DIRECT DEMONSTRATION OF THE CLONOGENIC POTENTIAL OF EVERY HUMAN PERIPHERAL-BLOOD T-CELL - CLONAL ANALYSIS OF HLA-DR EXPRESSION AND CYTOLYTIC ACTIVITY
DIRECT DEMONSTRATION OF THE CLONOGENIC POTENTIAL OF EVERY HUMAN PERIPHERAL-BLOOD T-CELL - CLONAL ANALYSIS OF HLA-DR EXPRESSION AND CYTOLYTIC ACTIVITY
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DOI：
10.1084/jem.157.2.743
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发表时间：
1983-01-01
影响因子：
15.3
通讯作者：
MINGARI, MC
中科院分区：
文献类型：
--
作者：
MORETTA, A;PANTALEO, G;MINGARI, MC
为了确定人外周血T细胞的克隆形成特性，开发了一种有限稀释微培养系统，该系统使用植物血凝素（PHA）作为T细胞激活剂，并使用来自PHA刺激的脾培养物的上清液作为T细胞生长因子的来源。在这些培养条件下能够广泛增殖的细胞频率在外周血单核细胞、E（红细胞）-玫瑰花结阳性和E -玫瑰花结阴性细胞群中分别为0.52 - 0.73、0.98 - 1.11和<0.02。在使用显微操作分离单个细胞的实验中，几乎所有T细胞的克隆形成潜能都得到了证实。在培养第14天，克隆大小在5 - 30×10⁴个细胞之间。使用相同的微培养系统来确定所有细胞毒性T淋巴细胞（CTL - P）的前体频率。通过一种依赖凝集素的⁵¹Cr释放试验评估，纯化T细胞群中的CTL - P频率在0.30 - 0.34之间。能够裂解[人白血病]K562靶细胞的T细胞前体频率在0.14 - 0.16之间。对单个克隆培养物的两种裂解活性的平行分析表明，50%表现出依赖凝集素裂解的克隆对K562靶细胞也有活性。通过流式细胞荧光测定评估，所有增殖的克隆都表达HLA - DR抗原，尽管程度不同。鉴于该培养系统的高克隆效率，现在似乎有可能确定T细胞群中各类功能细胞的前体频率。
To determine the clonogenic properties of human peripheral blood T cells, a limiting dilution microculture system using phytohemagglutinin (PHA) as T cell activator and supernatant from PHA-stimulated spleen cultures as a source of T cell growth factors was developed. The frequencies of cells capable of extensive proliferation under these culture conditions were 0.52-0.73, 0.98-1.11 and < 0.02 in peripheral blood mononuclear, E[erythrocyte]-rosette-positive and E-rosette-negative cell populations, respectively. The clonogenic potential of virtually all T cells was confirmed in experiments using single cells isolated by micromanipulation. Clone size ranged between 5-30 .times. 104 cells on day 14 of culture. The same microculture system was used to determine the precursor frequency of all cytolytic T lymphocytes (CTL-P). As assessed by a lectin-dependent 51Cr release assay, the CTL-P frequency in purified T cell populations ranged between 0.30-0.34. The precursor frequency of T cells capable of lysing [human leukemia] K562 target cells was between 0.14-0.16. Parallel analysis of individual clonal cultures for both lytic activities showed that 50% of the clones exhibiting lectin-dependent lysis were also active against K562 target cells. All of the proliferating clones expressed HLA-DR antigens, although to a varying degree, as assessed by flow cytofluorometry. Given the high cloning efficiency of this culture system, it now appears possible to determine the precursor frequencies of the various classes of functional cells in T cell populations.