免疫毒素去除脐血T细胞效率及对造血祖细胞的影响

Effects of immunotoxin on the depletion of T cells and on hematopoietic progenitor cells in human cord blood

目的探讨抗Ｔ细胞免疫毒素（ｉｍｍｕｎｏｔｏｘｉｎ，ＩＴ）体外对脐血Ｔ细胞的清除效率及对造血干／祖细胞的影响。方法（１）用碱性磷酸酶抗碱性磷酸酶桥联酶标技术法（ＡＰＡＡＰ）分别测定了脐血、骨髓、外周血标本ＣＤ＋５、ＣＤ＋８Ｔ细胞的百分率；（２）用单向混合淋巴细胞培养（ＭＬＣ）方法比较了外周血与脐血ＭＬＣ增殖反应；（３）分别用ＭＬＣ法和噻唑蓝（ＭＴＴ）比色法，观察了两组抗人白细胞分化抗原的单克隆抗体（ＣＤ＋５、ＣＤ＋８）与完整蓖麻毒素（Ｒｉｃｉｎ）偶联制备的两组免疫毒素（ＣＤ５：Ｒｉｃｉｎ、ＣＤ８：Ｒｉｃｉｎ）对脐血Ｔ细胞增殖反应及对Ｔ淋巴细胞转化功能抑制的影响；（４）用造血祖细胞培养的方法观察两组免疫毒素对脐血粒单系祖细胞（ＣＦＵ－ＧＭ）、早期红系祖细胞（ＢＦＵ－Ｅ）及混合系祖细胞（ＣＦＵ－Ｍｉｘ）的影响。结果（１）脐血含有一定比例的ＣＤ＋５及ＣＤ＋８Ｔ细胞亚群；（２）在ＭＬＣ中外周血与脐血对异源抗原的增殖反应相似；（３）ＣＤ５：Ｒｉｃｉｎ１０－１０～１０－８ｍｍｏｌ／Ｌ（Ｍ）、ＣＤ８：Ｒｉｃｉｎ在１０－９～１０－８ｍｍｏｌ／Ｌ浓度范围内均能有效降低脐血Ｔ淋巴细胞在ＭＬＣ中的增殖反应，并能有效抑制Ｔ淋巴细胞转?

Objective To study the selective toxicity of immunotoxin (IT) on T cells and to determine its effect on hematopoietic progenitor cells. Methods The percentage of CD + 5 and CD + 8 T cell subsets in cord blood (CB) and bone marrow (BM) as well as peripheral blood (PB) was measured by using alkaline phosphatase and monoclonal anti alkaline phosphatase (APAAP) technique. One way mixed lymphocyte culture (MLC) was performed to compare the proliferative response of CB with that of PB. The proliferative capability of cord blood T cells and T lymphocytes transformation capacity were evaluated in the presence of anti CD 8 or anti CD 5 immunotoxin by one way MLC and colorimetric MTT (tetrazolium) assay, respectively. The effects of IT on the growth of hematopoietic progenitor cells of colony forming unit granulocyte and macrophage (CFU GM), burst forming unit erythroid (BFU E) and multipotential hematopoietic progenitors (CFU Mix) from CB were estimated by colony forming assays. Results (1) A certain proportion of CD + 5 and CD + 8 T cells existed in CB. (2) The alloproliferative capacity of CB was the same as that of PB. (3) CD 5:Ricin at the dosage of 10 -10 ～10 -8 mmol/L (M) and CD 8:Ricin at the range of 10 -9 ～10 -8 M could effectively decrease both the proliferative capability of CB and T cell transformation. (4) At the range of 10 -10 ～10 -9 M, both kinds of IT didn′t affect the growth of hematopoietic progenitor cells. Conclusion Graft versus host disease (GVHD) might occur after cord blood transplantation (CBT). CD 5:Ricin and CD 8:Ricin might effictively deplete T cell and might potentially be used to decrease and prevent GVHD in HLA mismatched related and unrelated CBT.