Abstract
Lymphokine-activated killer (LAK) cells obtained from normal donors at various days of in vitro cultivation have been studied by several methods including scanning (SEM) and transmission (TEM) electron microscopy, immuno-electron microscopy, in situ hybridization and flow cytometric DNA measurements. In addition, the cytotoxic activity of LAK cells against several tumor cells was examined by 51Cr-release assay and by SEM and TEM. The LAK cells displayed a uniform ultrastructural appearance concerning surface structure and morphology of organelles. They contained typical lysosomal granules which by immuno-electron microscopy showed a specific localization of perforin I (PI). The presence of PI and granzymeA mRNA in the cytoplasm was confirmed by in situ hybridization using specific antisense probes. Frequency and increased of specific mRNA-containing cells was similar for both genes. Single LAK cells were further characterized by peculiar nuclear inclusion bodies (IB) which were presumably formed by trapped profiles of endoplasmic reticulum. Flow cytometric analysis revealed normal DNA content of LAK cells even after prolonged cultivation indicating that the IB were not associated with aneuploidy of the effector cells. The LAK cells were highly effective in lysing K562 and DAUDI cells as shown by 51Cr-release assay. They caused characteristic morphologic alterations of target cells similar to those found in cytotoxic T-lymphocyte (CTL) and NK-cell-mediated cytolysis. SEM and TEM studies on specimens prepared by routine procedures or by cryopreparation showed that the tumor cell membrane was the initial target for the LAK cell attack whereas other cell compartments were damaged only in advanced stages of cytolysis. Summarizing our study demonstrates that LAK cells have a characteristic ultrastructure which in some aspects differs from that of CTL and NK cells, and that LAK cells appear to destroy tumor cells by mechanisms similar to those of other cytotoxic effector cells.
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