Sarin A, Haddad E K, Henkart P A
Experimental Immunology Branch, National Cancer Institute, Bethesda, MD 20892-1360, USA.
J Immunol. 1998 Sep 15;161(6):2810-6.
Since the CTL secreted granule protease granzyme B can activate multiple target caspases, it has been proposed that this pathway is responsible for CTL-induced cytolysis of Fas-negative targets. However, target lysis via the granule exocytosis pathway is completely resistant to caspase inhibitors. To test the possibility that granzymes trigger a postcaspase cytoplasmic apoptotic pathway leading to lysis, we have examined the caspase dependence of several cytoplasmic changes associated with apoptotic death. Rapid prelytic phosphatidylserine externalization was induced in Jurkat target cells by both the Fas ligand (FasL)/Fas and the granule exocytosis effector pathways. This was specifically blocked by peptide ketone caspase inhibitors when induced by the former, but not by the latter, pathway. A rapid prelytic loss of target mitochondrial psi was also induced by both CTL effector pathways, and this was also specifically blocked by caspase inhibitors when induced by the FasL/Fas, but not by the granule exocytosis, pathway. Similarly, target membrane blebbing induced by CTL via the FasL/Fas, but not via the granule exocytosis, effector pathway was specifically blocked by caspase inhibitors. In contrast to the above nonnuclear damage, CTL-induced target staining by the lipid probe FM1-43 reflecting plasma membrane endocytosis was blocked by caspase inhibitors. Thus, when caspase activation is blocked, the granule exocytosis pathway triggers several parameters of target apoptotic damage in addition to lysis, suggesting that granzymes directly trigger a postcaspase cytoplasmic apoptotic death pathway.
由于细胞毒性T淋巴细胞（CTL）分泌的颗粒蛋白酶颗粒酶B可激活多种靶标半胱天冬酶，因此有人提出该途径负责CTL诱导的Fas阴性靶标的细胞溶解。然而，通过颗粒胞吐途径的靶标裂解对半胱天冬酶抑制剂完全耐药。为了测试颗粒酶触发导致裂解的半胱天冬酶后细胞质凋亡途径的可能性，我们研究了与凋亡死亡相关的几种细胞质变化的半胱天冬酶依赖性。Fas配体（FasL）/Fas和颗粒胞吐效应途径均可在Jurkat靶细胞中诱导快速的裂解前磷脂酰丝氨酸外化。当由前者诱导时，肽酮半胱天冬酶抑制剂可特异性阻断这种外化，但由后者途径诱导时则不能。CTL效应途径也可诱导靶标线粒体膜电位的快速裂解前丧失，当由FasL/Fas诱导时，半胱天冬酶抑制剂也可特异性阻断这种丧失，但颗粒胞吐途径则不能。同样，CTL通过FasL/Fas而非颗粒胞吐效应途径诱导的靶标膜泡化也可被半胱天冬酶抑制剂特异性阻断。与上述非核损伤相反，脂质探针FM1-43反映的CTL诱导的靶标染色（反映质膜内吞作用）被半胱天冬酶抑制剂阻断。因此，当半胱天冬酶激活被阻断时，颗粒胞吐途径除了裂解外还触发了靶标凋亡损伤的几个参数，这表明颗粒酶直接触发了半胱天冬酶后细胞质凋亡死亡途径。
