Parathyroid Hormone Stimulates Osteoblastic Expression of MCP-1 to Recruit and Increase the Fusion of Pre/Osteoclasts

Abstract

The clinical findings that alendronate blunted the anabolic effect of human parathyroid hormone (PTH) on bone formation suggest that active resorption is involved and enhances the anabolic effect. PTH signals via its receptor on the osteoblast membrane, and osteoclasts are impacted indirectly via the products of osteoblasts. Microarray with RNA from rats injected with human PTH or vehicle showed a strong association between the stimulation of monocyte chemoattractant protein-1 (MCP-1) and the anabolic effects of PTH. PTH rapidly and dramatically stimulated MCP-1 mRNA in the femora of rats receiving daily injections of PTH or in primary osteoblastic and UMR 106-01 cells. The stimulation of MCP-1 mRNA was dose-dependent and a primary response to PTH signaling via the cAMP-dependent protein kinase pathway in vitro. Studies with the mouse monocyte cell line RAW 264.7 and mouse bone marrow proved that osteoblastic MCP-1 can potently recruit osteoclast monocyte precursors and facilitate receptor activator of NF-κB ligand-induced osteoclastogenesis and, in particular, enhanced fusion. Our model suggests that PTH-induced osteoblastic expression of MCP-1 is involved in recruitment and differentiation at the stage of multinucleation of osteoclast precursors. This information provides a rationale for increased osteoclast activity in the anabolic effects of PTH in addition to receptor activator of NF-κB ligand stimulation to initiate greater bone remodeling. The clinical findings that alendronate blunted the anabolic effect of human parathyroid hormone (PTH) on bone formation suggest that active resorption is involved and enhances the anabolic effect. PTH signals via its receptor on the osteoblast membrane, and osteoclasts are impacted indirectly via the products of osteoblasts. Microarray with RNA from rats injected with human PTH or vehicle showed a strong association between the stimulation of monocyte chemoattractant protein-1 (MCP-1) and the anabolic effects of PTH. PTH rapidly and dramatically stimulated MCP-1 mRNA in the femora of rats receiving daily injections of PTH or in primary osteoblastic and UMR 106-01 cells. The stimulation of MCP-1 mRNA was dose-dependent and a primary response to PTH signaling via the cAMP-dependent protein kinase pathway in vitro. Studies with the mouse monocyte cell line RAW 264.7 and mouse bone marrow proved that osteoblastic MCP-1 can potently recruit osteoclast monocyte precursors and facilitate receptor activator of NF-κB ligand-induced osteoclastogenesis and, in particular, enhanced fusion. Our model suggests that PTH-induced osteoblastic expression of MCP-1 is involved in recruitment and differentiation at the stage of multinucleation of osteoclast precursors. This information provides a rationale for increased osteoclast activity in the anabolic effects of PTH in addition to receptor activator of NF-κB ligand stimulation to initiate greater bone remodeling. Parathyroid hormone (PTH) 2The abbreviations used are: PTH, parathyroid hormone; h, human; r, rat; RT, reverse transcription; ELISA, enzyme-linked immunosorbent assay; RANKL, receptor activator of NF-κB ligand; M-CSF, macrophage colony-stimulating factor; PKA, cAMP-dependent protein kinase; PKC, protein kinase C; PMA, phorbol 12-myristate 13-acetate; TRAP, tartrate-resistant acid phosphatase. 2The abbreviations used are: PTH, parathyroid hormone; h, human; r, rat; RT, reverse transcription; ELISA, enzyme-linked immunosorbent assay; RANKL, receptor activator of NF-κB ligand; M-CSF, macrophage colony-stimulating factor; PKA, cAMP-dependent protein kinase; PKC, protein kinase C; PMA, phorbol 12-myristate 13-acetate; TRAP, tartrate-resistant acid phosphatase. is a principal hormone regulating bone remodeling via its actions on both bone formation and bone resorption. The finding that the anabolic effect of human PTH-(1-34) on bone formation was blunted when bone resorption was inhibited indicated that active resorption is involved and enhances the in vivo anabolic effect of PTH (1Delmas P.D. Vergnaud P. Arlot M.E. Pastoureau P. Meunier P.J. Nilssen M.H. Bone (Elmsford). 1995; 16: 603-610Crossref PubMed Scopus (146) Google Scholar, 2Black D.M. Greenspan S.L. Ensrud K.E. Palermo L. McGowan J.A. Lang T.F. Garnero P. Bouxsein M.L. Bilezikian J.P. Rosen C.J. N. Engl. J. Med. 2003; 349: 1207-1215Crossref PubMed Scopus (1007) Google Scholar, 3Finkelstein J.S. Hayes A. Hunzelman J.L. Wyland J.J. Lee H. Neer R.M. N. Engl. J. Med. 2003; 349: 1216-1226Crossref PubMed Scopus (714) Google Scholar). However, the molecular basis of the osteoclast activation in this process has not been fully studied. In order to identify the key mediators for the anabolic effects of PTH, we performed microarray using RNA from the femora of 3-month-old Sprague-Dawley female rats injected daily with hPTH-(1-34) to increase bone formation or vehicle. This was compared with rats receiving hPTH-(1-34) in a catabolic protocol by continuous infusion. PTH-(1-34) regulated numerous genes (∼1,000), but differentially, in both regimes (see accompanying article, Ref. 4Li X. Liu H. Qin L. Tamasi J. Bergenstock M. Shapses S. Feyen J.H.M. Notterman D.A. Partridge N.C. J. Biol. Chem. 2007; 282: 33086-33097Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar). Notably, in the anabolic protocol, a number of cytokines and chemokines, RANKL, interleukin-6, CXCL1, and CCL2 (MCP-1), were highly induced. In contrast, none of these were significantly increased, by microarray, in the catabolic protocol. The chemokine, monocyte chemoattractant protein-1 (MCP-1, CCL2), was found to be the most highly stimulated gene from 14-day intermittent hPTH-(1-34) or hPTH-(1-31) administration (both cause over 100-fold changes). With continuous infusion of PTH, the MCP-1 mRNA was elevated less than 2-fold, and this difference indicates a potentially important role for MCP-1 in the anabolic effect of PTH. Chemokines are bioactive peptides that regulate leukocyte activation and migration. They are essential mediators of inflammation and are crucial for the control of viral infections. MCP-1 is one of the CC chemokines, one of the four main subfamilies of chemokines, consisting of C, CC, CXC, and CX3C, according to the location of the N-terminal four cysteine residues (5Sodhi A. Montaner S. Gutkind J.S. Nat. Rev. Mol. Cell Biol. 2004; 5: 998-1012Crossref PubMed Scopus (136) Google Scholar). As a potent chemokine for monocytes and macrophages, MCP-1 is implicated in the pathogenesis of diseases characterized by monocytic infiltrates (6Wise G.E. Frazier-Bowers S. D'Souza R.N. Crit. Rev. Oral. Biol. Med. 2002; 13: 323-334Crossref PubMed Scopus (226) Google Scholar). It can be commonly detected at the site of tooth eruption, rheumatoid arthritic bone degradation, and bacterially induced bone loss (6Wise G.E. Frazier-Bowers S. D'Souza R.N. Crit. Rev. Oral. Biol. Med. 2002; 13: 323-334Crossref PubMed Scopus (226) Google Scholar) due to stimulation by inflammatory mediators. MCP-1 is also expressed by mature osteoclasts, and its expression is stimulated by receptor activator of NF-κB ligand (RANKL) and regulated by nuclear factor κB (6Wise G.E. Frazier-Bowers S. D'Souza R.N. Crit. Rev. Oral. Biol. Med. 2002; 13: 323-334Crossref PubMed Scopus (226) Google Scholar, 7Ishida N. Hayashi K. Hoshijima M. Ogawa T. Koga S. Miyatake Y. Kumegawa M. Kimura T. Takeya T. J. Biol. Chem. 2002; 277: 41147-41156Abstract Full Text Full Text PDF PubMed Scopus (336) Google Scholar, 8Cappellen D. Luong-Nguyen N.H. Bongiovanni S. Grenet O. Wanke C. Susa M. J. Biol. Chem. 2002; 277: 21971-21982Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar). It is known that expression of MCP-1 is temporally and spatially associated with the recruitment of monocytes in both osseous inflammation and during developmentally regulated bone remodeling in the process of tooth eruption (9Graves D.T. Jiang Y. Valente A.J. Histol. Histopathol. 1999; 14: 1347-1354PubMed Google Scholar). In vitro studies have shown that osteoclasts in both mice and humans may form directly from precursor cell populations of monocytes and macrophages (10Udagawa N. Takahashi N. Akatsu T. Tanaka H. Sasaki T. Nishihara T. Koga T. Martin T.J. Suda T. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 7260-7264Crossref PubMed Scopus (818) Google Scholar). It is very likely that MCP-1 may play a role in hormone-stimulated bone remodeling through action on osteoclasts. However, the relationship between MCP-1 and PTH, one of the most effective hormones regulating bone remodeling, has never been investigated. Here we propose a model suggesting that PTH-induced osteoblastic expression of MCP-1 is involved in osteoclast recruitment and differentiation at the stage of fusion and multinucleation of osteoclast precursors. This hypothesis provides a rationale for increased osteoclast activity in the anabolic effect of PTH, apart from RANKL stimulation, to initiate greater bone remodeling in a transient time-dependent fashion. Collectively, our data strongly point toward MCP-1 being an important molecule for communication between osteoblasts and osteoclasts in the anabolic effect of PTH. Chemicals—Synthetic PTH-(1-34) (human), PTH-(3-34) (bovine), and PTH-(1-31) (human), were purchased from Bachem (Torrance, CA). Inhibitors H-89 and GF109203X were purchased from Calbiochem. Rat PTH-(1-34), 8-bromo-cAMP, PMA, and cycloheximide were obtained from Sigma. Recombinant rat/murine RANKL, M-CSF, MCP-1, and neutralizing anti-MCP-1 antibody were purchased from Chemicon (Temecula, CA). Penicillin/streptomycin was obtained from Invitrogen. In Vivo Treatment of PTH in Rats—Sprague-Dawley rats (3-month-old female, about 250 g) were purchased from Taconic Farms, Inc. (Germantown, NY). For the anabolic protocol, rats were injected subcutaneously with vehicle (0.9% saline solution) or hPTH-(1-34) (8 μg/100 g) and euthanized using at the indicated For the catabolic protocol, vehicle (0.9% saline solution) or hPTH-(1-34) in a of was at a of by continuous infusion of PTH-(1-34) was at μg/100 for the indicated to 3-month-old female Sprague-Dawley rats four The were by the and The primary from the were J. J. J. Bone (Elmsford). 1995; PubMed Scopus Google Scholar). of mRNA by or were at the indicated PTH As L. X. Partridge N.C. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google RNA was using by with the reverse was used to mRNA was performed on using a The used for the are in For cell was used an For was used an of for in a Cell 106-01 were in essential with and For osteoblastic were to to and to essential for the addition of PTH. Rat primary osteoblastic were obtained and D. PubMed Scopus Google Scholar). PTH these were for RAW 264.7 were at in essential with The were in in a osteoclasts were from bone marrow from femora and of bone marrow was from with and the marrow were by through a with an the cell were in and in at in were stimulated with the indicated of cytokines RANKL, from the of cytokines were or bone resorption H. K. P. T. S. S. Y. S.L. J. 2004; PubMed Scopus Google Scholar, Y. J. L. S.L. J. Biol. Chem. Full Text Full Text PDF PubMed Google Scholar). MCP-1 was purchased from to the protein of MCP-1 in cell were performed by the to of MCP-1 in UMR 106-01 cell that been in from to a were from for were in and the between the less than was performed A. P. A. A. J. PubMed Scopus Google Scholar, S. M. J. PubMed Scopus Google Scholar, L. K. J. Bone 1999; 14: PubMed Scopus Google Scholar). of MCP-1 or from UMR 106-01 cell with PTH-(1-34) for were to the of a with osteoclasts or the precursors from the with a cell P. X. H. P. Mol. Med. 2003; Google Scholar) and on the of the at in a were and the were from the of the were with and for by with for were osteoclasts. was by cell in of on the by using The were in from to a in the in this MCP-1 the by In to the of the effect of MCP-1, the the MCP-1 or was with of or anti-MCP-1 neutralizing antibody at for being used to of of MCP-1 with or RANKL and primary osteoclast was to cell for the indicated and osteoclast differentiation was by the activity of TRAP, a of osteoclasts, using acid and by the of osteoclasts K. 2002; PubMed Scopus Google Scholar, S. K. J. Bone PubMed Scopus Google Scholar). the were for and with and a for at by with a were to be using The number of and the number of cell in were The of osteoclasts were also Bone bone marrow were the The an bone in the form of that can be on a of was to the were with to cells. The were and by data are on a of performed on are shown in the or of and was used for PTH MCP-1 mRNA in Vivo and in microarray studies from our MCP-1 the most highly regulated gene in an anabolic PTH protocol in was not significantly stimulated in a catabolic protocol X. Liu H. Qin L. Tamasi J. Bergenstock M. Shapses S. Feyen J.H.M. Notterman D.A. Partridge N.C. J. Biol. Chem. 2007; 282: 33086-33097Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar). this by using RNA from the of the femora of rats that intermittent injections of PTH. As shown in daily injections of hPTH-(1-34) for and stimulated MCP-1 mRNA expression in this bone in rats the This very is a increase in the mRNA expression of MCP-1 one PTH the of MCP-1 mRNA with PTH in an increase of than with daily intermittent injections of PTH. In the PTH catabolic model hPTH-(1-34) was rats the MCP-1 mRNA was significantly increased in this of bone at and and but was never greater than of the vehicle that the stimulation of MCP-1 is an osteoblastic response to PTH, we the MCP-1 mRNA PTH-(1-34) administration in osteoblastic the UMR 106-01 cell and primary osteoblastic cells. The that PTH MCP-1 mRNA by in the UMR with the PTH showed that MCP-1 protein in UMR cell a PTH-(1-34) MCP-1 stimulation by PTH is also in primary osteoblastic the of osteoblasts in vivo by through and when in vitro. MCP-1 were stimulated at and to The stimulation by PTH is in the differentiation and the is in the It is that the is also to MCP-1, but these a in the of the femora PTH of MCP-1 via the and a in MCP-1 mRNA in UMR at of PTH when we performed a with PTH was to increase MCP-1 mRNA significantly effects were with PTH, and stimulation was to with PTH PTH signals through both and with its on the osteoblast S. J. Scholar, N.C. Martin T.J. 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PTH of MCP-1 is a primary UMR 106-01 were with in the or the of that cycloheximide effect on the PTH of MCP-1, suggesting that this process not protein this effect is a primary and UMR Cell on the expression of the chemokine and and the for not be detected by in osteoblastic not we the effect of MCP-1 on osteoclasts and precursors. macrophage RAW 264.7 were can cells. RAW to RANKL stimulation in vitro to osteoclasts a of fully osteoclasts H. D. A. L. L. T. S. C. S. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google Scholar). RAW not a factor in formation of osteoclast differentiation and P. X. H. P. Mol. Med. 2003; Google Scholar, H. D. A. L. L. T. S. C. S. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google Scholar, A. T. Y. H. T. H. T. K. H. K. Tanaka S. J. Bone 2002; PubMed Scopus Google the are with RANKL to mature osteoclasts. In both and UMR cell have strong effects on RAW RANKL we can that the effects of MCP-1 are by the of a neutralizing the MCP-1 antibody also significantly the of UMR cell we our using osteoclast precursor from mouse to the effect on primary Recombinant and UMR cell showed strong effects on primary osteoclast precursors the that MCP-1 has a strong effect on and is likely a factor in UMR migration. RAW and the from mouse bone were induced to by with RANKL for The osteoclasts were used in As shown in and UMR also a effect on these cells. the that MCP-1 has effects on both the mature osteoclast and its precursor cells. MCP-1 and found that MCP-1 stimulated osteoclast MCP-1 to the and RANKL of primary osteoclast for cytokines significantly increased mRNA expression and with MCP-1 of and The effect of MCP-1 to facilitate differentiation was with significantly increased number of in the fusion in osteoclasts and when compared with addition of MCP-1 in by using with mouse bone macrophages, we found MCP-1 significantly increased the at of both and and The activity was increased by and with addition of and of Parathyroid hormone is to have anabolic effects when injected in is a highly effective for this PTH not bone formation but also bone remodeling R.M. S. O. N. Engl. J. Med. PubMed Scopus Google Scholar). clinical and studies have shown that of bone can the anabolic response to PTH, suggesting that active bone resorption enhances the anabolic actions of PTH (1Delmas P.D. Vergnaud P. Arlot M.E. Pastoureau P. Meunier P.J. Nilssen M.H. Bone (Elmsford). 1995; 16: 603-610Crossref PubMed Scopus (146) Google Scholar, 2Black D.M. Greenspan S.L. Ensrud K.E. Palermo L. McGowan J.A. Lang T.F. Garnero P. Bouxsein M.L. Bilezikian J.P. Rosen C.J. N. Engl. J. Med. 2003; 349: 1207-1215Crossref PubMed Scopus (1007) Google Scholar, 3Finkelstein J.S. Hayes A. Hunzelman J.L. Wyland J.J. Lee H. Neer R.M. N. Engl. J. Med. 2003; 349: 1216-1226Crossref PubMed Scopus (714) Google Scholar). Bone resorption the activation of the to form mature osteoclasts. This expression of molecular and the increase in of osteoclasts. It is known that the PTH receptor on osteoblasts and has been that bone resorption is most likely through expressed by osteoblasts PTH play an important role in the differentiation of osteoclast in one is the of and the is between osteoclast and osteoblasts or osteoblastic cells. essential have been is the most important factor for the of to be for not of osteoclast but also differentiation mature osteoclasts and D. Luong-Nguyen N.H. Bongiovanni S. Grenet O. Wanke C. Susa M. J. Biol. Chem. 2002; 277: 21971-21982Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar, P. X. H. P. Mol. Med. 2003; Google Scholar). RANKL on osteoclast and to mature osteoclasts H. N. N. K. M. S. A. K. M. A. T. K. N. Takahashi N. Suda T. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). Parathyroid hormone increased and RANKL mRNA and protein expression by osteoblastic cells. As can of of the macrophage osteoclasts H. N. N. K. M. S. A. K. M. A. T. K. N. Takahashi N. Suda T. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, J. Bone PubMed Scopus Google Scholar). PTH stimulation, the in the of bone by osteoblast and bone macrophages be to This suggests that is essential for but PTH can regulate osteoclastogenesis via stimulation of RANKL of However, RANKL is a protein by the osteoblast that with its receptor on to these active osteoclasts and to initiate bone is essential that the osteoblast and osteoclast are in or a form of RANKL is and the remodeling T. A. S. Full Text Full Text PDF PubMed Scopus Google Scholar). cell is for RANKL stimulation of facilitate this process PTH stimulation, the osteoclasts or to RANKL on the osteoblast is of to initiate cell to osteoclast In the MCP-1 was to play an important role in this process by osteoclasts and precursors and osteoclastogenesis and fusion. Bone resorption is to the anabolic effects of PTH, but in the is also to have bone formation than resorption. the stimulation of bone resorption during the anabolic protocol be a transient osteoblastic bone formation to remodeling for osteoblasts. As is that the increase in osteoclast formation is directly to the of RANKL and a role H. N. N. K. M. S. A. K. M. A. T. K. N. Takahashi N. Suda T. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, X. D. H. J. J. PubMed Google Scholar). both in vitro and in vivo studies have that PTH osteoclast formation by of the S. PubMed Scopus Google Scholar). the PTH anabolic protocol, the hormone regulate bone in an intermittent fashion. osteoclasts through PTH osteoclasts to be and osteoblasts to It is important for the active osteoclast to to osteoblasts to to a in bone As the stimulation of RANKL by PTH administration is a transient response both in vitro and in The is for RANKL and the of increase in RANKL mRNA PTH administration is P. X. Bone (Elmsford). PubMed Scopus Google Scholar). The of the mRNA is PTH and to at in 3-month-old female rats with and 14-day intermittent administration X. Liu H. Qin L. Tamasi J. Bergenstock M. Shapses S. Feyen J.H.M. Notterman D.A. Partridge N.C. J. Biol. Chem. 2007; 282: 33086-33097Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar). the of MCP-1 is very to the of the of RANKL and with intermittent PTH It is that the stimulation of MCP-1 by PTH in vivo is a but transient this is very with its role for the anabolic effect of PTH. PTH-induced osteoblastic expression of MCP-1 can facilitate RANKL action in the of bone resorption by to the remodeling and osteoclast It is that with continuous RANKL mRNA are but the is also for MCP-1 we the catabolic effect is of elevated bone resorption due to the of RANKL with continuous infusion of PTH the of MCP-1 to facilitate the effect of the anabolic effect is of RANKL and MCP-1 is transient and bone is by increased bone It is very that is a and response to PTH in the anabolic protocol, and this may be a key difference the of intermittent continuous PTH. It is that a of osteoblasts is by this and these have a for anabolic bone from that of the at the of This suggests that PTH-induced osteoblastic expression of MCP-1 is involved in osteoclast and fusion of osteoclast precursors and provides a rationale for increased osteoclast activity in the anabolic effect of PTH in addition to the effects of RANKL to initiate bone remodeling. Our finding is with the from C.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, C.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar) that MCP-1 human osteoclast fusion. showed that RANKL, MCP-1 and can the differentiation of human to with the of osteoclasts C.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The difference between data and may be of the difference in the cell used human and the we were to mature osteoclasts with MCP-1, we that MCP-1 can recruit and osteoclasts and enhances the effects of RANKL on both of data suggest that MCP-1 has an important role during the activation of bone resorption. a was an essential of osteoclast and macrophage cell fusion and to have an essential role for osteoclast multinucleation in bone T. N. A. T. K. K. T. M. H. K. O. H. J. Med. 2004; PubMed Scopus Google Scholar, M. T. Y. K. N. N. K. K. T. K. Y. Takeya M. Y. Suda T. J. Med. PubMed Scopus Google Scholar). its ligand has not been is to that MCP-1 with the of both in regulating osteoclast fusion suggests that MCP-1 may with or the fusion for on its fusion The by C.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar) also suggests that MCP-1 is induced by RANKL during osteoclast that RANKL is to to the stimulation of MCP-1 mRNA during PTH intermittent administration for the The increase in MCP-1 mRNA is a primary response to PTH in osteoblastic in both MCP-1 and RANKL mRNA at PTH administration in and both to at the The RANKL mRNA for increased expressed RANKL protein to regulate the expression of the mRNA of MCP-1 is also at h, this is for an effect in the mRNA of MCP-1 is than at the be due to RANKL stimulation of the osteoclasts. However, the that MCP-1 mRNA is than that of the on the is to a MCP-1 may to process this of the of MCP-1, we to the bone and response to PTH of MCP-1 have been from mice for due to in monocyte recruitment L. J. S.L. C. J. Med. PubMed Scopus Google Scholar). was found that the of MCP-1 are elevated in human and associated with A. S. C. J. H. D. J. H. M.L. S. 13: PubMed Scopus Google Scholar, T. D. J. PubMed Scopus Google Scholar) through to play a role in the of macrophages D. J. S. K. J. PubMed Scopus Google Scholar). The stimulation of MCP-1 mRNA PTH in an anabolic protocol in the model is very our have found a difference in MCP-1 from rats injected with PTH compared with vehicle but not of the difference of mRNA not this is the of MCP-1 expression in bone the of It is also that the stimulation of MCP-1 by PTH, a stimulation, a of MCP-1 in these bone MCP-1 from the osteoblast PTH a of communication to on osteoclasts and its monocyte precursors. It is also that the to MCP-1 we mRNA expression of or by in osteoblastic 106-01 data not MCP-1 is to on the osteoblastic via an the osteoclast and monocytes are likely to be the main for MCP-1 in the of UMR with MCP-1 neutralizing antibody blunted and not its for likely in the of these and may to the of the effect was by an MCP-1 MCP-1 may be the main factor in UMR the of osteoclasts and its precursors. In shown in we propose a model that PTH the pathway is in in the expression of RANKL on the and of and precursor monocytes are to the remodeling site by MCP-1 to initiate the process of bone remodeling. the MCP-1 the fusion of the mature osteoclasts through a L. and for and the protocol for primary osteoclast