10498650

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4

Activated hepatic stellate cells (HSCs) are a potential source of gelatinase A, which accumulates in fibrotic livers. Progelatinase A activation requires its binding to a complex of membrane-type matrix metalloproteinase (MT-MMP) and tissue inhibitor of metalloproteinases (TIMP)-2. These studies examine gelatinase A, MT1-MMP, and TIMP-2 synthesis by HSCs during activation in vitro and the potential role of gelatinase A in promoting HSC proliferation. Gelatinase A, MT1-MMP, and TIMP-2 messenger RNA (mRNA) were all upregulated in HSCs activated on plastic over 5 to 14 days. Gelatinase A expression was maximal at 7 days of culture, coinciding with the peak of HSC proliferation and the onset of procollagen I and alpha-smooth muscle actin (alpha-SMA) mRNA expression. Active forms of gelatinase A of 62 kd and 66 kd were secreted by activated HSCs and reached a maximum of 12.1% of total enzyme in 14-day culture supernatants. Treatment of HSCs with concanavalin A (con A) induced activation of MT1-MMP and enhanced secretion of activated gelatinase A, which reached a maximum of 44.4% of the total enzyme secreted into culture supernatants using 30 microgram/mL con A. [(14)C]-gelatin degradation assays confirmed the presence of gelatinolytic activity in activated HSC supernatants, which reached a maximum level at 7 days of culture. Antisense oligonucleotide inhibition of endogenous progelatinase A production, or the MMP inhibitor 1,10-phenanthroline inhibited (3)H-thymidine incorporation into HSC DNA by greater than 50%. We conclude that HSCs produce progelatinase A during activation in vitro and activate this enzyme coincident with MT1-MMP and TIMP-2 synthesis. Gelatinase A activity is required for maximal proliferation of HSCs in vitro suggesting this metalloproteinase is an autocrine proliferation factor for HSCs.

eng

IM

MEDLINE

0270-9139

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NLM

977-86

pubmed-meshheading:10498650-Animals, pubmed-meshheading:10498650-Biocompatible Materials, pubmed-meshheading:10498650-Cell Division, pubmed-meshheading:10498650-Cells, Cultured, pubmed-meshheading:10498650-Collagen, pubmed-meshheading:10498650-Drug Combinations, pubmed-meshheading:10498650-Enzyme Activation, pubmed-meshheading:10498650-Enzyme Precursors, pubmed-meshheading:10498650-Gelatinases, pubmed-meshheading:10498650-Isoenzymes, pubmed-meshheading:10498650-Laminin, pubmed-meshheading:10498650-Liver, pubmed-meshheading:10498650-Male, pubmed-meshheading:10498650-Matrix Metalloproteinases, Membrane-Associated, pubmed-meshheading:10498650-Metalloendopeptidases, pubmed-meshheading:10498650-Proteoglycans, pubmed-meshheading:10498650-RNA, Messenger, pubmed-meshheading:10498650-Rats, pubmed-meshheading:10498650-Rats, Sprague-Dawley, pubmed-meshheading:10498650-Tissue Inhibitor of Metalloproteinase-2

Progelatinase A is produced and activated by rat hepatic stellate cells and promotes their proliferation.

Journal Article, Research Support, Non-U.S. Gov't