Interleukin 1 inhibitory activity secreted by a human myelomonocytic cell line (M20)

Abstract

Culture supernatants from a myelomonocytic cell line (M20) were found to inhibit interleukin 1 (IL 1) activity in vitro. The factor, isolated from these supernatants, inhibited augmentation of phytohemagglutinin response of mouse thymus cells induced by IL 1 derived from several established cell lines. Various IL 1-dependent activities such as lymphocyte and fibroblast proliferation in vitro were also inhibited by the factor. The factor did not inhibit IL 2-induced or other proliferative responses not related to IL 1. Preliminary biochemical characterization of the factor indicated that the activity resides in a protein with a molecular mass of 52 kDa.

In vivo anti inflammatory effects of the M20 IL-1 Inhibitor: I. Effects on acute inflammatory parameters

“…IL-1 Inhibitor was purified from the M20 cell line as described [19,22]. Crude supernatant of serum free culture media was concentrated by vacuum ultrafiltration using dialysis tubing, and purified on a HPLC-DEAE column.…”

“…The conditioned medium was also purified by molecular sieving on a Sephacryl S-300 (Pharmacia, Uppsala, Sweden) column, followed by isoelectric focusing (IEF) in free solution using a Rotofor cell (Bio Rad, Richmond, VA). Fractions were collected, dialyzed, bioassayed and the active peaks corresponding to a MW of 52 +-4 kDa with pI 4.1-4.2 were pooled and stored at -70 ~ This pooled peak revealed a single band on the SDS-PAGE [19,22,23]. The IL-1 Inhibitor preparations were routinely tested in vitro for their ability to inhibit IL-1 in the LAF/IL-1 mouse thymocyte assay [19,23].…”

“…Fractions were collected, dialyzed, bioassayed and the active peaks corresponding to a MW of 52 +-4 kDa with pI 4.1-4.2 were pooled and stored at -70 ~ This pooled peak revealed a single band on the SDS-PAGE [19,22,23]. The IL-1 Inhibitor preparations were routinely tested in vitro for their ability to inhibit IL-1 in the LAF/IL-1 mouse thymocyte assay [19,23]. The D10 assay and the fibroblast proliferation assay (1,2) were also used to test for IL-1 inhibitory activity.…”

“…We have previously described [19] an IL-1 inhibitor produced by a human myelomonocytic cell line [20]. The IL-1 inhibitory factor is secreted constitutively by the M20 cell line, which also produces IL-1, from which the IL-1 Inhibitor can be separated [21].…”

“…We partially purified the IL-1 Inhibitor and found it to be a protein of 52 -+ 4 kDa MW with a pI of 4.15 + 0.1 [22]. The M20 IL-1 Inhibitor has been shown to specifically inhibit IL-1 activities in vitro [19,23]. We have recently established that the M20 IL-1 Inhibitor is distinct from the IL-lra [24].…”