Antigen presentation and cytotoxic T lymphocyte killing studied in individual, living cells.
Antigen presentation and cytotoxic T lymphocyte killing studied in individual, living cells.
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在个体活细胞中研究抗原呈递和细胞毒性 T 淋巴细胞杀伤。
DOI：
10.1006/viro.1994.1298
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发表时间：
1994
期刊：
影响因子：
3.7
通讯作者：
Oldstone,M
中科院分区：
文献类型：
--
作者：
Hahn,K;DeBiasio,R;Tishon,A;Lewicki,H;Gairin,JE;LaRocca,G;Taylor,DL;Oldstone,M
通过使用视频增强微分干涉对比和多模式配置的荧光显微镜，分析个体活成纤维细胞和细胞毒性 T 淋巴细胞 (CTL) 克隆之间的相互作用。表达已知主要组织相容性复合体 I 等位基因（MC57：H-2b；Balb：H-2d）的成纤维细胞通过孵育或显微注射淋巴细胞脉络膜脑膜炎病毒的肽片段来致敏杀伤。先前对 CTL 克隆对这些肽和 MHC-I 等位基因的特异性的测定使我们能够研究 CTL 对成纤维细胞的杀伤，以及由于 CTL、靶标和/或肽的“不匹配”而导致的非致命性 CTL 与靶标的相互作用。在病毒肽特异性 MHC 限制性 CTL 杀伤过程中，观察到明显的形态变化（CTL 形状变化、CTL 细胞质中颗粒的运动以及靶细胞收缩和起泡）。当没有发生杀伤时，CTL 在靶细胞上进行长时间的、非随机的运动。同种反应性和病毒特异性CTL在杀伤过程中表现出相同的形态。为了进一步研究个体活细胞内的抗原呈递，用四甲基罗丹明共价标记了 LCMV 糖蛋白肽（aa 272-286，LSDSSGVENPGGYCL）。在 51 Cr 释放测定中，标记肽特异性诱导有效的 CTL 杀伤，但标记肽和未标记肽均未证明对未敏化靶标具有毒性。将标记的肽显微注射到成纤维细胞的细胞质中，导致 CTL 杀死这些细胞，但与 CTL 接触的附近未注射的细胞没有被杀死，表明杀伤是由于显微注射肽的呈递而不是细胞外肽与细胞表面 MHC 的结合。只有当与对肽和注射细胞的 MHC 等位基因特异的 CTL 结合时，注射肽的靶细胞才会被杀死。
Interactions between individual, living fibroblasts and cytotoxic T lymphocyte (CTL) clones were analyzed by using video-enhanced differential interference contrast and fluorescence microscopy in a multimode configuration. Fibroblasts expressing known major histocompatibility complex I alleles (MC57: H-2b; Balb: H-2d) were sensitized for killing by incubating or microinjecting them with peptide fragments of lymphocytic choriomeningitis virus. Previous determination of the CTL clones' specificity for these peptides and MHC-I alleles enabled us to study CTL killing of fibroblasts, and nonlethal CTL interaction with targets due to "mismatches" of the CTL, target, and/or peptide. During viral peptide-specific MHC-restricted CTL killing, distinct morphological alterations were observed (CTL shape changes, movements of granules in CTL cytoplasm, and target cell contraction and blebbing). When no killing occurred, CTL engaged in prolonged, nonrandom movement on the target cells. Alloreactive and virus-specific CTL displayed the same morphology during killing. To study antigen presentation further within individual, living cells, a LCMV glycoprotein peptide (aa 272-286, LSDSSGVENPGGYCL) was covalently labeled with tetramethylrhodamine. In51Cr release assays, the labeled peptide specifically induced potent CTL killing, but neither labeled nor unlabeled peptide proved toxic for unsensitized targets. Microinjection of the labeled peptide into the cytoplasm of fibroblast cells led to CTL killing of those cells, yet nearby uninjected cells contacted by CTL were not killed, indicating that killing was due to presentation of microinjected peptide rather than binding of extracellular peptide to cell surface MHC Peptide-injected target cells were killed only when combined with CTL specific for the peptide and for the MHC allele of the injected cell.