Elsevier

Bone

Volume 43, Issue 2, August 2008, Pages 264-273
Bone

Accentuated osteoclastic response to parathyroid hormone undermines bone mass acquisition in osteonectin-null mice

https://doi.org/10.1016/j.bone.2008.03.024Get rights and content

Abstract

Matricellular proteins play a unique role in the skeleton as regulators of bone remodeling, and the matricellular protein osteonectin (SPARC, BM-40) is the most abundant non-collagenous protein in bone. In the absence of osteonectin, mice develop progressive low turnover osteopenia, particularly affecting trabecular bone. Polymorphisms in a regulatory region of the osteonectin gene are associated with bone mass in a subset of idiopathic osteoporosis patients, and these polymorphisms likely regulate osteonectin expression. Thus it is important to determine how osteonectin gene dosage affects skeletal function. Moreover, intermittent administration of parathyroid hormone (PTH) (1–34) is the only anabolic therapy approved for the treatment of osteoporosis, and it is critical to understand how modulators of bone remodeling, such as osteonectin, affect skeletal response to anabolic agents. In this study, 10 week old female wild type, osteonectin-haploinsufficient, and osteonectin-null mice (C57Bl/6 genetic background) were given 80 μg/kg body weight/day PTH(1–34) for 4 weeks. Osteonectin gene dosage had a profound effect on bone microarchitecture. The connectivity density of trabecular bone in osteonectin-haploinsufficient mice was substantially decreased compared with that of wild type mice, suggesting compromised mechanical properties. Whereas mice of each genotype had a similar osteoblastic response to PTH treatment, the osteoclastic response was accentuated in osteonectin-haploinsufficient and osteonectin-null mice. Eroded surface and osteoclast number were significantly higher in PTH-treated osteonectin-null mice, as was endosteal area. In vitro studies confirmed that PTH induced the formation of more osteoclast-like cells in marrow from osteonectin-null mice compared with wild type. PTH treated osteonectin-null bone marrow cells expressed more RANKL mRNA compared with wild type. However, the ratio of RANKL:OPG mRNA was somewhat lower in PTH treated osteonectin-null cultures. Increased expression of RANKL in response to PTH could contribute to the accentuated osteoclastic response in osteonectin−/− mice, but other mechanisms are also likely to be involved. The molecular mechanisms by which PTH elicits bone anabolic vs. bone catabolic effects remain poorly understood. Our results imply that osteonectin levels may play a role in modulating the balance of bone formation and resorption in response to PTH.

Introduction

The “matricellular” protein family consists of extracellular matrix glycoproteins that play a predominant role in the modulation of cell–matrix interactions, as well as contributing to matrix organization. These proteins are expressed at high levels in development and in response to injury, generally induce cell de-adhesion, and interact with cell surface receptors, extracellular matrix, growth factors, cytokines and proteases [1]. Recent studies demonstrate that matricellular proteins play a unique role in the skeleton as regulators of bone remodeling. Matricellular proteins important in bone include osteopontin, bone sialoprotein, tenascin C, thrombospondins 1 and 2, and osteonectin or SPARC (secreted protein acidic and rich in cysteine; BM-40). These proteins interact with the extracellular matrix, regulating bone matrix deposition, assembly, and mineralization. Selected matricellular proteins have been shown to interact with growth factors, cytokines and proteases, modulating the binding of growth factors to receptors and promoting the activation of metalloproteinases (reviewed in [2], [3]). Further, the interaction of matricellular proteins with cell surface molecules, including integrins and their associated proteins, modulates signaling from down stream effectors [2], [4], [5]. The process of bone remodeling allows the skeleton to respond to mechanical and physiologic stresses, and matricellular proteins play an important role in skeletal remodeling induced by fracture repair, unloading, and estrogen depletion [2].

Although widely expressed in mammalian tissues, the matricellular protein osteonectin is the most abundant noncollagenous protein in bone [3], [6]. In the absence of osteonectin, mice develop progressive low turnover osteopenia. Trabecular bone volume is dramatically decreased, while cortical bone has compromised matrix quality [7], [8]. In vivo, osteonectin-null mice have decreased osteoblast and osteoclast numbers and surface, as well as decreased bone formation rate. Further, osteonectin suppresses adipogenesis, and in vitro studies indicate that osteonectin promotes osteoblastic commitment, differentiation, and survival [3], [9], [10].

We found that marrow stromal cells from wild type and osteonectin-null mice were not different in their ability to produce cAMP in response to parathyroid hormone (PTH) in vitro, however this response to PTH was attenuated in cultured osteonectin-null osteoblastic cells [10]. Nonetheless, the role of osteonectin in PTH-stimulated bone remodeling in vivo remains unknown. Since intermittent administration of the amino-terminal fragment of PTH(1–34) is the only anabolic therapy approved for the treatment of osteoporosis, it is critical to understand how modulators of bone remodeling, such as osteonectin, may affect skeletal response to anabolic agents [11].

Our previous studies of osteonectin-null mice showed that osteonectin is critical for normal bone remodeling. However, the impact of osteonectin-haploinsufficiency on skeletal biology has not yet been determined. It is known that osteonectin expression is decreased in osteoblasts from patients with osteogenesis imperfecta [12], [13]. Further, haplotypes consisting of 3 single nucleotide polymorphisms (SNPs) in the 3′ untranslated region (UTR) of the osteonectin gene have been associated with bone mass in a subset of idiopathic osteoporosis patients [14]. The 3′ UTR regulates mRNA stability, trafficking, and translation, and it is likely that these 3′ UTR SNPs differentially regulate osteonectin expression [15], [16]. Therefore, it is important to determine how osteonectin gene dosage affects skeletal function. In this study, we characterized the response of wild type (+/+), osteonectin-haploinsufficient (+/−) and osteonectin-null (−/−) mice to intermittent treatment with PTH(1–34). The goal of this study was to determine the impact of osteonectin gene dosage on the response of the skeleton to an anabolic therapy.

Section snippets

Animals

The osteonectin−/− mice used in these studies were back crossed 8 times into the C57/Bl6 genetic background [10]. Wild type mice of similar genetic background were also maintained. Osteonectin+/− mice were generated by crossing wild type and osteonectin−/− animals. Mice were maintained under standard non-barrier conditions and had access to mouse chow and water ad libitum. Female mice received intraperitoneal injections of calcien (10 mg/kg) and demeclocycline (30 mg/kg) 10 and 3 days prior to

BMD and BMC

At 10 weeks of age (i.e. base line), osteonectin−/− mice had significantly lower whole body BMD compared with osteonectin+/− or wild type mice (44.2 ± 0.3 vs. 45.9 ± 0.4 and 46.8 ± 0.4 mg/cm2, respectively; p  0.01). Wild type and osteonectin+/− mice treated with PTH for 4 weeks had a 13% increase in whole body BMD, whereas PTH treated osteonectin−/− mice had whole body BMD values similar to those observed in vehicle treated mice (Fig. 1A). However, significant increases in bone mineral content (BMC)

Discussion

This study provides 3 novel insights into the function of the matricellular protein osteonectin in the skeleton. First, we find that although young (i.e. 10–14 week old) osteonectin+/− mice have trabecular bone volume and cortical bone parameters similar to that of wild type mice, osteonectin gene dosage has a profound effect on bone microarchitecture. The connectivity density of trabecular bone in the vertebrae and femur of osteonectin+/− mice is substantially decreased compared with that of

Acknowledgments

We thank Dr B. Kream (University of Connecticut Health Center) and Dr. R.C. Pereira (University of California, Los Angeles) for thoughtful discussions of the data. We thank Dr. C Pilbeam (University of Connecticut Health Center) for assistance with the qRT-PCR analysis.

References (41)

  • P.G. Robey et al.

    Extracellular matrix and biomineraliztion of bone

  • C. Kessler et al.

    Increased notch 1 expression and attenuated stimulatory G protein coupling to adenylyl cyclase in osteonectin-null osteoblasts

    Endocrinology

    (2007)
  • A.D. Bradshaw et al.

    SPARC, a matricellular protein that functions in cellular differentiation and tissue response to injury

    J Clin Invest

    (2001)
  • A.M. Delany et al.

    Osteopenia and decreased bone formation in osteonectin-null mice

    J Clin Invest

    (2000)
  • A.L. Boskey et al.

    Infrared analysis of the mineral and matrix in bones of osteonectin-null mice and their wild type controls

    J Bone Miner Res

    (2003)
  • A.D. Bradshaw et al.

    SPARC-null mice exhibit increased adiposity without significant differences in overall body weight

    Proc Natl Acad Sci U S A

    (2003)
  • A.M. Delany et al.

    Osteonectin-null mutation compromises osteoblast formation, maturation, and survival

    Endocrinology

    (2003)
  • E. Canalis et al.

    Mechanisms of anabolic therapies for osteoporosis

    N Engl J Med

    (2007)
  • N.S. Fedarko et al.

    Extracellular matrix stoichiometry in osteoblasts from patients with osteogenesis imperfecta

    J Bone Miner Res

    (1995)
  • N.S. Fedarko et al.

    Extracellular matrix formation by osteoblasts from patients with osteogenesis imperfecta

    J Bone Miner Res

    (1992)
  • Cited by (0)

    This work was supported by the National Institute of Arthritis and Musculoskeletal and Skin Diseases, AR44877 (A. Delany).

    View full text