Prevention of Liver Fibrosis by the Purinoceptor Antagonist Pyridoxal-Phosphate-6-Azophenyl-2',4'-Disulfonate (PPADS)

Abstract

Background: Hepatic stellate cells (HSC) are important mediators of liver fibrosis. HSC express purinergic receptors for extracellular ATP that induce fibrogenesis. Pyridoxal-phosphate-6-azophenyl-2',4'-disulfonate (PPADS) is a highly bioavailable purinoceptor inhibitor. We sought to determine whether PPADS could prevent experimental liver fibrosis in rats. Materials and Methods: The effect of PPADS as an inhibitor of HSC purinoceptors was compared to the effect of suramin using confocal video microscopy. Rats were treated with CCl₄, dimethylnitrosamine, or common bile duct ligation in the presence or absence of PPADS. Fibrosis in liver sections was assessed using Trichrome and Sirius red stains. In HSC isolated from experimental animals, proliferation was determined by bromodeoxyuridine uptake, apoptosis was determined using Annexin V flow cytometry, and transcription of α(1)-procollagen and fibronectin were determined using quantitative RT-PCR. Results: Both PPADS and suramin inhibited HSC purinoceptor activation, but PPADS had a more durable effect. PPADS completely blocked the development of cirrhosis due to CCl₄ or dimethylnitrosamine but not due to bile duct ligation. PPADS inhibited HSC proliferation, but had no effect on HSC apoptosis. PPADS inhibited transcription of α(1)-procollagen and fibronectin by HSC. Conclusion: Blockade of purinergic receptors is a novel approach to prevention of non-biliary liver fibrosis. The primary action of PPADS is to inhibit HSC proliferation and fibrogenesis. Future design of purinergic receptor inhibitors may be an effective pharmacologic treatment to prevent liver fibrosis.