Thiele D L, Lipsky P E
Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas 75235.
J Immunol. 1992 Jun 15;148(12):3950-7.
Previous studies have demonstrated that the selective toxicity of leucyl-leucine methyl ester (Leu-Leu-OMe) for cytotoxic lymphocytes and myeloid cells is dependent on intracellular conversion to membranolytic metabolites by the acyl transferase activity of the granule enzyme dipeptidyl peptidase I (DPPI) that is enriched in these cells. The mechanism of cell death remained unclear, however, and was the subject of the experiments reported here. When human U937, HL60, or THP-1 myeloid tumor cell lines or murine CTLL-2 cells were treated with Leu-Leu-OMe, early release of both cytosolic 51Cr and soluble [3H]TdR labeled DNA fragments was observed, whereas antibody + C treatment of these cells caused only 51Cr release. Killing of U937 or THP-1 cells by incubation with the lysosomotropic amino acid methyl ester, Phe-OMe also induced only 51Cr release without evidence of DNA fragmentation. Preincubation with Zn2+, a known inhibitor of endonuclease activity prevented Leu-Leu-OMe-induced 51Cr or [3H]TdR release from these cell lines, but had no effect on antibody + C or Phe-OMe-induced 51Cr release. Zn2+ also prevented Leu-Leu-OMe-mediated killing of normal human CD16+ NK cells. Zn2+ had no inhibitory effect on Leu-Leu-OMe uptake or intracellular conversion to (Leu-Leu)n-OMe metabolites by these cell lines. Moreover, Zn2+ did not inhibit 51Cr release from nonnucleated E or nucleated U937 targets induced by extracellular production of DPPI-generated metabolites of Leu-Leu-OMe. Thus, killing of cytotoxic lymphocytes and myeloid cells by Leu-Leu-OMe appears to be dependent on generation of metabolites with membranolytic properties, but cell death induced by this process does not involve simple lysis of the plasma membrane. Rather, intracellular production of DPPI generated (Leu-Leu)n-OMe metabolites appears to trigger, an additional Zn(2+)-sensitive process that is associated with induction of apoptosis in cells with cytolytic potential.
先前的研究表明，亮氨酰 - 亮氨酸甲酯（Leu - Leu - OMe）对细胞毒性淋巴细胞和髓样细胞的选择性毒性取决于通过颗粒酶二肽基肽酶I（DPPI）的酰基转移酶活性在细胞内转化为膜溶解代谢物，DPPI在这些细胞中含量丰富。然而，细胞死亡的机制仍不清楚，这也是本文所报道实验的主题。当用人U937、HL60或THP - 1髓样肿瘤细胞系或小鼠CTLL - 2细胞用Leu - Leu - OMe处理时，观察到胞质51Cr和可溶性[3H]TdR标记的DNA片段均早期释放，而用抗体+C处理这些细胞仅导致51Cr释放。用溶酶体促渗氨基酸甲酯Phe - OMe孵育杀死U937或THP - 1细胞也仅诱导51Cr释放，而无DNA片段化的证据。用已知的核酸内切酶活性抑制剂Zn2+预孵育可阻止Leu - Leu - OMe诱导的这些细胞系中51Cr或[3H]TdR释放，但对抗体+C或Phe - OMe诱导的51Cr释放无影响。Zn2+也可阻止Leu - Leu - OMe介导的正常人CD16+NK细胞的杀伤。Zn2+对这些细胞系摄取Leu - Leu - OMe或细胞内转化为(Leu - Leu)n - OMe代谢物没有抑制作用。此外，Zn2+不抑制由Leu - Leu - OMe的DPPI产生的细胞外代谢物诱导的无核E或有核U937靶细胞的51Cr释放。因此，Leu - Leu - OMe对细胞毒性淋巴细胞和髓样细胞的杀伤似乎取决于具有膜溶解特性的代谢物的产生，但该过程诱导的细胞死亡并不涉及质膜的简单裂解。相反，DPPI产生的(Leu - Leu)n - OMe代谢物的细胞内产生似乎触发了一个额外的对Zn(2+)敏感的过程，该过程与具有细胞溶解潜能的细胞中凋亡的诱导有关。
