含高剂量CD34~+细胞的非血缘脐血移植治疗AML/MDS后原发植入失败（英文）

目的分析含高剂量CD34+细胞的非血缘脐血移植治疗急性粒细胞白血病合并骨髓增生综合征(AML/MDS)后发生原发植入失败的原因。方法 1例4岁女孩在四川大学华西第二医院儿科血液肿瘤科被诊断为AML/MDS。患儿经诱导及巩固治疗获完全缓解后,行无血缘人类白细胞抗原(HLA)部分不相合脐血移植。观察患儿术后造血重建及移植相关并发症情况。结果患儿术后发生原发植入失败,再次进行血缘间的半相合造血干细胞移植,期间患多药耐药鲍曼不动杆菌败血症,于第2次移植后7 d死于呼吸衰竭。结论含高剂量CD34+细胞脐血造血干细胞并不能抵消HLA配型不合的缺陷。患儿原发植入失败可能与患儿移植前存在长期血小板输注无效及潜在免疫异常,尤其是移植前产生抗-HLA供者特异性抗体有关。     

双份脐血移植后受者骨髓间充质干细胞嵌合情况

目的 研究双份脐血移植(DCBT)受者骨髓间充质干细胞(MSC)的嵌合状态.方法 急性粒细胞白血病M2a型男性患者1例,接受改良白消安环磷酰胺方案+抗胸腺细胞球蛋白(ATG)预处理,输注5个抗原(5/6)相合和4个抗原(4/6)相合的非血源脐血各1份,移植后19 d粒系造血重建.移植后87 d,采用密度梯度离心法分离DCBT后受者及正常供者的骨髓单个核细胞,分别培养MSC,用流式细胞术检测细胞表面标志,诱导其向成脂肪细胞和成骨细胞分化,应用逆转录聚合酶链反应法检测MSC表面造血及免疫相关分子的表达,短串联重复序列聚合酶链反应检测受者MSC、外周血、骨髓中供者细胞嵌合率.结果 移植后受者MSC与正常供者MSC具有相似的细胞形态、免疫表型以及分化潜能,均能表达白细胞介素6、干细胞因子、白血病抑制因子和粒-巨噬细胞集落刺激因子等造血及免疫相关分子的mRNA.DCBT后受者骨髓优势脐血嵌合度达96.4％,外周血嵌合度达95.7％,MSC的优势脐血嵌合度为5.4％,MSC中受者本身部分占94.6％.结论 DCBT后,受者造血重建仅来自于其中1份脐血.移植后骨髓MSC大部分来源受者本身,部分嵌合的供者MSC来源于植入的单份脐血.     

由脐血单个核细胞和富集的CD34+细胞扩增的造血干/祖细胞的生理功能比较

目的 探讨由脐血单个核细胞(MNC)和富集的CD34+细胞起始扩增所得的造血干/祖细胞在体内植入及造血重建的能力.方法 从人脐血中分离出MNC和CD34+细胞,在体外扩增7 d.将非肥胖糖尿病型重症联合免疫缺陷型(NOD/SCID)小鼠分为四组,在接受亚致死剂量铯源照射后,进行细胞移植,实验A组接受由MNC培养得到的CD34+细胞和CD34-细胞;实验B组接受由富集的CD34+细胞培养得到的CD34+细胞和CD34-细胞;阳性对照组接受从脐血新鲜分离的CD34+细胞和CD34-细胞;阴性对照组不接受细胞移植,仅输注相同体积的IMDM培养基.6周后处死存活的小鼠,取其骨髓、脾脏和外周血细胞,分别进行细胞表型分析、集落和人特异性基因的检测.结果 经过体外扩增,以富集的CD34+细胞起始培养者的细胞总扩增倍数为39.8倍,远高于以MNC为起始细胞者的1.88倍.移植6周后,所有接受细胞移植的小鼠均存活,存活小鼠的骨髓和脾脏细胞中均能检测到人源细胞(CD45+细胞)及人源的各系血细胞,实验A组各类细胞的含量稍高于实验B组,且接受细胞移植小鼠的骨髓和脾脏细胞中可检测出人特异的Alu序列.结论 与从脐血中新鲜分离的细胞相比,扩增后的造血干/祖细胞的体内植入能力有所下降,以MNC起始扩增的造血干/祖细胞体内植入能力优于以富集的CD34+细胞起始扩增者,但二者体内造血重建能力的差异不显著.     

非清髓性脐血移植治疗成人重型再生障碍性贫血

目的　研究与追踪观察无关供者脐血造血干细胞移植 (Allo CBSCT)治疗成人重型再生障碍性贫血 (SAA)后 ,脐血造血细胞的植入情况及疗效。方法　单份脐血 (3例 )和双份脐血 (3例 )移植治疗成人重型再生障碍性贫血患者共 6例。输入的脐血单个核细胞 (MNC)数为 (1.6～ 10 .7)×10 7/kg(按患者体重计 )。预处理方案由低剂量的环磷酰胺 (CTX)和抗淋巴细胞球蛋白 (ALG)组成。CTX总用量为 6 0mg/kg ;ALG为 12 0mg/kg。移植后供者细胞植入状态的检测方法采用微卫星DNA指纹法或萤光定量PCR分析。结果　 6例中有 5例移植后有供者细胞植入证据 ,均为供、受者细胞混合嵌合状态。 2例接受双份脐血移植者均仅显示单份脐血植入。迄今追踪时间平均 2 1个月 (7～ 5 0个月 ) ,4例造血完全恢复。 2例移植后早期死于重症感染 ,其中 1例曾有早期植入证据。结论　Allo CBSCT治疗成人重型再生障碍性贫血 ,可形成较长期的稳定供、受者造血细胞混合嵌合体。脐血可作为造血干细胞来源的一种选择。     

人脐血单个核细胞体外扩增后植入NOD/SCID小鼠重建多系造血

目的探讨人脐血单个核细胞（MNC）体外扩增后植入能力的改变，以及对NOD／SCID小鼠造血重建的影响。方法采用不同细胞因子组合对人脐血MNC进行短期无血清培养扩增，比较扩增后的效果及细胞凋亡的变化；将扩增6d后的人脐血MNC植入经半致死剂量照射的NOD／SCID小鼠体内，观察小鼠6周后的存活率和造血重建情况。结果人脐血MNC在干细胞因子（SCF）、酪氨酸激酶受体3配基（FL）、白细胞介素6（IL-6）和白细胞介素3（IL-3）细胞因子共同作用下，培养6～11）d获得最佳扩增效果，同时细胞表面膜联蛋白V（Annxin V）的表达明显减少；移植6周后有55．6％的小鼠存活，存活小鼠的骨髓、脾和胸腺细胞中均能检测出人类CD45、CD34、CD33、CD3和CDl9抗原，外周血提取的DNA可检测出人特异的Cart—Ⅰ基因和Alu基因。结论SCF＋FL＋IL-6＋IL-3细胞因子协同作用能有效扩增人脐血MNC，MNC扩增6d后可成功植入N（）D／SCID小鼠体内，并帮助小鼠重建多系造血。     

含高剂量CD34^＋细胞的非血缘脐血移植治疗AML/MDS后原发植入失败

目的分析含高剂量CD34^＋细胞的非血缘脐血移植治疗急性粒细胞白血病合并骨髓增生综合征（AML/MDS）后发生原发植入失败的原因。方法 1例4岁女孩在四川大学华西第二医院儿科血液肿瘤科被诊断为AML/MDS。患儿经诱导及巩固治疗获完全缓解后,行无血缘人类白细胞抗原（HLA）部分不相合脐血移植。观察患儿术后造血重建及移植相关并发症情况。结果患儿术后发生原发植入失败,再次进行血缘间的半相合造血干细胞移植,期间患多药耐药鲍曼不动杆菌败血症,于第2次移植后7 d死于呼吸衰竭。结论含高剂量CD34^＋细胞脐血造血干细胞并不能抵消HLA配型不合的缺陷。患儿原发植入失败可能与患儿移植前存在长期血小板输注无效及潜在免疫异常,尤其是移植前产生抗-HLA供者特异性抗体有关。     

脐血单个核细胞体外扩增的实验研究

目的 探讨含有细胞因子的无血清培养基对脐血单个核细胞(MNC)体外培养后的扩增情况和用于移植的安全性.方法 从新鲜脐血中分离出的MNC,在含细胞因子的无血清培养体系中培养.分别将培养前和培养第10天时的造血细胞进行细胞计数、细胞活力分析、集落分析、流式细胞仪检测表面标记、彗星试验分析DNA的损伤情况、无菌性分析及移植至NOD/SCID小鼠体内等项研究.结果 经过体外短期培养,脐血中MNC、CD34+、CD133+、CD34+CXCR4+及CD34+ VLA-4+细胞扩增倍数均比培养前增高(P<0.05);半固体培养基可支持多系集落的生长;培养前和培养第10天时脐血细胞DNA损伤率均低于5%;无菌性分析提示细胞未受污染.将体外扩增后的脐血造血细胞移植入NOD/SCID小鼠体内,与新鲜脐血移植相比,小鼠的存活时间及植入能力的差异均无统计学意义(P>0.05).结论 脐血造血细胞体外扩增是解决脐血造血细胞数量不足的有效方法.脐血造血细胞经短期培养能为造血干细胞移植提供安全而具植入能力的造血细胞.     

体外扩增后的脐血巨核细胞促进血小板在NOD/SCID小鼠体内的植入

目前,脐血造血干细胞移植已成为治疗血液系统疾病的一种主要手段[1].但脐血移植的植入速度慢,尤其是血小板的恢复时间延长,阻碍其广泛应用.非肥胖糖尿病型重症联合免疫缺陷(NOD/SCID)小鼠不仅存在T、B淋巴细胞功能缺陷,而且自然杀伤(NK)细胞功能低下,巨噬细胞功能基本缺失,无溶血性补体,故成为我们研究脐血体外扩增后造血细胞移植的有效动物模型.     

造血干细胞移植的临床应用

造血干细胞移植（HSCT）是指将各种来源的正常造血干细胞包括骨髓干细胞、外周血干细胞或脐带血干细胞在患者接受超剂量化疗或放疗后，通过静脉输注植入患者体内，重建患者由于各种原因被摧毁或已衰竭的造血及免疫功能。HSCT的理论基础是造血干细胞具有自我更新及分化成熟为各种血细胞和免疫活性细胞的能力。HSCT不仅重建了患者的造血功能，亦重建了患者的免疫功能。     

HLA半相合造血干细胞移植进展

HLA半相合造血干细胞移植因为植入率低,移植物抗宿主病重,免疫重建慢,感染发生率高等并发症,主要用于无相合供者的高危白血病患者。但最近随着基础和临床研究的进展,在移植物处理、免疫耐受诱导、移植后过继免疫治疗等方面取得长足的进步,其总的生存率已经与无关供者的相合造血干细胞移植相似。本文就最近的一些进展做一综述。     

人脐血造血干/祖细胞的磁力搅拌悬浮培养及移植实验

目的 探讨磁搅拌大规模培养体系对人脐血造血祖细胞的扩增效果以及扩增的人造血祖细胞植入动物体内后的造血重建情况.方法 从新鲜抗凝脐血中分离出单个核细胞(MNC),以添加干细胞因子、酪氨酸激酶受体3配基及血小板生成素的无血清培养体系进行培养.静态扩增组的细胞置于T25培养瓶中培养,磁搅拌悬浮扩增组(磁搅拌扩增组)的细胞采用Celstir装置进行培养,培养体系为50～100 ml.培养7 d后进行细胞计数、集落培养检测和细胞表面分子表达的测定.以不进行培养者为对照组.非肥胖糖尿病重症联合免疫缺陷(NOD/SCID)小鼠在接受2.5 Gy的亚致死剂量X射线照射后分别从尾静脉输入上述静态扩增组、磁搅拌扩增组和对照组的MNC(5×106个),另设不移植的空白对照组.观察小鼠的存活情况,6周后处死存活小鼠,检测骨髓细胞中CD34+细胞、CD3+细胞、CD19+细胞、CD33+细胞及CD45+细胞的含量以及人特异的Cart-Ⅰ和Alu基因的表达.结果 经过7天的培养,磁搅拌扩增组的造血祖细胞扩增倍数为(2.8±0.45)倍,明显高于静态扩增组的(2.1±0.48)倍(P<0.01).磁搅拌扩增组形成的红系集落、粒-巨噬细胞集落数均明显高于静态扩增组(P<0.05).静态扩增组扩增后的CD34+细胞、CD34+CD38-细胞和CD133+细胞含量均高于磁搅拌扩增组(P<0.05),而CD184+细胞和CD62L+细胞含量低于磁搅拌扩增组(P<0.01).移植后6周,对照组、静态扩增组和磁搅拌扩增组分别有3、4、5只小鼠存活,三组间两两比较,6周存活率的差异无统计学意义(P>0.05).存活6周的小鼠,其骨髓中能检人特异性CD34+细胞,以及CD3+细胞、CD19+细胞、CD33+细胞及CD45+细胞,也检测到人Alu基因和Cart-Ⅰ基因的表达.结论 磁搅拌培养能大规模扩增脐带血造血祖细胞,扩增的细胞能植入x射线照射的NOD/SCID小鼠,并重建其多系造血.     

体外扩增的脐血细胞移植于BALB/C小鼠的实验研究

目的　探讨经体外扩增的脐血造血细胞对小鼠造血重建功能的影响 ,寻求脐血用于成人移植的可行方法。方法　将经致死剂量γ射线照射后的BALB/C小鼠 ,随机分为 4组 ,每组 10只。A组 :每只移植 2 .5× 10 6个新鲜脐血细胞 ;B组 :每只分别移植 1.2 5× 10 6个新鲜脐血细胞和 1.2 5×10 6个扩增培养 1周的脐血细胞 ;C组 :每只移植 2 .5× 10 6个扩增培养 1周的脐血细胞 ;D组 :每只仅输注 0 .5ml生理盐水。比较各组造血重建的差别。结果　在脐血细胞移植后第 13d ,A组存活 5只(5 0 % ) ,B组 8只 (80 % ) ,C组 6只 (6 0 % ) ,D组全部死亡。B组小鼠的死亡率最低。脐血细胞移植后 14d ,小鼠的WBC已开始回升 ,B组与C组的恢复明显较A组快 ,P <0 .0 5。结论　混合移植新鲜的和体外培养 1周的脐血造血细胞 ,可能是脐血移植的较好办法。     

Host marrow has a competitive advantage that limits donor hematopoietic repopulation in mixed xenogeneic chimeras

Abstract: In mixed xenogeneic (rat/mouse) chimeras, in which both donor and host hematopoietic cells coexist, donor hematopoiesis eventually declines, despite apparent immunological tolerance of the host to the donor. To explain this observation, we hypothesized that donor hematopoietic cells might have a competitive disadvantage in a xenogeneic environment. Since SCID (severe combined immunodeficient) bone marrow cells (BMC) are unable to produce functional B and T lymphocytes, they might compete for myeloid but not lymphocytic reconstitution of the animals, whereas marrow from the normal congenic strain, C.B-17, would compete for repopulation of both lineages. We therefore evaluated the capacity of SCID and C.B-17 mouse marrow to inhibit repopulation by rat BMC in irradiated SCID mouse recipients. Nine weeks after administration of T-cell depleted (TCD) F344 rat BMC alone to 3 Gy-irradiated SCID mice, rat cells of both the myeloid and the lymphoid lineages were readily detectable among peripheral blood leukocytes. Rat plus SCID mouse BMC reconstitution resulted in a marked reduction in rat myeloid repopulation, without affecting rat lymphoid repopulation. Rat plus TCD normal C.B-17 mouse BMC, on the other hand, completely inhibited repopulation of all rat lineages by 9 weeks. Since mouse marrow inhibited only the repopulation by rat cells of the lineages that the mouse marrow was itself capable of generating, we conclude that physiologic factors, such as species specificity or selectivity of cytokines, adhesion molecules, and other molecules important for hematopoiesis, provide a competitive advantage to host marrow over xenogeneic hematopoietic cells. This physiologic barrier must be overcome in addition to immunologic barriers if bone marrow transplantation (BMT) is to provide a feasible approach to inducing xenograft tolerance. In addition, we describe the development of a chronic graft-vs.-host-disease (GVHD)-like syndrome in SCID mouse recipients of TCD rat marrow.     

Successful multilineage engraftment of human cord blood cells in pigs after in utero transplantation

BACKGROUND: Successful engraftment of human hematopoietic stem and progenitor cells (HSPCs) in a large animal may serve not only as a model to study human hematopoiesis but also as a bioreactor to expand human HSPCs in vivo. The aim of this study was to accomplish xenotransplantation of human HSPCs into pig. METHODS: Total mononuclear or CD34-positive HSPCs obtained from human cord blood were xenotransplanted percutaneously under an ultrasonographic guidance into preimmune pig fetuses. Peripheral blood and bone marrow (BM) cells of recipient pigs were collected and analyzed for the presence of human cells by a polymerase chain reaction to detect human specific Alu sequence on DNA extracted from those cells. Fluorescence-activated cell sorting (FACS) analysis was also performed to detect human hematopoietic cells. RESULTS: Transplantation of human cord blood cells into pig fetuses aged less than 52 days postcoitus resulted in a good engraftment rate. In one case, engraftment was detected up to 315 days posttransplantation by polymerase chain reaction. Human hematopoietic cells were detectable also by FACS in peripheral blood and BM. Furthermore, human CD34+ HSPCs were also observed in the BM of recipients. Those CD34+ cells in BM were sorted by FACS and subjected to further analyses. First, in vitro colony formation assay resulted in formations of multilineage colonies. Second, when they were transplanted into an immunodeficient mouse they were engrafted in the mouse. CONCLUSIONS: These data indicate an engraftment of human HSPCs in pig BM. In utero transplantation of human HSPCs into a preimmune pig fetus is useful to establish a pig reproducing human hematopoiesis.     

人脐血造血干/祖细胞的生物反应器大规模扩增及移植实验

目的 利用生物反应器大规模扩增人脐血造血干/祖细胞,并通过动物移植实验检验该方法的有效性.方法 采集抗凝脐血10份,分离出单个核细胞(MNC),分别进行生物反应器扩增培养和静态扩增培养.检测扩增前后细胞表面CD34、CD38、CD133、CD184和CD62L分子的表达,并进行造血干/祖细胞集落的培养.取非肥胖糖尿病重症联合免疫缺陷小鼠,以X射线照射后,分为4组,其中MNC组小鼠注射未经扩增培养的MNC;静态扩增组小鼠注射经过静态扩增培养的细胞;反应器扩增组小鼠注射经过生物反应器扩增培养的细胞;空白对照组小鼠注射生理盐水.移植后6周处死存活小鼠,收集骨髓细胞,检测其中CD45+、CD3+、CD19+和CD33+细胞的含量以及人特异的Cart-Ⅰ和Alu基因的表达.结果 生物反应器扩增前MNC为(1.2～2.8)×108个,扩增后为(3.7～12.6)×108个,扩增后的细胞数明显高于静态扩增培养者(P<0.01).经生物反应器扩增后所形成的红系集落形成单位、粒-巨噬细胞集落形成单位数明显高于经静态扩增者(P<0.05).移植6周后,空白对照组小鼠均死亡,MNC组存活率为35%,静态扩增组存活率为30%,反应器扩增组存活率为62.9%,后者明显高于前二者(P<0.05).各组存活小鼠骨髓细胞中均检测到Alu基因和Cart-Ⅰ基因的表达以及人源CD33+、CD45+、CD3+及CD19+细胞.结论 利用生物反应器可大规模扩增人脐血造血干/祖细胞,所得细胞能植入小鼠体内,并能获得造血功能重建.     

Gene delivery using human cord blood-derived CD34+cells into inflamed glomeruli in NOD/SCID mice

BACKGROUND: Bone marrow reconstitution using genetically-modified hematopoietic stem cells has been reported to confer resistance to inflammation and prevent renal injury in glomerulonephritis. Although this strategy has potentials for clinical use, taking hematopoietic stem cells from bone marrow is highly stressful for patients. In this regard, umbilical cord blood may be a useful alternative and, therefore, we focused on their suitability as a source of hematopoietic stem cells for transplantation-based therapy for glomerulonephritis. METHODS: CD34+ cells were obtained from human umbilical cord blood, retrovirally transduced with human beta-glucuronidase (HBG) gene, and transplanted into nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice. After confirming the successful chimerism, these mice were treated with lipopolysaccharide (LPS), and local HBG expression in glomeruli was examined using immunohistochemical analysis, HBG bioassay, and Western blot analysis. RESULTS: Clonogenic assay showed that 88.4 +/- 5.9% burst-forming unit-erythroid (BFU-E), 79.7 +/- 11.4% in colony-forming unit-macrophage (CFU-M), and 81.1 +/- 14.1% in colony-forming unit-granulocyte (CFU-G), respectively, possessed the transgene after transfection, suggesting that precommited cells were susceptible to retroviral infection. Flow cytometric analysis revealed that 24.1 +/- 14.5% of bone marrow cells in these chimera mice expressed human lymphocyte antigen (HLA) 8 weeks after transplantation. Also, clonogenic assay showed that a sustained engraftment of human hematopoietic cells expressed HBG. CD14-positive cells were recruited into the glomeruli upon LPS treatment and they secreted bioactive HBG, suggesting that cord blood-derived CD34+cells may differentiate into monocyte lineage while maintaining the expression of the transgene. CONCLUSION: These data indicate that umbilical cord blood cells can be utilized as a source of hematopoietic stem cells for the transplantation-based therapy of glomerulonephritis.     

Donor-specific growth factors promote swine hematopoiesis in severe combined immune deficient mice

Abstract: Induction of a state of mixed hematopoietic chimerism following bone marrow transplantation is associated with donor-specific tolerance in the concordant xenogeneic rat-to-mouse species combination. We are now attempting to induce such tolerance in a discordant species combination, pig to mouse. Our initial studies showed that non-immune physiologic factors limited the level of swine hematopoietic reconstitution in severe combined immune deficient (SCID) mice. We have now examined the ability of swine-specific growth factors, interleukin-3 (IL-3), granulocyte macrophage-colony stimulating factor (GM-CSF), and stem cell factor (SCF) to enhance repopulation by swine bone marrow cells in SCID recipients. The results indicate that swine IL-3 promotes pig hematopoiesis in SCID mouse recipients. The percentage of swine class I⁺ cells in bone marrow, spleen, and peripheral blood was markedly increased by a 3-week treatment course with porcine IL-3. In longer-term studies, the effect of IL-3 was further enhanced by combining it with porcine GM-CSF. Almost all repopulating porcine cells expressed a swine myeloid marker. Colony-forming assays showed a correlation of the number of pig-specific colonies with the number of swine cells detected by flow cytometry in transplanted-SCID bone marrow recipients. Porcine CD2⁺ cells which did not express CD4 or CDS coreceptors were also detected in SCID mouse recipients of pig bone marrow, and their numbers were also increased by swine cytokine treatment. Swine IgG, but not B cells were detected in SCID recipients at 3 and 6 weeks following bone marrow transplantation, and declined over time, suggesting that mature B cells engrafted, but that de novo B lymphopoiesis did not occur in these mice. Thus, our study demonstrates that donor-specific growth factors can help to overcome the physiologic barrier to xenogeneic hematopoiesis in the discordant pig to mouse species combination.     

Engraftment of human T, B and NK cells in CB.17 SCID/beige mice by transfer of human spleen cells

Models of severe combined immuno-deficient (SCID) mice reconstituted with a competent human immune system represent a valuable tool for the study of human immune responses in vivo. Reconstitution with human cells can be achieved using large numbers of peripheral blood lymphocytes, but levels of engraftment are poor and graft versus host disease (GVHD) frequently occurs. SCID/beige mice are at the same time deficient for adaptive and innate immunity and the objective of this study was to develop a safe and efficient way to achieve human lymphocyte engraftment in these mice using human spleen cells. After institutional authorisations and informed consent of relatives, a piece of spleen was obtained from cadaveric organ donors and the splenocytes were isolated and cryopreserved for later use. Single intraperitoneal injections of 5-100 x10(6) splenocytes were performed into SCID/beige mice. Reconstitution of a human immune system was monitored weekly by the presence of human cells and IgG in peripheral blood. The mice were sacrificed 4 weeks after the injection and the engraftment in lymphoid organs was studied. A reproducible reconstitution was obtained with intraperitoneal injection of 30-40 x10(6) spleen cells. Human T, B and NK cells as well as human IgG were present in peripheral blood. In lymphoid tissues, the same lymphocytic subpopulations were detected and in addition some antigen presenting cells. The reconstitution was functional because graft rejection was observed after transplantation of human allogeneic tissues. When less than 30 x10(6) cells were injected, the reconstitution was variable. When more than 40 x10(6) cells were injected, GVHD occurred with increasing frequency. In conclusion, we show that intraperitoneal injection of 30-40 x10(6) human splenocytes into SCID/beige mice induces a quick and functional engraftment of human T, B and NK cells with no risk of GVHD. This model may be used to study human transplantation immunobiology in vivo.     

T cells from newborn humans are fully capable of developing into cytotoxic T lymphocyte effector cells in adoptive hosts

BACKGROUND: The finding of reduced incidence of graft-versus-host disease (GVHD) associated with cord blood transplantation, compared to unrelated allogeneic bone marrow, could be related to the lower number of T cells infused in the cord blood (CB) inoculum or it might represent an intrinsic property of CB T cells. We investigated the in vivo function of human cord blood mononuclear cells (CBMC) after their adoptive transfer into lethally irradiated BALB/c radioprotected with severe combined immunodeficiency (SCID) mouse bone marrow. METHODS: The ability of human CBMC to engraft and produce antigen-specific alloreactive cytotoxic T lymphocytes (CTLs) and antibodies was determined by FACS, 51Chromium-release assay, and ELISA. RESULTS: Recipients of human CBMC showed engraftment of high levels of both CD14+ and CD3+ cells. Human cells recovered from the peritoneal cavity of chimeric mice, 1 week after immunization with irradiated allogeneic cells, showed adult-level human CTL response against the immunizing cells, without further stimulation in vitro. In contrast, immunization with the tetanus (TT) antigen did not lead to the generation of anti-TT immunoglobulins (Ig) in the cord blood chimera. Furthermore, whereas the addition of purified adult T cells to the cord blood inoculum resulted in the enhancement of human Ig production of both IgG and IgM subclasses, it could not induce antigen-specific antibodies after immunization. CONCLUSION: This report demonstrates in mice for the first time the generation of classical human alloreactive CTLs, derived from cord blood cells. The alloreactivity exhibited by the cord blood mononuclear cells is not different from that displayed by cells originating from adult blood. Any reduction in the observed GVHD associated with cord blood transplants might therefore represent a quantitative difference in the total number of T cells infused.