Muscle aspartyl protease (cathepsin D) activity: detection using a chromophoric substrate and relation to wasting in DBA/2 mice implanted with leukemic L1210 tumor cells.
The relationship between skeletal muscle aspartyl protease activity (APA) and wasting was investigated in male DBA/2 mice inoculated with L1210 tumor cells. Using the peptidic substrate H-Pro-Thr-Glu-Phe-Phe(NO2)-Arg-Leu-OH, which is specific for aspartyl proteases, proteases, proteolytic activity was detected in a number of tissues including muscle by using a crude extraction procedure for isolation of lysosomal enzymes. Biochemical characterization and increased muscle levels following either fasting or injection of endotoxin (ETX) suggest that the enzyme is likely cathepsin D. The wasting syndrome accompanying the tumor was measured by comparing the weight of the skinned hind limb in treated and control animals. DBA/2 mice inoculated intraperitoneally with L1210 cells developed multiple solid tumors in the peritoneum and ascites; maximal tumor burden was reached by 16 days. There was a significant reduction in hind limb weight (16 +/- 2%; mean +/- SE) and significant increase (31 +/- 8%) in muscle APA associated with the development of ascites and solid tumors. Plasma APA activity was substantially increased (240 +/- 33%), while liver and spleen APA were increased (10-20%) but not significantly. Chronic pepstatin administration, 30 mg.kg-1.day-1, for 7 days concurrent with the initiation of observable ascites and solid tumor formation (7 days post-inoculation), completely inhibited hind limb weight loss and alleviated the tumor-dependent increase of APA in both plasma and muscle without altering tumor development. Delaying the administration of pepstatin by 3 days resulted in less of an inhibition (33 +/- 13%) of hind limb weight loss. Thus, cathepsin D or a similar aspartyl protease appears to be of key importance in the wasting syndrome associated with cachexia.
Bolger
G T
GT
Department of Pharmacology, Bio-Méga/Boehringer Ingelheim Research Inc., Laval, QC, Canada.
Jaramillo
J
J
D000818:Animals; D002100:Cachexia / Q000473:pathology; D002402:Cathepsin D / Q000378:metabolism*; D004731:Endotoxins / Q000633:toxicity; D004926:Escherichia coli; D006614:Hindlimb / Q000473:pathology; D007939:Leukemia L1210 / Q000473:pathology*; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D051379:Mice; D008811:Mice, Inbred DBA; D018482:Muscle, Skeletal / Q000201:enzymology*; D009368:Neoplasm Transplantation; D009929:Organ Size / Q000187:drug effects / Q000502:physiology; D010436:Pepstatins / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology
D004731:Endotoxins; D008070:Lipopolysaccharides; D010436:Pepstatins; D011480:Protease Inhibitors; C014195:Streptomyces pepsin inhibitor; C044101:endotoxin, Escherichia coli; D002402:Cathepsin D; C031375:pepstatin