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。vbl.使用 抗T细胞受体和A细胞的交联型杂交抗体 靶膜抗原，有效裂解红细胞和 用克隆的CTL可以达到有核的靶细胞。脾T细胞 细胞或胸腺细胞不会引起可检测到的靶点溶解。 令人惊讶的是，克隆的辅助T细胞也杀死了红细胞靶标 但当被这种方法触发时，不会有核目标。这个 这两个效应器的红细胞溶解的生理学似乎是 类似于经典的CTL介导的对有核目标的杀伤。 对LGL颗粒溶细胞素的作用机理进行了分析。 通过利用观察到的低pH和低pH 离子强度(以等渗蔗糖为单位)在下一步阻断裂解 细胞溶血素与靶细胞结合。在洗去所有未捆绑的 细胞溶血素是一种稳定的中间体，当 重新悬浮在含有中性pH的等渗盐水的缓冲液中。 这一裂解步骤需要钙通过以下两种途径之一 中间队形。结合步骤需要低钙 离子强度，但在低pH值下不会。结合的细胞溶血素可以 被蛋白酶消化或被抗细胞溶血素中和 抗体。当有核肿瘤细胞暴露于细胞溶血素时 在亚致死剂量下，它们显示出膜损伤的证据为 由它们快速的膜去极化和离子损失来表示 和小记号笔。然而，这种亚致命性的伤害是通过 几分钟内的肿瘤细胞，如修复所示 膜电位和钠的正态分布 钾。我们发现直径为3-15的聚苯乙烯微珠 涂有抗T3抗体和抗靶抗体的MU将触发 CTL杀死正常不能识别的细胞。这种裂解似乎 是旁观者裂解的特例，这是典型的 观察CTL细胞毒作用。我们已经任命了一位新将军 用靶标记法测量细胞毒性的方法 荧光染料BCECF。这应该会取代标准的51Cr. 用于多种目的的释放方法。

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.