Virchows Archiv B Cell Pathol (1993) 63:181-189 Virchows Arch& B CellPathology Including Molecular Pathology 9 Springer-Verlag 1993 Heparins modulate extracellular matrix and protein synthesis of cultured rat mesangial cells Albert Wolthuis 1, Adriana Boes ~, Jo H.M. Berden 2, and Joris Grond ~ Department of Pathology,University of Groningen, and 2 Department of Nephrology,Universityof Nijmegen,The Netherlands ReceivedAugust31 / AcceptedDecember15, 1992 Summary. Heparins blunt the development of glomeru- losclerosis in several disease models in the rat and this protective effect may be related to suppression of glo- merular cell proliferation. In this study the direct effect of heparins on another key event in glomerulosclerosis, extracellular matrix (ECM) deposition, was examined. Standard heparin (hep) and non-anticoagulant N-desul- fated acetylated heparin (DSA-hep) significantly re- duced the fibronectin content in the conditioned media of subconfluent, confluent, and supraconfluent rat glo- merular mesangial cells (MCs) in culture, as assessed by a sandwich ELISA technique. Both heparins signifi- cantly increased the amount of cell-associated fibronec- tin in sparse and subconfluent MCs. DSA-hep, but not hep, increased the fibronectin content of ECM formed by confluent and supraconfluent MCs. Using 3H-proline pulse-labeling, Hep and DSA-hep were found to signifi- cantly decrease cell-associated collagen in subconfluent but not in confluent MCs. No effects were seen on newly synthesized collagen secreted into the culture medium. Neither hep nor DSA-hep affected total protein synthe- sis, studied by metabolic labeling with 35S-methionine. High resolution 2-D electrophoresis (molecular weight range, 120 to l0 Kd; isoelectric interval, 5.0 to 7.0) re- vealed one particular intracellular protein (molecular weight 54 Kd, pI 5.91) which was consistently overex- pressed by MCs exposed to DSA-hep but underex- pressed in hep. Both heparins affected an identical set of another 19 different intracellular MC proteins (over-/ underexpression or shift to higher molecular weights). In conclusion, the present data demonstrate the pro- found direct metabolic effects of hep and DSA-hep. In addition to their antiproliferative activity, heparins may also affect the course of glomerular disease in-vivo by direct modulation of ECM and protein synthesis of MCs. Correspondence to." A. Wolthuis, Department of Pathology, Univer- sity of Groningen, Oostersingel 63, NL-9713 EZ Groningen, The Netherlands Key words: Heparin - Mesangial cells - Extracellular Matrix - Proliferation - Biosynthesis Introduction The end stage of various chronic renal diseases is charac- terized by progressive glomerular and tubulo-interstitial scarring. In the glomerulus, epithelial cell injury with the formation of adhesions to Bowman's capsule, me- sangial cell (MC) hyperplasia, and an excess accumula- tion of extracellular matrix (ECM) progressively obliter- ates the capillary network with loss of filtration surface (Rennke and Klein 1989). During the last decade much attention has been focused on dietary and pharmacolog- ic maneuvers to interfere with this process of glomerular obsolescence (Klahr et al. 1988). A number of studies have indicated that heparin and heparin derivates have beneficial effects on the develop- ment of vascular and glomerular sclerosis (Hirsh 1991). Heparin remarkably suppressed mesangial cell prolifera- tion in Habu snake venom-induced glomerular injury (Coffey and Karnovsky 1985). Heparin was found to protect remnant glomeruli from the development of scle- rosis after renal ablation or cortical infarction (Olson 1984; Purkerson et al. 1982), to blunt the progression of glomerular disease in rats with chronic nephrosis (Diamond and Karnovsky 1986), and to mitigate glo- merulonephritis (Naparstek et al. 1990) and proteinuria (Berden et al. 1990) in MRL/1 mice with SLE glomerulo- nephritis. The N-desulfated/acetylated derivate of hepar- in, which is devoid of anticoagulant activity, was as ef- fective as intact heparin in preventing obsolescence of remnant nephrons after ablation (Purkerson et al. 1988). In vitro, heparin was found to be a potent growth inhibi- tor of cultured vascular smooth muscle cells (Castellot et al. 1985a; Reilly et al. 1989) as well as glomerular epithelial cells (Adler 1988), and MCs (Castellot et al. 1985b; Groggel et al. 1990). The protective effect of he- parins in glomerular disease may therefore be related to mitigation of glomerular cell proliferation.