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.