16% of colonyforming units (CFU) were retained respectively in the femoral bone marrow and spleen of the recipients. After 24 h, 1–2% and 21–26% of CFU respectively remained in these organs. If 2.5×10⁶–1.7×10⁷ bone marrow cells were transplanted, the fractionf in the bone marrow and spleen of the recipients (under saturation conditions) was unchanged during the 24-h period. The value for CFU settling in the spleen was 20–22%, whereas for CFU settling in the femoral marrow the value off was 2.4–2.7%. It is postulated that the same number of CFU settles in the whole volume of the bone marrow as in the whole spleen. The total number of CFU settling in the spleen and in the whole volume of the bone marrow averages 40–50%. The fate of the remaining CFU is unknown.Presented by Academician of the Academy of Medical Sciences of the USSR P. D. Gorizontov.Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 7, pp. 827–829, July, 1976.     

Donor bone marrow infusion in deceased and living donor renal transplantation

The infusion and persistence in a transplant recipient of donor-derived bone marrow cells (DBMC) of multi-lineage can lead to a state of permanent chimerism. In solid vascular organ transplantation, the donor bone marrow lineage cells can even be derived from the transplant organ, and these cells can be detected in very small numbers in the recipient. This has been called microchimerism. Much controversy has developed with respect to the function of chimeric cells in organ transplantation. One idea is that the occurrence of these donor cells found in microchimerism in the recipient are coincidental and have no long-term beneficial effect on engraftment. A second and opposing view, is that these donor cells have immunoregulatory function that affect both the acute and chronic phases of the recipient anti-donor responses. It follows that detecting quantitative changes in chimerism might serve as an indication of the donor-specific alloimmune or regulatory response that could occur in concert with or independent of other adaptive immune responses. The latter, including autoimmune native disease, need to be controlled in the transplant organ. The safety and immune tolerance potential of DBMC infusion with deceased and living donor renal transplants was evaluated in a non-randomized trial at this center and compared with non-infused controls given identical immunosuppression. Overall DBMC infusions were well tolerated by the recipients. There were no complications from the infusion(s), no episodes of graft-vs-host disease (GVHD) and no increase infections or other complications. In the deceased DBMC-kidney trial, actuarial graft survival at 5 years was superior especially when graft survival was censored for recipient death. Acute rejections were significant reduced in patients given two DBMC infusions, and chronic rejection was dramatically reduced in all DBMC treated patients. The most interesting finding was that the degree of microchimerism slowly increased over the years the DBMC group that had exhibited no rejection episodes. In the DBMC-living related trial, the incidence of acute rejection did not differ between groups. However, DBMC chimerism in recipient iliac crest marrow had increased more rapidly than might be predicted from results previously seen in the cadaver group, despite four times fewer DBMC infused, with the generation of T- regulartory cells in-vitro assays.     

Self-tolerance to host and donor following HLA-mismatched bone marrow transplantation

The transplantation of T cell-depleted HLA-haploidentical bone marrow can correct the severe combined immunodeficiency disease (SCID) caused by the inherited absence of T lymphocytes. Despite a different environment, no severe graft-vs.-host reaction occurred and engrafted T lymphocytes became functional. We have studied tolerance of engrafted T lymphocytes to donor and host HLA antigens in four SCID patients who have been transplanted with bone marrow from one of their HLA-haploidentical parents. Graft-vs.-host reaction was prevented by T cell depletion of infused bone marrow using E rosetting and by in vivo administration of cyclosporine A. Subsequent to bone marrow transplantation (BMT), the engrafted T lymphocytes were shown to be unresponsive in vitro towards host cells collected prior to BMT. Generally, this tolerance could not be explained by a suppressive mechanism. Nevertheless, in one patient suppressive cells were found transiently. In contrast to the early appearance of a tolerance towards host, a reactivity of engrafted donor cells towards donor was always observed within the first 300 days post-grafting. This autoreactivity was mediated by T cells of donor origin and its targets were HLA class II molecules (at least HLA-DR and DQ). The progressive disappearance of this autoreactivity was correlated with the engraftment of Ia-positive cells (monocytes plus B lymphocytes) of donor origin and the achievement of complete immunological reconstitution. In the patient showing the strongest autoreactivity, a donor-specific T cell line has been grown which was shown to specifically inhibit the proliferative response of donor lymphocytes. Concomittantly, the immunological reconstitution remains poor in this patient. These data suggest that tolerance to HLA class II molecules is dependent on the presence of the relevant HLA class II molecule-expressing cells allowing the elimination or the suppression of T lymphocytes specifically directed at these molecules.     

Tolerance of engrafted donor T cells following bone marrow transplantation for severe combined immunodeficiency

Patients transplanted for the treatment of severe combined immunodeficiency (SCID) frequently develop a unique state of split lymphoid chimerism. Such patients have T cells of donor origin, and non-T cells which are predominantly or exclusively of host origin. We have studied the reactivity of engrafted donor T cells to host and/or donor antigens in 12 patients transplanted for SCID, focusing on the characteristics of the tolerance to host and/or donor MHC antigens observed in nine of these patients who were recipients of T-cell-depleted, haploidentical parental bone marrow. In both proliferative and cytolytic assays, engrafted, donor-derived T cells were shown to be selectively nonreactive to histoincompatible host cells. This tolerance could not be ascribed to cells with suppressive activity in the engrafted T-cell population. T cells from a subset of patients, however, exhibited proliferative but not cytolytic reactivity to donor peripheral blood mononuclear cells. The responding cells were shown to be donor-derived CD3+ cells and were predominantly reactive to B-cell fractions from the donor. Two patients who received transplants from each parent in sequence engrafted T cells from one parent and had non-T cells of host, paternal, and maternal origin. The engrafted T cells proliferated weakly to B cells from the other parent, but were tolerant in cytolytic assays. Donor anti-donor reactivity was seen only in haploidentical split chimeras who had not been treated with cytotoxic drugs prior to T-cell engraftment. This proliferative reactivity toward donor may be due to an absence of donor derived Ia+ antigen presenting cells resident in the thymus of SCID patients at the time when the T-cell repertoire is being shaped.     

Amotosalen-treated donor T cells have polyclonal antigen-specific long-term function without graft-versus-host disease after allogeneic bone marrow transplantation.

We have previously shown that amotosalen HCl (S-59 psoralen)-treated donor splenocytes, which have limited proliferative capacity in vitro, can protect major histocompatibility complex-mismatched bone marrow transplant (BMT) recipients from lethal murine cytomegalovirus infection without causing graft-versus-host disease. In this study, we further investigated the effects of amotosalen-treated donor T cells on immune reconstitution after allogeneic BMT. We were surprised to find that amotosalen-treated donor T cells persisted long-term in vivo, comprising 6% to 10% on average of the T-cell compartment of transplant recipients at 4 months after transplantation. Donor T cells derived from amotosalen-treated splenocytes were predominantly polyclonal CD44 hi/int CD8 + memory T cells and were functionally active, synthesizing interferon gamma in response to stimulation with murine cytomegalovirus antigen. Amotosalen-treated donor T cells, reisolated from BMT recipients' spleens >/=4 months after transplantation, proliferated in vitro, thus indicating repair of amotosalen-mediated DNA cross-links. Compared with infusion of untreated donor splenocytes, amotosalen-treated cells enhanced thymopoiesis by bone marrow-derived stem cells in BMT recipients. However, amotosalen treatment abrogated the thymopoietic activity of lymphoid progenitor cells among the donor splenocytes. Thus, infusion of amotosalen-treated donor T cells produced rapid immune reconstitution after major histocompatibility complex-mismatched BMT by transferring long-lived polyclonal memory T cells with antiviral activity and also by enhancing bone marrow-derived thymopoiesis. This is a novel approach to adoptive immunotherapy in allogeneic BMT.