Biochimica et Biophysica Acta, 1053 (1990) 135-143 135 Elsevier BBAMCR 12734 Proteoglycans synthesized in vitro by nude and normal mouse peritoneal macrophages Vera L. Petricevich * and Yara M. Michelacci Escola Paulista de Medicina, Departamento de Bioqulmica, Sao Paulo (Brazil) (Received 30 November 1989) Key words: Proteoglycan synthesis; Maerophage;(Mouse peritoneum) Peritoneal macrophages from nude mice were found to be functionally similar to 'activated' macrophages from normal mice. The objective of the present study was to characterize the proteoglycans synthesized and secreted in vitro by peritoneal macrophages isolated from nude and normal Balb/c mice and to investigate the relationship between macrophage 'activation' and changes in the proteoglycan patterns. Macrophages obtained by peritoneal lavage were seeded in Petri dishes. After 2 h incubation at 37 ° C, the adherent cells (macrophages) were exposed to [35S]sulphate for the biosynthetic labelling of proteoglycans. After incubation, the cell and medium fractions were collected and analysed for proteoglycans and glycosaminoglycans. The glycosaminoglycans were identified and characterized by a combination of agarose gel electrophoresis and enzymatic degradation with specific mucopolysaccharidases. It was shown that 3/4 of the total 35S-labelled glycosaminoglycans were in the extracellular compartment after 24-48 h. The macrophages synthesized dermatan sulphate (68%), chondroitin sulphate (7%) and heparan sulphate (25%). Both cell and medium fractions of normal and nude mouse macrophages contained glycosaminoglycans with the same ratios, although the nude mouse macrophages synthesized 2-fold less glycosaminoglycans than the normal mouse macrophages. Lower levels of 3SS-proteoglycans were also obtained from in vitro 'activated' macrophages, but the ratios of dermatan sulphate: chondroitin sulphate: heparan sulphate were altered in these cells as compared to the control. Furthermore, all the 3SS-macromolecules found in the extracellular compartment of nude and normal control cells were of proteoglycan nature, in contrast to the medium fractions of 'activated' macrophages, which contain both intact proteoglycans and 'free' glycosaminoglycan chains. These results indicate that, at least as regards the proteoglycans and glycosaminogly- cans, the nude mouse macrophages are not identical to the 'activated' macrophages from normal mice. Introduction Macrophages have been shown to be involved in different homeostatic mechanisms and pathological events and may be engaged in complex interactions with lymphocytes, granulocytes, tumour cells and plate- lets, as well as with the complement, coagulation and kinin systems [1]. For years investigators focused on Abbreviations: GAG, glycosaminoglycan(s); ADi4S, 2-acetamido-2- deoxy-3-O-(4-enepyranosyluronie aeid)-4-O-sulfo-D-galactopyranose; ADi6S, 2-acetamido-2-deoxy-3-O-(4-enepyranosyluronic acid)-6-O- sulfo-D-galactopyranose. * Permanent address: V.L. Petricevich,Laboratorio Especial de Im- unologia Viral, Instituto Butantan, Av. Vital Brasil, 1500 - 05505 SAo Paulo, Brazil. Correspondence: Y.M. Miehelacei,Escola Paulista de Medieina, De- partamento de Bioquimica,Caixa Postal 20372, 04023, Silo Paulo, SP, Brazil. phagocytosis and intracellular digestion of particles by macrophages but, in the past few years, it was shown that macrophages are also secretory cells [1]. The secre- tory repertoire of the macrophages also includes proteo- glycans. Proteoglycans are glycoconjugates that consist of a protein core with covalently linked glycosaminoglycan side-chains [2]. They are ubiquitous compounds found in a wide variety of vertebrate and invertebrate tissues [3] and are actively synthesized by cells in culture [4,5]. The proteoglycans have been shown to be present on the plasma membrane and extracellular matrix [6,7] and exhibit a peculiar structural variability according to the tissue and species of origin [2,7]. Alterations of the differentiative or functional state of cells are often reflected in quantitative or qualitative changes in the proteoglycan patterns [8-11]. Kolset et al. [12-14] have shown that human mono- cytes maintained in vitro on plastic or glass substrates differentiate into 'macrophage-like' cells and switch the 0167-4889/90/$03.50 © 1990 Elsevier SciencePublishers B.V. (BiomedicalDivision)