Limiting dilution analysis of the frequency of human T cells and large granular lymphocytes proliferating in response to interleukin 2. I. The effect of lectin on the proliferative frequency and cytotoxic activity of cultured lymphoid cells.
Limiting dilution analysis of the frequency of human T cells and large granular lymphocytes proliferating in response to interleukin 2. I. The effect of lectin on the proliferative frequency and cytotoxic activity of cultured lymphoid cells.
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人类 T 细胞和大颗粒淋巴细胞响应白细胞介素 2 增殖频率的有限稀释分析。 I. 凝集素对培养淋巴细胞增殖频率和细胞毒活性的影响。
DOI：
10.4049/jimmunol.130.2.687
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发表时间：
1983
影响因子：
4.4
通讯作者：
G. Bonnard
中科院分区：
文献类型：
--
作者：
B. Vose;G. Bonnard
介导NK和小T细胞的人外周血LGL通过在不连续Percoll梯度上的密度沉降以高纯度（98%，通过形态学）分离。通过有限稀释分析确定这些亚群在含有IL 2的无凝集素条件培养基存在下的增殖频率。LGL显示增殖细胞前体的频率比小T细胞高20倍（分别为1/200和1/4970）。通过对K562的细胞毒性的平行测试，证实了在IL 2中从LGL制剂扩增的细胞的NK样性质。而T细胞微培养物显示对K562无溶解活性（细胞毒性前体频率小于1/10，000），LGL培养物显示细胞毒性前体频率（1/170）与增殖前体频率相当。两种应答细胞类型均未对NK不敏感小鼠淋巴瘤RL雄性1或异源细胞产生可检测的溶解活性。在培养开始时，PHA的存在仅对LGL增殖产生最小影响（2微克/ml PHA时升至1/74）。相比之下，小T细胞显示出增殖频率的剂量依赖性增加，在2 μ g/ml PHA下达到1/11，提供PBMC、单核细胞或LGL形式的辅助细胞，但不存在T细胞。在大量培养物中证实了在IL 2中扩增的LGL和小T细胞的细胞毒活性。LGL-CLC对NK敏感细胞系和大多数新鲜分离的同种异体人肿瘤靶标显示出高裂解活性。T细胞-CLC显示出对细胞系靶标（K562、Raji、L1210、RL雄性1）几乎没有活性，但对一些新鲜肿瘤细胞具有裂解作用。这些数据建立了人LGL在IL 2依赖性培养物中生长的最佳条件，并表明CLC溶解同种异体人肿瘤的主要贡献者可能是NK细胞。这些数据表明，大量的活化的T细胞不能在体内检测到，在体外诱导IL 2受体的凝集素/抗原是必要的抗原反应性T细胞系的建立。相反，一部分LGL似乎是自发激活的，对IL 2依赖性生长敏感。因此，在没有刺激的情况下，在IL 2存在下培养未分级淋巴细胞可能选择具有NK活性的LGL生长。
Human peripheral blood LGL that mediated NK and small T cells were isolated in high purity (98% by morphology) by density sedimentation on discontinuous Percoll gradients. The proliferative frequency of these subpopulations in the presence of lectin-free conditioned media containing IL 2 was determined by limiting dilution analysis. LGL showed a 20-fold greater frequency of proliferative cell precursors than small T cells (1/200 and 1/4970, respectively). The NK-like nature of cells expanded from LGL preparations in IL 2 was confirmed by parallel testing of the cytotoxicity against K562. Whereas T cell microcultures showed no lytic activity against K562 (cytotoxic precursor frequency less than 1/10,000), LGL cultures showed frequencies of cytotoxic precursors (1/170) comparable to those of proliferative precursors. Neither responder cell type gave rise to detectable lytic activity against NK-insusceptible mouse lymphoma RL male 1 or alloblasts. LGL proliferation was only minimally affected by the presence of PHA at the onset of culture (rise to 1/74 with 2 micrograms/ml PHA). By contrast, small T cells showed a dose-dependent increase of proliferative frequency, to reach 1/11 with 2 micrograms/ml PHA, provided accessory cells in the form of PBMC, monocytes, or LGL but not T cells were present. The cytotoxic activity of LGL and small T cells expanded in IL 2 was confirmed in bulk cultures. LGL-CLC showed high lytic activity against NK-susceptible cell lines and a majority of freshly isolated allogeneic human tumor targets. T cell-CLC showed little activity against cell line targets (K562, Raji, L1210, RL male 1) but were lytic for some fresh tumor cells. These data establish optimal conditions for the growth of human LGL in IL 2-dependent culture and suggest that a major contributor to lysis of allogeneic human tumors by CLC is likely to be NK cells. The data indicate that large numbers of activated T cells cannot be detected in vivo and that in vitro induction of IL 2 receptors by lectin/antigen is necessary for the establishment of antigen-reactive T cell lines. In contrast, a proportion of LGL appear to be spontaneously activated and susceptible to IL 2-dependent growth. Thus, in the absence of stimulation, culture of unfractionated lymphoid cells in the presence of IL 2 is likely to select for the growth of LGL with NK activity.