Bone Anabolism Research Articles

Enhancement of osteoblast differentiation using poly(ethylene sodium phosphate)

Biodegradable polyphosphoesters (PPEs) are of increasing interest due to their promising biomedical applications. Polyphosphodiesters (PPDEs), formed by polyphosphotriester (PPTE) dealkylation, have bone-targeting properties in vivo and therefore are promising polymer drug candidates for bone disease treatment. However, their effects on osteoblasts are still unclear. This study prepared two polymer structures, poly(methyl ethylene phosphate) (PMP) and poly(ethylene sodium phosphate) (PEP•Na), as PPTE and PPDE models, respectively, and investigated their effects on mouse osteoblastic cell (MC3T3-E1) differentiation. Results showed that PMP is inert toward osteoblast differentiation. In contrast, PEP•Na enhanced alkaline phosphatase (ALP) synthesis, matrix mineralization, and osteoblast-related gene expression. PEP•Na also enhanced Wnt signaling pathway–dependent Alp expression, in addition to its own internalization into the cytosol during 3 days of differentiation culture. These results showed that dealkylated PPEs are a polymer drug candidate for osteoporosis treatment, not only because of their bone-targeting properties but also because of the controllable effects of bone anabolism via osteoblast differentiation enhancement.

Glutamine Metabolism in Osteoprogenitors Is Required for Bone Mass Accrual and PTH-Induced Bone Anabolism in Male Mice.

Skeletal homeostasis critically depends on the proper anabolic functioning of osteolineage cells. Proliferation and matrix synthesis are highly demanding in terms of biosynthesis and bioenergetics, but the nutritional requirements that support these processes in bone-forming cells are not fully understood. Here, we show that glutamine metabolism is a major determinant of osteoprogenitor function during bone mass accrual. Genetic inactivation of the rate-limiting enzyme glutaminase 1 (GLS1) results in decreased postnatal bone mass, caused by impaired biosynthesis and cell survival. Mechanistically, we uncovered that GLS1-mediated glutamine catabolism supports nucleotide and amino acid synthesis, required for proliferation and matrix production. In addition, glutamine-derived glutathione prevents accumulation of reactive oxygen species and thereby safeguards cell viability. The pro-anabolic role of glutamine metabolism was further underscored in a model of parathyroid hormone (PTH)-induced bone formation. PTH administration increases glutamine uptake and catabolism, and GLS1 deletion fully blunts the PTH-induced osteoanabolic response. Taken together, our findings indicate that glutamine metabolism in osteoprogenitors is indispensable for bone formation. © 2020 American Society for Bone and Mineral Research (ASBMR).

Inhibition of miR-29-3p isoforms via tough decoy suppresses osteoblast function in homeostasis but promotes intermittent parathyroid hormone-induced bone anabolism.

miRNAs play a vital role in post-transcriptional regulation of gene expression in osteoblasts and osteoclasts, and the miR-29 family is expressed in both lineages. Using mice globally expressing a miR-29-3p tough decoy, we demonstrated a modest 30-60% decrease all three miR-29-3p isoforms: miR-29a, miR-29b, and miR-29c. While the miR-29-3p decoy did not impact osteoclast number or function, the tough decoy decreased bone formation in growing mice, which led to decreased trabecular bone volume in mature animals. These data support previous in vitro studies suggesting that miR-29-3p is a positive regulator of osteoblast differentiation. In contrast, when mice were treated with intermittent parathyroid hormone (PTH1-34), inhibition of miR-29-3p augmented the effect of PTH on cortical bone anabolism, increased bone formation rate and osteoblast surface, and increased levels of Ctnnb1/βcatenin mRNA, which is a miR-29 target. These findings highlight differences in the mechanisms controlling basal level bone formation and bone formation induced by intermittent PTH. Overall, the global miR-29-3p tough decoy model represents a modest loss-of-function, which could be a relevant tool for assessing the possible impact of systemically administered miR-29-3p inhibitors. Our studies provide a potential rationale for co-administration of PTH1-34 and miR-29-3p inhibitors, to boost bone formation in severely affected osteoporosis patients, particularly in the cortical compartment.

Development of cyclic peptides with potent in vivo osteogenic activity through RaPID-based affinity maturation

Osteoporosis is caused by a disequilibrium between bone resorption and bone formation. Therapeutics for osteoporosis can be divided into antiresorptives that suppress bone resorption and anabolics which increase bone formation. Currently, the only anabolic treatment options are parathyroid hormone mimetics or an anti-sclerostin monoclonal antibody. With the current global increases in demographics at risk for osteoporosis, development of therapeutics that elicit anabolic activity through alternative mechanisms is imperative. Blockade of the PlexinB1 and Semaphorin4D interaction on osteoblasts has been shown to be a promising mechanism to increase bone formation. Here we report the discovery of cyclic peptides by a novel RaPID (Random nonstandard Peptides Integrated Discovery) system-based affinity maturation methodology that generated the peptide PB1m6A9 which binds with high affinity to both human and mouse PlexinB1. The chemically dimerized peptide, PB1d6A9, showed potent inhibition of PlexinB1 signaling in mouse primary osteoblast cultures, resulting in significant enhancement of bone formation even compared to non-Semaphorin4D-treated controls. This high anabolic activity was also observed in vivo when the lipidated PB1d6A9 (PB1d6A9-Pal) was intravenously administered once weekly to ovariectomized mice, leading to complete rescue of bone loss. The potent osteogenic properties of this peptide shows great promise as an addition to the current anabolic treatment options for bone diseases such as osteoporosis.

Traumatic joint injury induces acute catabolic bone turnover concurrent with articular cartilage damage in a rat model of posttraumatic osteoarthritis.

Assess acute alterations in bone turnover, microstructure, and histomorphometry following noninvasive anterior cruciate ligament rupture (ACLR). Twelve female Lewis rats were randomized to receive noninvasive ACLR or Sham loading (n = 6/group). In vivo μCT was performed at 3, 7, 10, and 14 days postinjury to quantify compartment-dependent subchondral (SCB) and epiphyseal trabecular bone remodeling. Near-infrared (NIR) molecular imaging was used to measure in vivo bone anabolism (800 CW BoneTag) and catabolism (Cat K 680 FAST). Metaphyseal bone remodeling and articular cartilage morphology was quantified using ex vivo μCT and contrast-enhanced µCT, respectively. Calcein-based dynamic histomorphometry was used to quantify bone formation. OARSI scoring was used to assess joint degeneration, and osteoclast number was quantified on TRAP stained-sections. ACLR induced acute catabolic bone remodeling in subchondral, epiphyseal, and metaphyseal compartments. Thinning of medial femoral condyle (MFC) SCB was observed as early as 7 days postinjury, while lateral femoral condyles (LFCs) exhibited SCB gains. Trabecular thinning was observed in MFC epiphyseal bone, with minimal changes to LFC. NIR imaging demonstrated immediate and sustained reduction of bone anabolism (~15%-20%), and a ~32% increase in bone catabolism at 14 days, compared to contralateral limbs. These findings were corroborated by reduced bone formation rate and increased osteoclast numbers, observed histologically. ACLR-injured femora had significantly elevated OARSI score, cartilage thickness, and cartilage surface deviation. ACL rupture induces immediate and sustained reduction of bone anabolism and overactivation of bone catabolism, with mild-to-moderate articular cartilage damage at 14 days postinjury.

Bone Marrow Reconversion With Reambulation: A Prospective Clinical Trial.

In a prospective clinical trial, 20 healthy men participated in a 60-day, 6-degree head-down tilt bed rest study. Serial 3-T magnetic resonance (MR) imaging measures of the lumbar spine were performed at baseline, after 57 days of bed rest, and at 30, 360, and 720 days of reambulation (100 MR imaging scans). Proton density with and without fat saturation, 2-point Dixon, and single-voxel MR spectroscopy techniques were used to assess bone marrow composition (300 measures). Erythropoiesis was measured using hematocrit, reticulocyte, and ferritin. Also, participants randomly received either a nutritional intervention composed of polyphenols, omega-3, vitamin E, and selenium or a normal diet. Thirty days of reambulation after 60 days of bed rest caused a marked decrease of the mean lumbar vertebral fat fraction (VFF) (-9.2 ± 1.6 percentage points, -8.0 ± 1.3 percentage points, and -12.7 ± 1.2 percentage points compared with baseline using proton density, Dixon, MR spectroscopy, respectively; all 3, P < 0.05). Reambulation also decreased the fat saturation index (-5.3 ± 1.1 percentage points compared with baseline; P < 0.05). These coincided with lower hematocrit and ferritin and with increased reticulocytes at reambulation day 13 compared with baseline (all 3, P < 0.05). After 57 days of bed rest, the VFF was unchanged from baseline (all 3 MR techniques, P > 0.05); reambulation for 2 years returned the lumbar VFF to baseline values. This longitudinal trial established that 30 days of reambulation after 60 days of bed rest constituted a powerful stimulus for bone marrow reconversion. In this model, the enhanced erythropoiesis coupled with preferential consumption of fatty acids from regulated marrow adipose tissue to supply energy for erythropoiesis and bone anabolism may explain the lumbar vertebrae reconversion. These results will help interpreting bone marrow signal in ambulatory patients after long periods of bed rest.

LINC00968 promotes osteogenic differentiation in vitro and bone formation in vivo via regulation of miR-3658/RUNX2

Osteogenic differentiation of dental pulp stem cells (DPSCs) is considered as a promising strategy in posterior maxilla tooth implantation. Information on the function and mechanisms of long non-coding RNAs (lncRNAs) in osteogenic differentiation of DPSCs is growing, however, the mechanism of LINC00968 and miR-3658 in regulating osteogenic differentiation of DPSCs still needs to be explored. In this study, the LINC00968 and miR-3658 expression level was upregulated and downregulated in DPSCs and peri-implantitis DPSCs (pDPSCs) treated with bone morphogenic protein (BMP)2, respectively. Moreover, the effects of LINC00968 and miR-3658 on BMP2-induced osteogenic differentiation of DPSCs in vitro using Alizarin Red S staining, alkaline phosphatase (ALP) activity, quantitative real time PCR and Western blot assays showed that overexpression of LINC00968 significantly promoted mineralized bone matrix, alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2), and osterix (OSX) expression levels for osteogenic differentiation of DPSCs and pDPSCs; and overexpression of miR-3658 showed an opposite result that inhibited osteogenic differentiation of DPSCs and pDPSCs. Luciferase reporter assay showed that luciferase activities of LINC00968-WT reporter and RUNX2-WT reporter were strongly suppressed by miR-3658 overexpression. In addition, the miR-3658 upregulation interfered ectopic bone formation in vivo stimulated by LINC00968. In general, we had identified a novel molecular pathway involving LINC00968/miR-3658/RUNX2 during DPSCs and pDPSCs differentiation into osteoblasts, which might facilitate bone anabolism.

Tissue Engineering and Regeneration Therapy: A Boon for Periodontal Research

Complete regeneration of the periodontium is the aim of any successful periodontal treatment. RecentTechnologies have come up with many advances like cell-based therapy, peptides, genetic transfer,scaffolds, bone anabolics, and above all lasers. Four basic elements are required for periodontal repair andregeneration that includes adequate blood supply and wound stability, a source of bone and ligament formingcells, a supporting scaffold, and growth factors to regulate cell migration, synthesis and angiogenesis forrevascularization of the site. The newer technologies not only offer unique opportunities but also enhancethe predictability of regenerative procedures.

Glutamine metabolism in osteoprogenitors governs bone mass accrual and PTH-induced bone anabolism

Glutamine metabolism in osteoprogenitors governs bone mass accrual and PTH-induced bone anabolism

PTH-induced bone anabolism promotes systemic breast cancer growth and metastasis

PTH-induced bone anabolism promotes systemic breast cancer growth and metastasis

A Neutralizing Antibody Targeting Oxidized Phospholipids Promotes Bone Anabolism in Chow-Fed Young Adult Mice.

ABSTRACTOxidized phospholipids containing phosphocholine (OxPL) are pro‐inflammatory lipid peroxidation products that bind to scavenger receptors (SRs), such as Scarb1, and toll‐like receptors (TLRs). Excessive OxPL, as found in oxidized low‐density lipoprotein (OxLDL), overwhelm these defense mechanisms and become pathogenic in atherosclerosis, nonalcoholic steatohepatitis (NASH), and osteoporosis. We previously reported that the innate IgM natural antibody E06 binds to OxPL and neutralizes their deleterious effects; expression of the single‐chain (scFv) form of the antigen‐binding domain of E06 (E06‐scFv) as a transgene increases trabecular bone in male mice. We show herein that E06‐scFv increases trabecular and cortical bone in female and male mice by increasing bone formation and decreasing osteoblast apoptosis in vivo. Homozygous E06‐scFv mice have higher bone mass than hemizygous, showing a dose effect of the transgene. To investigate how OxPL restrain bone formation under physiologic conditions, we measured the levels of SRs and TLRs that bind OxPL. We found that osteoblastic cells primarily express Scarb1. Moreover, OxLDL‐induced apoptosis and reduced differentiation were prevented in bone marrow–derived or calvaria‐derived osteoblasts from Scarb1 knockout mice. Because Scarb1‐deficient mice are reported to have high bone mass, our results suggest that E06 may promote bone anabolism in healthy young mice, at least in part, by neutralizing OxPL, which in turn promote Scarb1‐mediated apoptosis of osteoblasts or osteoblast precursors. © 2020 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR)..

Gut Microbiota Manipulation Promotes Bone Homeostasis in Obese Mice Through Regulatory T-Cell Differentiation

A better understanding of the role of the gut microbiome in regulating skeletal metabolism has recently been reported. Manipulation of the gut microbiome with probiotics can prevent pathologic bone loss. However, the effects of probiotics/gut microbiota on immunomodulatory effect and skeletal homeostasis in the high fat diet (HFD)-linked obesity remains to be explored. Here, we examined the impact of supplementation with probiotics (VSL#3) on immune function and bone homeostasis in HFD mice. µ-CT and mechanical testing revealed that probiotics induced a significant increase of trabecular bone volume and bone mechanical strength in obese mice. Butyrate produced in the gut following probiotics treatment, or butyrate injection directly to obese mice, prevented gut inflammation, improved gut-barrier integrity, and induced the expansion of bone marrow (BM) regulatory T (Treg) cells. Mechanistically, probiotics treatment promoted the Kdm6b/Jmjd3 histone demethylase dependent Tfam promoter demethylation by inhibiting the H3K27me3 epigenetic mark in Treg cells, which increases Tfam expression. Reduced Tfam expression in Treg cells, were found to secrete greater concentration of mtDNA and modulate paracrine signaling. BMMSCs expressed DEC205 that bound to mtDNA and contributed to BMMSCs inflammation. Further, Tfam-transgenic mice, fed with HFD, prevented the obesity-linked reduction in Treg cells and mtDNA-DEC205 dependent BMMSC inflammation. Moreover, fecal transplantation from healthy, probiotic-treated donor mice is sufficient to improve gut barrier permeability and bone loss, suggesting that the probiotics mediated changes in the gut microbiome and may be a novel agent to regulate bone anabolism via the gut-immune axis.

Frequent administration of abaloparatide shows greater gains in bone anabolic window and bone mineral density in mice: A comparison with teriparatide.

Abaloparatide (ABL) is a novel 34-amino acid peptide analog of parathyroid hormone-related protein. In clinical studies, although ABL showed a greater bone mineral density (BMD) increase than teriparatide (TPTD, human parathyroid hormone 1-34), the responses of ABL to bone formation and resorption markers were weaker, making it difficult to understand the relationship between the bone anabolic window (increase in bone formation versus resorption) and bone mass. In the present study, the effects of ABL and TPTD were compared in mice. Given that the rate of bone turnover is higher in rodents than in humans, the comparison was made with several administration regimens providing equivalent daily dosages: once daily (QD, 30μg/kg every 24h), twice daily (BID, 15μg/kg every 12h), or three times a day (TID, 10μg/kg every 8h). Frequent administration of ABL showed higher BMD with enhancement of trabecular and cortical bone mass and structures than that of TPTD, consistent with the clinical results seen with once daily administration. ABL increased bone formation marker levels more than TPTD with more frequent regimens, while bone resorption marker levels were not different between ABL and TPTD in all regimens. Analysis of bone histomorphometry and gene expression also suggested that ABL increased bone formation more than TPTD, while the effect on bone resorption was almost comparable between ABL and TPTD. The bone anabolic windows calculated from bone turnover markers indicated that ABL enhanced the anabolic windows more than TPTD, leading to a robust increase in BMD. The mechanism by which ABL showed a better balance of bone turnover was suggested to be partly due to the enhanced remodeling-based bone formation involved in Ephb4. Taken together, our findings would help elucidate the mechanism by which ABL shows excellent BMD gain and reduction of fractures in patients with osteoporosis.

Mechano-adaptive Responses of Alveolar Bone to Implant Hyper-loading in a pre-clinical in vivo model.

Oral implants transmit biting forces to peri-implant bone. In turn, those forces subject peri-implant bone to mechanical stresses and strains. Here, our objective was to understand how peri-implant bone responded to conditions of normal versus hyper-loading in a mouse model. Sixty-six mice were randomly assigned to 2 groups; both groups underwent bilateral maxillary first molar extraction followed by complete healing. Titanium alloy implants were placed in healed sites and positioned below the occlusal plane. After osseointegration, a composite crown was affixed to the implant so masticatory loading would ensue. In controls, the remaining dentition was left intact but in the hyper-loaded (test) group, the remaining molars were extracted. 3D finite element analysis (FEA) calculated peri-implant strains resulting from normal and hyper-loading. Peri-implant tissues were analyzed at multiple time points using micro-computed tomography (µCT) imaging, histology, enzymatic assays of bone remodeling, and vital dye labeling to evaluate bone accrual. Compared to controls, hyper-loaded implants experienced a 3.6-fold increase in occlusal force, producing higher peri-implant strains. Bone formation and resorption were both significantly elevated around hyper-loaded implants, eventually culminating in a significant increase in peri-implant bone volume/total volume (BV/TV). In our mouse model, masticatory hyper-loading of an osseointegrated implant was associated with increased peri-implant strain, increased peri-implant bone remodeling, and a net gain in bone deposition. Hyper-loading results in bone strain with catabolic and anabolic bone responses, leading to a net gain in bone deposition.

Uncaria tomentosa reduces osteoclastic bone loss in vivo

BackgroundThe genus Uncaria (Rubiaceae) has several biological properties significant to human health. However, the mechanisms underlying the protective effect of this plant on bone diseases are uncertain. PurposeThe present study investigated the role of Uncaria tomentosa extract (UTE) on alveolar bone loss in rats and on osteoclastogenesis in vitro. MaterialsUTE was characterized by an Acquity UPLC (Waters) system, coupled to an Electrospray Ionization (ESI) interface and Quadrupole/Flight Time (QTOF, Waters) Mass Spectrometry system (MS). The effect of UTE treatment for 11 days on the ligature-induced bone loss was assessed focusing on several aspects: macroscopic and histological analysis of bone loss, neutrophil and osteoclast infiltration, and anabolic effect. The effect of UTE on bone marrow cell differentiation to osteoclasts was assessed in vitro. ResultsThe analysis of UTE by UPLC-ESI-QTOF-MS/MS identified 24 compounds, among pentacyclic or tetracyclic oxindole alkaloids and phenols. The administration of UTE for 11 days on ligature-induced rat attenuated the periodontal attachment loss and alveolar bone resorption. It also diminished neutrophil migration to the gingiva tissue, demonstrated by a lower level of MPO. UTE treatment also decreased the level of RANKL/OPG ratio, the main osteoclast differentiation-related genes, followed by reduced TRAP-positive cell number lining the alveolar bone. Additionally, the level of bone-specific alkaline phosphatase, an anabolic bone marker, was elevated in the plasma of UTE treated rats. Next, we determined a possible direct effect of UTE on osteoclast differentiation in vitro. The incubation of primary osteoclast with UTE decreased RANKL-induced osteoclast differentiation without affecting cell viability. This effect was supported by downregulation of the nuclear factor activated T-cells, cytoplasmic 1 expression, a master regulator of osteoclast differentiation, and other osteoclast-specific activity markers, such as cathepsin K and TRAP. ConclusionUTE exhibited an effective anti-resorptive and anabolic effects, which highlight it as a potential natural product for the treatment of certain osteolytic diseases, such as periodontitis.

Bone-Targeting Systems to Systemically Deliver Therapeutics to Bone Fractures for Accelerated Healing.

Compared with the current standard of implanting bone anabolics for fracture repair, bone fracture-targeted anabolics would be more effective, less invasive, and less toxic and would allow for control over what phase of fracture healing is being affected. We therefore sought to identify the optimal bone-targeting molecule to allow for systemic administration of therapeutics to bone fractures. We found that many bone-targeting molecules exist, but most have been developed for the treatment of bone cancers, osteomyelitis, or osteoporosis. There are a few examples of bone-targeting ligands that have been developed for bone fractures that are selective for the bone fracture over the body and skeleton. Acidic oligopeptides have the ideal half-life, toxicity profile, and selectivity for a bone fracture-targeting ligand and are the most developed and promising of these bone fracture-targeting ligands. However, many other promising ligands have been developed that could be used for bone fractures.

Lactate Dehydrogenase Inhibition With Oxamate Exerts Bone Anabolic Effect.

Cellular bioenergetics is a promising new therapeutic target in aging, cancer, and diabetes because these pathologies are characterized by a shift from oxidative to glycolytic metabolism. We have previously reported such glycolytic shift in aged bone as a major contributor to bone loss in mice. We and others also showed the importance of oxidative phosphorylation (OxPhos) for osteoblast differentiation. It is therefore reasonable to propose that stimulation of OxPhos will have bone anabolic effect. One strategy widely used in cancer research to stimulate OxPhos is inhibition of glycolysis. In this work, we aimed to evaluate the safety and efficacy of pharmacological inhibition of glycolysis to stimulate OxPhos and promote osteoblast bone-forming function and bone anabolism. We tested a range of glycolytic inhibitors including 2-deoxyglucose, dichloroacetate, 3-bromopyruvate, and oxamate. Of all the studied inhibitors, only a lactate dehydrogenase (LDH) inhibitor, oxamate, did not show any toxicity in either undifferentiated osteoprogenitors or osteoinduced cells in vitro. Oxamate stimulated both OxPhos and osteoblast differentiation in osteoprogenitors. In vivo, oxamate improved bone mineral density, cortical bone architecture, and bone biomechanical strength in both young and aged C57BL/6J male mice. Oxamate also increased bone formation by osteoblasts without affecting bone resorption. In sum, our work provided a proof of concept for the use of anti-glycolytic strategies in bone and identified a small molecule LDH inhibitor, oxamate, as a safe and efficient bone anabolic agent. © 2020 American Society for Bone and Mineral Research (ASBMR).

Bone's Response to Mechanical Loading in Aging and Osteoporosis: Molecular Mechanisms.

Mechanotransduction is pivotal in the maintenance of homeostasis in different tissues and involves multiple cell signaling pathways. In bone, mechanical stimuli regulate the balance between bone formation and resorption; osteocytes play a central role in this regulation. Dysfunctions in mechanotransduction signaling or in osteocytes response lead to an imbalance in bone homeostasis. This alteration is very relevant in some conditions such as osteoporosis and aging. Both are characterized by increased bone weakness due to different causes, for example, the increase of osteocyte apoptosis that cause an alteration of fluid space, or the alteration of molecular pathways. There are intertwined yet very different mechanisms involved among the cell-intrinsic effects of aging on bone, the cell-intrinsic and tissue-level effects of estrogen/androgen withdrawal on bone, and the effects of reduced mechanical loading on bone, which are all involved to some degree in how aged bone fails to respond properly to stress/strain compared to younger bone. This review aims at clarifying how the cellular and molecular pathways regulated and induced in bone by mechanical stimulation are altered with aging and in osteoporosis, to highlight new possible targets for antiresorptive or anabolic bone therapeutic approaches.

Mimicking the Organic and Inorganic Composition of Anabolic Bone Enhances Human Mesenchymal Stem Cell Osteoinduction and Scaffold Mechanical Properties.

Engineered bone graft designs have been largely inspired by adult bone despite functionally significant differences from the composition of anabolic bone in both the mineralized and non-mineralized fractions. Specifically, anabolic bone contains hydroxyapatite with ionic substitutions that facilitate bone turnover and relatively rare collagens type VI and XII that are important for normal bone development. In this work, human mesenchymal stem cells (hMSCs) were cultured in lyophilized collagen type I scaffolds mineralized with hydroxyapatite containing Mg2+ substitutions, then induced to deposit an extracellular matrix (ECM) containing collagens VI and XII by exposure to GW9662, a PPARγ inhibitor. Delivery of GW9662 was accomplished through either Supplemented Media or via composite microspheres embedded in the scaffolds for localized delivery. Furthermore, hMSCs and scaffolds were cultured in both static and perfuse conditions to investigate the interaction between GW9662 treatment and perfusion and their effects on ECM deposition trends. Perfusion culture enhanced cell infiltration into the scaffold, deposition of collagen VI and XII, as well as osteogenic differentiation, as determined by gene expression of osteopontin, BMP2, and ALP. Furthermore, scaffold mineral density and compressive modulus were increased in response to both GW9662 treatment and perfusion after 3 weeks of culture. Local delivery of GW9662 with drug-eluting microspheres had comparable effects to systemic delivery in the perfusate. Together, these results demonstrate a strategy to create a scaffold mimicking both organic and inorganic characteristics of anabolic bone and its potential as a bone graft.

Characterization of a pluripotent stem cell-derived matrix with powerful osteoregenerative capabilities

Approximately 10% of fractures will not heal without intervention. Current treatments can be marginally effective, costly, and some have adverse effects. A safe and manufacturable mimic of anabolic bone is the primary goal of bone engineering, but achieving this is challenging. Mesenchymal stem cells (MSCs), are excellent candidates for engineering bone, but lack reproducibility due to donor source and culture methodology. The need for a bioactive attachment substrate also hinders progress. Herein, we describe a highly osteogenic MSC line generated from induced pluripotent stem cells that generates high yields of an osteogenic cell-matrix (ihOCM) in vitro. In mice, the intrinsic osteogenic activity of ihOCM surpasses bone morphogenic protein 2 (BMP2) driving healing of calvarial defects in 4 weeks by a mechanism mediated in part by collagen VI and XII. We propose that ihOCM may represent an effective replacement for autograft and BMP products used commonly in bone tissue engineering.