Phorbol myristate acetate-protein complex mimics bioactivity of human IL-1. 1. Direct evidence that PMA temporarily enters and is released from cellular compartment during superinduction protocol.
The K-562 cell line was treated with the superinduction protocol involving phorbol myristate acetate (PMA) for production of human interleukin 1 (IL-1). The resultant dialyzed K-562 superinduced supernatants contained potent IL-1-like activity when tested in 4 bioassays for IL-1. However, these K-562 SIS failed to induce fever in rabbits, the IL-1 like activity was not inhibited by an anti-human IL-1 antiserum in the murine thymocyte (LAF) assay, and the IL-1-like activity did not adhere to a specific immunoadsorbent column. Together with recent evidence that PMA may contaminate supernatants produced by the superinduction protocol and can mimic IL-1 bioactivity (9,10), the results of these novel methods suggested that IL-1 was not the bioactive moiety in K-562 SIS. To further examine the source of K-562 SIS IL-1-like activity, the active K-562 SIS were fractionated over Sephadex G-50, and the large molecular weight component (approximately 50,000) was found to express potent LAF activity yet remained nonpyrogenic. 3H-PMA was added to the superinduction protocol as a tracer for PMA migration. The 3H-PMA provided new and direct evidence that PMA adhered to or entered K-562 cells and adhered to plastic culture flasks during the serum free wash steps of the superinduction, and that PMA was released by K-562 cells into the serum containing supernatants during the incubation phase of the protocol. A 3H-PMA component of K-562 SIS co-migrated on gel filtration with the large molecular weight proteins which expressed LAF activity, and is a nondialyzable contaminant of superinduction supernatants. This PMA/protein complex is the main and perhaps the sole mediator of four distinct biological activities ascribed to IL-1.
D000818:Animals; D001667:Binding, Competitive; D002451:Cell Compartmentation / Q000187:drug effects*; D002460:Cell Line; D015232:Dinoprostone; D006801:Humans; D007106:Immune Sera / Q000494:pharmacology; D007375:Interleukin-1 / Q000096:biosynthesis / Q000276:immunology / Q000502:physiology*; D007941:Leukemia P388 / Q000276:immunology / Q000378:metabolism*; D004915:Leukemia, Erythroblastic, Acute / Q000032:analysis / Q000276:immunology / Q000378:metabolism*; D007942:Leukemia, Experimental / Q000378:metabolism*; D008213:Lymphocyte Activation / Q000187:drug effects; D008264:Macrophages / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D009363:Neoplasm Proteins / Q000008:administration & dosage / Q000032:analysis / Q000378:metabolism; D010704:Phorbols / Q000494:pharmacology*; D010835:Phytohemagglutinins / Q000494:pharmacology; D011458:Prostaglandins E / Q000378:metabolism; D011749:Pyrogens / Q000008:administration & dosage; D011817:Rabbits; D013601:T-Lymphocytes / Q000276:immunology; D013755:Tetradecanoylphorbol Acetate / Q000378:metabolism / Q000494:pharmacology*
D007106:Immune Sera; D007375:Interleukin-1; D009363:Neoplasm Proteins; D010704:Phorbols; D010835:Phytohemagglutinins; D011458:Prostaglandins E; D011749:Pyrogens; D015232:Dinoprostone; D013755:Tetradecanoylphorbol Acetate