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Studies on the Mechanism of Lymphocyte-Mediated Cytolysis

X. Enucleated Cells As Targets for Cytotoxic Attack¹

From the Department of Medicine and Microbiology of the Johns Hopkins University School of Medicine and the O'Neill Memorial Research Laboratories of the Good Samaritan Hospital, Baltimore, Maryland

Abstract

These studies were addressed to the hypothesis that interactions between a target cell's plasma membrane and its nucleus might be of significance in determining the susceptibility of the cell to lysis by cytotoxic lymphocytes. To this end, P815 mastocytoma cells were enucleated by centrifugation through a discontinuous Ficoll gradient containing 20 µg/ml cytochalasin B. Two types of vesicles were isolated, both enclosed by plasma membrane: one fraction, termed cytoplasts, contained significant quantities of the cytoplasmic marker lactate dehydrogenase, but lacked nuclei by both biochemical and phase-contrast microscopic criteria. The other type, karyoplasts, contained nuclei. Cytoplasts displayed surface H-2 antigens but "capped" these molecules poorly; karyoplasts showed a "capping" capacity similar to that of intact cells. Despite a deficient capacity to "cap" surface H-2, cytoplasts were susceptible to the action of cytotoxic T cells. There was no indication that cytoplasts were more resistant to lysis than either karyoplasts or intact cells. Moreover, sources of NK, K, and lectin-dependent cytotoxic cells were all capable of inducing specific ⁵¹Cr release from both cytoplast and karyoplast preparations.

Thus, in keeping with our earlier suggestion that the target cell plays a passive role in the lytic process, these studies show that possession of a nucleus is not a prerequisite for susceptibility to cell-mediated cytoxicity.

Footnotes

¹ This work was supported by Grants AI 10280 and AI 13894 from the National Institute of Allergy and Infectious Disease. This is communication No. 302 from the O'Neill Memorial Research Laboratories.

² Supported by Medical Scientist Training Program 5T32 GM 07309.

³ Current address, Basic Immunology Program, Fred Hutchinson Cancer Research Center, 1124 Columbia St., Seattle, Washington 98104.