Localization of CD44, the hyaluronate receptor; on the plasma membrane of osteocytes and osteoclasts in rat tibiae

Nakamura, Hiroaki; Kenmotsu, Shin-ichi; Sakai, Haruna; Ozawa, Hiroki

doi:10.1007/bf00307793

Cited by 53 publications

(41 citation statements)

References 46 publications

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“…6) indicates that, in the femoral growth plate, CD44 is present on the surfaces of cells in the zones of erosion and ossification, but not in other regions. (32) In the femoral metaphysis, osteoclasts, osteocytes, and some osteoblasts are CD44 + , findings consistent with those of Nakamura et al (32) and others. (33,34) Whereas many CD44 isoforms are expressed in other cell types, (35) the standard form (85 kDa) of CD44 is the only form expressed in osteoblasts (our data) (36) and in osteoclasts.…”

Section: Discussionsupporting

confidence: 85%

Hyaluronan Increases RANKL Expression in Bone Marrow Stromal Cells Through CD44

Cao

Singleton

Majumdar

et al. 2005

Journal of Bone and Mineral Research

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HA activates CD44 to stimulate RANKL expression in bone marrow stromal cells. HA stimulation of RANKL is blocked by anti-CD44 antibody and is absent in cells from CD44 −/− mice. CD44 −/− mice exhibit thicker cortical bone and a smaller medullary cavity, but indices of bone resorption are not affected.Introduction: Hyaluronan (HA), the major nonprotein glycosaminoglycan component of the extracellular matrix in mammalian bone marrow, functions in part through its receptor, CD44, to stimulate a series of intracellular signaling events that lead to cell migration, adhesion, and activation. To determine whether HA activation of CD44 influences RANKL and osteoprotegerin (OPG) expression and whether CD44 is functionally important in bone metabolism, we studied whole bone and bone marrow stromal cells (BMSCs) from wildtype and CD44 −/− mice. Materials and Methods: BMSCs from wildtype and CD44−/− mice at 7 weeks of age were cultured and treated with either HA or anti-CD44 antibody. The levels of mRNA of RANKL, OPG, CD44, alkaline phosphatase (ALP), osteocalcin (OC), and ␣I collagen (COLL) were determined by quantitative real-time RT-PCR. Levels of RANKL and CD44 protein were measured by immunoblotting, and expression of CD44 in whole bone was determined by immunohistochemical staining. Double immunofluorescence staining and confocal microscopy were used to study colocalization of Cbfa1, CD44, and HA. Tibias were imaged using CT, and cancellous and cortical parameters were measured. Osteoblast and osteoclast surface in the distal femoral metaphysis and osteoclast on the endocortical surface at the tibio-fibular junction were measured using quantitative histomorphometry. Differences were analyzed using ANOVA and the Newman-Keuls test. Results: Addition of HA dose-dependently increased RANKL mRNA (3.6-fold) and protein (3-fold) levels in BMSCs. Stimulation of RANKL by HA could be blocked with anti-CD44 antibody. Treatment of cells with HA or anti-CD44 antibody had no significant effect on OPG mRNA levels. Both CD44 and HA localized on the plasma membrane in cells expressing Cbfa1. HA localization on the cell membrane disappeared when cells were preincubated with anti-CD44 antibody. Compared with control mice, cortical bone of CD44 −/− was thicker, and medullary area was smaller at both 7 and 17 weeks, but at 7 weeks, indices of bone resorption were normal. At 17 weeks of age, tibial mass of CD44 −/− mice was higher than control mice. CD44 −/− animals expressed less RANKL in whole bone (−30%) and in BMSCs (−50%). Cells from CD44 −/− animals failed to respond to either HA or CD44 antibody treatment. Conclusions: HA can increase RANKL expression in BMSCs through CD44.

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Section: Discussionsupporting

confidence: 85%

Hyaluronan Increases RANKL Expression in Bone Marrow Stromal Cells Through CD44

Cao

Singleton

Majumdar

et al. 2005

Journal of Bone and Mineral Research

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“…Immunocytochemical analyses of non-permeabilized osteoclasts have revealed the localization of CD44 and ␤ 3 on the surface (72). Colocalization of moesin and CD44 at the basolateral plasma membrane was also shown by electron microscopy analyses (30). In Madin-Darby canine kidney cells, endogenous and transfected CD44 are localized to the basolateral plasma membrane (73).…”

Section: Effects Of Rho and Rok Inhibitors On In Vitro Bone Resorptiomentioning

confidence: 96%

Rho-dependent Rho Kinase Activation Increases CD44 Surface Expression and Bone Resorption in Osteoclasts

Chellaiah

Biswas

Rittling

et al. 2003

Journal of Biological Chemistry

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Osteoclasts from osteopontin-efficient mice exhibit decreased CD44 surface expression. Osteopontin (OPN)/ ␣ v ␤ 3 generated Rho signaling pathway is required for the surface expression of CD44. In this work we show the Rho effector, Rho kinase (ROK-␣), to be a potent activator of CD44 surface expression. ROK-␣ activation was associated with autophosphorylation, leading to its translocation to the plasma membrane, as well as its association with CD44. ROK-␣ promoted CD44 surface expression through phosphorylation of CD44 and ezrinradixin-moesin (ERM) proteins and CD44⅐ERM⅐actin complex formation. Osteoclasts from OPN؊/؊ mice exhibited an ϳ55-60% decrease in basal level ROK-␣ phosphorylation as compared with wild type osteoclasts. Furthermore, RhoVal-14 transduction was only partially effective in stimulating ROK-␣/CD44 phosphorylation, as well as CD44 surface expression, in these osteoclasts. Studies on the inhibition of Rho by C3 transferase or ROK-␣ by the specific inhibitor, Y-27632, showed a decrease in the phosphorylation mediated by ROK-␣ and CD44 surface expression. Neutralizing antibodies to ␣ v , ␤ 3 , or CD44 inhibited the migration and bone resorption of wild type osteoclasts. However, only anti-␣ v or -␤ 3 antibodies blocked OPN-induced phosphorylation of ROK-␣, CD44, and the ERM proteins. Our results strongly suggest a role for ROK-␣ in ␣ v ␤ 3 -mediated Rho signaling, which is required for the phosphorylation events and CD44 surface expression. The functional deficiencies in the Rho effector(s) because of the lack of OPN were associated with decreased CD44 surface expression and hypomotility in the OPN؊/؊ osteoclasts. Finally, we find that cooperativity exists between ␣ v ␤ 3 and CD44 for osteoclast motility and bone resorption.Osteopontin (OPN), 1 a major component of protein complexes organized around mineralization foci, is a phosphorylated acidic glycoprotein synthesized by many cell types throughout the body in response to stress or injury (1-3). Although osteopontin serves many functions in bone cells besides regulating mineralization of the matrix, regulation of osteoclast motility and resorption is perhaps its most important. Osteopontin is a ligand for several cell surface adhesion receptors, such as, and ␣ 9 ␤ 1 (reviewed in Refs. 4 and 5). The hyaluronic acid receptor, CD44, is also a receptor for OPN (6). CD44 has been implicated in chemotaxis mediated by OPN (6, 7), as well as in bone resorption (8). CD44 expression was reported in osteoclasts of human and rat bone sections (9 -11). Weber et al. (6), have shown that the binding of soluble OPN to fibroblasts transfected with CD44 cDNA is blocked by anti-CD44 antibody. Moreover, anti-CD44 or anti-OPN antibodies inhibited the migration of CD44-transfected fibroblasts toward OPN in Boyden chambers (7), suggesting CD44-OPN interaction and a role in chemotactic function. Expression of OPN and CD44 was increased in migrating fetal fibroblasts (12), and the colocalization of CD44 and OPN was prominent at the leading edge of migrating osteoclast...

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“…A proteoglycan matrix fills the pericellular space between the cell membrane of the osteocyte process and the canalicular wall; glycosaminoglycan side chains of the pericellular matrix are assumed to have a 7-nm spacing typical of the glycocalyx on endothelial cells (19). Transverse tethering elements (i.e., core proteins of the pericellular proteoglycan matrix) are linked physically to cross-filaments via transmembrane molecules (e.g., CD44) (8,10) and also are arranged in a double-helix pattern of cross-filaments along the axial direction of the canaliculus. A local conical canalicular projection takes the place of one otherwise transverse tethering element, and the width of its base is 75 nm as shown in Fig.…”

Section: Theoretical Modelmentioning

confidence: 99%

“…The latter study also provided key input data for a greatly refined three-dimensional theoretical model by Han et al (9). Although both models elegantly showed that very small whole-tissue strains would be amplified 10-fold or more at the cellular level because of the tensile forces in the transverse tethering elements, the molecular mechanism for initiating intracellular signaling was hard to identify because none of the likely molecules in the tethering complex [i.e., proteoglycans, hyaluronic acid, or CD44 (8,[10][11][12]] are known mediators of mechanically induced cell signaling. In this paper, we propose a paradigm for cellular-level strain amplification by integrin-based focal attachment complexes along osteocyte cell processes.…”

mentioning

confidence: 99%

A model for the role of integrins in flow induced mechanotransduction in osteocytes

Wang

McNamara

Schaffler

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

223

217

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A fundamental paradox in bone mechanobiology is that tissuelevel strains caused by human locomotion are too small to initiate intracellular signaling in osteocytes. A cellular-level strainamplification model previously has been proposed to explain this paradox. However, the molecular mechanism for initiating signaling has eluded detection because none of the molecules in this previously proposed model are known mediators of intracellular signaling. In this paper, we explore a paradigm and quantitative model for the initiation of intracellular signaling, namely that the processes are attached directly at discrete locations along the canalicular wall by ␤3 integrins at the apex of infrequent, previously unrecognized canalicular projections. Unique rapid fixation techniques have identified these projections and have shown them to be consistent with other studies suggesting that the adhesion molecules are ␣v␤3 integrins. Our theoretical model predicts that the tensile forces acting on the integrins are <15 pN and thus provide stable attachment for the range of physiological loadings. The model also predicts that axial strains caused by the sliding of actin microfilaments about the fixed integrin attachments are an order of magnitude larger than the radial strains in the previously proposed strain-amplification theory and two orders of magnitude greater than whole-tissue strains. In vitro experiments indicated that membrane strains of this order are large enough to open stretch-activated cation channels.bone mechanotransduction ͉ integrin attachments ͉ osteocyte cell process ͉ strain amplification ͉ bone fluid flow A fundamental paradox in bone biology is that tissue-level strains, which rarely exceed 0.1% in vivo (1, 2), are too small to initiate intracellular signaling in bone cells in vitro (3,4), where the necessary strains (typically 1.0%) would cause bone fracture. Osteocytes, the most abundant cells in adult bone, are widely believed to be the primary sensory cells for mechanical loading because of their ubiquitous distribution throughout the bone tissue and their dendritic interconnections with both neighboring osteocytes and osteoblasts (5, 6), but osteocytes also require high local strains for mechanical stimulation. You et al. (7) developed an intuitive strain-amplification model to explain this paradox wherein osteocyte processes are attached to the canalicular wall by transverse tethering elements in the pericellular matrix. According to this model, the drag generated by load-induced fluid flow through the pericellular matrix would create tensile forces along the transverse elements supporting the pericellular matrix. These resulting tensions then were transmitted by transmembrane proteins to the central actin filament bundle in the osteocyte cell process leading to circumferential expansion of the cell process. The basic structural features in this model, the transverse tethering elements, and the organization of the actin filament bundle in the dendritic cell process were shown experimentally by You et...

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Localization of CD44, the hyaluronate receptor; on the plasma membrane of osteocytes and osteoclasts in rat tibiae

Cited by 53 publications

References 46 publications

Hyaluronan Increases RANKL Expression in Bone Marrow Stromal Cells Through CD44

Hyaluronan Increases RANKL Expression in Bone Marrow Stromal Cells Through CD44

Rho-dependent Rho Kinase Activation Increases CD44 Surface Expression and Bone Resorption in Osteoclasts

A model for the role of integrins in flow induced mechanotransduction in osteocytes

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