Proliferation and transformation of cultured liver fat-storing cells (perisinusoidal lipocytes) under conditions of beta-D-xyloside-induced abrogation of proteoglycan synthesis.
Fat-storing cells (perisinusoidal lipocytes, Ito cells) are the major connective tissue-producing cell type in liver. In areas of necroinflammation the cells proliferate and transform into desmin and smooth muscle alpha-actin-positive myofibroblast-like cells which synthesize a broad spectrum of significant amounts of collagens, proteoglycans, and matrix glycoproteins. Available data suggest a central role for these cells in the pathogenesis of fibrosis. Beta-D-Xyloside, an artificial initiation site for galactose-linked glycosaminoglycans, thereby uncoupling the synthesis of core protein and GAG, was used as a probe to study main cellular functions under conditions of abrogated proteoglycan synthesis. The exposure for 48 hr of fat-storing cells to p-nitrophenyl beta-D-xyloside (PNP-Xyl) increased dose-dependently the synthesis of [35S]sulfate-labeled medium GAG. Maximum stimulation of fivefold above normal was reached at 1.0 mM PNP-Xyl. Higher concentrations of PNP-Xyl progressively decreased the stimulatory effect on GAG synthesis. The relative composition of GAG in medium (60% chondroitin sulfate, 34% dermatan sulfate), at the cell surface, and intracellularly (mainly heparan sulfate) was not changed significantly by PNP-Xyl. The amounts of intracellular and cell surface-bound GAG were reduced by 40 and 30%, respectively, by PNP-Xyl leading to a depletion of heparan sulfate at the cell surface. Pulse-chase experiments revealed that xyloside-initiated GAG were secreted immediately after synthesis into the medium. GAG synthesized in the presence of 1 and 5 mM PNP-Xyl were free of core protein, and the molecular size of the GAG chains was smaller than that of GAG obtained from beta-eliminated proteoglycans synthesized in control cultures. At concentrations above 3 mM PNP-Xyl generated a dose-dependent inhibition of cell proliferation, which was at any stage of culture fully reversible upon removal of the drug. Viability and general protein synthesis were not reduced, but fat-storing cell transformation and deposition of matrix glycoproteins were retarded. Only a very small fraction of drug-treated cells (5 mM PNP-Xyl) did express on the 11th culture day smooth muscle iso-alpha-actin- and desmin-containing cytoskeletal filaments, which are important indicators of transformation into myofibroblast-like cells. Furthermore, the synthesis of hyaluronan and the expression of immunostained fibronectin, laminin, and tenascin were reduced in cultures exposed to 5 mM PNP-Xyl. The described cellular functions were not affected by exposure of fat-storing cells to p-nitrophenyl beta-D-galactoside.(ABSTRACT TRUNCATED AT 400 WORDS)
Gressner
A M
AM
Department of Clinical Chemistry, Philipps University, Marburg, Germany.
D000199:Actins / Q000378:metabolism; D000818:Animals; D015816:Cell Adhesion Molecules, Neuronal / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D003599:Cytoskeleton / Q000187:drug effects / Q000648:ultrastructure; D003893:Desmin / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005109:Extracellular Matrix / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005353:Fibronectins / Q000378:metabolism; D006025:Glycosaminoglycans / Q000378:metabolism; D006027:Glycosides / Q000494:pharmacology*; D006820:Hyaluronic Acid / Q000378:metabolism; D007797:Laminin / Q000378:metabolism; D008055:Lipids / Q000032:analysis*; D008099:Liver / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism; D046911:Macromolecular Substances; D008297:Male; D011509:Proteoglycans / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D019063:Tenascin
D000199:Actins; D015816:Cell Adhesion Molecules, Neuronal; D003893:Desmin; D016326:Extracellular Matrix Proteins; D005353:Fibronectins; D006025:Glycosaminoglycans; D006027:Glycosides; D007797:Laminin; D008055:Lipids; D046911:Macromolecular Substances; D011509:Proteoglycans; D019063:Tenascin; C010807:xylosides; D006820:Hyaluronic Acid; D003094:Collagen