A peptidyl derivative structurally based on the inhibitory center of cystatin C inhibits bone resorption in vitro

Citation
L. Johansson et al., A peptidyl derivative structurally based on the inhibitory center of cystatin C inhibits bone resorption in vitro, BONE, 26(5), 2000, pp. 451-459
Citations number
48
Categorie Soggetti
Endocrynology, Metabolism & Nutrition","da verificare
Journal title
BONE
ISSN journal
87563282 → ACNP
Volume
26
Issue
5
Year of publication
2000
Pages
451 - 459
Database
ISI
SICI code
8756-3282(200005)26:5<451:APDSBO>2.0.ZU;2-R
Abstract
Human cystatin C is a cysteine proteinase inhibitor belonging to the cystat in superfamily, which previously has been shown to inhibit bone resorption in bone organ culture, The aminoterminal segment, Arg(8)-Leu(9)-Va(10)-Gly( 11) (RLVG), of the single polypeptide chain of cystatin C constitutes an es sential part of its inhibitory center, In the present study, the effect of benzyloxycarbonyl-Arg(8)-Leu(9)-Val(10)-Gly(11)-diazomethane (Z- RLVG-CHN2) on bone resorption in vitro was compared with the effects of cystatin C an d calcitonin, Bone resorption was assessed by the release of Ca-45 and H-3 from mouse calvarial bones prelabeled with [Ca-45]CaCl2 and [H-3]-proline, respectively. Z-RLVG-CHN2 concentration-dependently inhibited the release o f Ca-45 and H-3 in bones stimulated by parathyroid hormone (PTH), with half -maximal inhibition obtained at 1 mu mol/L. The inhibitory actions of Z-RLV G-CHN2 and cystatin C were persistent, whereas action induced initially by calcitonin was lost with time. The inhibition caused by Z-RLVG-CHN2 and cys tatin C on PTH-stimulated Ca-45 release was observed after 6 h, whereas inh ibition by calcitonin was seen already after 2 h, In contrast, the inhibito ry effects of Z-RLVG-CHN2 and cystatin C, as well as that of calcitonin, on H-3 release was seen already after 2 h, Z-RLVG-CHN2, in which the reactive carboxyterminal diazomethane was substituted by nonreactive groups [-OH, - NH2, or -N(CH3)(2)], resulted in peptidyl derivatives, which, in contrast t o Z-RLVG-CHN2 and cystatin C, inhibited neither cysteine proteinases nor bo ne resorption, In contrast to wild-type cystatin C, recombinant human cysta tin C with Gly substitutions for residues Arg(8), Leu(9), Val(10), and Trp( 106), and with low or nonexistent affinity for cysteine proteinases, did no t display any inhibitory effect on bone resorption, These data strongly ind icate that Z-RLVG-CHN2 inhibits bone resorption in vitro by a mechanism tha t seems primarily to be due to an inhibition of bone matrix degradation via cysteine proteinases. The data also corroborate the hypothesis that cystat in C inhibits bone resorption by virtue of its cysteine proteinase inhibito ry capacity. (C) 2000 by Elsevier Science Inc, All rights reserved.