TARGET CELL DAMAGE BY IMMUNE MECHANISMS

我们检测了细胞毒性淋巴细胞的杀伤能力 哺乳动物的红细胞作为简单的靶细胞。 使用 针对T细胞受体的交联杂合抗体和 靶向膜抗原，有效溶解红细胞和 使用克隆的CTL可以获得有核的靶。 脾T 细胞或胸腺细胞不会引起可检测的靶溶解。 令人惊讶的是，克隆的辅助性T细胞也能杀死红细胞靶细胞。 但当由该方法触发时不具有核的目标。 的 两种效应物溶解红细胞的生理学似乎 类似于经典的CTL介导的有核靶的杀伤。 分析了LGL颗粒溶细胞素的溶细胞机理 通过利用低pH值和低pH值的观察， 离子强度（在等渗蔗糖中）在以下步骤阻断裂解 溶细胞素结合靶细胞。 在洗掉所有未结合的 细胞溶解素是一种稳定的中间体， 重悬于含有中性pH等渗盐水的缓冲液中。 通过以下任一途径，该裂解步骤需要钙 中级编队 结合步骤需要低浓度的钙。 结合的溶细胞素可以在离子强度下，但不能在低pH下。 被蛋白酶消化或被抗溶细胞素中和 抗体的 当有核肿瘤细胞暴露于溶细胞素时 在亚致死剂量下，它们显示出膜损伤的证据， 通过它们的快速膜去极化和离子损失来指示 和小标记。 然而，这种亚致死性损伤是由 肿瘤细胞在几分钟内，如恢复所示， 膜电位和正常分布的钠和 钾 我们发现直径为3-15微米的聚苯乙烯珠粒 涂有抗T3和抗靶抗体的mu将触发 CTL杀死通常不被识别的细胞。 这种溶解似乎 是旁观者溶解的一个特例， 在CTL细胞毒性中观察到。 我们培养了一位新将军 使用靶标记的细胞毒性测量方法 荧光染料BCECF。 这将取代标准的51 Cr 用于多种用途的释放方法。

We have examined the ability of cytotoxic lymphocytes to killing of mammalian red blood cells as simple target cells. Using crosslinked hybrid antibodies against the T cell receptor and a target membrane antigen, efficient lysis of both red cell and nucleated targets could be achieved using cloned CTL. Splenic T cells or thymocytes do not cause detectable target lysis. Surprisingly, cloned helper T cells also killed red cell targets but not nucleated targets when triggered by this method. The physiology of red cell lysis by both effectors appears to be similar to classical CTL-mediated killing of nucleated targets. The mechanism of lysis by LGL granule cytolysin as been analyzed by taking advantage of the observation that both low pH and low ionic strength (in isotonic sucrose) block lysis at a step after cytolysin binding to target cells. After washing off all unbound cytolysin a stable intermediate is formed which will lyse when resuspended in a buffer containing isotonic saline at neutral pH. Calcium is required for this lytic step by either route of intermediate formation. The binding step requires calcium in low ionic strength, but not at low pH. The bound cytolysin can be digested off by proteases or neutralized by anti-cytolysin antibodies. When nucleated tumor cells are exposed to cytolysin at sub-lethal doses, they show evidence of membrane injury as indicated by their rapid membrane depolarization and loss of ions and small markers. However, this sublethal injury is repaired by the tumor cells within a few minutes, as shown by the restoration of membrane potential and normal distribution of sodium and potassium. We have found that polystyrene beads of diameter 3-15 mu coated with both anti-T3 and anti-target antibody will trigger CTL to kill cells not normally recognized. This lysis appears to be a special case of bystander lysis, which is classically not observed in CTL cytotoxicity. We have developed a new general method for measurement of cytotoxicity using target labeling by the fluorescent dye BCECF. This should replace the standard 51Cr release method for many purposes.