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Hypoxia-inducible factor regulates osteoclast-mediated bone resorption: role of angiopoietin-like 4.

Knowles HJ, Cleton-Jansen AM, Korsching E, Athanasou NA - FASEB J. (2010)

Bottom Line: Normoxic inducers of HIF (CoCl(2), desferrioxamine, and l-mimosine) and 100 ng/ml ANGPTL4 stimulated osteoclastic resorption 2- to 3-fold in assays of lacunar dentine resorption, without affecting osteoclast viability.In the osteoblastic Saos2 cell line, ANGPTL4 caused a dose-dependent increase in proliferation (P<0.01, 100 ng/ml) and, at lower doses (1-25 ng/ml), mineralization.These results demonstrate that HIF is sufficient to enhance osteoclast-mediated bone resorption and that ANGPTL4 can compensate for HIF-1α deficiency with respect to stimulation of osteoclast activity and also augments osteoblast proliferation and differentiation.

View Article: PubMed Central - PubMed

Affiliation: Botnar Research Centre, University of Oxford, Nuffield Orthopaedic Centre, Headington, Oxford, OX3 7LD, UK. helen.knowles@ndorms.ox.ac.uk

ABSTRACT
Hypoxia and the hypoxia-inducible factor (HIF) transcription factor regulate angiogenic-osteogenic coupling and osteoclast-mediated bone resorption. To determine how HIF might coordinate osteoclast and osteoblast function, we studied angiopoietin-like 4 (ANGPTL4), the top HIF target gene in an Illumina HumanWG-6 v3.0 48k array of normoxic vs. hypoxic osteoclasts differentiated from human CD14(+) monocytes (14.3-fold induction, P<0.0004). ANGPTL4 mRNA and protein were induced by 24 h at 2% O(2) in human primary osteoclasts, monocytes, and osteoblasts. ANGPTL4 protein was observed by immunofluorescence in osteoclasts and osteoblasts in vivo. Normoxic inducers of HIF (CoCl(2), desferrioxamine, and l-mimosine) and 100 ng/ml ANGPTL4 stimulated osteoclastic resorption 2- to 3-fold in assays of lacunar dentine resorption, without affecting osteoclast viability. Isoform-specific HIF-1α small interfering RNA ablated hypoxic induction of ANGPTL4 and of resorption, which was rescued by addition of exogenous ANGPTL4 (P<0.001). In the osteoblastic Saos2 cell line, ANGPTL4 caused a dose-dependent increase in proliferation (P<0.01, 100 ng/ml) and, at lower doses (1-25 ng/ml), mineralization. These results demonstrate that HIF is sufficient to enhance osteoclast-mediated bone resorption and that ANGPTL4 can compensate for HIF-1α deficiency with respect to stimulation of osteoclast activity and also augments osteoblast proliferation and differentiation.

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ANGPTL4 and the osteoblastic phenotype. A) Proliferation of Saos2 cells over a 10-d period in the presence of 1–100 ng/ml ANGPTL4. ANGPTL4 at 100 ng/ml (■——■) induced significant osteoblast proliferation compared with the untreated control (×——×). ○– – –○, 1 ng/ml ANGPTL4; ▵– – –▵, 5 ng/ml ANGPTL4; ◊– – –◊, 25 ng/ml ANGPTL4. **P < 0.01 vs. untreated control. B) Visualization of Saos2 mineralization with Alizarin red. Quantification at d 7 (open bars) and d 14 (shaded bars) showed the most significant effect at 5 ng/ml ANGPTL4. *P < 0.05, **P < 0.01 vs. no ANGPTL4 control.
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Figure 8: ANGPTL4 and the osteoblastic phenotype. A) Proliferation of Saos2 cells over a 10-d period in the presence of 1–100 ng/ml ANGPTL4. ANGPTL4 at 100 ng/ml (■——■) induced significant osteoblast proliferation compared with the untreated control (×——×). ○– – –○, 1 ng/ml ANGPTL4; ▵– – –▵, 5 ng/ml ANGPTL4; ◊– – –◊, 25 ng/ml ANGPTL4. **P < 0.01 vs. untreated control. B) Visualization of Saos2 mineralization with Alizarin red. Quantification at d 7 (open bars) and d 14 (shaded bars) showed the most significant effect at 5 ng/ml ANGPTL4. *P < 0.05, **P < 0.01 vs. no ANGPTL4 control.

Mentions: Effects of exogenous ANGPTL4 on the osteoblast phenotype were therefore investigated. ANGPTL4 at 100 ng/ml significantly enhanced osteoblast proliferation in comparison with that in untreated control cells (Fig. 8A). Lower concentrations of ANGPTL4 (1–25 ng/ml) induced osteoblast mineralization (Fig. 8B).


Hypoxia-inducible factor regulates osteoclast-mediated bone resorption: role of angiopoietin-like 4.

Knowles HJ, Cleton-Jansen AM, Korsching E, Athanasou NA - FASEB J. (2010)

ANGPTL4 and the osteoblastic phenotype. A) Proliferation of Saos2 cells over a 10-d period in the presence of 1–100 ng/ml ANGPTL4. ANGPTL4 at 100 ng/ml (■——■) induced significant osteoblast proliferation compared with the untreated control (×——×). ○– – –○, 1 ng/ml ANGPTL4; ▵– – –▵, 5 ng/ml ANGPTL4; ◊– – –◊, 25 ng/ml ANGPTL4. **P < 0.01 vs. untreated control. B) Visualization of Saos2 mineralization with Alizarin red. Quantification at d 7 (open bars) and d 14 (shaded bars) showed the most significant effect at 5 ng/ml ANGPTL4. *P < 0.05, **P < 0.01 vs. no ANGPTL4 control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2992372&req=5

Figure 8: ANGPTL4 and the osteoblastic phenotype. A) Proliferation of Saos2 cells over a 10-d period in the presence of 1–100 ng/ml ANGPTL4. ANGPTL4 at 100 ng/ml (■——■) induced significant osteoblast proliferation compared with the untreated control (×——×). ○– – –○, 1 ng/ml ANGPTL4; ▵– – –▵, 5 ng/ml ANGPTL4; ◊– – –◊, 25 ng/ml ANGPTL4. **P < 0.01 vs. untreated control. B) Visualization of Saos2 mineralization with Alizarin red. Quantification at d 7 (open bars) and d 14 (shaded bars) showed the most significant effect at 5 ng/ml ANGPTL4. *P < 0.05, **P < 0.01 vs. no ANGPTL4 control.
Mentions: Effects of exogenous ANGPTL4 on the osteoblast phenotype were therefore investigated. ANGPTL4 at 100 ng/ml significantly enhanced osteoblast proliferation in comparison with that in untreated control cells (Fig. 8A). Lower concentrations of ANGPTL4 (1–25 ng/ml) induced osteoblast mineralization (Fig. 8B).

Bottom Line: Normoxic inducers of HIF (CoCl(2), desferrioxamine, and l-mimosine) and 100 ng/ml ANGPTL4 stimulated osteoclastic resorption 2- to 3-fold in assays of lacunar dentine resorption, without affecting osteoclast viability.In the osteoblastic Saos2 cell line, ANGPTL4 caused a dose-dependent increase in proliferation (P<0.01, 100 ng/ml) and, at lower doses (1-25 ng/ml), mineralization.These results demonstrate that HIF is sufficient to enhance osteoclast-mediated bone resorption and that ANGPTL4 can compensate for HIF-1α deficiency with respect to stimulation of osteoclast activity and also augments osteoblast proliferation and differentiation.

View Article: PubMed Central - PubMed

Affiliation: Botnar Research Centre, University of Oxford, Nuffield Orthopaedic Centre, Headington, Oxford, OX3 7LD, UK. helen.knowles@ndorms.ox.ac.uk

ABSTRACT
Hypoxia and the hypoxia-inducible factor (HIF) transcription factor regulate angiogenic-osteogenic coupling and osteoclast-mediated bone resorption. To determine how HIF might coordinate osteoclast and osteoblast function, we studied angiopoietin-like 4 (ANGPTL4), the top HIF target gene in an Illumina HumanWG-6 v3.0 48k array of normoxic vs. hypoxic osteoclasts differentiated from human CD14(+) monocytes (14.3-fold induction, P<0.0004). ANGPTL4 mRNA and protein were induced by 24 h at 2% O(2) in human primary osteoclasts, monocytes, and osteoblasts. ANGPTL4 protein was observed by immunofluorescence in osteoclasts and osteoblasts in vivo. Normoxic inducers of HIF (CoCl(2), desferrioxamine, and l-mimosine) and 100 ng/ml ANGPTL4 stimulated osteoclastic resorption 2- to 3-fold in assays of lacunar dentine resorption, without affecting osteoclast viability. Isoform-specific HIF-1α small interfering RNA ablated hypoxic induction of ANGPTL4 and of resorption, which was rescued by addition of exogenous ANGPTL4 (P<0.001). In the osteoblastic Saos2 cell line, ANGPTL4 caused a dose-dependent increase in proliferation (P<0.01, 100 ng/ml) and, at lower doses (1-25 ng/ml), mineralization. These results demonstrate that HIF is sufficient to enhance osteoclast-mediated bone resorption and that ANGPTL4 can compensate for HIF-1α deficiency with respect to stimulation of osteoclast activity and also augments osteoblast proliferation and differentiation.

Show MeSH
Related in: MedlinePlus