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Isolation and characterization of a novel liver‐derived immunoinhibitory factor
Author(s) -
Tzung ShiePon,
Gaines Katherine C.,
Henderson Mark,
Smith Terry J.,
Cohen Stefan A.
Publication year - 1991
Publication title -
hepatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.488
H-Index - 361
eISSN - 1527-3350
pISSN - 0270-9139
DOI - 10.1002/hep.1840140522
Subject(s) - chromatofocusing , biochemistry , cytosol , biology , isoelectric focusing , microbiology and biotechnology , in vitro , pronase , gel electrophoresis , arginase , glycine , isoelectric point , amino acid , arginine , enzyme , trypsin
Cytosolic extracts prepared from perfused whole liver or purified hepatocytes of C57BL/6 mice inhibited interleukin‐2– and concanavalin A–induced spleen cell proliferation in vitro . In contrast, cytosolic extracts from purified nonparenchymal liver cells had no effect. Arginase and very‐low‐density lipoprotein were previously identified as two immunoinhibitory substances present in liver cytosolic extracts. We demonstrated, however, that inhibitory activity remained after removal of very‐low‐density lipoprotein and arginase from liver cytosolic extract by repeated ultracentrifugation and gel filtration chromatography, respectively, suggesting the presence of another inhibitor. Further purification by anion‐exchange chromatography and chromatofocusing led to the isolation of a novel liverderived immunoinhibitory factor. This liver‐derived immunoinhibitory factor is sensitive to pronase digestion and heat and acid treatment; it has an estimated isoelectric point of 8.25. The M r of liver‐derived immunoinhibitory factor is 28 kD as estimated from its migration on sodium dodecyl sulfate–polyacrylamide gel electrophoresis, which is identical under both reducing and nonreducing conditions, indicating a monomeric nature of this protein. Amino acid composition analysis discloses that liver‐derived immunoinhibitory factor is relatively rich in glycine and proline residues. Interleukin‐2–induced spleen cell proliferation in vitro is inhibited by this liver‐derived immunoinhibitory factor, with a 50% inhibitory dose of 1.4 nmol/L. Furthermore, the biological activity of the liver‐derived immunoinhibitory factor is not confined to mouse spleen cells, since the growth of B16 mouse melanoma and H35 rat hepatoma cells is also inhibited. A comparison with other liver‐derived immunoinhibitors reported previously supports our claim that the liver‐derived immunoinhibitory factor is a novel inhibitory protein. (H EPATOLOGY 1991;14:888–894).