Lysosomes and Fas-mediated liver cell death.
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 contribute to the permeabilization of the outer mitochondrial membrane; they could also activate effector caspases. The present study aims at testing whether 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 aFas (anti-Fas antibody). 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 analysing 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 microg of aFas causes a rapid and large increase in DEVDase activity and in unsedimentable sulfite cytochrome c reductase. This modifies neither the proportion of unsedimentable lysosomal enzyme in the homogenates nor the behaviour of lysosomes in centrifugation. Experiments performed with a lower dose of aFas (5 microg) indicate that unsedimentable lysosomal hydrolase activity increases in the homogenate after injection but with a marked delay with respect to the increase in DEVDase activity and in unsedimentable sulfite cytochrome c reductase. Comparative experiments ex vivo performed with Jurkat cells show an increase in unsedimentable lysosomal hydrolases, but much later than caspase 3 activation, and a release of dipeptidyl peptidase III and DEVDase into culture medium. It is proposed that the weakening of lysosomes observed after aFas treatment in vivo and ex vivo results from a necrotic process that takes place late after initiation of apoptosis.
Wattiaux
Robert
R
Laboratoire de Chimie Physiologique, URPhiM, FUNDP (Facultés Universitaires Notre-Dame de la Paix), 61 rue de Bruxelles, 5000 Namur, Belgium. robert.wattiaux@fundp.ac.be
Wattiaux-de Coninck
Simone
S
Thirion
Jacqueline
J
Gasingirwa
Mańe-Christine
MC
Jadot
Michel
M
D000818:Animals; D053148:Caspase 3 / Q000378:metabolism; D016923:Cell Death; D002458:Cell Fractionation; D005260:Female; D022781:Hepatocytes / Q000166:cytology / Q000502:physiology; D006801:Humans; D019169:Jurkat Cells; D008099:Liver / Q000166:cytology* / Q000502:physiology; D008247:Lysosomes / Q000502:physiology*; D051379:Mice; D008815:Mice, Inbred Strains; D019014:fas Receptor / Q000502:physiology*
D019014:fas Receptor; D053148:Caspase 3