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Osteoblast recruitment and bone formation enhanced by cell matrix-associated heparin-binding growth-associated molecule (HB-GAM).

Imai S, Kaksonen M, Raulo E, Kinnunen T, Fages C, Meng X, Lakso M, Rauvala H - J. Cell Biol. (1998)

Bottom Line: We show here that heparin-binding growth-associated molecule (HB-GAM), an extracellular matrix-associated protein that enhances migratory responses in neurons, is prominently expressed in the cell matrices that act as target substrates for bone formation.The HB-GAM transgenic mice develop a phenotype characterized by an increased bone thickness.HB-GAM may thus play an important role in bone formation, probably by mediating recruitment and attachment of osteoblasts/osteoblast precursors to the appropriate substrates for deposition of new bone.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Shiga University of Medical Science, Shiga-ken, 520-2192, Japan. simai@belle.shiga-med.ac.jp

ABSTRACT
Bone has an enormous capacity for growth, regeneration, and remodeling. This capacity is largely due to induction of osteoblasts that are recruited to the site of bone formation. The recruitment of osteoblasts has not been fully elucidated, though the immediate environment of the cells is likely to play a role via cell- matrix interactions. We show here that heparin-binding growth-associated molecule (HB-GAM), an extracellular matrix-associated protein that enhances migratory responses in neurons, is prominently expressed in the cell matrices that act as target substrates for bone formation. Intriguingly, N-syndecan, which acts as a receptor for HB-GAM, is expressed by osteoblasts/osteoblast precursors, whose ultrastructural phenotypes suggest active cell motility. The hypothesis that HB-GAM/N-syndecan interaction mediates osteoblast recruitment, as inferred from developmental studies, was tested using osteoblast-type cells that express N-syndecan abundantly. These cells migrate rapidly to HB-GAM in a haptotactic transfilter assay and in a migration assay where HB-GAM patterns were created on culture wells. The mechanism of migration is similar to that previously described for the HB-GAM-induced migratory response of neurons. Our hypothesis that HB-GAM/N-syndecan interaction participates in regulation of osteoblast recruitment was tested using two different in vivo models: an adjuvant-induced arthritic model and a transgenic model. In the adjuvant-induced injury model, the expression of HB-GAM and of N-syndecan is strongly upregulated in the periosteum accompanying the regenerative response of bone. In the transgenic model, the HB-GAM expression is maintained in mesenchymal tissues with the highest expression in the periosteum. The HB-GAM transgenic mice develop a phenotype characterized by an increased bone thickness. HB-GAM may thus play an important role in bone formation, probably by mediating recruitment and attachment of osteoblasts/osteoblast precursors to the appropriate substrates for deposition of new bone.

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Recruitment of UMR-106 cells using the pattern of  UV-irradiated and intact HB-GAM. (a) A schematic representation of the experiment. The substrate is first coated with HB-GAM,  covered with a conventional EM grid, and irradiated with UV  that denatures the uncovered HB-GAM. (b) Cells are evenly  plated, and there is no apparent migration 30 min after plating  the cells. (c) Cells migrating from the denatured HB-GAM to the  intact HB-GAM at 3 h after plating. (d) The cells have migrated  and spread on the intact HB-GAM at 7 h after plating.
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Figure 4: Recruitment of UMR-106 cells using the pattern of UV-irradiated and intact HB-GAM. (a) A schematic representation of the experiment. The substrate is first coated with HB-GAM, covered with a conventional EM grid, and irradiated with UV that denatures the uncovered HB-GAM. (b) Cells are evenly plated, and there is no apparent migration 30 min after plating the cells. (c) Cells migrating from the denatured HB-GAM to the intact HB-GAM at 3 h after plating. (d) The cells have migrated and spread on the intact HB-GAM at 7 h after plating.

Mentions: Since cells may attach nonspecifically to polycationic proteins, another cell recruitment assay was designed. Irradiation with UV light has a protein-denaturing effect, but it is not expected to destroy the polycationic nature of the protein. UV-irradiation has been previously used to create patterned substrates from laminin (Hammerback et al., 1985) and from HB-GAM (Rauvala et al., 1994). UMR-106 cells were clearly recruited to the intact HB-GAM from the UV-irradiated HB-GAM at the concentration of 50 μg/ml (Fig. 4, b–d), suggesting that the intact molecular structure of HB-GAM is a prerequisite for the recruitment of the UMR-106 cells. Cell migration was inactive at lower concentration of HB-GAM, and the cells did not migrate at all when the substrate was not previously coated with HB-GAM (data not shown). The N-syndecan–negative control cells, N18 neuroblastoma cells and 3T3 fibroblasts, were not recruited to the intact HB-GAM (data not shown).


Osteoblast recruitment and bone formation enhanced by cell matrix-associated heparin-binding growth-associated molecule (HB-GAM).

Imai S, Kaksonen M, Raulo E, Kinnunen T, Fages C, Meng X, Lakso M, Rauvala H - J. Cell Biol. (1998)

Recruitment of UMR-106 cells using the pattern of  UV-irradiated and intact HB-GAM. (a) A schematic representation of the experiment. The substrate is first coated with HB-GAM,  covered with a conventional EM grid, and irradiated with UV  that denatures the uncovered HB-GAM. (b) Cells are evenly  plated, and there is no apparent migration 30 min after plating  the cells. (c) Cells migrating from the denatured HB-GAM to the  intact HB-GAM at 3 h after plating. (d) The cells have migrated  and spread on the intact HB-GAM at 7 h after plating.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2132954&req=5

Figure 4: Recruitment of UMR-106 cells using the pattern of UV-irradiated and intact HB-GAM. (a) A schematic representation of the experiment. The substrate is first coated with HB-GAM, covered with a conventional EM grid, and irradiated with UV that denatures the uncovered HB-GAM. (b) Cells are evenly plated, and there is no apparent migration 30 min after plating the cells. (c) Cells migrating from the denatured HB-GAM to the intact HB-GAM at 3 h after plating. (d) The cells have migrated and spread on the intact HB-GAM at 7 h after plating.
Mentions: Since cells may attach nonspecifically to polycationic proteins, another cell recruitment assay was designed. Irradiation with UV light has a protein-denaturing effect, but it is not expected to destroy the polycationic nature of the protein. UV-irradiation has been previously used to create patterned substrates from laminin (Hammerback et al., 1985) and from HB-GAM (Rauvala et al., 1994). UMR-106 cells were clearly recruited to the intact HB-GAM from the UV-irradiated HB-GAM at the concentration of 50 μg/ml (Fig. 4, b–d), suggesting that the intact molecular structure of HB-GAM is a prerequisite for the recruitment of the UMR-106 cells. Cell migration was inactive at lower concentration of HB-GAM, and the cells did not migrate at all when the substrate was not previously coated with HB-GAM (data not shown). The N-syndecan–negative control cells, N18 neuroblastoma cells and 3T3 fibroblasts, were not recruited to the intact HB-GAM (data not shown).

Bottom Line: We show here that heparin-binding growth-associated molecule (HB-GAM), an extracellular matrix-associated protein that enhances migratory responses in neurons, is prominently expressed in the cell matrices that act as target substrates for bone formation.The HB-GAM transgenic mice develop a phenotype characterized by an increased bone thickness.HB-GAM may thus play an important role in bone formation, probably by mediating recruitment and attachment of osteoblasts/osteoblast precursors to the appropriate substrates for deposition of new bone.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Shiga University of Medical Science, Shiga-ken, 520-2192, Japan. simai@belle.shiga-med.ac.jp

ABSTRACT
Bone has an enormous capacity for growth, regeneration, and remodeling. This capacity is largely due to induction of osteoblasts that are recruited to the site of bone formation. The recruitment of osteoblasts has not been fully elucidated, though the immediate environment of the cells is likely to play a role via cell- matrix interactions. We show here that heparin-binding growth-associated molecule (HB-GAM), an extracellular matrix-associated protein that enhances migratory responses in neurons, is prominently expressed in the cell matrices that act as target substrates for bone formation. Intriguingly, N-syndecan, which acts as a receptor for HB-GAM, is expressed by osteoblasts/osteoblast precursors, whose ultrastructural phenotypes suggest active cell motility. The hypothesis that HB-GAM/N-syndecan interaction mediates osteoblast recruitment, as inferred from developmental studies, was tested using osteoblast-type cells that express N-syndecan abundantly. These cells migrate rapidly to HB-GAM in a haptotactic transfilter assay and in a migration assay where HB-GAM patterns were created on culture wells. The mechanism of migration is similar to that previously described for the HB-GAM-induced migratory response of neurons. Our hypothesis that HB-GAM/N-syndecan interaction participates in regulation of osteoblast recruitment was tested using two different in vivo models: an adjuvant-induced arthritic model and a transgenic model. In the adjuvant-induced injury model, the expression of HB-GAM and of N-syndecan is strongly upregulated in the periosteum accompanying the regenerative response of bone. In the transgenic model, the HB-GAM expression is maintained in mesenchymal tissues with the highest expression in the periosteum. The HB-GAM transgenic mice develop a phenotype characterized by an increased bone thickness. HB-GAM may thus play an important role in bone formation, probably by mediating recruitment and attachment of osteoblasts/osteoblast precursors to the appropriate substrates for deposition of new bone.

Show MeSH
Related in: MedlinePlus