THE CYTOLYTIC PROTEIN OF HUMAN-LYMPHOCYTES RELATED TO THE 9TH COMPONENT (C9) OF HUMAN-COMPLEMENT - ISOLATION FROM ANTI-CD3-ACTIVATED PERIPHERAL-BLOOD MONONUCLEAR-CELLS
THE CYTOLYTIC PROTEIN OF HUMAN-LYMPHOCYTES RELATED TO THE 9TH COMPONENT (C9) OF HUMAN-COMPLEMENT - ISOLATION FROM ANTI-CD3-ACTIVATED PERIPHERAL-BLOOD MONONUCLEAR-CELLS
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DOI：
10.1073/pnas.84.8.2426
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发表时间：
1987-04-01
影响因子：
11.1
通讯作者：
MULLEREBERHARD, HJ
中科院分区：
文献类型：
--
作者：
ZALMAN, LS;MARTIN, DE;MULLEREBERHARD, HJ
最近从白介素2依赖培养的人大颗粒淋巴细胞中分离出一种70 kDa的通道形成蛋白。该蛋白被证明与补体的第九组分(C9)具有免疫化学相关性，因此被命名为C9相关蛋白(C9RP)。利用开发的从大颗粒淋巴细胞中分离C9RP的方法--即用Sepharose连接的抗人C9亲和层析法，现在已经从OKT3激活的人外周血单核细胞中分离出一种细胞溶解蛋白。从1次中获得19~40微克的活性蛋白。109个人外周血单个核细胞在OKT3(细胞表面抗原T3的单抗)作用下培养3d。在此期间，细胞内的细胞毒蛋白含量显著增加，表明OKT3诱导了蛋白质的从头合成。用NaDodSO4/PAGE测定其相对分子质量为70 kDa。经EL ISA检测，大颗粒淋巴细胞分离蛋白和C9RP与抗C9RP反应程度相同。以K-562或M21人黑色素瘤细胞为靶细胞，该分离蛋白在5 mM钙离子存在下的细胞毒活性与C9RP相当。从OKT3激活前去除CD16+细胞的外周血单核细胞中分离出相同的细胞溶解蛋白，该细胞主要由CD4+和CD8+T淋巴细胞组成。这些结果表明，OKT3激活的细胞毒性T淋巴细胞的细胞溶解蛋白与白细胞介素2刺激的大颗粒淋巴细胞的C9RP是一致的。
A 70-kDa channel-forming protein has recently been isolated from human large granular lymphocytes maintained in interleukin-2-dependent culture. The protein was shown to be immunochemically related to the ninth component of complement (C9) and was therefore designated C9-related protein (C9RP). Using the procedure that was developed for the isolation of C9RP from large granular lymphocytes-i.e., affinity chromatography employing anti-human C9 linked to Sepharose, a cytolytic protein has now been isolated from OKT3-activated human peripheral blood mononuclear cells. Nineteen to 40 .mu.g of active protein was obtained from 1 .times. 109 human peripheral blood mononuclear cells after the cells were cultured for 3 days with OKT3 (monoclonal antibody to cell surface antigen T3). During this period, a marked increment occurred in the amount of the cytotoxic protein contained per cell, indicating that OKT3 induced de novo synthesis of the protein. By NaDodSO4/PAGE the molecular mass was determined to be 70 kDa. By ELISA the isolated protein and C9RP of large granular lymphocytes reacted to the same extent with anti-C9RP. Using K-562 or M21 human melanoma cells as targets, the cytotoxic activity of the isolated protein, in the presence of 5 mM Ca2+, was comparable to that of C9RP. The same cytolytic protein was isolated from peripheral blood mononuclear cells that were depleted of CD16+ cells prior to OKT3 activation and that consisted primarily of CD4+ and CD8+ T lymphocytes. These results suggest that the cytolytic protein of OKT3-activated cytotoxic T lymphocytes is identical with C9RP of interleukin-2-stimulated large granular lymphocytes.