ACTIVATION AND TRIGGERING OF CYTOXIC CELLS

细胞毒性细胞的激活和触发

• 批准号: 2463758
• 负责人: D M SEGAL
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2463758
• 关键词: CD16 molecule; CD44 molecule; T lymphocyte; antitumor antibody; biological signal transduction; breast neoplasms; cell adhesion molecules; cell mediated cytotoxicity; cytolysis; interleukin 2; laboratory mouse; leukocyte activation /transformation; natural killer cells; neoplasm /cancer immunology; neoplastic growth; protein tyrosine kinase

We have previously used redirected cytotoxicity experiments to study and define triggering molecules on cytotoxic cells. While the TcR and FcR are the principal triggering molecules on human leukocytes, we have recently found that adhesion molecules can also serve as cytotoxic triggers on some cell types. In this project we have used anti-CD44 containing bispecific antibodies to show that CD44 can trigger cytotoxic responses in activated NK cells and in fresh PMN. In NK cells, we found that CD44 becomes a cytotoxic trigger after activation with IL-2. Since CD44 is present before and after activation, we asked what causes activation. No changes in CD44 isoform or protein molecular weight were detected by PCR and immunoprecipitation analysis but activation did require protein transcription. A tyrosine- phosphorylated protein was found to co-precipitate with CD44 from activated NK cells. By adding inhibitors during the effector phase of lysis, we found that PI 3-kinase is required for CD44-dependent lysis and that protein kinase C and the cytoskeleton modulate lysis. No increase in intracellular Ca2+ or release of BLT esterase was observed as a result of CD44 crosslinking, whereas CD16 (Fc RIII) induced both these responses. In addition to CD44, we have found that CD38, CD69, and CD56 are also cytotoxic triggers on activated NK cells. In PMN, CD44 triggers a cytotoxic response as measured in an 18 hr assay. Unlike NK cells, fresh, unactivated PMN mediate this response. PMN recognize and kill target cells coated with hyaluronic acid (HA), the principal ligand of CD44, and this activity is blocked by anti-CD44 antibodies. This is in contrast to NK cells that do not lyse HA coated targets. Thus CD44 and perhaps other adhesion molecules, appear to be important to PMN and NK cell cytotoxic function. In other studies involving effector cell activation, we found that in mice bearing mammary tumors, there is a progressive loss in cytolytic function of T cells that correlates with tumor growth. In looking for the source of this phenomenon, we found that there is a selective loss in STAT5 protein and RNA during tumor growth. Other STAT proteins were unchanged during tumor growth. STAT5 is used in the signaling of some cytokines, including IL-2, 3,5,7,9 and others, and thus may be a target for regulating immune function in vivo.

我们以前使用重定向细胞毒性实验来研究 并定义细胞毒性细胞上的触发分子。 虽然TcR和 FcR是人类白细胞上的主要触发分子， 最近发现，粘附分子也可以作为细胞毒性， 会触发某些细胞类型 在这个项目中，我们使用了抗CD 44 含有双特异性抗体，以显示CD 44可以触发 在活化的NK细胞和新鲜PMN中的细胞毒性应答。 在NK 细胞，我们发现CD 44在活化后成为细胞毒性触发因子， IL-2。 由于CD 44在活化前后都存在，我们问 是什么导致了激活 CD 44亚型或蛋白无变化 采用PCR和免疫沉淀法检测其分子量 但激活需要蛋白质转录。 酪氨酸- 发现磷酸化的蛋白质与来自 激活NK细胞。 通过在效应期加入抑制剂， 我们发现PI 3-激酶是CD 44依赖性裂解所必需的 并且蛋白激酶C和细胞骨架调节溶解。 没有 观察到细胞内Ca 2+增加或BLT酯酶释放 作为CD 44交联的结果，而CD 16（Fc RIII）诱导了 这些回应。 除CD 44外，我们还发现CD 38，CD 69， 和CD 56也是活化NK细胞上的细胞毒性触发物。 在PMN中， 如在18小时测定中测量的，CD 44触发细胞毒性应答。 与NK细胞不同，新鲜的未活化的PMN介导这种反应。 PMN 识别并杀死涂有透明质酸（HA）的靶细胞， CD 44的主要配体，并且该活性被抗CD 44阻断 抗体的 这与不裂解HA包被的NK细胞相反， 目标的 因此，CD 44和其他粘附分子似乎是 对PMN和NK细胞的细胞毒功能很重要。 在其他涉及效应细胞激活的研究中，我们发现， 在携带乳腺肿瘤的小鼠中， 与肿瘤生长相关的T细胞功能。 在寻找 这种现象的来源，我们发现有一个选择性的损失 STAT 5蛋白和RNA在肿瘤生长过程中的作用。 其他STAT蛋白是 在肿瘤生长过程中没有变化。 STAT 5被用于一些细胞的信号传导中。 细胞因子，包括IL-2、3、5、7、9等，因此可以是靶点 用于调节体内免疫功能。