Osteoblast–Osteoclast Coculture Amplifies Inhibitory Effects of <scp>FG</scp>‐4592 on Human Osteoclastogenesis and Reduces Bone Resorption

Hulley, P A; Papadimitriou-Olivgeri, Ioanna; Knowles, Helen J.

doi:10.1002/jbm4.10370

Cited by 17 publications

(16 citation statements)

References 52 publications

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“…This differentiation was confirmed by changes in cellular morphology (significant increase in multinucleation, ruffled border, and cell diameter) and by increased expression of TRAP, a typical OC marker [55] . Unlike other different hydrogel systems for culturing OC (such as gellan gum, alginate or hyaluronic acid hydrogels [63] , [64] , [65] ), our system resembles the typical OC-bone interface. In fact, despite its limitations (e.g.…”

Section: Discussionmentioning

confidence: 99%

Hydroxyapatite-decorated Fmoc-hydrogel as a bone-mimicking substrate for osteoclast differentiation and culture

et al. 2022

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Hydrogels are water-swollen networks with great potential for tissue engineering applications. However, their use in bone regeneration is often hampered due to a lack of materials’ mineralization and poor mechanical properties. Moreover, most studies are focused on osteoblasts (OBs) for bone formation, while osteoclasts (OCs), cells involved in bone resorption, are often overlooked. Yet, the role of OCs is pivotal for bone homeostasis and aberrant OC activity has been reported in several pathological diseases, such as osteoporosis and bone cancer. For these reasons, the aim of this work is to develop customised, reinforced hydrogels to be used as material platform to study cell function, cell-material interactions and ultimately to provide a substrate for OC differentiation and culture. Here, Fmoc-based RGD-functionalised peptide hydrogels have been modified with hydroxyapatite nanopowder (Hap) as nanofiller, to create nanocomposite hydrogels. Atomic force microscopy showed that Hap nanoparticles decorate the peptide nanofibres with a repeating pattern, resulting in stiffer hydrogels with improved mechanical properties compared to Hap- and RGD-free controls. Furthermore, these nanocomposites supported adhesion of Raw 264.7 macrophages and their differentiation in 2D to mature OCs, as defined by the adoption of a typical OC morphology (presence of an actin ring, multinucleation, and ruffled plasma membrane). Finally, after 7 days of culture OCs showed an increased expression of TRAP, a typical OC differentiation marker. Collectively, the results suggest that the Hap/Fmoc-RGD hydrogel has a potential for bone tissue engineering, as a 2D model to study impairment or upregulation of OC differentiation. Statement of significance Altered osteoclasts (OC) function is one of the major cause of bone fracture in the most commonly skeletal disorders (e.g. osteoporosis). Peptide hydrogels can be used as a platform to mimic the bone microenvironment and provide a tool to assess OC differentiation and function. Moreover, hydrogels can incorporate different nanofillers to yield hybrid biomaterials with enhanced mechanical properties and improved cytocompatibility. Herein, Fmoc-based RGD-functionalised peptide hydrogels were decorated with hydroxyapatite (Hap) nanoparticles to generate a hydrogel with improved rheological properties. Furthermore, they are able to support osteoclastogenesis of Raw264.7 cells in vitro as confirmed by morphology changes and expression of OC-markers. Therefore, this Hap-decorated hydrogel can be used as a template to successfully differentiate OC and potentially study OC dysfunction.

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Section: Discussionmentioning

confidence: 99%

Hydroxyapatite-decorated Fmoc-hydrogel as a bone-mimicking substrate for osteoclast differentiation and culture

et al. 2022

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“…It is of interest that the effect of HIF-2α siRNA was different when osteoclastogenesis was performed in 3D, rather than 2D, culture; resulting in an increase, rather than a reduction, in the number of osteoclasts formed. It is not uncommon for cells cultured in 3D to exhibit altered sensitivity to cytokines, inhibitors or drugs 33,38,39 or a stronger differentiation response 38,40 , but we can find no other reports of a reversal in the direction of effect of a stimulus due to 3D culture. It is possible www.nature.com/scientificreports/ that this relates to the microenvironment-and tissue-specific nature of the regulation and phenotypic effects of HIF-2α 4,5 and is an area of considerable interest for future study.…”

Section: Discussionmentioning

confidence: 62%

“…3 b). We recently reported that osteoclastogenesis can be differentially affected in 3D versus 2D culture 33 . In contrast to standard culture conditions, HIF-2α siRNA caused a moderate increase in osteoclast formation when cells were cultured in 3D collagen gels (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…We recently showed that the PHD enzyme inhibitor FG-4592 stabilises HIF-1α and HIF-2α, inhibits the differentiation of human monocytes into osteoclasts and stimulates osteoclast-mediated bone resorption. However, co-culture with osteoblasts amplified the inhibition of osteoclastogenesis and dampened the increase in bone resorption 33 . This study further opens the door to understanding the differential effects of HIF-1α and HIF-2α on osteoclast biology but acknowledges the complexities that will be involved in studying these differential effects in vivo.…”

Section: Discussionmentioning

confidence: 99%

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Distinct roles for the hypoxia-inducible transcription factors HIF-1α and HIF-2α in human osteoclast formation and function

Knowles

2020

Sci Rep

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Bone homeostasis is maintained by a balance between osteoblast-mediated bone formation and osteoclast-driven bone resorption. Hypoxia modulates this relationship partially via direct and indirect effects of the hypoxia-inducible factor-1 alpha (HIF-1α) transcription factor on osteoclast formation and bone resorption. Little data is available on the role(s) of the HIF-2α isoform of HIF in osteoclast biology. Here we describe induction of HIF-1α and HIF-2α during the differentiation of human CD14+ monocytes into osteoclasts. Knockdown of HIF-1α did not affect osteoclast differentiation but prevented the increase in bone resorption that occurs under hypoxic conditions. HIF-2α knockdown did not affect bone resorption but moderately inhibited osteoclast formation. Growth of osteoclasts in 3D gels reversed the effect of HIF-2α knockdown; HIF-2α siRNA increasing osteoclast formation in 3D. Glycolysis is the main HIF-regulated pathway that drives bone resorption. HIF knockdown only affected glucose uptake and bone resorption in hypoxic conditions. Inhibition of glycolysis with 2-deoxy-d-glucose (2-DG) reduced osteoclast formation and activity under both basal and hypoxic conditions, emphasising the importance of glycolytic metabolism in osteoclast biology. In summary, HIF-1α and HIF-2α play different but overlapping roles in osteoclast biology, highlighting the importance of the HIF pathway as a potential therapeutic target in osteolytic disease.

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“…As a potent HIF-α stabilizer, roxadustat was found to promote fracture healing by stimulating the proliferation and migration of bone marrow mesenchymal stem cells, in which activation of the HIF-1α pathway was shown to be critical [ 15 ]. Moreover, roxadustat inhibited osteoclast differentiation and reduced bone resorption in vitro [ 16 ]. However, the effects of roxadustat on osteoblast differentiation and bone remodeling remain unknown.…”

Section: Introductionmentioning

confidence: 99%

Roxadustat promotes osteoblast differentiation and prevents estrogen deficiency-induced bone loss by stabilizing HIF-1α and activating the Wnt/β-catenin signaling pathway

Zhu

et al. 2022

J Orthop Surg Res

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Background Osteoporosis is a very common skeletal disorder that increases the risk of fractures. However, the treatment of osteoporosis is challenging. Hypoxia-inducible factor-1α (HIF-1α) plays an important role in bone metabolism. Roxadustat is a novel HIF stabilizer, and its effects on bone metabolism remain unknown. This study aimed to investigate the effects of roxadustat on osteoblast differentiation and bone remodeling in an ovariectomized (OVX) rat model. Methods In vitro, primary mouse calvarial osteoblasts were treated with roxadustat. Alkaline phosphatase (ALP) activity and extracellular matrix mineralization were assessed. The mRNA and protein expression levels of osteogenic markers were detected. The effects of roxadustat on the HIF-1α and Wnt/β-catenin pathways were evaluated. Furthermore, osteoblast differentiation was assessed again after HIF-1α expression knockdown or inhibition of the Wnt/β-catenin pathway. In vivo, roxadustat was administered orally to OVX rats for 12 weeks. Then, bone histomorphometric analysis was performed. The protein expression levels of the osteogenic markers HIF-1α and β-catenin in bone tissue were detected. Results In vitro, roxadustat significantly increased ALP staining intensity, enhanced matrix mineralization and upregulated the expression of osteogenic markers at the mRNA and protein levels in osteoblasts compared with the control group. Roxadustat activated the HIF-1α and Wnt/β-catenin pathways. HIF-1α knockdown or Wnt/β-catenin pathway inhibition significantly attenuated roxadustat-promoted osteoblast differentiation. In vivo, roxadustat administration improved bone microarchitecture deterioration and alleviated bone loss in OVX rats by promoting bone formation and inhibiting bone resorption. Roxadustat upregulated the protein expression levels of the osteogenic markers, HIF-1α and β-catenin in the bone tissue of OVX rats. Conclusion Roxadustat promoted osteoblast differentiation and prevented bone loss in OVX rats. The use of roxadustat may be a new promising strategy to treat osteoporosis.

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Osteoblast–Osteoclast Coculture Amplifies Inhibitory Effects of FG‐4592 on Human Osteoclastogenesis and Reduces Bone Resorption

Cited by 17 publications

References 52 publications

Hydroxyapatite-decorated Fmoc-hydrogel as a bone-mimicking substrate for osteoclast differentiation and culture

Hydroxyapatite-decorated Fmoc-hydrogel as a bone-mimicking substrate for osteoclast differentiation and culture

Distinct roles for the hypoxia-inducible transcription factors HIF-1α and HIF-2α in human osteoclast formation and function

Roxadustat promotes osteoblast differentiation and prevents estrogen deficiency-induced bone loss by stabilizing HIF-1α and activating the Wnt/β-catenin signaling pathway

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