Pepstatin A, an Aspartic Proteinase Inhibitor, Suppresses RANKL-Induced Osteoclast Differentiation

• Published in: Journal of biochemistry (Tokyo) 2006-03, Vol.139 (3), p.583-590
• Main Authors: Yoshida, Hajime, Okamoto, Kuniaki, Iwamoto, Tsutomu, Sakai, Eiko, Kanaoka, Kazuhiro, Hu, Jin-Ping, Shibata, Mitsue, Hotokezaka, Hitoshi, Nishishita, Kazuhisa, Mizuno, Akio, Kato, Yuzo
• Format: Article
• Language: English
• Subjects: acid phosphatase; Animals; Animals, Newborn; Aspartic Acid Endopeptidases - antagonists & inhibitors; aspartic proteinase; cathepsin; Cell Differentiation - drug effects; ERK; extracellular signal-regulated kinase; inhibitor of NF-κB; IκB; JNK; Jun N-terminal kinase; M-CSF; macrophage colony–stimulating factor; Male; MAPK; Mice; mitogen-activated protein kinase; NFATc1; nuclear factor of activated T cells c1; OPG; osteoclast; Osteoclasts - cytology; Osteoclasts - drug effects; osteoprotegerin; pepstatin A; Pepstatins - pharmacology; phosphatidylinositol-3 kinase; PI3K; Protease Inhibitors - pharmacology; RANK; RANK Ligand - physiology; RANKL; receptor activator of NF-κB; receptor activator of NF-κB ligand; reverse transcription–polymerase chain reaction; RT-PCR; tartrate-resistant; TNF; TNF receptor-associated factor; TRAF; TRAP; tumor necrosis factor
• ISSN: 0021-924X; 1756-2651
• DOI: 10.1093/jb/mvj066

Pepstatin A is well known to be an inhibitor of aspartic proteinases such as pepsin, cathepsins D and E. Except for its role as a proteinase inhibitor, however, the pharmacological action of pepstatin A upon cells remain unclear. In this study, we found that pepstatin A suppressed receptor activator of NF-[kappa]B ligand (RANKL)-induced osteoclast differentiation. Pepstatin A suppressed the formation of multinuclear osteoclasts dose-dependently. This inhibition of the formation only affected osteoclast cells, i.e., not osteoblast-like cells. Furthermore, pepstatin A also suppressed differentiation from pre-osteoclast cells to mononuclear osteoclast cells dose-dependently. This inhibition seems to be independent of the activities of proteinases such as cathepsin D, because the formation of osteoclasts was not suppressed with the concentration that inhibited the activity of cathepsin D. Cell signaling analysis indicated that the phosphorylation of ERK was inhibited in pepstatin A-treated cells, while the phosphorylation of I[kappa]B and Akt showed almost no change. Furthermore, pepstatin A decreased the expression of nuclear factor of activated T cells c1 (NFATc1). These results suggest that pepstatin A suppresses the differentiation of osteoclasts through the blockade of ERK signaling and the inhibition of NFATc1 expression.