RANK Ligand and Osteoprotegerin
Paracrine Regulators of Bone Metabolism and Vascular Function
In 1997, investigators isolated a secreted glycoprotein that blocked osteoclast differentiation from precursor cells, prevented osteoporosis (decreased bone mass) when administered to ovariectomized rats, and resulted in osteopetrosis (increased bone mass) when overexpressed in transgenic mice. Since then, the isolation and characterization of the protein named osteoprotegerin (OPG) has stimulated much work in the fields of endocrinology, rheumatology, and immunology. OPG functions as a soluble decoy receptor for receptor activator of nuclear factor-κB ligand (RANKL, or OPG ligand) and shares homologies with other members of the tumor necrosis factor receptor superfamily. OPG acts by competing with the receptor activator of nuclear factor-κB, which is expressed on osteoclasts and dendritic cells for specifically binding to RANKL. RANKL is crucially involved in osteoclast functions and bone remodeling as well as immune cell cross-talks, dendritic cell survival, and lymph node organogenesis. More recently, emerging evidence from in vitro studies and mouse genetics attributed OPG an important role in vascular biology. In fact, OPG could represent the long sought-after molecular link between arterial calcification and bone resorption, which underlies the clinical coincidence of vascular disease and osteoporosis, which are most prevalent in postmenopausal women and elderly people.
Osteoprotegerin (OPG) was isolated independently by two laboratories,1,2⇓ and synonyms such as osteoclastogenesis inhibitory factor (OCIF),3 TNF receptor-related molecule-1 (TR1), 4 or follicular dendritic cell–derived receptor-1 (FDCR-1)5 have been coined. According to the American Society for Bone and Mineral Research Committee, the term osteoprotegerin (OPG) is now being recommended.6 The mouse7 and the human8 OPG genes have been cloned and characterized, and the human OPG gene represents a single-copy gene that contains 5 exons and spans 29 kb of the human genome located on chromosome 8.9 Murine OPG gene expression starts between days 8 and 9 during embryogenesis.7 Of note, the human OPG promoter sequence harbors binding elements for the osteoblast-specific transcription factor cbfa-1, which was found to increase OPG gene transcription.10
OPG is a member of the tumor necrosis factor receptor (TNFR) superfamily, and it represents a secretory basic glycoprotein that exists in a 60-kd monomeric form and a disulfide-linked homodimeric form of 120 kd.11 It has also been detected in a cell surface–associated form with some cell types,5 although sequence analysis failed to detect a classical hydrophobic transmembrane domain, which is typical for all other members of the TNFR superfamily.11 The molecule is composed of 401 amino acid residues as deduced from cDNA nucleotide sequencing with a signal peptide of 21 amino acids.3 OPG consists of 7 structural domains, of which the amino-terminal cysteine-rich domains 1 to 4 share some features with the extracellular domains of other members of the TNFR family (Figure 1).12
Mutation analyses have been used for functional characterization of OPG. Domains 1 to 4 are sufficient for conferring osteoclastogenesis inhibitory activity, which can be demonstrated by carboxy-terminal truncation mutants.11 Of note, monomeric and dimeric OPG were indistinguishable in their specific activity to inhibit osteoclastogenesis.13 The carboxy-terminal portion of the protein with domains 5 and 6 contains two death domain homologous regions, motifs that are found in the cytoplasmic region of mediators of apoptosis such as TNFR 1, DR3, CD95/Fas, or TNF-related apoptosis-inducing ligand (TRAIL) receptors.14–16⇓⇓ In fact, domains 5 and 6 of OPG have been demonstrated to transduce an apoptotic signal when expressed as an OPG/Fas fusion protein in which the transmembrane region of Fas is inserted between domains 4 and 5 of OPG.11 However, death domain-containing members of the TNFR family are also able to stimulate alternative signaling pathways, thus preventing rather than triggering apoptosis.17 Finally, domain 7 harbors a heparin-binding region, a common feature of peptide growth factors and signal molecules,18–20⇓⇓ as well as an unpaired cysteine residue required for disulfide bond formation and dimerization (Figure 1).11,13⇓
Expression and Regulation of OPG
OPG is produced by a variety of tissues including the cardiovascular system (heart, arteries, veins), lung, kidney, intestine, and bone, as well as hematopoietic and immune cells.1,5,21⇓⇓ The expression and production of the protein is modulated by various cytokines, peptides, hormones, and drugs. Cytokines, including TNF-α, interleukin (IL)-1α, IL-18, transforming growth factor (TGF)-β, bone morphogenetic proteins, and steroid hormones such as 17β-estradiol are known to up-regulate OPG mRNA levels.22–29⇓⇓⇓⇓⇓⇓⇓ In contrast, glucocorticoids (known to promote bone resorption) and the immunosuppressant cyclosporine A (which has the propensity to cause osteoporosis and vascular disease), parathyroid hormone (PTH), prostaglandin E2, and basic fibroblast growth factor all suppress the expression of OPG.30–35⇓⇓⇓⇓⇓ Moreover, tensional force applied to bone surface is followed by enhanced OPG mRNA synthesis,36 whereas expression of OPG by bone marrow cells declines with aging,37 thus implicating OPG as a potential mediator of immobilization and senile osteoporosis.
Role of RANKL and OPG in Bone Metabolism
The RANKL gene encodes a protein of 316 amino acids with a molecular mass of 38 kd, of which the extracellular domains self-associate as a trimer. Its expression is also modulated by various cytokines (IL-1, IL-6, IL-11, TNF-α), glucocorticoids, and PTH.38 RANKL is produced by osteoblastic lineage cells and activated T cells and promotes osteoclast formation, fusion, differentiation, activation, and survival, leading to enhanced bone resorption and bone loss.39,40⇓ Except for a primary secreted form produced by T cells and some cancer cell lines, RANKL exists either in a cell-bound form or a truncated ectodomain variant derived from enzymatic cleavage of the cellular form by a TNF-α–converting enzyme-like protease (TACE) (Figure 2).41 RANKL stimulates its specific receptor RANK, which is expressed by a restricted number of cell types, including progenitor and mature osteoclasts, activated T cells, and myeloid-derived dendritic cells (DCs) (Figure 2).42–46⇓⇓⇓⇓ RANK activation by RANKL initiates intracellular signaling cascades that involve c-Jun, NF-κB, and serine/threonine kinase Akt/PKB pathways.47
The biological effects of OPG are opposite of the RANKL-mediated effects, because OPG acts as a soluble inhibitor that prevents RANKL interaction and subsequent stimulation with its receptor, RANK (Figure 2).48 Therefore, mice with excessive or defective production of RANKL, RANK, or OPG display both extremes of skeletal phenotypes, ie, osteoporosis (OPG knockout) and osteopetrosis (OPG transgenic, RANKL knockout, RANK knockout).1,49–51⇓⇓⇓ In conclusion, RANKL, RANK, and OPG represent a novel cytokine network and act as key regulators of bone metabolism and osteoclast biology (for review, see Suda et al52 and Teitelbaum53).
OPG and the Immune System
A number of studies have highlighted the involvement of OPG and its cognate ligand, RANKL in immune responses. Binding of RANKL to RANK augments DC survival via Bcl-xL induction, enhances the immunostimulatory capacity of DCs, and modulates activated T cells.54–58⇓⇓⇓⇓ Thus, in addition to its osteotropic effects, RANKL exerts important immunomodulatory functions, as evident from the phenotype of RANKL-deficient mice, which display lymph node agenesis as well as impaired splenic structures and Peyer’s patches.51 Interestingly, deletion of the RANK gene leads to the exact phenocopy of RANKL-deficient mice,59 thus establishing the importance of the RANKL-RANK pathway in immune regulation. By studying OPG-deficient mice, OPG was found to be critically involved in B cell maturation and the generation of efficient antibody responses.60 Furthermore, bone marrow–derived DCs are more potent in stimulating allogeneic T cells in OPG-deficient mice as compared with DCs from wild-type mice.60 Of note, ligation of CD40 up-regulates OPG expression by B cells and DCs.5
An important aspect of OPG function in the immune system is related to the cytotoxic ligand TRAIL, a potent activator of apoptosis of susceptible cells following binding to death domain–containing receptors.61 OPG is able to bind TRAIL and thereby inhibits TRAIL-induced apoptosis of cells.62 Vice versa, TRAIL can block the inhibitory activity of OPG on osteoclastogenesis.62
RANKL and OPG as Potential Mediators of Arterial Calcification
The first evidence for an involvement of OPG in arterial calcification was derived from OPG knockout mice, which displayed osteoporosis and arterial calcification of the media of the aorta and the renal arteries.49 In contrast, delivery of transgenic OPG from mid-gestation through adulthood could rescue OPG-deficient mice from development of mineral deposits in the vascular system.63 Interestingly, affected vessel sites in the animal model resemble those in patients with arterial calcifications.64 OPG expression can be demonstrated in the media of great arteries,1 and different vascular cell types such as coronary smooth muscle cells32 and endothelial cells65 have been implicated as cellular sources and targets of vascular OPG production (Figure 3). In endothelial cells, OPG has been demonstrated to act as an autocrine survival factor.65 In contrast, RANKL and RANK transcripts could only be demonstrated in calcified arterial lesions of OPG-deficient mice but not in wild-type mice.63 In addition, RANKL and RANK have not been shown to be directly involved in human vascular diseases. Moreover, the cellular basis and the molecular mechanisms for the vascular effects of OPG are elusive. The study by Min and co-workers63 reported multinucleated osteoclast-like cells in the calcified vascular lesions of OPG-deficient mice with the concomitant detection of RANK transcripts, which are indicative of the osteoclastic and the DC lineage.42,44⇓
The hypothesis that the RANKL/OPG system could link osteoporosis and arterial calcification is underlined by the high clinical prevalence and coincidence of arterial calcification and cardiovascular disease in postmenopausal women and elderly people with o steoporosis.66–68⇓⇓ Interestingly, a recent study in elderly women found a significant correlation of elevated OPG serum levels and cardiovascular mortality.69 Similarly, an earlier study detected increased serum concentrations of OPG in osteoporotic and postmenopausal women as compared with age-matched women without osteoporosis, and OPG levels were highest in those with the highest bone turnover and the most severe osteoporosis.70 The seeming paradox of increased serum levels of OPG in patients with active osteoporosis and vascular disease has been interpreted as an incomplete regulatory mechanism to counteract disease progression.
Finally, OPG (used in concentrations known to block bone resorption) was found to inhibit warfarin- and vitamin D–induced vascular calcification in rats in vivo,71 which was similar to the effects of bisphosphonates, another established drug class known to inhibit osteoclastic bone resorption and bone loss (Figure 3).72
The novel TNF superfamily members RANKL and OPG are essential paracrine mediators of bone metabolism and immune functions and have been clearly implicated in various skeletal and immune disorders and diseases at the interface between bone metabolism and the immune system such as rheumatoid arthritis. In the vascular system, OPG is produced by smooth muscle and endothelial cells in vitro and acts as a survival factor for endothelial cells. One recent study demonstrated RANKL and OPG immunoreactivity in the nondiseased vessel wall and in early atherosclerotic lesions in human tissues, whereas in advanced calcified lesions, only RANKL was detected in the extracellular matrix surrounding calcium deposits.73 Moreover, systemic administration of OPG has been found to prevent vitamin D–induced vascular calcification in rodents. Finally, a recent clinical study has demonstrated that OPG administration is a safe and effective therapy in reducing biochemical markers of bone turnover in postmenopausal women.74 In summary, emerging evidence indicates that OPG is not merely a protective factor for bone, but may, in fact, act as a protective factor for the vascular system. Further studies are required to assess the contribution of RANKL and OPG in vascular disease, to analyze their role as biochemical markers of vascular diseases, and to evaluate the potential of OPG as a new “vasculoprotegrin” in therapeutic studies.
This work was supported by grants Ho 1875/2-1 (Deutsche Forschungsgemeinschaft) and 10-1697-Ho 1 (Deutsche Krebshilfe), Bonn, Germany to L.C. Hofbauer.
Received September 5, 2001; revision accepted January 23, 2002.
- ↵Simonet WS, Lacey DL, Dunstan CR, Kelley M, Chang M-S, Lüthy R, Nguyen HQ, Wooden S, Bennett L, Boone T, Shimamoto G, DeRose M, Eliott R, Colombero A, Tan H-L, Trail G, Sullivan J, Davy E, Bucay N, Renshaw-Gegg L, Hughes TM, Hill D, Pattison W, Campbell P, Sander S, Van G, Tarpley J, Derby P, Lee R, Amgen EST Program, Boyle WJ. Osteoprotegerin: a novel secreted protein involved in the regulation of bone density. Cell. 1997; 89: 159–161.
- ↵Yasuda H, Shima N, Nakagawa N, Mochizuki SI, Yano K, Fujise N, Sato Y, Goto M, Yamaguchi K, Kuriyama M, Kanno T, Murakami A, Tsuda E, Morinaga T, Higashio K. Identity of osteoclastogenesis inhibitory factor (OCIF) and osteoprotegerin (OPG): a mechanism by which OPG/OCIF inhibits osteoclastogenesis in vitro. Endocrinology. 1998; 139: 1329–1337.
- ↵Kwon BS, Wang S, Udagawa N, Haridas V, Lee ZH, Kim KK, Oh KO, Greene J, Li Y, Su J, Gentz R, Aggarwal BB, Ni J. TR1, a new member of the tumor necrosis factor receptor family, induces fibroblast proliferation and inhibits osteoclastogenesis and bone resorption. FASEB J. 1998; 12: 845–854.
- ↵Yun TJ, Chaudhary PM, Shu GL, Frazer JK, Ewings MK, Schwartz SM, Pascual V, Hood LE, Clark EA. OPG/FDCR-1, a TNF receptor family member, is expressed in lymphoid cells and is up-regulated by ligating CD40. J Immunol. 1998; 161: 6113–6121.
- ↵Thirunavukkarasu K, Halladay DL, Miles RR, Yang X, Galvin RJ, Chandrasekhar S, Martin TJ, Onyia JE. The osteoblast-specific transcription factor Cbfa1 contributes to the expression of osteoprotegerin, a potent inhibitor of osteoclast differentiation and function. J Biol Chem. 2000; 275: 25163–25172.
- ↵Yamaguchi K, Kinosaki M, Goto M, Kobayashi F, Tsuda E, Morinaga T, Higashio K. Characterization of structural domains of human osteoclastogenesis inhibitory factor. J Biol Chem. 1998; 273: 5117–5123.
- ↵Itoh N, Nagata S. A novel protein domain required for apoptosis: mutational analysis of human Fas antigen. J Biol Chem. 1993; 268: 10932–10937.
- ↵Chinnaiyan AM, O’Rourke K, Yu GL, Lyons RH, Garg M, Duan DL, Xing L, Gentz R, Ni J, Dixit VM. Signal transduction by DR3, a death domain–containing receptor related to TNFR-1 and CD95. Science. 1996; 274: 990–992.
- ↵Walczak H, Degli-Esposti MA, Johnson RS, Smolak PJ, Waugh JY, Boiani N, Timour MS, Gerhart MJ, Schooley KA, Smith CA, Goodwin RG, Rauch CT. TRAIL-R2: a novel apoptosis-mediating receptor for TRAIL. EMBO J. 1997; 16: 5386–5397.
- ↵Lee SY, Kaufmann DR, Mora AL, Santana A, Boothby M, Choi Y. Stimulus-dependent synergism of the antiapoptotic tumor necrosis factor receptor-associated factor 2 (TRAF2) and nuclear factor-κB pathways. J Exp Med. 1998; 188: 1381–1384.
- ↵Leung DW, Cachianes G, Kuang WJ, Goeddel DV, Ferrara N. Vascular endothelial growth factor is a secreted angiogenic mitogen. Science. 1989; 246: 1306–1309.
- ↵Eriksson AE, Cousens LS, Weaver LH, Matthews BW. Three-dimensional structure of human basic fibroblast growth factor. Proc Natl Acad Sci U S A. 1991; 88: 3441–3445.
- ↵Zhang JD, Cousens LS, Barr PJ, Sprang SR. Three-dimensional structure of human basic fibroblast growth factor, a structural homolog of interleukin 1β. Proc Natl Acad Sci U S A. 1991; 88: 3446–3450.
- ↵Tan KB, Harrop J, Reddy M, Young P, Terrett J, Emery J, Moore G, Truneh A. Characterization of a novel TNF-like ligand and recently described TNF ligand and TNF receptor superfamily genes and their constitutive and inducible expression in hematopoietic and non-hematopoietic cells. Gene. 1997; 204: 35–46.
- ↵Takai H, Kanematsu M, Yano K, Tsuda E, Higashio K, Ikeda K, Watanabe K, Yamada Y. Transforming growth factor-β stimulates the production of osteoprotegerin/osteoclastogenesis inhibitory factor by bone marrow stromal cells. J Biol Chem. 1998; 273: 27091–27096.
- ↵Wan M, Shi X, Feng X, Cao X. Transcriptional mechanisms of bone morphogenetic protein-induced osteoprotegerin gene expression. J Biol Chem. 2001; 276: 10119–10125.
- ↵Collin-Osdoby P, Rothe L, Anderson F, Nelson M, Maloney W, Osdoby P. Receptor activator of NF-κB and osteoprotegerin expression by human microvascular cells, regulation by inflammatory cytokines, and role in human osteoclastogenesis. J Biol Chem. 2001; 276: 20659–20672.
- ↵Vidal NO, Brandstrom H, Jonsson KB, Ohlsson C. Osteoprotegerin mRNA is expressed in primary human osteoblast-like cells: down-regulation by glucocorticoids. J Endocrinol. 1998; 159: 191–195.
- ↵Hofbauer LC, Gori F, Riggs BL, Lacey DL, Dunstan CR, Spelsberg TC, Khosla S. Stimulation of osteoprotegerin ligand and inhibition of osteoprotegerin production by glucocorticoids in human osteoblastic lineage cells: potential paracrine mechanisms of glucocorticoid-induced osteoporosis. Endocrinology. 1999; 140: 4382–4389.
- ↵Hofbauer LC, Shui C, Riggs BL, Dunstan CR, Spelsberg TC, O’Brien T, Khosla S. Effects of immunosuppressants on receptor activator of NF-κB ligand and osteoprotegerin production by human osteoblastic and coronary smooth muscle cells. Biochem Biophys Res Commun. 2001; 280: 334–339.
- ↵Onyia JE, Miles RR, Yang X, Halladay DL, Hale J, Glasebrook A, McClure D, Seno G, Churgay L, Chandrasekhar S, Martin TJ. In vivo demonstration that human parathyroid hormone 1-38 inhibits the expression of osteoprotegerin in bone with the kinetics of an immediate early gene. J Bone Miner Res. 2000; 15: 863–871.
- ↵Nakagawa N, Yasuda H, Yano K, Mochizuki S, Kobayashi N, Fujimoto H, Shima N, Morinaga T, Chikazu D, Kawaguchi H, Higashio K. Basic fibroblast growth factor induces osteoclast formation by reciprocally regulating the production of osteoclast differentiation factor and osteoclastogenesis inhibitory factor in mouse osteoblastic cells. Biochem Biophys Res Commun. 1999; 265: 158–163.
- ↵Kobayashi Y, Hashimoto F, Miyamoto H, Kanaoa K, Miyazaki-Kawashita Y, Nakashima T, Shibata M, Kobayashi K, Kato Y, Sakai H. Force-induced osteoclast apoptosis in vivo is accompanied by elevation in transforming growth factor-β and osteoprotegerin expression. J Bone Miner Res. 2000; 15: 1924–1934.
- ↵Lacey DL, Timms E, Tan HL, Kelly MJ, Dunstan CR, Burgess T, Elliott R, Colombero A, Elliott G, Scully S, Hsu H, Sullivan J, Hawkins N, Davy E, Capparelli C, Eli A, Qian YX, Kaufman S, Sarosi I, Shalhoub V, Senaldi G, Guo J, Delaney J, Boyle WJ. Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell. 1998; 93: 3597–3602.
- ↵Kong YY, Feige U, Sarosi I, Bolon B, Tafuri A, Morony S, Capparelli C, Li J, Elliott R, McCabe S, Wong T, Campagnuolo G, Moran E, Bogoch ER, Van G, Nguyen LT, Ohashi PS, Lacey DL, Fish E, Boyle WJ, Penninger JM. Activated T cells regulate bone loss and joint destruction in adjuvant arthritis through osteoprotegerin ligand. Nature. 1999; 402: 304–309.
- ↵Lum L, Wong BR, Josien R, Becherer JD, Erdjument-Bromage H, Schlöndorff J, Tempst P, Choi Y, Blobel CP. Evidence for a role of a tumor necrosis factor-α (TNF-α)–converting enzyme-like protease in shedding of TRANCE, a TNF family member involved in osteoclastogenesis and dendritic cell survival. J Biol Chem. 1999; 274: 13613–13618.
- ↵Hsu H, Lacey DL, Dunstan CR, Solovyev I, Colombero A, Timms E, Tan HL, Elliott G, Kelley MJ, Sarosi I, Wang L, Xia XZ, Elliott R, Chiu L, Black T, Scully S, Capparelli C, Morony S, Shimamoto G, Bass MB, Boyle WJ. Tumor necrosis factor receptor family member RANK mediates osteoclast differentiation and activation induced by osteoprotegerin ligand. Proc Natl Acad Sci U S A. 1999; 96: 3540–3545.
- ↵Green EA, Flavell RA. TRANCE-RANK, a new signal pathway involved in lymphocyte development and T cell activation. J Exp Med. 1999; 189: 1017–1020.
- ↵Yasuda H, Shima N, Nakagawa N, Yamaguchi K, Kinosaki M, Mochizuki SI, Tomoyasu A, Yano K, Goto M, Murakami A, Tsuda E, Morinaga T, Higashio K, Udagawa N, Takahashi N, Suda T. Osteoclast differentiation factor is a ligand for osteoprotegerin/osteoclastogenesis-inhibitory factor and is identical to TRANCE/RANKL. Proc Natl Acad Sci U S A. 1998; 95: 3597–3602.
- ↵Bucay N, Sarosi I, Dunstan CR, Morony S, Tarpley J, Capparelli C, Scully S, Tan HL, Xu W, Lacey DL, Boyle WJ, Simonet WS. Osteoprotegerin-deficient mice develop early onset osteoporosis and arterial calcification. Genes Dev. 1998; 12: 1260–1268.
- ↵Mizuno A, Amizuka N, Irie K, Murakami A, Fujise N, Kanno T, Sato Y, Nakagawa N, Yasuda H, Mochizuki S, Gomibuchi T, Yano K, Shima N, Washida N, Tsuda E, Morinaga T, Higashio K, Ozawa H. Severe osteoporosis in mice lacking osteoclastogenesis inhibitory factor/osteoprotegerin. Biochem Biophys Res Commun. 1998; 247: 610–615.
- ↵Kong YY, Yoshida H, Sarosi I, Tan HL, Timms E, Capparelli C, Morony S, Oliveira-dos-Santos AJ, Van G, Itie A, Khoo W, Wakeham A, Dunstan CR, Lacey DL, Mak TW, Boyle WJ, Penninger JM. OPGL is a key regulator of osteoclastogenesis, lymphocyte development and lymph-node organogenesis. Nature. 1999; 397: 315–323.
- ↵Teitelbaum SL. Bone resorption by osteoclasts. Science. 2000; 289: 1504–1508.
- ↵Wong BR, Josien R, Choi Y. TRANCE is a TNF family member that regulates dendritic cell and osteoclast function. J Leukoc Biol. 1999; 65: 715–724.
- ↵Wong BR, Josien R, Lee SY, Sauter B, Li HL, Steinman RM, Choi Y. TRANCE, a new TNF family member predominantly expressed in T cells, is a dendritic cell-specific survival factor. J Exp Med. 1997; 186: 2075–2080.
- ↵Josien R, Wong BR, Li HL, Steinman RM, Choi Y. TRANCE, a TNF family member, is differentially expressed on T cell subsets and induces cytokine production in dendritic cells. J Immunol. 1999; 162: 2562–2568.
- ↵Josien R, Li HL, Ingulli E, Sarma S, Wong BR, Vologodskaia M, Steinman RM, Choi Y. TRANCE, a TNF family member, enhances the longevity and adjuvant properties of dendritic cells in vivo. J Exp Med. 2000; 191: 495–502.
- ↵Bachmann MF, Wong BR, Josien R, Steinman RM, Oxenius A, Choi Y. TRANCE, a TNF family member critical for CD40 ligand-independent T helper cell activation. J Exp Med. 1999; 189: 1025–1031.
- ↵Dougall WC, Glaccum M, Charrier K, Rohrbach K, Brasel K, De Smedt T, Daro E, Smith J, Tometsko ME, Maliszewski CR, Armstrong A, Shen V, Bain S, Cosman D, Anderson D, Morrissey PJ, Peschon JJ, Schuh J. RANK is essential for osteoclast and lymph node development. Genes Dev. 1999; 13: 2412–2424.
- ↵Yun TJ, Tallquist MD, Aicher A, Rafferty KL, Marshall AJ, Moon JJ, Ewings ME, Mohaupt M, Herring SW, Clark EA. Osteoprotegerin, a crucial regulator of bone metabolism, also regulates B cell development and function. J Immunol. 2001; 166: 1482–1491.
- ↵Degli-Esposti M. To die or not to die: the quest of the TRAIL receptors. J Leukoc Biol. 1999; 65: 535–542.
- ↵Emery JG, Mc Donnell P, Brigham Burke M, Deen KC, Lyn S, Silverman C, Dul E, Appelbaum ER, Eichman C, DiPrinzio R, Dodds RA, James IE, Rosenberg M, Lee JC, Young PR. Osteoprotegerin is a receptor for the cytotoxic ligand TRAIL. J Biol Chem. 1998; 273: 14363–14367.
- ↵Min H, Morony S, Sarosi I, Dunstan CR, Capparelli C, Scully S, Van G, Kaufman S, Kostenuik PJ, Lacey DL, Boyle WJ, Simonet WS. Osteoprotegerin reverses osteoporosis by inhibiting endosteal osteoclasts and prevents vascular calcification by blocking a process resembling osteoclastogenesis. J Exp Med. 2000; 192: 463–474.
- ↵Gayard P, Garcier JM, Boire JY, Ravel A, Perez N, Privat C, Lucien P, Viallet JF, Boyer L. Spiral CT quantification of aorto-renal calcification and its use in the detection of atheromatous renal artery stenosis: a study in 42 patients. Cardiovasc Intervent Radiol. 2000; 23: 17–21.
- ↵Malyankar UM, Scatena M, Suchland KL, Yun TJ, Clark EA, Giachelli CM. Osteoprotegerin is an αvβ3-induced, NF-κB-dependent survival factor for endothelial cells. J Biol Chem. 2000; 275: 20959–20962.
- ↵Hak AE, Pols HA, van Hemert AM, Hofman A, Witteman JC. Progression of aortic calcification is associated with metacarpal bone loss during menopause: a population-based longitudinal study. Arterioscler Thromb Vasc Biol. 2000; 20: 1926–1931.
- ↵Yano K, Tsuda E, Washida N, Kobayashi F, Goto M, Harada A, Ikeda K, Higashio K, Yamada Y. Immunological characterization of circulating osteoprotegerin/osteoclasto-genesis inhibitory factor: increased serum concentrations in postmenopausal women with osteoporosis. J Bone Miner Res. 1999; 14: 518–527.
- ↵Price PA, June HH, Buckley JR, Williamson MK. Osteoprotegerin inhibits artery calcification induced by warfarin and by vitamin D. Arterioscler Thromb Vasc Biol. 2001; 21: 1610–1616.
- ↵Price PA, Faus SA, Williamson MK. Bisphosphonates alendronate and ibandronate inhibit artery calcification at doses comparable to those that inhibit bone resorption. Arterioscler Thromb Vasc Biol. 2001; 21: 817–824.
- ↵Dhore CR, Cleutjens JP, Lutgens E, Cleutjens KB, Geusens PP, Kitslaar PJ, Tordoir JH, Spronk HM, Vermeer C, Daemen MJ. Differential expression of bone matrix regulatory proteins in human atherosclerotic plaques. Arterioscler Thromb Vasc Biol. 2001; 21: 1998–2003.