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High glucose inhibits receptor activator of nuclear factor‑κB ligand-induced osteoclast differentiation via downregulation of v‑ATPase V0 subunit d2 and dendritic cell‑specific transmembrane protein.

Xu J, Yue F, Wang J, Chen L, Qi W - Mol Med Rep (2014)

Bottom Line: The expression levels of Atp6V0d2 and DC‑STAMP are regulated by NFATc1 and c‑fos, and are required for osteoclast fusion, which is important for osteoclast maturation.To the best of our knowledge, the present study demonstrated for the first time that high glucose decreased the gene expression of ATP6v0d2 and DC‑STAMP in RAW264.7 cells mediated by RANKL.Therefore, the suppression of pre‑osteoclast or osteoclast fusion may be essential for the inhibition of osteoclast differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology, Qilu Hospital, Shandong University, Jinan, Shandong 250012, P.R. China.

ABSTRACT
The balance between bone formation and resorption is compromised in diabetes, which may contribute to the high risk of fractures in diabetic patients. However, the mechanism by which high glucose affects bone turnover remains to be elucidated. The present study demonstrated that high glucose inhibited receptor activator of nuclear factor‑κB ligand (RANKL)‑induced osteoclastogenesis. In order to examine the mechanism involved in the inhibition of osteoclastogenesis, the present study examined several key molecules involved in osteoclast differentiation, including v‑ATPase V0 subunit d2 (Atp6V0d2), dendritic cell‑specific transmembrane protein (DC-STAMP), c‑fos and nuclear factor of activated T cells c1 (NFATc1). The expression levels of Atp6V0d2 and DC‑STAMP are regulated by NFATc1 and c‑fos, and are required for osteoclast fusion, which is important for osteoclast maturation. To the best of our knowledge, the present study demonstrated for the first time that high glucose decreased the gene expression of ATP6v0d2 and DC‑STAMP in RAW264.7 cells mediated by RANKL. Therefore, the suppression of pre‑osteoclast or osteoclast fusion may be essential for the inhibition of osteoclast differentiation.

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Effect of high glucose on osteoclast formation in RAW264.7 cells. The Raw264.7 cells were seeded at a density of 2×104 cells/well in 24-well plates and cultured with RANKL and different doses (5.6 and 20.2 mM) of D-glucose or mannitol for 4 days. (A) High glucose culture with 20.2 mM D-glucose significantly decreased the number of TRAP-positive multinucleated cells compared with the control culture in 5.6 mM D-glucose (**P<0.01; n=4). No significant differences were observed in the osmotic control culture compared with the control culture. Data are expressed as the mean ± standard deviation. (B) Images of the TRAP-positive multinucleated cells were captured under a microscope (magnification, ×200). RANKL, receptor activator of nuclear factor-κB ligand; TRAP, tartrate-resistant acid phosphatase.
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f1-mmr-11-02-0865: Effect of high glucose on osteoclast formation in RAW264.7 cells. The Raw264.7 cells were seeded at a density of 2×104 cells/well in 24-well plates and cultured with RANKL and different doses (5.6 and 20.2 mM) of D-glucose or mannitol for 4 days. (A) High glucose culture with 20.2 mM D-glucose significantly decreased the number of TRAP-positive multinucleated cells compared with the control culture in 5.6 mM D-glucose (**P<0.01; n=4). No significant differences were observed in the osmotic control culture compared with the control culture. Data are expressed as the mean ± standard deviation. (B) Images of the TRAP-positive multinucleated cells were captured under a microscope (magnification, ×200). RANKL, receptor activator of nuclear factor-κB ligand; TRAP, tartrate-resistant acid phosphatase.

Mentions: To examine the effect of high glucose on RANKL-induced osteoclastogenesis, the RAW264.7 cells were incubated with RANKL and different doses of glucose. TRAP staining was then performed to determine the number of osteoclast-like cells. As shown in Fig. 1, RANKL induced the formation of TRAP-positive multinucleated osteoclast-like cells; however, the increase of TRAP-positive multinucleated cells in the control cultures (5.6 mM D-glucose) was inhibited by the addition of 20.2 mM D-glucose. By contrast, no inhibition of RANKL-induced osteoclastogenesis were observed in the cultures incubated with mannitol (osmotic control). TRAP-positive cells appeared dark red in the cytoplasm as shown in the representative images of TRAP-positive multinucleated cells in the different treatment groups in Fig. 1. In order to determine whether the inhibitory effect of high glucose on RANKL-induced osteoclastogenesis was due to its effect on cell growth, a cell proliferation assay was performed. At glucose concentrations of 20.0 mM, high glucose increased cell growth (Fig. 2).


High glucose inhibits receptor activator of nuclear factor‑κB ligand-induced osteoclast differentiation via downregulation of v‑ATPase V0 subunit d2 and dendritic cell‑specific transmembrane protein.

Xu J, Yue F, Wang J, Chen L, Qi W - Mol Med Rep (2014)

Effect of high glucose on osteoclast formation in RAW264.7 cells. The Raw264.7 cells were seeded at a density of 2×104 cells/well in 24-well plates and cultured with RANKL and different doses (5.6 and 20.2 mM) of D-glucose or mannitol for 4 days. (A) High glucose culture with 20.2 mM D-glucose significantly decreased the number of TRAP-positive multinucleated cells compared with the control culture in 5.6 mM D-glucose (**P<0.01; n=4). No significant differences were observed in the osmotic control culture compared with the control culture. Data are expressed as the mean ± standard deviation. (B) Images of the TRAP-positive multinucleated cells were captured under a microscope (magnification, ×200). RANKL, receptor activator of nuclear factor-κB ligand; TRAP, tartrate-resistant acid phosphatase.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4262508&req=5

f1-mmr-11-02-0865: Effect of high glucose on osteoclast formation in RAW264.7 cells. The Raw264.7 cells were seeded at a density of 2×104 cells/well in 24-well plates and cultured with RANKL and different doses (5.6 and 20.2 mM) of D-glucose or mannitol for 4 days. (A) High glucose culture with 20.2 mM D-glucose significantly decreased the number of TRAP-positive multinucleated cells compared with the control culture in 5.6 mM D-glucose (**P<0.01; n=4). No significant differences were observed in the osmotic control culture compared with the control culture. Data are expressed as the mean ± standard deviation. (B) Images of the TRAP-positive multinucleated cells were captured under a microscope (magnification, ×200). RANKL, receptor activator of nuclear factor-κB ligand; TRAP, tartrate-resistant acid phosphatase.
Mentions: To examine the effect of high glucose on RANKL-induced osteoclastogenesis, the RAW264.7 cells were incubated with RANKL and different doses of glucose. TRAP staining was then performed to determine the number of osteoclast-like cells. As shown in Fig. 1, RANKL induced the formation of TRAP-positive multinucleated osteoclast-like cells; however, the increase of TRAP-positive multinucleated cells in the control cultures (5.6 mM D-glucose) was inhibited by the addition of 20.2 mM D-glucose. By contrast, no inhibition of RANKL-induced osteoclastogenesis were observed in the cultures incubated with mannitol (osmotic control). TRAP-positive cells appeared dark red in the cytoplasm as shown in the representative images of TRAP-positive multinucleated cells in the different treatment groups in Fig. 1. In order to determine whether the inhibitory effect of high glucose on RANKL-induced osteoclastogenesis was due to its effect on cell growth, a cell proliferation assay was performed. At glucose concentrations of 20.0 mM, high glucose increased cell growth (Fig. 2).

Bottom Line: The expression levels of Atp6V0d2 and DC‑STAMP are regulated by NFATc1 and c‑fos, and are required for osteoclast fusion, which is important for osteoclast maturation.To the best of our knowledge, the present study demonstrated for the first time that high glucose decreased the gene expression of ATP6v0d2 and DC‑STAMP in RAW264.7 cells mediated by RANKL.Therefore, the suppression of pre‑osteoclast or osteoclast fusion may be essential for the inhibition of osteoclast differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Endocrinology, Qilu Hospital, Shandong University, Jinan, Shandong 250012, P.R. China.

ABSTRACT
The balance between bone formation and resorption is compromised in diabetes, which may contribute to the high risk of fractures in diabetic patients. However, the mechanism by which high glucose affects bone turnover remains to be elucidated. The present study demonstrated that high glucose inhibited receptor activator of nuclear factor‑κB ligand (RANKL)‑induced osteoclastogenesis. In order to examine the mechanism involved in the inhibition of osteoclastogenesis, the present study examined several key molecules involved in osteoclast differentiation, including v‑ATPase V0 subunit d2 (Atp6V0d2), dendritic cell‑specific transmembrane protein (DC-STAMP), c‑fos and nuclear factor of activated T cells c1 (NFATc1). The expression levels of Atp6V0d2 and DC‑STAMP are regulated by NFATc1 and c‑fos, and are required for osteoclast fusion, which is important for osteoclast maturation. To the best of our knowledge, the present study demonstrated for the first time that high glucose decreased the gene expression of ATP6v0d2 and DC‑STAMP in RAW264.7 cells mediated by RANKL. Therefore, the suppression of pre‑osteoclast or osteoclast fusion may be essential for the inhibition of osteoclast differentiation.

Show MeSH
Related in: MedlinePlus