Interleukin-6 production and secretion in human melanoma cell lines: regulation by interleukin-1.

These investigations were designed to test the hypothesis that exogenous and/or endogenous interleukin-1 (IL-1) regulates interleukin-6 (IL-6) production in human melanoma cell lines. Ten cell lines were examined for IL-1 and IL-6 expression. Six of these 10 lines constitutively expressed detectable IL-6 mRNA by RT-PCR; three of these six cell lines also produced intracellular and secreted IL-6 as evidenced by positive reaction for IL-6 using immunohistochemistry staining and ELISA methods; three others produced only intracellular IL-6. Addition of exogenous IL-1 alpha was shown to have the following effects on IL-6 production: first, de novo induction of detectable IL-6 intracellular protein and secreted IL-6 in a cell line void of either; second, stimulation of IL-6 secretion in all three cell lines producing only intracellular IL-6 protein; and third, quantitative enhancement of IL-6 secretion in cell lines that constitutively secreted IL-6. Three of the 10 lines which secreted IL-6 also constitutively secreted IL-1 alpha. Experiments employing the IL-1 receptor antagonist confirmed an extracellular receptor-mediated role of IL-1 in regulation of IL-6 production in such cells. These results indicate that IL-1 can regulate IL-6 in human melanoma cells; however, heterogeneity in the constitutive expression as well as variability in response exists with respect to these two cytokines.

Protein tyrosine kinase inhibition blocks granule exocytosis and cytolytic function of lymphokine-activated killer cells.

Short-term pretreatment of human lymphokine-activated killer cells (LAK) with the protein tyrosine kinase-specific inhibitor Herbimycin A (Herb A) blocked cytotoxic function against the NK-resistant (LAK-sensitive) tumor targets, SK-Mel-1 (human melanoma) and Daudi (human lymphoma). Greater than 50% inhibition of LAK activity was observed after a 2.5-h pretreatment with 0.125 microgram/ml (ca. 0.2 microM) Herb A. Inhibition of LAK occurred over a time interval in which LAK were not dependent upon IL-2 for maintenance of killing function, supporting the conclusion that the drug interfered with mobilization of cytotoxic function. Conjugate formation between LAK and tumor targets was unaffected by Herb A, indicating that inhibition was occurring at a post-binding step. Granule exocytosis as measured by BLT-esterase release was detected from LAK after coincubation with tumor targets, and was inhibited by Herb A pretreatment. The majority of LAK killing was dependent upon extracellular calcium, supporting the hypothesis that granule exocytosis rather than Fas ligand was the principal pathway leading to target cell death. The data suggest that protein tyrosine kinases play a pivotal role in LAK cytolytic function by regulating granule exocytosis.