Abstract
Previous studies have indicated that endotoxin and other bacterial and protozoal products can stimulate macrophages to produce a factor that can suppress the activity of the enzyme lipoprotein lipase (LPL), in vivo and in vitro. In the present report we describe the purification of this factor, cachectin, to apparent homogeneity from the conditioned medium of endotoxin-stimulated RAW 264.7 cells. The isolated protein has an isoelectric point of 4.7 and a subunit molecular weight of 17,000. Although cachectin's isoelectric point and molecular weight are similar to those described for interleukin 1, pure cachectin has no leukocyte-activating factor (LAF) activity. Cachectin at a concentration of 10(-11) M has the ability to suppress the LPL activity of the 3T3-L1 adipocyte cell line by 80%. Binding studies using radio- labeled cachectin and 3T3-L1 adipocytes and C2 myotubules revealed approximately 10(4) high-affinity receptors per cell on both cell types (Ka, 3 X 10(9]. Cachectin receptors were also present on liver membranes but were absent on erythrocytes and lymphocytes. The isolation of cachectin and characterization of its receptor should facilitate further investigations into the role of cachectin and other macrophage mediators in the metabolic derangements that occur during infection and cachexia.
Full Text
The Full Text of this article is available as a PDF (871.0 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Fraker P. J., Speck J. C., Jr Protein and cell membrane iodinations with a sparingly soluble chloroamide, 1,3,4,6-tetrachloro-3a,6a-diphrenylglycoluril. Biochem Biophys Res Commun. 1978 Feb 28;80(4):849–857. doi: 10.1016/0006-291x(78)91322-0. [DOI] [PubMed] [Google Scholar]
- Francis R. T., Jr, Davie J. R., Sayre M., Rocha E., Ziemer F., Riedel G. Efficient method for visualization and isolation of proteins resolved in polyacrylamide gels. J Chromatogr. 1984 Aug 17;298(1):115–121. doi: 10.1016/s0021-9673(01)92699-8. [DOI] [PubMed] [Google Scholar]
- Green H., Meuth M. An established pre-adipose cell line and its differentiation in culture. Cell. 1974 Oct;3(2):127–133. doi: 10.1016/0092-8674(74)90116-0. [DOI] [PubMed] [Google Scholar]
- Hotez P. J., Le Trang N., Fairlamb A. H., Cerami A. Lipoprotein lipase suppression in 3T3-L1 cells by a haematoprotozoan-induced mediator from peritoneal exudate cells. Parasite Immunol. 1984 May;6(3):203–209. doi: 10.1111/j.1365-3024.1984.tb00793.x. [DOI] [PubMed] [Google Scholar]
- Kawakami M., Cerami A. Studies of endotoxin-induced decrease in lipoprotein lipase activity. J Exp Med. 1981 Sep 1;154(3):631–639. doi: 10.1084/jem.154.3.631. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kawakami M., Pekala P. H., Lane M. D., Cerami A. Lipoprotein lipase suppression in 3T3-L1 cells by an endotoxin-induced mediator from exudate cells. Proc Natl Acad Sci U S A. 1982 Feb;79(3):912–916. doi: 10.1073/pnas.79.3.912. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lachman L. B., Hacker M. P., Handschumacher R. E. Partial purification of human lymphocyte-activating factor (LAF) by ultrafiltration and electrophoretic techniques. J Immunol. 1977 Dec;119(6):2019–2023. [PubMed] [Google Scholar]
- Lomedico P. T., Gubler U., Hellmann C. P., Dukovich M., Giri J. G., Pan Y. C., Collier K., Semionow R., Chua A. O., Mizel S. B. Cloning and expression of murine interleukin-1 cDNA in Escherichia coli. 1984 Nov 29-Dec 5Nature. 312(5993):458–462. doi: 10.1038/312458a0. [DOI] [PubMed] [Google Scholar]
- Mackall J. C., Student A. K., Polakis S. E., Lane M. D. Induction of lipogenesis during differentiation in a "preadipocyte" cell line. J Biol Chem. 1976 Oct 25;251(20):6462–6464. [PubMed] [Google Scholar]
- Nilsson-Ehle P., Schotz M. C. A stable, radioactive substrate emulsion for assay of lipoprotein lipase. J Lipid Res. 1976 Sep;17(5):536–541. [PubMed] [Google Scholar]
- Ralph P., Nakoinz I. Antibody-dependent killing of erythrocyte and tumor targets by macrophage-related cell lines: enhancement by PPD and LPS. J Immunol. 1977 Sep;119(3):950–954. [PubMed] [Google Scholar]
- Raschke W. C., Baird S., Ralph P., Nakoinz I. Functional macrophage cell lines transformed by Abelson leukemia virus. Cell. 1978 Sep;15(1):261–267. doi: 10.1016/0092-8674(78)90101-0. [DOI] [PubMed] [Google Scholar]
- Rouzer C. A., Cerami A. Hypertriglyceridemia associated with Trypanosoma brucei brucei infection in rabbits: role of defective triglyceride removal. Mol Biochem Parasitol. 1980 Oct;2(1):31–38. doi: 10.1016/0166-6851(80)90046-8. [DOI] [PubMed] [Google Scholar]
- Wray W., Boulikas T., Wray V. P., Hancock R. Silver staining of proteins in polyacrylamide gels. Anal Biochem. 1981 Nov 15;118(1):197–203. doi: 10.1016/0003-2697(81)90179-2. [DOI] [PubMed] [Google Scholar]