Lysosomes and Fas-mediated liver cell death

Robert Wattiaux (1) , Simone Wattiaux-de Coninck , Jacqueline Thirion , Marie Christine Gasingirwa , Michel Jadot

1 Chimie Physiologique

A number of studies, mostly performed « ex vivo », suggest that lysosomes are involved in apoptosis as a result of a release of their cathepsins into the cytosol. These enzymes could then be contributing to the permeabilization of the outer mitochondrial membrane, they could also activate effector caspases. This work aims at testing if the membrane of liver lysosomes is disrupted during Fas-mediated cell-death of hepatocytes «in vivo », a process implicated in several liver pathologies. Apoptosis was induced by injecting mice with antiFas antibody (aFas). The state of lysosomes was assessed by determining the proportion of lysosomal enzymes ({beta}-galactosidase, {beta}-glucuronidase, cathepsin C and cathepsin B) present in homogenate supernatants, devoid of intact lysosomes and by analyzing the behaviour in differential and isopycnic centrifugation of {beta}-galactosidase. Apoptosis was monitored by measuring caspase-3 activity (DEVDase) and the release of sulfite cytochrome c reductase, an enzyme located in the mitochondrial intermembrane space . Results show that an injection of 10 microgram aFas causes a rapid and large increase of DEVDase activity and of unsedimentable sulfite cytochrome c reductase. This neither modifies the proportion of unsedimentable lysosomal enzyme in the homogenates nor the behavior of lysosomes in centrifugation. Experiments performed with a lower dose of aFas (5 microgram) indicate that lysosomal hydrolase unsedimentable activity increases in the homogenate after injection but with a marked delay with respect to the increase of DEVDase activity and of unsedimentable sulfite cytochrome c reductase. Comparative experiments «ex vivo» performed with Jurkat cells show an increase of unsedimentable lysosomal hydrolases but much later than caspase-3 activation and a release of dipeptidylpeptidase III and DEVDase into culture medium. It is proposed that the fragilisation of lysosomes observed after aFas treatment «in vivo» and «ex vivo» results from a necrotic process that takes place late after initiation of apoptosis.

Mots clés: Life Sciences

Robert Wattiaux, Simone Wattiaux-de Coninck, Jacqueline Thirion, Marie Christine Gasingirwa, Michel Jadot. Lysosomes and Fas-mediated liver cell death. Biochemical Journal, 2006, 403 (1), pp.89-95. ⟨10.1042/BJ20061738⟩. ⟨hal-00478700⟩