Bone Morphogenic Protein Research Articles

Characterization of the osteogenic differentiation capacity of human bone cells on hybrid β-TCP/ZrO2 structures

This study deals with synthetic biomaterials that are biocompatible and mechanically stable to serve as a potential bone graft for improved mandibular reconstruction. To fabricate novel hybrid scaffolds containing beta-tricalcium phosphate (β-TCP) and zirconia (ZrO2), 3D printing was combined with Freeze Foaming to achieve a certain porosity, mimicking cancellous bone. The aim was to characterize the differentiation capacity of human pre-osteoblasts on the hybrid scaffolds. Although a good biocompatibility was demonstrated for the tested components of the hybrid scaffold, pure β-TCP foams showed the best results regarding osteogenic differentiation and pro-inflammatory processes. To enhance the osteoinductive properties of the β-TCP foam, the structures were also biofunctionalized with bone morphogenic protein 2 (BMP–2) and its effect was analyzed either on single cell cultures of pre-osteoblasts or pre-osteoblasts directly co-cultured with human peripheral blood mononuclear cells (PBMCs). The latter served to analyze the induction of bone remodelling processes. The immobilization of BMP-2 on scaffolds and its biological accessibility could be demonstrated, however, the biofunctionalization did not result in an enhanced differentiation capacity and bone remodeling processes of either pre-osteoblasts or directly co-cultured pre-osteoblasts and PBMCs.

Single-cell RNA sequencing analysis identifies one subpopulation of endothelial cells that proliferates and another that undergoes the endothelial-mesenchymal transition in regenerating pig hearts.

Background: In our previous work, we demonstrated that when newborn pigs undergo apical resection (AR) on postnatal day 1 (P1), the animals' hearts were completely recover from a myocardial infarction (MI) that occurs on postnatal day 28 (P28); single-nucleus RNA sequencing (snRNAseq) data suggested that this recovery was achieved by regeneration of pig cardiomyocyte subpopulations in response to MI. However, coronary vasculature also has a key role in promoting cardiac repair. Method: Thus, in this report, we used autoencoder algorithms to analyze snRNAseq data from endothelial cells (ECs) in the hearts of the same animals. Main results: Our results identified five EC clusters, three composed of vascular ECs (VEC1-3) and two containing lymphatic ECs (LEC1-2). Cells from VEC1 expressed elevated levels of each of five cell-cyclespecific markers (Aurora Kinase B [AURKB], Marker of Proliferation Ki-67 [MKI67], Inner Centromere Protein [INCENP], Survivin [BIRC5], and Borealin [CDCA8]), as well as a number of transcription factors that promote EC proliferation, while (VEC3 was enriched for genes that regulate intercellular junctions, participate in transforming growth factor β (TGFβ), bone morphogenic protein (BMP) signaling, and promote the endothelial mesenchymal transition (EndMT). The remaining VEC2 did not appear to participate directly in the angiogenic response to MI, but trajectory analyses indicated that it may serve as a reservoir for the generation of VEC1 and VEC3 ECs in response to MI. Notably, only the VEC3 cluster was more populous in regenerating (i.e., ARP1MIP28) than non-regenerating (i.e., MIP28) hearts during the 1-week period after MI induction, which suggests that further investigation of the VEC3 cluster could identify new targets for improving myocardial recovery after MI. Histological analysis of KI67 and EndMT marker PDGFRA demonstrated that while the expression of proliferation of endothelial cells was not significantly different, expression of EndMT markers was significantly higher among endothelial cells of ARP1MIP28 hearts compared to MIP28 hearts, which were consistent with snRNAseq analysis of clusters VEC1 and VEC3. Furthermore, upregulated secrete genes by VEC3 may promote cardiomyocyte proliferation via the Pi3k-Akt and ERBB signaling pathways, which directly contribute to cardiac muscle regeneration. Conclusion: In regenerative heart, endothelial cells may express EndMT markers, and this process could contribute to regeneration via a endothelial-cardiomyocyte crosstalk that supports cardiomyocyte proliferation.

Signalling interaction between β-catenin and other signalling molecules during osteoarthritis development.

Osteoarthritis (OA) is the most prevalent disorder of synovial joint affecting multiple joints. In the past decade, we have witnessed conceptual switch of OA pathogenesis from a 'wear and tear' disease to a disease affecting entire joint. Extensive studies have been conducted to understand the underlying mechanisms of OA using genetic mouse models and ex vivo joint tissues derived from individuals with OA. These studies revealed that multiple signalling pathways are involved in OA development, including the canonical Wnt/β-catenin signalling and its interaction with other signalling pathways, such as transforming growth factor β (TGF-β), bone morphogenic protein (BMP), Indian Hedgehog (Ihh), nuclear factor κB (NF-κB), fibroblast growth factor (FGF), and Notch. The identification of signalling interaction and underlying mechanisms are currently underway and the specific molecule(s) and key signalling pathway(s) playing a decisive role in OA development need to be evaluated. This review will focus on recent progresses in understanding of the critical role of Wnt/β-catenin signalling in OA pathogenesis and interaction of β-catenin with other pathways, such as TGF-β, BMP, Notch, Ihh, NF-κB, and FGF. Understanding of these novel insights into the interaction of β-catenin with other pathways and its integration into a complex gene regulatory network during OA development will help us identify the key signalling pathway of OA pathogenesis leading to the discovery of novel therapeutic strategies for OA intervention.

Serum Follicular Fluid Growth Differentiation Factor 9 (GDF-9) and Bone Morphogenic Protein 15 (BMP-15) as Markers of Ovarian Reserve

Serum Follicular Fluid Growth Differentiation Factor 9 (GDF-9) and Bone Morphogenic Protein 15 (BMP-15) as Markers of Ovarian Reserve

Stiffness and BMP-2 Mimetic Peptide Jointly Regulate the Osteogenic Differentiation of Rat Bone Marrow Stromal Cells in a Gelatin Cryogel.

Both biochemical and mechanical cues could regulate the function of stem cells, but the interaction mechanism of their signaling pathway remains unclear, especially in the three-dimensional (3D) culture mode. Higher matrix stiffness promotes osteogenic differentiation of stem cells, and bone morphogenic protein-2 (BMP-2) has been clinically applied to promote bone regeneration. Here, the crosstalk of extracellular mechanical signals on BMP-2 signaling was investigated in rat bone marrow stromal cells (rMSCs) cultured inside cryogels with interconnective pores. Stiff cryogel independently promoted osteogenic differentiation and enhanced the autocrine secretion of BMP-2, thus stimulating increased phosphorylation levels of the Smad1/5/8 complex. BMP-2 mimetic peptide (BMMP) and high cryogel stiffness jointly guided the osteogenic differentiation of rMSCs. Inhibition of rho-associated kinase (ROCK) by Y-27632 or inhibition of nonmuscle myosin II (NM II) by blebbistatin showed that osteogenesis induction by BMP-2 signaling, as well as autocrine secretion of BMP-2 and phosphorylation of the Smad complex, requires the involvement of cytoskeletal tension and ROCK pathway signaling. An interconnective microporous cryogel scaffold promoted rMSC osteogenic differentiation by combining matrix stiffness and BMMP, and it accelerated critical cranial defect repair in the rat model.

Intravitreal injection of Noggin ameliorates retinal degeneration in a mouse model of retinitis pigmentosa

Purpose: To determine the therapeutic efficacy of intravitreal injections of Noggin in a rodent model of retinitis pigmentosa (RP). Noggin is expressed by Myo/Nog cells which are critical for eye morphogenesis and have been shown to be both neuroprotective and a source of contractile myofibroblasts in ocular diseases. Noggin is a bone morphogenic protein (BMP) inhibitor. BMPs play important roles in eye development but the strength of this signalling pathway must be carefully regulated. Furthermore, BMP levels increase in the central nervous system under stress and injury.Methods: Male and female C3H/HEJ mice (n = 30) exhibiting Pde6b gene mutation, considered a model of RP, were divided into 3 experimental groups (n = 10 each). At week 2.5 post birth, mice received either a bilateral, single intravitreal injection (1 μL) of phosphate buffer saline (PBS; Control 1), mouse serum albumin (MSA; Control 2), or human recombinant Noggin (25 ng; Treatment). At weeks 3 and 5, electroretinograms (ERGs) were performed to assess retinal function, and mice were sacrificed. Then, eyes were enucleated, fixed, frozen, cryosectioned, and immunolabelled to evaluate apoptosis (TUNEL), Myo/Nog cells (BAI1), and neuroretinal thickness.Results: Apoptosis was significantly decreased in the Noggin treated group compared to controls (PBS and MSA) at both time points. Noggin treatment also resulted in increased thickness of the neuroretina. The number of Noggin producing Myo/Nog cells was higher in the retinas of all the control groups compared to the Noggin‐injected group. ERGs showed significant improvement of the A‐ and B‐waves in Noggin injected mice when compared to control groups at 3‐ and 5‐weeks post‐birth.Conclusions: Intravitreal injection of Noggin in a RP rodent model improved retinal function and decreased apoptosis and retinal thinning. Myo/Nog cells increase in number in response to stress and injury. The decrease in the number of Myo/Nog cells in response to Noggin treatment further supports the conclusion that this BMP inhibitor reduced tissue damage. These results suggest that inhibition of BMP signalling may be a therapeutic approach to reducing retinal degeneration in RP.

LAPONITE® nano-silicates potentiate the angiogenic effects of FG-4592 and osteogenic effects of BMP-2.

LAPONITE®-based drug delivery systems offer many advantages due to the unique ionic and physical properties of LAPONITE®. The high ionicity and large surface area of LAPONITE® nanoparticles enable the intercalation and dissolution of biomolecules. In this study, we explored the potential of LAPONITE® as a carrier for FG-4592 to support angiogenesis and as a carrier for bone morphogenic protein-2 (BMP-2) to support osteogenesis. Interestingly, we found that LAPONITE® promoted the FG-4592 induced upregulation of vascular endothelial growth factor (VEGF) gene expression of human umbilical cord endothelial cells (HUVECs). Additionally, we observed that LAPONITE® could provide a sustained release of BMP-2 and significantly potentiate the osteogenic effects of BMP-2 on adipose derived mesenchymal stem cells (AMSCs). Overall, current findings on the LAPONITE®-drug/protein model system provide a unique way to potentiate the angiogenic activities of FG-4592 on HUVECs and osteogenic effects of BMP-2 on AMSCs for tissue engineering application. Future studies will be directed towards gaining a deeper understanding of these effects on a co-culture system of HUVECs and AMSCs.

Improvements in Quality, Safety and Costs Associated with Use of Implant Registries Within a Health System

Clinical quality registries (CQRs) are intended to enhance quality, safety, and cost reduction using real-world data for a self-improving health system. Starting in 2001, Kaiser Permanente established several medical device CQRs as a quality improvement initiative. This report examines the contributions of these CQRs on improvement in health outcomes, changes in clinical practice, and cost-effectiveness over the past 20 years. Eight implant registries were instituted with standardized collection from the electronic health record and other institutional data sources of patient characteristics, medical comorbidities, implant attributes, procedure details, surgical techniques, and outcomes (including complications, revisions, reoperations, hospital readmissions, and other utilization measures). A rigorous quality control system is in place to improve and maintain the quality of data. Data from the Implant Registries form the basis for multiple quality improvement and patient safety initiatives to minimize variation in care, promote clinical best practices, facilitate recalls, perform benchmarking, identify patients at risk, and construct reports about individual surgeons. Following the inception of the Implant Registries, there was an observed (1) reduction in opioid utilization following orthopedic procedures, (2) reduction in use of bone morphogenic protein during lumbar fusion allowing for cost savings, (3) reduction in allograft for anterior cruciate ligament reconstruction and subsequent decrease in organizationwide revision rates, (4) cost savings through expansion of same-day discharge programs for joint arthroplasty, (5) increase in the use of cement fixation in the hemiarthroplasty treatment of hip fracture, and (6) organizationwide discontinuation of an endograft device associated with a higher risk for adverse outcomes following endovascular aortic aneurysm repair. The use of Implant Registries within our health system, along with clinical leadership and organizational commitment to a learning health system, was associated with improved quality and safety outcomes and reduced costs. The exact mechanisms by which such registries affect health outcomes and costs require further study.

The Combined Effects of RhBMP-2 and Systemic RANKL Inhibitor in Patients With Bone Density Loss Undergoing Posterior Lumbar Interbody Fusion: A Retrospective Observational Analysis With Propensity Score Matching.

The risks of nonunion and subsidence are high in patients with bone density loss undergoing spinal fusion surgery. The internal application of recombinant human bone morphogenic protein 2 (rhBMP-2) in an interbody cage improves spinal fusion; however, related complications have been reported. Denosumab, a human monoclonal antibody targeting the receptor activator of nuclear factor kappa B ligand (RANKL), hinders osteoblast differentiation and function. Therefore, this study aimed to observe the combined effect of the local application of rhBMP-2 in a lumbar cage and systemic RANKL inhibition on postoperative spinal fusion in patients with bone density loss undergoing posterior lumbar interbody fusion (PLIF). This retrospective observational study included 251 consecutive patients with spinal stenosis who underwent PLIF at a single center between 2017 and 2021. Clinical outcomes were assessed, and radiographic evaluations included lumbar flexion, extension, range of motion, and subsidence. Statistical analyses were conducted to identify the combined effect of the treatment and the subsidence and spinal fusion status. One hundred patients were included in the final analysis. Denosumab treatment significantly reduced the rate of osteolysis (p = 0.013). When denosumab was administered in combination with rhBMP-2, the fusion status remained similar; however, the incidences of postoperative osteolysis and postoperative oozing day decreased. The combined use of rhBMP-2 and RANKL inhibition in patients with bone density loss can enhance bone formation after PLIF with fewer complications than rhBMP-2 alone.

ACVR1: A Novel Therapeutic Target to Treat Anemia in Myelofibrosis.

Activin receptor type I (ACVR1) is a transmembrane kinase receptor belonging to bone morphogenic protein receptors (BMPs). ACVR1 plays an important role in hematopoiesis and anemia via the BMP6/ACVR1/SMAD pathway, which regulates expression of hepcidin, the master regulator of iron homeostasis. Elevated hepcidin levels are inversely associated with plasma iron levels, and chronic hepcidin expression leads to iron-restricted anemia. Anemia is one of the hallmarks of myelofibrosis (MF), a bone marrow (BM) malignancy characterized by BM scarring resulting in impaired hematopoiesis, splenomegaly, and systemic symptoms. Anemia and red blood cell transfusions negatively impact MF prognosis. Among the approved JAK inhibitors (ruxolitinib, fedratinib, momelotinib, and pacritinib) for MF, momelotinib and pacritinib are preferably used in cytopenic patients; both agents are potent ACVR1 inhibitors that suppress hepcidin expression via the BMP6/ACVR1/SMAD pathway and restore iron homeostasis/erythropoiesis. In September 2023, momelotinib was approved as a treatment for patients with MF and anemia. Zilurgisertib (ACVR1 inhibitor) and DISC-0974 (anti-hemojuvelin monoclonal antibody) are evaluated in early phase clinical trials in patients with MF and anemia. Luspatercept (ACVR2B ligand trap) is assessed in transfusion-dependent MF patients in a registrational phase 3 trial. Approved ACVR1 inhibitors and novel agents in development are poised to improve the outcomes of anemic MF patients.

Bone Regeneration Potential of Periodontal Ligament Stem Cells in Combination with Cold Atmospheric Plasma-Pretreated Beta-Tricalcium Phosphate: An In Vivo Assessment

In regenerative bone tissue medicine, combining artificial bone substitutes with progenitor cells is a prospective approach. Surface modification via cold atmospheric plasma (CAP) enhances biomaterial–cell interactions, which are crucial for successful bone regeneration. Using a rabbit calvarial critical-size defect model, we assessed the use of CAP-pretreated beta-tricalcium phosphate (β-TCP), alone or with periodontal ligament stem cells (PDLSCs), for bone regeneration. Histological and histomorphometric analyses at two and four weeks revealed significantly improved bone regeneration and reduced inflammation in the CAP-treated β-TCP with PDLSCs compared to β-TCP alone. Immunohistochemical analysis also showed an increase in the bone healing markers, including bone morphogenic proteins 2 and 4, runt-related transcription factor 2, collagen-1, and osteonectin, after two and four weeks in the CAP-treated β-TCP implants with PDLSC. This in vivo study demonstrates for the first time the superior bone regenerative capacity of CAP-pretreated β-TCP seeded with PDLSCs, highlighting the therapeutic potential of this combined approach in osteoregeneration.

The influence of Simvastatin carried by Chitosan nanoparticle on bone regeneration using Masson’s Trichrome histochemical stain

Background: Due to the complicated and time-consuming physiological procedure of bone healing, certain graft materials have been frequently used to enhance the reconstruction of the normal bone architecture. However, owing to the limitations of these graft materials, some pharmaceutical alternatives are considered instead. Chitosan is a biopolymer with many distinguishing characteristics that make it one of the best materials to be used as a drug delivery system for simvastatin. Simvastatin is a cholesterol lowering drug, and an influencer in bone formation process, because it stimulates osteoblasts differentiation, bone morphogenic protein 2, and vascular endothelial growth factor. Objectives: histological, histochemical and histomorphometrical analyses were carried out to evaluate the effect of local application of chitosan simvastatin nanoparticles (ChSimN) on bone healing. Materials and Methods: New Zealand rabbits (n=14) were used in this study. Two defects were made: one on the right side (the experimental side) that received ChSimN and the other one on the left side (the control side), which left to heal spontaneously. Seven rabbits were sacrificed after 2 weeks of the experiments, while the others after 4 weeks. Bone samples were taken for histological and histomorphometric study after the sacrifice. Results: The histological study, using both H&E and Masson’s Trichrome stain, revealed that the ChSimN group recorded an increased amount of bone formation at both time points. Histomorphometrical analysis recorded a significant increment in bone marrow and trabecular areas in the ChSimN group. Conclusion: ChSimN had a pronounced effect on bone formation.

Role of endothelial PDGFB in arterio-venous malformations pathogenesis

Arterial-venous malformations (AVMs) are direct connections between arteries and veins without an intervening capillary bed. Either familial inherited or sporadically occurring, localized pericytes (PCs) drop is among the AVMs’ hallmarks. Whether impaired PC coverage triggers AVMs or it is a secondary event is unclear. Here we evaluated the role of the master regulator of PC recruitment, Platelet derived growth factor B (PDGFB) in AVM pathogenesis. Using tamoxifen-inducible deletion of Pdgfb in endothelial cells (ECs), we show that disruption of EC Pdgfb-mediated PC recruitment and maintenance leads to capillary enlargement and organotypic AVM-like structures. These vascular lesions contain non-proliferative hyperplastic, hypertrophic and miss-oriented capillary ECs with an altered capillary EC fate identity. Mechanistically, we propose that PDGFB maintains capillary EC size and caliber to limit hemodynamic changes, thus restricting expression of Krüppel like factor 4 and activation of Bone morphogenic protein, Transforming growth factor β and NOTCH signaling in ECs. Furthermore, our study emphasizes that inducing or activating PDGFB signaling may be a viable therapeutic approach for treating vascular malformations.

Suppression of BMP signaling restores mitral cell development impaired by FGF signaling deficits in mouse olfactory bulb

Fibroblast growth factors (FGFs) and bone morphogenic proteins (BMPs) play various important roles in the development of the central nervous system. However, the roles of FGF and BMP signaling in the development of the olfactory bulb (OB) are largely unknown. In this study, we first showed the expression of FGF receptors (FGFRs) and BMP receptors (BMPRs) in OB RGCs, radial glial cells (RGCs) in the developing OB, which generate the OB projection neurons, mitral and tufted cells. When the FGF signaling was inhibited by a dominant-negative form of FGFR1 (dnFGFR1), OB RGCs accelerated their state transition to mitral cell precursors without affecting their transcription cascade and fate. However, the mitral cell precursors could not radially migrate to form the mitral cell layer (MCL). In addition, FGF signaling inhibition reduced the expression of a BMP antagonist, Noggin, in the developing OB. When BMP signaling was suppressed by the ectopic expression of Noggin or a dominant-negative form of BMPR1a (dnBMPR1a) in the developing OB, the defect in MCL formation caused by the dnFGFR1 was rescued. However, the dnBMPR1a did not rescue the accelerated state transition of OB RGCs. These results demonstrate that FGF signaling is important for OB RGCs to maintain their self-renewal state and MCL formation. Moreover, the suppression of BMP signaling is required for mitral cells to form the MCL. This study sheds new light on the roles of FGFs and BMPs in OB development.

Age‐Related Enhanced BMP Signaling Elicits APOE‐Isoform Dependent Effects in IPSC Model of Alzheimer’s Disease

AbstractBackgroundAging is the strongest risk factor for developing Alzheimer’s Disease (AD), and APOE is the largest genetic risk factor for developing AD. Bone morphogenic protein 4 (BMP4) expression and subsequent signaling increases 10‐fold with age and is increased in AD patients compared to age‐matched controls. Enhanced BMP signaling in aged and AD brains suggest a potential molecular cascade involved in AD pathogenesis. A potential interaction exists between increased BMP signaling across lifespan APOE expression that contribute to risk of AD.MethodAD patient derived iPSC’s (APOE3/E4 genotype) and isogenic controls (APOE3/E3 genotype) were differentiated into astrocytes and neurons. Cells were grown for 35 days before treatment with BMP4 or vehicle for 3 days prior to fixation, protein/media collection, or preparation for single cell RNA sequencing (scRNAseq). For coculture studies, neurons and astrocytes of the same patient background were differentiated separately prior to combined plating and BMP treatment. Cocultures were in matched (APOE3/E4 neurons with APOE3/E4 astrocytes) or mismatched (APOE3/E3 neurons with APOE3/E4 astrocytes) conditions to understand the non‐cell autonomous effects of APOE genotype.ResultUpon BMP treatment, APOE3/E4 compared to APOE3/E3 astrocytes exhibit accumulation of lipid droplets with changes in lipid composition. Increased free cholesterol levels as well as triglyceride accumulation is observed in APOE3/E4 astrocytes and is further exacerbated with BMP treatment. RNA‐sequencing analysis reveals marked decreased expression in genes related to oxidative phosphorylation and increased cholesterol synthesis and metabolism in APOE3/E4 astrocytes that is modulated by BMP treatment. In a non‐cell autonomous fashion, APOE3/E3 neurons, when paired with APOE3/E4 astrocytes, express exacerbated pTau levels and aberrant metabolic processes in a BMP‐dependent manner.ConclusionOur results indicate an interaction between genetic and aging factors in cellular processes relevant to AD. Enhanced BMP signaling confers aberrant lipid composition and metabolic processing in APOE3/E4 astrocytes when compared to APOE3/E3. Subsequent BMP‐dependent non‐cell autonomous effects in APOE3/E3 neurons are observed when cocultured with APOE3/E4 astrocytes. This data suggests age‐associated increase in BMP signaling may modulate APOE isoform‐dependent metabolic processes and increase risk for APOE‐mediated pathological cascades.

Inhibition of BMP signaling delays tauopathy and cognitive deficits in P301S mouse model

AbstractBackgroundAging is the highest risk factor for sporadic Alzheimer’s disease (AD). Expression of bone morphogenic protein 4 (BMP4) increases 10‐fold with age, while expression of the BMP antagonist, noggin, decreases. We have observed that activating BMP signaling in induced human neurons increased levels of phosphorylated tau. The increase of p‐tau has been correlated with the activated non‐canonical BMP pathway leading to p38 activity. We hypothesized that modulating BMP signaling in an AD mouse model of tau would rescue disease phenotype.MethodP301S (PS19) mice were injected with lentivirus overexpressing noggin (LV‐noggin) or lentivirus mcherry (LV‐mcherry) control in the dentate gyrus. Cognitive behavioral testing was run at 7 months of age and brain tissue was collected for protein and immunofluorescent imaging.ResultWe observe that levels of BMP4, as well as BMP signaling, are increased in the brains of PS19 mice compared to littermate wildtype mice. Analysis of noggin injected PS19 brains show downregulation of BMP signaling in the DG shown by decreased pSMAD1/5/8, as well as non‐canonical signaling through p38 kinase activity. Inhibition of BMP signaling through overexpression of noggin improves cognitive ability shown through Barnes maze testing. Downregulation of BMP signaling in P301S mice led to a decrease in tau phosphorylation seen both in immunoblotting, as well as immunocytochemical analysis in the DG.ConclusionPS19 model of tauopathy show increased levels of BMP signaling. We have shown that increased levels of BMP signaling lead to increased tau phosphorylation and reduced degradation of p‐tau. We show that by inhibiting BMP signaling in vivo we can delay cognitive behavioral deficits seen in these mice by decreasing tau phosphorylation. This suggests that the age‐related increase in BMP signaling may participate in the initiation and progression of AD pathology and help to explain why age is the largest risk factor for AD.

Selective Inhibition of Repulsive Guidance Molecule C/Hemojuvelin (RGMc/HJV) with SRK-256 to Mobilize Iron in Functional Iron Deficiency

Functional iron deficiency is a serious condition marked by serum iron restriction in the presence of adequate iron stores that can lead to anemia, worsening quality of life, and leading to poor patient outcomes. Functional iron deficiency is characterized by abnormally high levels of hepcidin given serum iron status; it is frequently observed in diseases with chronic inflammation, such as anemia of chronic disease, chronic kidney disease, and myelofibrosis. Current treatments to address this patient population involve non-specifically targeting hepcidin and come with risks of off-target side effects. The secretion of hepcidin from the liver hepatocytes is linked to circulating serum iron levels and is controlled by the Bone Morphogenic Protein (BMP) 2/6 signaling pathway. Repulsive Guidance Molecule C or Hemojuvelin (RGMc/HJV) is a BMP 2/6 co-receptor in liver hepatocytes. Human genetic mutations in this gene result in juvenile hemochromatosis and genetic mouse studies have demonstrated that RGMc/HJV has a specific role in maintaining systemic iron homeostasis through modulation of hepcidin expression. We have generated a highly selective, fully human antibody (SRK-256) against RGMc/HJV which binds to the target with 39 pM affinity. SRK-256 has no detectable binding to RGMa or RGMb, thus avoiding potential side effects of targeting these gene family members with key physiological roles outside of iron metabolism. A high-resolution crystal structure (2.2 Å) elucidates not only the mechanism of action of SRK-256 directly competing for BMP binding to RGMc via contacts within the BMP binding domain, but also identifies binding to regions within this domain highly unique to RGMc over RGMa and RGMb, thus supporting the selectivity of the molecule. This mechanism of action is corroborated by in vitro assay data demonstrating that SRK-256 blocks signaling by BMP 2, 4, and 6. Pharmacodynamic and pharmacokinetic data supportive of a monthly subcutaneous dosing regimen in human patients has been generated in cynomolgus macaques, where SRK-256 demonstrates a dose-proportional duration of effect on suppression of hepcidin expression and elevation of transferrin saturation and achieves ~95% bioavailability via subcutaneous dosing. Furthermore, in the rat peptidoglycan-polysaccharide (PGPS) model of anemia of chronic disease, we have shown that in contrast to treatment with erythropoiesis stimulating agent alone (EPO, serum iron 47.7 ± 9.8 µg/dL), SRK-256 can restore serum iron to normal levels (Healthy control, serum iron 231.9 ± 12.0 µg/dL; SRK-256, serum iron 248.7 ± 60.9 µg/dL; and SRK-256+EPO, serum iron 277.6 ± 92.9 µg/dL) in the context of inflammation-driven, iron-restricted anemia (Figure 1). SRK-256 represents a potent and selective potential therapy for the treatment of patients with iron-restricted anemias across diseases of chronic inflammation.

Interpenetrating alginate network as drug delivery matrix: Effects on protein stability and release.

The study investigates the rheological properties and protein release capacity of a uniform hydrogel composed of sodium alginate (SA) and poloxamer (P407). The hydrogel is prepared through the sustained release of calcium ions, resulting in a reinforced and homogeneous interpenetrating networks (IPNs) of SA and P407 polymeric chains. By adjusting the amount of crosslink agent, the hydrogel exhibites an adjustable dissolution ratio and adaptable gelling time. Moreover, the composite showed a well-structured network and superior mechanical strength, enabling the sustained release of both calcium ions and Soybean Trypsin Inhibitor (STI) protein, a model of Bone Morphogenic Protein (BMP). Importantly, the protein release kinetic can be tuned based on the SA content in the polymeric blend, highlighting the versatile nature of this hydrogel for drug delivery purposes.

Multifactorial Comparative Analysis of Platelet-Rich Plasma and Serum Prepared Using a Commercially Available Centrifugation Kit.

Background Platelet-rich plasma (PRP) is an autologous product prepared by centrifuging whole blood. PRP is reported to havehigh tissue repair potential and anti-inflammatory properties. Recently, PRP has become a potential treatment option for osteoarthritis, contributing to pain relief and locomotive improvement. However, the underlying therapeutic mechanisms and key biochemical factors in PRP remain unclear. This study aimed to estimate the major factors for tissue repair involved in PRP treatment by comparing between serum and PRP prepared from the same patients using the Luminex assay. Methodology Blood samples were collected from nine healthy volunteers, and serum and PRP were prepared. PRP was prepared using a PEAK©︎PRP SYSTEM kit of DePuy Synthes Mitek Sports Medicine (Raynham, Massachusetts, USA), which is a commercially available PRP preparation kit. The white blood cell count, hemoglobin level, and platelet count were automatically measured for both whole blood and PRP in the hospital's clinical laboratory using the XE-5000™ Automated Hematology System (Sysmex, Kobe, Japan). Comparative analysis of biological factors was then performed using the Luminex assay on serum and PRP. Results PRP was found to have significantly higher white blood cell and platelet counts and lower hemoglobin levels than whole blood. Furthermore, PRP contained significantly higher levels of various factors, includinginterleukin (IL)-1ra, IL-10, IL-13, C-C motif chemokine ligand (CCL)-2, CCL3, CCL4, CCL8, CCL13, CCL21, C-X-C motif chemokine ligand (CXCL)-10, matrixmetalloproteinase(MMP)-3, MMP-9, cluster of differentiation (CD) 40 ligand, vascular endothelial growth factor (VEGF), VEGF-C, platelet-derived growth factor (PDGF)-AB, PDGF-BB, and bone morphogenic protein (BMP)-2. Additionally, IL-1ra and IL-4 showed significant correlations with white blood cell counts in PRP, whereas VEGF had a significant correlation with platelet counts. Conclusions PRP contains various factors in higher quantities than serum. Specifically, the notable increase in the anti-inflammatory cytokine IL-1ra is suggested to play a key role as a major therapeutic mechanism of PRP.

The importance of BMPs and TGF-βs for endochondral bone repair – A longitudinal study in hip arthroplasty patients

IntroductionOsseointegration of hip implants, although a decade-long process, shows striking similarities with the four major phases of endochondral bone repair. In the current study we investigated the spatiotemporal involvement of bone morphogenic proteins (BMPs) and transforming growth factor betas (TGF-βs) throughout the process of bone repair leading to successfully osseointegrated hip implants. Materials and methodsTwenty-four patients that had undergone primary total hip arthroplasty (THA) due to one-sided osteoarthritis (OA) were investigated during a period of 18 years (Y) with repeated measurements of plasma biomarkers as well as clinical and radiological variables. All implants were clinically and radiographically well anchored throughout the follow-up. Eighty-one healthy donors divided in three gender- and age-matched groups and twenty OA patients awaiting THA, served as controls. Plasma was analyzed for BMP-1, -2, -3, -4, -6, -7 -9 and TGF-β1, -β2, -β3 by use of a high-sensitivity and wide dynamic range electrochemiluminescence technique allowing for detection of minor changes. ResultsSpatiotemporal changes during the follow-up are presented in the context of the four phases of endochondral bone repair shown in earlier studies and transposed to the current study based on similarities in biomarker responses. Phase 1: Primary proinflammatory phase lasting from surgery until day 7, Phase 2: Chondrogenic phase from day 7 until 18 months postsurgery, Phase 3: Secondary proinflammatory and cartilage remodeling phase lasting from 18 months until 7Y, Phase 4: coupled bone remodeling from 7Y until 18Y postsurgery. BMP-1 increased sharply shortly after surgery and remained significantly above healthy during the chondrocyte recruitment, proliferation, and hypertrophy phases with a subsequent return to control level at 5Y postsurgery. BMP-2 was above healthy controls before surgery and 1 day after surgery before decreasing to control level and remaining there throughout the follow-up. BMP-3 was at control level from presurgery until 6M after surgery when it increased to a peak at 2Y during the cartilage hypertrophy phase followed by a gradual decrease to control level at 10Y during the phase of bone formation. In the following, BMP-3 decreased below controls to a nadir 15Y postsurgery during coupled bone remodeling. BMP-4 was at control level from presurgery until 10Y postsurgery when it increased to a sharp peak at 15Y after surgery followed by a return to the level of healthy at 18Y. BMP-6 did not differ from healthy during the follow-up. BMP-7 was at control level from presurgery until 1Y postsurgery before gradually increasing to a peak at 10Y during the early phase of osteogenesis with a gradual return to control level at 18Y during the phase of coupled bone remodeling. BMP-9 was above OA before surgery followed by a decrease to basal level on day 1 after surgery and a renewed increase to a plateau above controls lasting from 6 W until returning to the level of healthy at 18Y postsurgery, i.e., throughout the phases of cartilage formation, cartilage hypertrophy and remodeling, bone formation and coupled bone remodeling. TGF-β1 was above controls presurgery before decreasing to baseline shortly after surgery followed by a renewed increase at 6 M to a peak at 2Y during cartilage hypertrophy/remodeling followed by a gradual return to baseline at 10Y during early osteoblastogenesis. TGF-β2 was at control level from presurgery until the phase of cartilage remodeling at 5Y when it increased sharply to a peak at 7Y with a gradual return to baseline at 18Y postsurgery. TGF-β3 remained at control level throughout the study. ConclusionThis study shows that the involvement of BMPs and TGF-βs in endochondral bone repair is a process of stepwise recruitment of individual biomarkers characterized by distinct, yet overlaping, spatiotemporal patterns that extend from the early phase of pre-chondrocyte recruitment until the late phase of coupled bone remodeling.