Increased activities of cathepsin B and other lysosomal hydrolases in fibroblasts and bone tissue cultured in the presence of cysteine proteinases inhibitors.
Leupeptin is an established, reversible inhibitor of cathepsin B, a lysosomal cysteine proteinase. Yet, in rat fibroblasts as well as in foetal mouse calvaria, we observed an increase of the activity of cathepsin B in homogenates of cells and tissue harvested after culture in the presence of leupeptin. This effect was also seen for other lysosomal hydrolases, namely sphingomyelinase, N-acetyl-beta-glucosaminidase, arylsulphatase A and phospholipase A1 in fibroblasts, and beta-glucuronidase in mouse calvaria. In calvaria, antipain, another reversible cysteine proteinase inhibitor, caused a similar effect, whereas E-64, an irreversible inhibitor, was consistently inhibitory of the cathepsin B activity; yet it also caused an increase of beta-glucuronidase activity. The effect of leupeptin in fibroblasts was dose and time-dependent, required the continuous presence of the inhibitor, and was not dependent from protein synthesis. Actually, addition of cycloheximide caused a severe loss of activity of cathepsin B and of sphingomyelinase. In the presence of both cycloheximide and leupeptin, however, these two activities were retained to a value corresponding to that found in excess in cells cultivated with leupeptin alone. The data therefore suggests that leupeptin exerts the effects described in this paper by preventing the degradation of cathepsin B, sphingomyelinase and probably several other lysosomal hydrolases by cysteine proteinases. We therefore propose that cysteine proteinases play a key role in the control of the steady-state levels of these enzymes in normal conditions.
Montenez
J P
JP
Laboratoire de Chimie Physiologique (Groupe de Pharmacologie Cellulaire et Moléculaire, Université Catholique de Louvain, Brussels, Belgium.
Delaissé
J M
JM
Tulkens
P M
PM
Kishore
B K
BK
D000818:Animals; D001862:Bone Resorption; D002401:Cathepsin B / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D046508:Culture Techniques; D003513:Cycloheximide / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D006867:Hydrolases / Q000187:drug effects / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology*; D051379:Mice; D010281:Parathyroid Hormone / Q000037:antagonists & inhibitors / Q000494:pharmacology; D014176:Protein Biosynthesis; D011506:Proteins / Q000187:drug effects; D051381:Rats; D017208:Rats, Wistar; D012886:Skull / Q000187:drug effects* / Q000196:embryology; D013997:Time Factors
D007976:Leupeptins; D010281:Parathyroid Hormone; D011506:Proteins; D003513:Cycloheximide; D006867:Hydrolases; D002401:Cathepsin B; C032854:leupeptin