Adiponectin's anti-fibrogenic action on hepatic stellate cells: stimulation of TIMP-1 and the mediation of HSC apoptosis, proliferation and invasion

Adiponectin (ADN), an adipocytokine with anti-inflammatory, anti-atherogenic and insulin sensitising properties induces an anti-fibrogenic HSC profile in vitro. ADN has been previously reported by us to inhibit proliferation, migration and to induce apoptosis in activated HSCs. Adiponectin binds to the AdipoR1 and -R2 receptors and signals through the AMP- activated protein kinase (AMPK). We find that adiponectin through AMPK stimulates HSCs to release the matrix metalloproteinase inhibitor, TIMP-1. Traditionally, TIMP-1 has been associated with the progression of liver fibrosis. Thus, the induction of TIMP-1 by adiponectin would appear to be counter-intuitive to its published anti-fibrotic effects. Therefore, the aim of this study was to determine the functional significance of adiponectin-stimulated TIMP-1 on activated HSCs and their role in liver fibrosis. Methods: Activated primary rat HSCs were treated with ADN for between 4 and 48 h. To determine the signalling pathways that mediate ADN’s stimulation of TIMP-1, cells were treated with the AMPK antagonists, Compound C or AraA. HSC proliferation and apoptosis was assessed by BrdU incorporation and Annexin V binding (FACS), respectively. TIMP-1 protein levels were determined by ELISA and HSC invasion assays were conducted according to a modified migration assay using transwell membranes (Boyden chambers) coated with rat tail collagen I. Results: ADN treatment for 4–48 h resulted in approximately a threefold increase in TIMP-1 levels. The application of ADN for 24 h concurrently with Compound C or AraA reduced TIMP-1 levels. Treatment of activated HSCs with ADN for 48 h resulted in a 20% increase in apoptosis as measured by Annexin V binding and a 50% reduction in proliferation. Concurrent treatment of ADN with a TIMP-1 blocking antibody resulted in a further 30% increase in Annexin V binding and a further reduction in proliferation. Furthermore, ADN treatment of activated HSCs resulted in a 50% reduction in invasion. Similarly, the concurrent treatment of ADN with a TIMP-1 blocking antibody reversed the inhibitory effects of ADN on HSC invasion. Conclusion: Adiponectin stimulates TIMP-1 levels in activated HSCs through an AMPK-dependent mechanism. TIMP-1 mediates ADN- associated reductions in HSC invasion and modulates its effects on HSC apoptosis and proliferation. These data suggest novel roles for TIMP-1 in the context of liver fibrosis. We are now exploring the association of adiponectin and TIMP-1 in mouse models of liver fibrosis.

Journal of Gastroenterology and Hepatology

Journal of Gastroenterology and Hepatology

25

1440-1746

N/A

Suppl. 3

1