A Cysteine Protease Inhibitor Prevents Suspension-Induced Declines in Bone Weight and Strength in Rats.

Abstract

In this study, we examined the effects of a potent cysteine protease inhibitor, N-(L-3-transcarboxyoxirane-2-cabonyl)-L-leucine-4-aminobutylamide (E-64a), on bone weight and strength in tail-suspended rats. We first administered a vehicle or 4 or 8 mg/rat of E64a to rats fed with a low calcium diet for 7 wks to determine effective doses of E-64a on bone resorption in vivo. Femoral cathepsin K-like activity and serum hydroxyproline level in rats fed with a low calcium diet were significantly higher than those in rats fed with a standard diet. The intraperitoneal injection of 8 mg/rat of E-64a to rats decreased their serum calcium and hydroxyproline concentrations after 3 to 6 hrs in parallel with changes in femoral cathepsin K-like activity, while 4 mg/rat of E-64a had weaker effects on these parameters. Based on these results, we injected 8 mg/rat of E-64a to tail-suspended rats twice a day for 2 wks and compared the results with those of treatment with 1 mg/rat of etidronate, a bisphosphonate, twice a week. In tailsuspended rats, femoral weight and strength, assessed by three-point bending test, significantly decreased from Day 5 to 21, while femoral cathepsin K-like activity and serum calcium and hydroxyproline concentrations did not change. E-64a inhibited femoral cathepsin K-like activity in tail-suspended rats, but etidronate did not. E64a as well as etidronate significantly prevented the suspension-induced declines in bone weight and strength. However, more frequent injection and higher doses were required for E-64a to exhibit significant efficacy of antiresorption, compared with those of etidronate. Our results suggest that a cysteine protease inhibitor could improve suspension-induced osteopenia by inhibiting cathepsin K-like activity in bone; however, it needs several improvements in the effect as a clinical drug.

Mice Lacking Cathepsin K Maintain Bone Remodeling but Develop Bone Fragility Despite High Bone Mass

“…We observed the hallmark osteopetrosis seen in the pyknodysostosis patients (8,9) and other CatK-deficient mice (5)(6)(7) and mimicked by cysteine proteinase inhibitors like E-64. (19) We also found CatK-deficient mice to have shallow resorption lacunae containing a layer of demineralized matrix like that observed in vivo (6) and recapitulated by CatK-deficient osteoclasts in vitro. (5) However, in the face of these indicators of diminished bone resorption, we also found elevations in both osteoclast number and surface in cancellous bone (confirming observations of Kiviranta et al) (7) and an unexpected dramatic increase in intracortical resorption.…”

“…The CatK‐deficient mice in this study exhibited both expected and unusual features in their long bones. We observed the hallmark osteopetrosis seen in the pyknodysostosis patients (8,9) and other CatK‐deficient mice (5–7) and mimicked by cysteine proteinase inhibitors like E‐64 (19) . We also found CatK‐deficient mice to have shallow resorption lacunae containing a layer of demineralized matrix like that observed in vivo (6) and recapitulated by CatK‐deficient osteoclasts in vitro (5) .…”

Immunomodulatory activity of a chymotrypsin inhibitor from Momordica cochinchinensis seeds

“…Several protease inhibitors have been shown to act on bone marrow cells. For example, specific cysteine protease inhibitors could prevent bone metastasis [23]. Different metalloproteinases were found in murine cartilages and the tissue metalloproteinase inhibitors could be used to prevent the degradation of cartilages [19].…”