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Dimethyloxalylglycine prevents bone loss in ovariectomized C57BL/6J mice through enhanced angiogenesis and osteogenesis.

Peng J, Lai ZG, Fang ZL, Xing S, Hui K, Hao C, Jin Q, Qi Z, Shen WJ, Dong QN, Bing ZH, Fu DL - PLoS ONE (2014)

Bottom Line: However, there were no significant differences in serum CTX or in the number of tartrate-resistant acid phosphatase-stained cells between DMOG-treated OVX mice and OVX mice.Western blot results showed that DMOG administration partly rescued the decrease in HIF-1α and β-catenin expression following ovariectomy.Collectively, these results indicate that DMOG prevents bone loss due to ovariectomy in C57BL/6J mice by enhancing angiogenesis and osteogenesis, which are associated with activated HIF-1α and Wnt/β-catenin signaling pathways.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Institute of Traumatology and Orthopaedics, Shanghai Key Laboratory for Prevention and Treatment of Bone and Joint Diseases with Integrated Chinese-Western Medicine, Ruijin Hospital, Jiao Tong University School of Medicine, Shanghai, China.

ABSTRACT
Hypoxia-inducible factor 1-α (HIF-1α) plays a critical role in angiogenesis-osteogenesis coupling during bone development and bone regeneration. Previous studies have shown that 17β-estradiol activates the HIF-1α signaling pathway and that mice with conditional activation of the HIF-1α signaling pathway in osteoblasts are protected from ovariectomy (OVX)-induced bone loss. In addition, it has been shown that hypoxia facilitates the osteogenic differentiation of mesenchymal stem cells (MSCs) and modulates Wnt/β-catenin signaling. Therefore, we hypothesized that activation of the HIF-1α signaling pathway by hypoxia-mimicking agents would prevent bone loss due to estrogen deficiency. In this study, we confirmed the effect of dimethyloxalylglycine (DMOG), a hypoxia-mimicking agent, on the HIF-1α signaling pathway and investigated the effect of DMOG on MSC osteogenic differentiation and the Wnt/β-catenin signaling pathway. We then investigated the effect of DMOG treatment on OVX-induced bone loss. Female C57BL/6J mice were divided into sham, OVX, OVX+L-DMOG (5 mg/kg/day), and OVX+H-DMOG (20 mg/kg/day) groups. At sacrifice, static and dynamic bone histomorphometry were performed with micro computed tomography (micro-CT) and undecalcified sections, respectively. Bone strength was assessed with the three-point bending test, and femur vessels were reconstructed and analyzed by micro-CT. Serum vascular endothelial growth factor (VEGF), osteocalcin, and C-terminal telopeptides of collagen type(CTX) were measured by ELISA. Tartrate-resistant acid phosphatase staining was used to assess osteoclast formation. Alterations in the HIF-1α and Wnt/β-catenin signaling pathways in the bone were detected by western blot. Our results showed that DMOG activated the HIF-1α signaling pathway, which further activated the Wnt/β-catenin signaling pathway and enhanced MSC osteogenic differentiation. The micro-CT results showed that DMOG treatment improved trabecular bone density and restored the bone microarchitecture and blood vessels in OVX mice. Bone strength was also partly restored in DMOG-treated OVX mice. Dynamic bone histomorphometric analysis of the femur metaphysic revealed that DMOG increased the mineralizing surface, mineral apposition rate, and bone formation rate. The serum levels of VEGF and osteocalcin were higher in DMOG-treated OVX mice. However, there were no significant differences in serum CTX or in the number of tartrate-resistant acid phosphatase-stained cells between DMOG-treated OVX mice and OVX mice. Western blot results showed that DMOG administration partly rescued the decrease in HIF-1α and β-catenin expression following ovariectomy. Collectively, these results indicate that DMOG prevents bone loss due to ovariectomy in C57BL/6J mice by enhancing angiogenesis and osteogenesis, which are associated with activated HIF-1α and Wnt/β-catenin signaling pathways.

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Effect of 17β-estradiol and DMOG on HIF-1α signaling pathway.(A) 17β-estradiol stabilized HIF-1α under normal oxygen pressure. (B) 17β-estradiol upregulated VEGF protein expression. (C) DMOG enhanced HIF-1α protein expression. (D–E) DMOG upregulated VEGF expression at the mRNA and protein levels. P<0.05 for comparisons among the groups designated with an asterisk.
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pone-0112744-g001: Effect of 17β-estradiol and DMOG on HIF-1α signaling pathway.(A) 17β-estradiol stabilized HIF-1α under normal oxygen pressure. (B) 17β-estradiol upregulated VEGF protein expression. (C) DMOG enhanced HIF-1α protein expression. (D–E) DMOG upregulated VEGF expression at the mRNA and protein levels. P<0.05 for comparisons among the groups designated with an asterisk.

Mentions: Consistent with previous reports, our western blot analysis showed that 17β-estradiol stabilized HIF-1α under normal oxygen tension (Fig. 1A). Furthermore, 17β-estradiol increased the expression of VEGF protein (control vs. 10−9E2P = 0.001<0.05, control vs. 10−7E2P = 0.000<0.05, control vs. 10−5E2P = 0.000<0.05), the main downstream angiogenic target of the HIF-1α pathway, in a dose-dependent manner (Fig. 1B). Similarly, DMOG enhanced HIF-1α protein expression under normal oxygen tension and upregulated VEGF expression at the mRNA and protein levels (Fig. 1C–E).


Dimethyloxalylglycine prevents bone loss in ovariectomized C57BL/6J mice through enhanced angiogenesis and osteogenesis.

Peng J, Lai ZG, Fang ZL, Xing S, Hui K, Hao C, Jin Q, Qi Z, Shen WJ, Dong QN, Bing ZH, Fu DL - PLoS ONE (2014)

Effect of 17β-estradiol and DMOG on HIF-1α signaling pathway.(A) 17β-estradiol stabilized HIF-1α under normal oxygen pressure. (B) 17β-estradiol upregulated VEGF protein expression. (C) DMOG enhanced HIF-1α protein expression. (D–E) DMOG upregulated VEGF expression at the mRNA and protein levels. P<0.05 for comparisons among the groups designated with an asterisk.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231053&req=5

pone-0112744-g001: Effect of 17β-estradiol and DMOG on HIF-1α signaling pathway.(A) 17β-estradiol stabilized HIF-1α under normal oxygen pressure. (B) 17β-estradiol upregulated VEGF protein expression. (C) DMOG enhanced HIF-1α protein expression. (D–E) DMOG upregulated VEGF expression at the mRNA and protein levels. P<0.05 for comparisons among the groups designated with an asterisk.
Mentions: Consistent with previous reports, our western blot analysis showed that 17β-estradiol stabilized HIF-1α under normal oxygen tension (Fig. 1A). Furthermore, 17β-estradiol increased the expression of VEGF protein (control vs. 10−9E2P = 0.001<0.05, control vs. 10−7E2P = 0.000<0.05, control vs. 10−5E2P = 0.000<0.05), the main downstream angiogenic target of the HIF-1α pathway, in a dose-dependent manner (Fig. 1B). Similarly, DMOG enhanced HIF-1α protein expression under normal oxygen tension and upregulated VEGF expression at the mRNA and protein levels (Fig. 1C–E).

Bottom Line: However, there were no significant differences in serum CTX or in the number of tartrate-resistant acid phosphatase-stained cells between DMOG-treated OVX mice and OVX mice.Western blot results showed that DMOG administration partly rescued the decrease in HIF-1α and β-catenin expression following ovariectomy.Collectively, these results indicate that DMOG prevents bone loss due to ovariectomy in C57BL/6J mice by enhancing angiogenesis and osteogenesis, which are associated with activated HIF-1α and Wnt/β-catenin signaling pathways.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Institute of Traumatology and Orthopaedics, Shanghai Key Laboratory for Prevention and Treatment of Bone and Joint Diseases with Integrated Chinese-Western Medicine, Ruijin Hospital, Jiao Tong University School of Medicine, Shanghai, China.

ABSTRACT
Hypoxia-inducible factor 1-α (HIF-1α) plays a critical role in angiogenesis-osteogenesis coupling during bone development and bone regeneration. Previous studies have shown that 17β-estradiol activates the HIF-1α signaling pathway and that mice with conditional activation of the HIF-1α signaling pathway in osteoblasts are protected from ovariectomy (OVX)-induced bone loss. In addition, it has been shown that hypoxia facilitates the osteogenic differentiation of mesenchymal stem cells (MSCs) and modulates Wnt/β-catenin signaling. Therefore, we hypothesized that activation of the HIF-1α signaling pathway by hypoxia-mimicking agents would prevent bone loss due to estrogen deficiency. In this study, we confirmed the effect of dimethyloxalylglycine (DMOG), a hypoxia-mimicking agent, on the HIF-1α signaling pathway and investigated the effect of DMOG on MSC osteogenic differentiation and the Wnt/β-catenin signaling pathway. We then investigated the effect of DMOG treatment on OVX-induced bone loss. Female C57BL/6J mice were divided into sham, OVX, OVX+L-DMOG (5 mg/kg/day), and OVX+H-DMOG (20 mg/kg/day) groups. At sacrifice, static and dynamic bone histomorphometry were performed with micro computed tomography (micro-CT) and undecalcified sections, respectively. Bone strength was assessed with the three-point bending test, and femur vessels were reconstructed and analyzed by micro-CT. Serum vascular endothelial growth factor (VEGF), osteocalcin, and C-terminal telopeptides of collagen type(CTX) were measured by ELISA. Tartrate-resistant acid phosphatase staining was used to assess osteoclast formation. Alterations in the HIF-1α and Wnt/β-catenin signaling pathways in the bone were detected by western blot. Our results showed that DMOG activated the HIF-1α signaling pathway, which further activated the Wnt/β-catenin signaling pathway and enhanced MSC osteogenic differentiation. The micro-CT results showed that DMOG treatment improved trabecular bone density and restored the bone microarchitecture and blood vessels in OVX mice. Bone strength was also partly restored in DMOG-treated OVX mice. Dynamic bone histomorphometric analysis of the femur metaphysic revealed that DMOG increased the mineralizing surface, mineral apposition rate, and bone formation rate. The serum levels of VEGF and osteocalcin were higher in DMOG-treated OVX mice. However, there were no significant differences in serum CTX or in the number of tartrate-resistant acid phosphatase-stained cells between DMOG-treated OVX mice and OVX mice. Western blot results showed that DMOG administration partly rescued the decrease in HIF-1α and β-catenin expression following ovariectomy. Collectively, these results indicate that DMOG prevents bone loss due to ovariectomy in C57BL/6J mice by enhancing angiogenesis and osteogenesis, which are associated with activated HIF-1α and Wnt/β-catenin signaling pathways.

Show MeSH
Related in: MedlinePlus