Interactions between the bone matrix proteins osteopontin and bone sialoprotein and the osteoclast integrin alpha v beta 3 potentiate bone resorption

Methylmethacrylate (MMA) embedding of undecalcified bone is routinely employed for histomorphometric analyses. Although MMA-embedded bone has been used for immunolabeling at the light microscopic level after removal of the resin, there are no such reports for electron microscopy. The aim of the present study was to determine whether MMA embedding can be used for ultrastructural immunolabeling and how it compares to LR White (LRW), an acrylic resin frequently used for immunocytochemistry of bone. Rat tibiae were fixed by vascular perfusion with aldehyde and embedded either in MMA or LRW resin. Thin sections were processed for postembedding protein A-gold immunolabeling with antibodies to rat bone sialoprotein (BSP) and osteopontin (OPN). The density of gold particles over bone was quantified. The density and distribution of immunolabeling for BSP and OPN respectively, were comparable between MMA and LRW. These results indicate that MMA performs as well as LRW for the ultrastructural immunolabeling of noncollagenous bone matrix proteins.

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Peptidomimetic antagonists of alphavbeta3 inhibit bone resorption by inhibiting osteoclast bone resorptive activity, not osteoclast adhesion to bone

Journal of Endocrinology ◽

10.1677/joe.0.1650587 ◽

2000 ◽

Vol 165 (3) ◽

pp. 587-598 ◽

Cited By ~ 39

Author(s):

CP Carron ◽

DM Meyer ◽

VW Engleman ◽

JG Rico ◽

PG Ruminski ◽

...

Keyword(s):

Extracellular Matrix ◽

Bone Resorption ◽

Bone Matrix ◽

Matrix Proteins ◽

Integrin Receptors ◽

Or Efficiency ◽

Inhibit Bone Resorption ◽

Coated Surfaces ◽

Alpha V Beta 3 ◽

Resorptive Activity

Osteoclasts are actively motile on bone surfaces and undergo alternating cycles of migration and resorption. Osteoclast interaction with the extracellular matrix plays a key role in the osteoclast resorptive process and a substantial body of evidence suggests that integrin receptors are important in osteoclast function. These integrin receptors bind to the Arg-Gly-Asp (RGD) sequence found in a variety of extracellular matrix proteins and it is well established that the interaction of osteoclast alpha v beta 3 integrin with the RGD motif within bone matrix proteins is important in osteoclast-mediated bone resorption. In this study, we characterized the effects of two synthetic peptidomimetic antagonists of alpha v beta 3, SC-56631 and SC-65811, on rabbit osteoclast adhesion to purified matrix proteins and bone, and on bone resorption in vitro. SC-56631 and SC-65811 are potent inhibitors of vitronectin binding to purified alpha v beta 3. Both SC-56631 and SC-65811 inhibited osteoclast adhesion to osteopontin- and vitronectin-coated surfaces and time-lapse video microscopy showed that osteoclasts rapidly retract from osteopontin-coated surfaces when exposed to SC-56631 and SC-65811. SC-56631 and SC-65811 blocked osteoclast-mediated bone resorption in a dose-responsive manner. Further analysis showed that SC-65811 and SC-56631 reduced the number of resorption pits produced per osteoclast and the average pit size. SC-65811 was a more potent inhibitor of bone resorption and the combination of reduced pit number and size led to a 90% inhibition of bone resorption. Surprisingly, however, osteoclasts treated with SC-65811, SC-56631 or the disintegrin echistatin, at concentrations that inhibit bone resorption did not inhibit osteoclast adhesion to bone. These results suggest that alphavbeta3 antagonists inhibited bone resorption by decreasing osteoclast bone resorptive activity or efficiency but not by inhibiting osteoclast adhesion to bone per se.

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Gene expression for bone matrix proteins during bone resorption in calcium-deficient rats.

The Japanese Journal of Pharmacology ◽

10.1016/s0021-5198(19)50795-8 ◽

1994 ◽

Vol 64 ◽

pp. 290

Author(s):

Keiichi Ohya ◽

Maki Sumikawa ◽

Naoya Arai ◽

Masashi Tatematsu ◽

Hideaki Ogura

Keyword(s):

Gene Expression ◽

Bone Resorption ◽

Bone Matrix ◽

Matrix Proteins ◽

Bone Matrix Proteins

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Bone matrix proteins in osteogenesis and remodelling in the neonatal rat mandible as studied by immunolocalization of osteopontin, bone sialoprotein, α2HS-glycoprotein and alkaline phosphatase

Archives of Oral Biology ◽

10.1016/0003-9969(94)00144-z ◽

1995 ◽

Vol 40 (2) ◽

pp. 145-155 ◽

Cited By ~ 39

Author(s):

Gerald J. Pinero ◽

Mary C. Farach-Carson ◽

Robert E. Devoll ◽

Jane E. Aubin ◽

Jan C. Brunn ◽

...

Keyword(s):

Alkaline Phosphatase ◽

Bone Matrix ◽

Bone Sialoprotein ◽

Matrix Proteins ◽

Neonatal Rat ◽

Bone Matrix Proteins ◽

Rat Mandible

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Effect of protein restriction on the messenger RNA contents of bone-matrix proteins, insulin-like growth factors and insulin-like growth factor binding proteins in femur of ovariectomized rats

British Journal Of Nutrition ◽

10.1079/bjn19960188 ◽

1996 ◽

Vol 75 (6) ◽

pp. 811-823 ◽

Cited By ~ 10

Author(s):

Yusuke Higashi ◽

Asako Takenaka ◽

Shin-Ichiro Takahashi ◽

Tadashi Noguchi

Keyword(s):

Dietary Protein ◽

Type I Collagen ◽

Control Diet ◽

Bone Matrix ◽

Protein Restriction ◽

Matrix Proteins ◽

Type I ◽

Bone Matrix Proteins ◽

Mrna Content ◽

Insulin Like Growth Factors

It has been reported that loss of ovarian oestrogen after menopause or by ovariectomy causes osteoporosis. In order to elucidate the effect of dietary protein restriction on bone metabolism after ovariectomy, we fed ovariectomized young female rats on a casein-based diet (50g/kg diet (protein restriction) or 200g/kg diet (control)) for 3 weeks and measured mRNA contents of bone-matrix proteins such as osteocalcin, osteopontin and α1 type I collagen, insulin-like growth factors (IGF) and IGF-binding proteins (IGFBP) in femur. Ovariectomy decreased the weight of fat-free dry bone and increased urinary excretion of pyridinium cross-links significantly, although dietary protein restriction did not affect them. Neither ovariectomy nor protein restriction affected the content of mRNA of osteopontin and osteocalcin; however, ovariectomy increased and protein restriction extensively decreased the α1 type I collagen mRNA content in bone tissues. Ovariectomy increased IGF-I mRNA only in the rats fed on the control diet. Conversely, protein rest riction increased and ovariectomy decreased the IGF-II mRNA content in femur. Furthermore, the contents of IGFBP-2, IGFBP-4 and IGFBP-5 mRNA increased, but the content of IGFBP-3 mRNA decreased in femur of the rats fed on the protein-restricted diet. In particular, ovariectomy decreased the IGFBP-2 mRNA content in the protein-restricted rats and the IGFBP-6 mRNA content in the rats fed on the control diet. These results clearly show that the mRNA for some of the proteins which have been shown to be involved in bone formation are regulated by both quantity of dietary proteins and ovarian hormones.

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