Evidence that anti-asialo GM1 in vivo improves engraftment of T cell-depleted bone marrow in hybrid recipients

Engraftment of T cell-depleted bone marrow was studied in a P----F1 murine bone marrow transplant model which features long-term stability of mixed chimerism of donor (B6) and host (B6AF1) cells after BMT. We report that a polyclonal antibody to asialo GM1 (anti-ASGM1) given in vivo after transplant was able to increase long-term donor bone marrow engraftment. In vivo anti-ASGM1 eliminated NK activity but did not affect the generation of cytotoxic T cells nor did it stimulate hematopoiesis in vitro. Anti-Thy 1.2, a pan-T cell monoclonal antibody, had no effect on donor engraftment. We conclude that ASGM1+ cells with NK activity inhibit the long-term engraftment of bone marrow stem cells in this model and that antibodies to NK cells can be used in vivo as an effective component of the transplant conditioning regimen.     

Long-term donor-specific tolerance in rat cardiac allografts by intrabone marrow injection of donor bone marrow cells

BACKGROUND: Donor-specific central tolerance in cardiac allograft can be induced by hematopoietic chimerism via conventional intravenous bone marrow transplantation (IV-BMT). However, there are problems with IV-BMT, such as the risk of graft failure and of the toxicity from conditioning regimens. METHODS: A new method for heart transplantation is presented. This method consists of administration of fludarabine phosphate (50 mg/kg) and fractionated low-dose irradiation (3.5 Gyx2 or 4.0 Gyx2), followed by intrabone marrow injection of whole bone marrow cells (IBM-BMT) plus heterotopic heart transplantation. RESULTS: Cardiac allografts with IBM-BMT were accepted and survived long-term (>10 months) showing neither acute rejection nor chronic rejection including cardiac allograft vasculopathy by such conditioning regimens. In contrast, cardiac allografts with conventional IV-BMT were rejected within 1 month after the treatment with irradiation of 3.5 Gyx2 or within 3 months after the treatment with irradiation of 4.0 Gyx2. Macrochimerism (>70%) was favorably established and stably maintained by IBM-BMT but not IV-BMT. Low levels of transient mixed chimerism (<7%) were induced by IV-BMT with fludarabine plus 4.0 Gyx2, but the chimerism was lost within 1 month after the treatment. CONCLUSIONS: These findings indicate that IBM-BMT is a feasible strategy for the induction of persistent donor-specific tolerance, enables the use of reduced radiation doses as conditioning regimens, and obviates the need for immunosuppressants.     

骨髓输注和抗淋巴细胞球蛋白诱导对同种异体移植肾特异性免疫无反应性的临床研究

国外使用抗淋巴细胞球蛋白（ＡＬＧ）和输入供体特有骨髓（ＢＭ）诱导同种移植物的耐受力，并建立了动物模型。我们将这研究用于诱导特异性免疫无反应性病例２０例（骨髓组），并随机抽样２２例尸体肾移植患者作为对照组。骨髓组接受常规三联免疫抑制治疗，在１４天从ＡＬＧ诱导期后，停用１周，于第７天输入冷藏骨髓，随访５～４１个月。结果：１～６个月发生排斥反应２例（２／２０），均用大剂量激素逆转；另１例合并败血性肺炎死亡。对照组采用三联常规免疫抑制治疗剂量较骨髓组大，但不输入骨髓，结果同期发生排斥反应６例（６／２２），后期排斥反应２例，其中４例因排斥反应未能逆转而移植肾失功，有１例因合并心衰死亡。从两组结果分析，人的一年成活率两组无明显差别。移植肾一年成活率则骨髓组高于对照组（Ｐ＜０．０１），排斥反应发生率骨髓组明显低于对照组（Ｐ＜０．０５），且骨髓组常规免疫抑制剂用量较对照组小。     

Engraftment following T cell-depleted bone marrow transplantation. III. Differential effects of increased total-body irradiation on semiallogeneic and allogeneic recipients

Three different doses of total-body irradiation (TBI) (1100, 1300, 1500 cGy) have been analyzed as conditioning regimens for semiallogeneic B6AF1 (H-2b X H-2a) and allogeneic A/J (H-2a) recipients of T cell-depleted C57BL6 (H-2b) bone marrow transplants. Recipient survival and engraftment of both donor erythrocytes and lymphocytes were examined in each group. The large majority of allogeneic mice prepared with 1100 cGy rejected their grafts, which resulted in poor survival (less than 30%); improved survival (up to 80%) and complete donor engraftment were noted as the TBI dose was increased. By contrast, survival in semiallogeneic B6AF1 recipients was independent of TBI dose and was greater than 80% in all groups. Outright failure of marrow grafts (less than 10% donor hematopoiesis) did not occur in these recipients, but mixed chimerism (simultaneous occurrence of both donor and host cells) was frequently observed at lower TBI doses. Complete (greater than 90%) donor engraftment was noted for erythrocytes but not for lymphocytes. Possible mechanisms accounting for these differences between semiallogeneic and allogeneic recipients of marrow transplants are discussed.     

钻孔复合浓缩骨髓对兔结构性骨移植愈合的影响

目的 观察异体骨钻孔并复合自体浓缩骨髓移植对兔骨缺损的修复和重建能力,为临床结构性骨移植提供新方法.方法 将36只日本大耳白兔随机分成三组,每组12只,制成左侧股骨中段3 cm缺损,分别采用不同方式处理过的同种异体骨进行移植,A组:植入未打孔异体骨+骨泥;B组:植入打孔异体骨+骨泥;C组:植入打孔异体骨+骨泥浓缩骨髓复合物.术后3个月分别行x线片、组织学观察、骨密度测定和生物力学测试. 结果 ①放射学检查:A组仅在供.宿结合部可见有骨痂包绕,异体骨周围无明显骨痂;B组及C组整个移植骨段周围均可见明显骨痂,C组骨痂延伸范围更广.②组织学观察:三组异体骨供.宿结合部均完全吸收,被大量骨痂取代且塑形良好;中间横断面A组异体骨周围可见少量内骨痂,B组可见少量内骨痂及外骨痂且被大量纤维结缔组织所分隔;C组异体骨可见大量内外骨痂生长,髓腔内充满骨母细胞,外骨痂骨小梁已相互融合形成编织网状骨,但与异体骨间仍被纤维结缔组织间断分隔.③平均骨密度值测定:C组高于A、B组及对侧正常股骨,差异有统计学意义(P＜0.05).④力学性能检测:A、B、C三组间最大载荷比较,差异无统计学意义(P＞0.05),A、B、C组均低于对侧正常股骨(P＜0.05). 结论 钻孔并复合浓缩骨髓能促进大段移植骨的早期活化,提高新骨产量.     

Antibody-induced rejection of pig proislet xenografts in CD4+ T cell-depleted diabetic mice

Reversal of diabetes in mice was achieved following in vivo depletion of host CD4+ T cells and transplantation of xenogeneic fetal pig proislets (pancreatic islet precursors). These procedures resulted in xenograft tolerance since established pig proislet xenografts were not rejected by antipig antibodies produced in the host, and rejection was not induced following the administration of donor major histocompatibility complex--specific pig lymphocytes. Proislet xenografts were rejected following the administration of donor MHC-specific hyper-immune antipig PBL serum raised in normal mice. Although established proislet xenografts in anti-CD4-treated mice are sensitive to antibody-mediated destruction, such hosts are unable to produce an antibody response that leads to graft rejection. The study indicates that the mechanism of preventing xenograft rejection by anti-CD4 treatment in vivo involves not only initial CD4+ T cell depletion but also quantitative and/or qualitative modulation of a CD4+ T cell-dependent antibody response. As a consequence, an apparent state of xenograft tolerance is produced.     

Impact of donor bone marrow on survival of composite tissue allografts

Composite tissue allotransplants (CTA) involves transplantation of various tissues including vessels, nerves, skin, and immune cells and bears significant antigenic load. Different immunosuppressive protocols are used for experimental and clinical CTA. Immunosuppressive agents maintain survival of the different components of composite tissue allografts. However, the potential side effects of chronic immunosuppression currently limit the widespread application of CTA transplants. Bone marrow therapy in many tolerance induction protocols therefore provides a guide to reaching the target of permanent immunotolerance. Multiple studies suggest that bone marrow is immunomodulatory and may facilitate allograft acceptance.In this review, bone marrow-based therapy protocols of experimental and clinical models are presented in composite tissue transplantation.     

淋巴因子激活的骨髓细胞对小鼠同种异体骨髓移植的影响

以致死性剂量照射的成年小鼠为受鼠进行同种异体骨髓移植（Ａｌｌｏ－ＢＭＴ），实验组小鼠观察期间平均存活天数较对照组明显延长（Ｐ＜０．００１），应用肿瘤移植及单向混合淋巴细胞培养（ＭＬＣ）二项指标证实Ａｌｌｏ－ＢＭＴ的部分植活。实验结果表明，淋巴因子激活的骨髓细胞（ＬＡＢＭＣ）可有效预防致死性移植物抗宿主病（ＧＶＨＤ），且不影响供髓植活。     

Induction of donor-specific tolerance to rat cardiac allografts by intrathymic inoculation of bone marrow.

BACKGROUND. Induction of donor-specific tolerance to tissue or organ allografts can readily be achieved by administration of allogeneic bone marrow to neonatal rodents; however, in adult recipients induction of transplantation tolerance by this strategy generally requires intensive cytoablative conditioning. Described here is a novel method of promoting transplantation tolerance that involves inoculation of donor bone marrow into the thymus of transiently immunosuppressed adult recipients. METHODS. Prospective Wistar-Furth recipients were inoculated with allogeneic Lewis bone marrow cells (BMCs) either intrathymically or intravenously in conjunction with a single dose of antilymphocyte serum 2 to 3 weeks before receiving donor-strain cardiac allografts. Recipients were monitored for graft survival and examined for presence of hematopoietic chimerism. RESULTS. Intrathymic but not intravenous inoculation of donor BMCs led to permanent survival of donor-strain cardiac allografts, whereas third-party Dark agouti cardiac allografts were rejected promptly. Persistence of donor chimerism was demonstrated in the thymus of Wistar-Furth recipients of intrathymic Lewis BMCs for as long as 3 weeks after BMC inoculation. CONCLUSIONS. Intrathymic inoculation of BMCs concurrently with a single dose of antilymphocyte serum induces donor-specific unresponsiveness to rat cardiac allografts. The unresponsiveness may be the result of deletion or functional inactivation of alloreactive clones maturing in a thymus bearing donor alloantigen. Intrathymic inoculation of BMCs deserves further evaluation as a possible clinical strategy for the induction of transplantation tolerance.     

Development of stable mixed T cell chimerism and transplantation tolerance without immune modulation in recipients of vascularized bone marrow allografts

A consistent majority (62.5%) of immunologically unmodified rat recipients transplanted with vascularized hind-limb bone marrow allografts across a semiallogeneic transplant barrier developed tolerance with absence of graft-versus-host disease. A minority of recipients (37.5%) demonstrated lethal GVHD. Transplantation tolerance in the majority was associated with the induction of stable low-level mixed T cell chimerism, including donor CD5+, CD4+, and CD8+ lymphocytes. Chimeras were specifically immune nonresponsive to host alloantigenic determinants. These results emphasized a potentially important mechanism for low-level stable mixed lymphoid chimerism (SMLC) in tolerance induction, independent of immune suppressive effects due to irradiation or immunopharmacologic intervention. These vascularized bone marrow transplantation (VBMT) results may establish the experimental foundation for a novel approach to stem cell transfer and bone marrow transplantation.     

尼古丁对小鼠骨髓细胞微核影响的实验研究

目的:探讨香烟成分尼古丁对骨髓细胞微核产生的影响.方法:建立小鼠骨髓嗜多染红细胞微核实验模型,将小鼠随机分为尼古丁(低、中、高剂量)实验组、顺铂对照组、乙醇对照组、生理盐水对照组,每组10只,采用腹腔注射给药,油镜下检测骨髓中嗜多红染细胞(PCE)微核情况及绘制量效关系曲线.结果:尼古丁低剂量组和乙醇对照组的微核发生率分别为(1.90±0.88)％和(0.70±0.22)％,两组间有显著差异(P＜0.05).采用不同剂量尼古丁染毒小鼠,其骨髓细胞微核率呈递增趋势.结论:尼古丁在嗜多染红细胞微核实验中具有明显的致突作用.