J. C. Hiserodt

Laminin production by murine melanoma cells: possible involvement in cell motility

Three lines of B16 melanoma cells (B16-F1, B16-F10 and B16-BL6) were examined for motility in the micropore filter assay and for synthesis in culture of the basal lamina glycoprotein laminin. All three lines synthesized laminin as judged by the incorporation of [35S]methionine into immunoreactive laminin and secreted (or shed) laminin into the culture medium as indicated by biosynthetic labeling studies and enzyme-linked immunosorbent assays. Immunoreactive laminin was also seen on the surface of the cells as indicated by immunofluorescence staining and by complement-mediated killing. Analysis of [35S]methionine-labeled laminin immunoprecipitates by sodium dodecylsulfatepolyacrylamide gel electrophoresis (SDS-PAGE) both with and without reduction of intersubunit disulfide bonds revealed that all three cell lines produced a similar array of laminin forms, and that the Mr=950kD laminin molecule (but not the uncombined subunits) was secreted into the culture medium. Laminin biosynthesis appeared to be limited by the availability of the Mr=400kD A subunit as shown by the intracellular accumulation of excess B subunit in the form of uncombined B subunit (Mr =200kD) and as a disulfide-linked B dimer (Mr=400 kD). The motility of all three cell lines was stimulated four- to five-fold by the addition of either exogenous laminin from the EHS sarcoma or culture medium from the B16 cells containing the secreted laminin. The stimulated motility was inhibited by antilaminin serum. These observations suggest that the laminin synthesized by the B16 melanoma cells themselves may facilitate their motility.

A Unified Theory for the Mechanism of the Lethal Hit by Cytotoxic T Lymphocytes (CTL) and Natural Killer (NK) Cells as Defined by Antisera Capable of Blocking the Lethal Hit Stage of Cytotoxicity

Understanding the mechanisms in which cytotoxic lymphocytes (particularly cytotoxic T lymphocytes (CTL) and natural killer (NK) cells) mediate the lysis of target cells has been a fundamental problem in molecular immunology. While these reactions involve complex biochemical and physiological processes the mechanism of cytotoxicity has been resolved into several operationally identifiable stages (1–4). In the most simplistic scheme, the effector lymphocyte must recognize and bind to the appropriate target. Subsequent to binding, the killer cell undergoes of series of physiological responses collectively termed activation. It is during this activation phase that the killer cell initiates infliction of the lethal hit on the target. Once the lethal hit has been completed the killer cell can detach and recycle to other targets. Target cells having received the lethal hit are “programmed to lyse” and will rapidly do so during the final stage of the lytic reaction known as the killer cell independent stage.