Osteogenic effect of poly(lactic-co-glycolic acid) microcapsules with different molecular weights encapsulating bone morphogenetic protein 2.

OBJECTIVES: This study aimed to explore the effects of bone morphogenetic protein 2 (BMP-2) encapsula-ted in poly(lactic-co-glycolic acid) (PLGA) microcapsules with different molecular weights on the osteogenic ability of osteoblasts. METHODS: PLGA microcapsules with different molecular weights (12 000, 30 000) encapsulating BMP-2, were prepared using a dual-channel microinjection pump. The morphology and structure of the microcapsules were characterized by optical microscopy and scanning electron microscopy. The sustained-release performance of the microcapsules was characterized by phosphate buffered saline immersion method. The cell compatibility of the microcapsules was detected by the Calcein-AM/PI staining and CCK-8 method. The chemotactic effect of BMP-2-encapsulated microcapsules on MC3T3-E1 cells after 48 h of treatment was detected by the Transwell assay. The alkaline phosphatase activity assay and Alizarin Red S staining were used to characterize the effect of microcapsules on the osteogenic ability of MC3T3-E1 cells. RESULTS: Both types of microcapsules with different molecular weights exhibited smooth surfaces, as well as uniform and good cell compatibility. The chemotactic effect of the 12 000 microcapsules was outstanding. The 30 000 microcapsules had a longer sustained-release time, and the initial burst release was reduced by approximately 25% compared with the 12 000 microcapsules. In addition, 30 000 microcapsules performed better in long-term osteogenesis induction than 12 000 microcapsules. CONCLUSIONS: In this study, the release of BMP-2 is regulated by adjusting the molecular weight of PLGA, and the results indicate that 30 000 microcapsules can better induce the long-term osteogenic ability of MC3T3-E1 cells.

The effects of BMP2 and the mechanisms involved in the invasion and angiogenesis of IDH1 mutant glioma cells.

PURPOSE: This study investigated the effect of an isocitrate dehydrogenase 1 (IDH1) mutation (mutIDH1) on the invasion and angiogenesis of human glioma cells. METHODS: Doxycycline was used to induce the expression of mutIDH1 in glioma cells. Transwell and wound healing assays were conducted to assess glioma cell migration and invasion. Western blotting and cell immunofluorescence were used to measure the expression levels of various proteins. The influence of bone morphogenetic protein 2 (BMP2) on invasion, angiogenesis-related factors, BMP2-related receptor expression, and changes in Smad signaling pathway-related proteins were evaluated after treatment with BMP2. Differential gene expression and reference transcription analysis were performed. RESULTS: Successful infection with recombinant lentivirus expressing mutIDH1 was demonstrated. The IDH1 mutation promoted glioma cell migration and invasion while positively regulating the expression of vascularization-related factors and BMP2-related receptors. BMP2 exhibited a positive regulatory effect on the migration, invasion, and angiogenesis of mutIDH1-glioma cells, possibly mediated by BMP2-induced alterations in Smad signaling pathway-related factors.After BMP2 treatment, the differential genes of MutIDH1-glioma cells are closely related to the regulation of cell migration and cell adhesion, especially the regulation of Smad-related proteins. KEGG analysis confirmed that it was related to BMP signaling pathway and TGF-ß signaling pathway and cell adhesion. Enrichment analysis of gene ontology and genome encyclopedia further confirmed the correlation of these pathways. CONCLUSION: Mutation of isocitrate dehydrogenase 1 promotes the migration, invasion, and angiogenesis of glioma cells, through its effects on the BMP2-driven Smad signaling pathway. In addition, BMP2 altered the transcriptional patterns of mutIDH1 glioma cells, enriching different gene loci in pathways associated with invasion, migration, and angiogenesis.

A Narrative Review on Recombinant Human Bone Morphogenetic Protein 2: Where Are We Now?

Spinal fusion is a prevalent surgical intervention for degenerative spinal diseases, with increasing demand driven by ageing populations. The coexistence of multiple chronic conditions, termed multimorbidity, often complicates surgical outcomes, making advanced bone grafts crucial for successful fusions. This paper reviews the development, clinical application, and controversies surrounding the use of recombinant human bone morphogenetic protein-2 (rhBMP-2) in spinal fusion surgeries. A comprehensive narrative review was conducted, focusing on literature from January 1980 to January 2024, sourced from PubMed and Google Scholar. Studies included those examining rhBMP-2 specifically in spinal fusion contexts, excluding other bone morphogenetic proteins (BMPs) and non-spinal applications. This review presents an overarching synopsis of rhBMP-2, its development history and clinical efficacy, the emergence of side effects, and evolving patterns of clinical use. As discussed in this review, clinical practice has adjusted usage and dosages to mitigate adverse effects, yet the need for safer delivery mechanisms persists. rhBMP-2 remains a potent osteoinductive agent with comparable fusion success, as measured by radiographic fusion and good clinical outcomes, to autologous grafts but poses unique risks. This review sets out how further research is essential to optimise the delivery of rhBMP-2 to reduce side effects. Enhanced understanding and innovation of spatio-temporal presentation relative to endogenous BMP could significantly improve patient outcomes in spinal fusion surgeries. The review contributes to the growing body of literature on the use of rhBMP-2 in spine surgery and discusses changing patterns of clinical use over time.

Glycinamide Facilitates Nanocomplex Formation and Functions Synergistically with Bone Morphogenetic Protein 2 to Promote Osteoblast Differentiation In Vitro and Bone Regeneration in a Mouse Calvarial Defect Model.

BACKGROUND: This study aimed to identify glycine analogs conducive to the formation of cell-absorbable nanocomplexes, enhancing collagen synthesis and subsequent osteogenesis in combination with BMP2 for improved bone regeneration. METHODS: Glycine and its derivatives were assessed for their effects on osteogenic differentiation in MC3T3-E1 cells and human bone marrow mesenchymal stem cells (BMSCs) under osteogenic conditions or with BMP2. Osteogenic differentiation was assessed through alkaline phosphatase staining and real-time quantitative polymerase chain reaction (RT-qPCR). Nanocomplex formation was examined via scanning electron microscopy, circular dichroism, and ultraviolet-visible spectroscopy. In vivo osteogenic effects were validated using a mouse calvarial defect model, and bone regeneration was evaluated through micro-computed tomography and histomorphometric analysis. RESULTS: Glycine, glycine methyl ester, and glycinamide significantly enhanced collagen synthesis and ALP activity in conjunction with an osteogenic medium (OSM). GA emerged as the most effective inducer of osteoblast differentiation marker genes. Combining GA with BMP2 synergistically stimulated ALP activity and the expression of osteoblast markers in both cell lines. GA readily formed nanocomplexes, facilitating cellular uptake through strong electrostatic interactions. In an in vivo calvarial defect mouse model, the GA and BMP2 combination demonstrated enhanced bone volume, bone volume/tissue volume ratio, trabecular numbers, and mature bone formation compared to other combinations. CONCLUSION: GA and BMP2 synergistically promoted in vitro osteoblast differentiation and in vivo bone regeneration through nanocomplex formation. This combination holds therapeutic promise for individuals with bone defects, showcasing its potential for clinical intervention.

Activation of the BMP2/SMAD4 signaling pathway for enhancing articular cartilage regeneration of mesenchymal stem cells utilizing chitosan/alginate nanoparticles on 3D extracellular matrix scaffold.

This study investigated the efficacy of using chitosan/alginate nanoparticles loaded with recombinant human bone morphogenetic-2 (rhBMP-2) and SMAD4 encoding plasmid to enhance the chondrogenesis of human bone marrow mesenchymal stem cells (hBM-MSCs) seeded on an extracellular matrix (ECM). The research treatments included the stem cells treated with the biological cocktail (BC), negative control (NC), hBM-MSCs with chondrogenic medium (MCM), hBM-MSCs with naked rhBMP-2 and chondrogenic medium (NB/C), and hBM-MSCs with naked rhBMP-2 and chondrogenic medium plus SMAD4 encoding plasmid transfected with polyethyleneimine (PEI) (NB/C/S/P). The cartilage differentiation was performed with real-time quantitative PCR analysis and alizarin blue staining. The data indicated that the biological cocktail (BC) exhibited significantly higher expression of cartilage-related genes compared to significant differences with MCM and negative control (NC) on chondrogenesis. In the (NB/C/S/P), the expression levels of SOX9 and COLX were lower than those in the BC group. The expression pattern of the ACAN gene was similar to COL2A1 changes suggesting that it holds promising potential for cartilage regeneration.

Tooth movement toward alveolar bone grafting with rhBMP-2 in a child with unilateral cleft lip and palate: Case report with long-term follow-up.

The secondary alveolar bone grafting procedure is typically recommended during the late mixed dentition phase, prior to the eruption of the permanent canine, in patients with cleft lip and palate. The anatomical and functional adaptations observed in the grafted area allow spontaneous migration and eruption of the adjacent maxillary canine. An alveolar bone graft can be performed using autogenous bone or recombinant human bone morphogenetic protein-2 (rhBMP-2). Employing rhBMP-2 in a collagen membrane eliminates the need for a donor site, thus reducing surgical morbidity. This paper aims to present a case involving complete orthodontic rehabilitation with a three-year follow-up of a male patient with a unilateral complete cleft lip and palate, posterior and anterior crossbite, where grafting was performed with rhBMP-2 at a single centre. Orthodontic intervention began at 8 years of age with rapid maxillary expansion, followed by facemask therapy. The alveolar bone grafting procedure was performed using rhBMP-2 in a collagen membrane, according to the surgical protocol developed by the Oslo team. Comprehensive orthodontic treatment started 15 months post bone grafting, during which the maxillary permanent lateral incisor distal to the alveolar cleft was successfully moved mesially into the grafted region. This intervention resulted in adequate occlusal and periodontal outcomes. The alveolar graft with rhBMP-2 produced adequate and stable alveolar bone formation, facilitating tooth eruption, orthodontic movement, and stability at the cleft site.

Potentiating effect of AMD3100 on bone morphogenetic protein-2 induced bone regeneration.

BACKGROUND: AMD3100, a CXCR4 antagonist, is currently prescribed for activating the mobilization of hematopoietic stem cells. Recently, AMD3100 was shown to potentiate bone morphogenetic protein-2 (BMP-2)-induced bone formation by stimulating the trafficking of mesenchymal cells. However, optimization of the strategic combination of AMD3100 and BMP-2 has not yet been clearly established. The purpose of this study was to evaluate the effect of AMD3100 on BMP-2-induced bone regeneration in vitro and in a mouse calvarial defect healing model. METHODS: In vitro osteoblastic differentiation and cell migration after sequential treatments with AMD3100 and BMP-2 were analyzed by alkaline phosphatase (ALP) activity, ALP staining, and calcium accumulation. Migration capacity was evaluated after treating mesenchymal cells with AMD3100 and/or BMP-2. A critical-size calvarial defect model was used to evaluate bone formation after sequential or continuous treatment with AMD3100 and BMP-2. The degree of bone formation in the defect was analyzed using micro-computed tomography (micro-CT) and histological staining. RESULTS: Compared with single treatment using either AMD3100 or BMP-2 alone, sequential treatment with AMD3100 followed by BMP-2 on mesenchymal cells increased osteogenic differentiation. Application of AMD3100 and subsequent BMP-2 significantly activated cell migration on mesenchymal cell than BMP-2 alone or AMD3100 alone. Micro-CT and histomorphometric analysis showed that continuous intraperitoneal (IP) injection of AMD3100 resulted significantly increased new bone formation in BMP-2 loaded scaffold in calvarial defect than control groups without AMD3100 IP injection. Additionally, both single IP injection of AMD3100 and subsequent BMP-2 injection to the scaffold in calvarial defect showed pronounced new bone formation compared to continuous BMP-2 treatment without AMD3100 treatment. CONCLUSION: Our data suggest that single or continuous injection of AMD3100 can potentiate BMP-2-induced osteoblastic differentiation and bone regeneration. This strategic combination of AMD3100 and BMP-2 may be a promising therapy for bone regeneration.

Glycosaminoglycans exhibit distinct interactions and signaling with BMP2 according to their nature and localization.

The role of glycosaminoglycans (GAGs) in modulating bone morphogenetic protein (BMP) signaling represents a recent and underexplored area. Conflicting reports suggest a dual effect: some indicate a positive influence, while others demonstrate a negative impact. This duality suggests that the localization of GAGs (either at the cell surface or within the extracellular matrix) or the specific type of GAG may dictate their signaling role. The precise sulfation patterns of heparan sulfate (HS) responsible for BMP2 binding remain elusive. BMP2 exhibits a preference for binding to HS over other GAGs. Using well-characterized biomaterials mimicking the extracellular matrix, our research reveals that HS promotes BMP2 signaling in the extracellular space, contrary to chondroitin sulfate (CS), which enhances BMP2 bioactivity at the cell surface. Further observations indicate that a central IdoA (2S)-GlcNS (6S) tri-sulfated motif within HS hexasaccharides enhances binding. Nevertheless, BMP2 exhibits a degree of adaptability to various HS sulfation types and sequences. Molecular dynamic simulations attribute this adaptability to the BMP2 N-terminal end flexibility. Our findings illustrate the complex interplay between GAGs and BMP signaling, highlighting the importance of localization and specific sulfation patterns. This understanding has implications for the development of biomaterials with tailored properties for therapeutic applications targeting BMP signaling pathways.

Chitosan and hyaluronic acid based injectable dual network hydrogels - Mediating antimicrobial and inflammatory modulation to promote healing of infected bone defects.

In bone defects, infections lead to excessive inflammation, increased bacterial, and bone lysis, resulting in irregular wounds that hinder new bone regeneration. Injectable bioactive materials with adequate antimicrobial activity and strong osteogenic potential are urgently required to remedy irregular defects, eradicate bacteria, and facilitate the generation of new bone tissue. In this research, injectable dual-network composite hydrogels consisting of sulfated chitosan, oxidized hyaluronic acid, ß-sodium glycerophosphate, and CuSr doped mesoporous bioactive glass loaded with bone morphogenetic protein (CuSrMBGBMP-2) were utilized for the first time to treat infectious bone defects. Initially, the hydrogel was injected into the wound at 37 °C with minimal invasion to establish a stable state and prevent hydrogel loss. Subsequently, sulfated chitosan eliminated bacteria at the wound site and facilitated cell proliferation with oxidized hyaluronic acid. Additionally, CuSrMBGBMP-2 strengthened antibacterial properties, regulated inflammatory reactions, promoted angiogenesis and osteogenic differentiation, addressing the deficiency in late-stage osteogenesis. Specifically, the injectable dual-network hydrogel based on chitosan and hyaluronic acid is minimally invasive, offering antibacterial, anti-inflammatory, pro-angiogenic, and bone regeneration properties. Therefore, this hydrogel with injectable dual network properties holds great promise for the treatment of bone infections in the future.

Development of gelatin-methacryloyl composite carriers for bone morphogenetic Protein-2 delivery: A potential strategy for spinal fusion.

To reduce the risk of nonunion after spinal fusion surgery, the in situ transplantation of bone marrow mesenchymal stem cells (BMSCs) induced toward osteogenic differentiation by bone morphogenetic protein-2 (BMP2) has been proven effective. However, the current biological agents used for transplantation have limitations, such as a short half-life and low bioavailability. To address this, our study utilized a safe and effective gelatin-methacryloyl (GelMA) as a carrier for BMP2. In vitro, experiments were conducted to observe the ability of this composite vehicle to induce osteogenic differentiation of BMSCs. The results showed that the GelMA hydrogel, with its critical properties and controlled release performance of BMP2, exhibited a slow release of BMP2 over 30 days. Moreover, the GelMA hydrogel not only enhanced the proliferation activity of BMSCs but also significantly promoted their osteogenic differentiation ability, surpassing the BMP2 effects. To investigate the potential of the GelMA-BMP2 composite vehicle, a rabbit model was employed to explore its ability to induce in situ intervertebral fusion by BMSCs. Transplantation experiments in rabbits demonstrated the effective induction of intervertebral bone fusion by the GelMA-BMP2-BMSC composite vehicle. In conclusion, the GelMA-BMP2-BMSC composite vehicle shows promising prospects in preclinical translational therapy for spinal intervertebral fusion. It addresses the limitations of current biological agents and offers a controlled release of BMP2, enhancing the proliferation and osteogenic differentiation of BMSCs.

Fibronectin/α5 Integrin Contribute to Hypertension-Associated Arterial Ageing and Calcification through Affecting BMP2/MGP Imbalance and Enhancing Vascular Smooth Muscle Cell Phenotypic Transformation.

INTRODUCTION: Hypertension can accelerate and aggravate the process of arterial ageing and calcification. However, the mechanism behind has yet to be well elucidated. METHODS: Here, we monitored the dynamic changes of fibronectin (FN)/α5 integrin, bone morphogenetic protein 2/matrix Gla protein (BMP2/MGP), and Runx2 in the aorta of spontaneously hypertensive rats (SHRs) and thoracic aortic vascular smooth muscle cells (VSMCs), also the phenotypic transformation of VSMCs during the process of arterial ageing and calcification. Further, study on arterial ageing and calcification through antagonist experiments at the molecular level was explored. RESULTS: We found extracellular FN and its α5 integrin receptor expressions were positively associated with arterial ageing and calcification in SHR during ageing, as well in VSMCs from SHR in vitro. Integrin receptor inhibitor of GRGDSP would delay this arterial ageing and calcification process. Moreover, the elevated FN and α5 integrin receptor expression evoked the disequilibrium of BMP2/MGP, where the expression of BMP2, a potent osteogenic inducer, increased while MGP, a calcification inhibitor, decreased. Furthermore, it was followed by the upregulation of Runx2 and the phenotypic transformation of VSMCs from the contractile phenotype into the osteoblast-like cells. Notably, BMP2 antagonist of rmNoggin was sufficient to ameliorate the ageing and calcification process of VSMCs and exogenous BMP2-adding accelerate and aggregate the process. CONCLUSION: Our study revealed that hypertension-associated arterial ageing and calcification might be a consequence that hypertension up-regulated FN and its high binding affinity integrin α5 receptor in the aortic wall, which in turn aggravated the imbalance of BMP2/MGP, promoted the transcription of Runx2, and induced the phenotypic transformation of VSMCs from the contractile phenotype into the osteoblast-like cells. Our study would provide insights into hypertension-associated arterial ageing and calcification and shed new light on the control of arterial calcification, especially for those with hypertension.

Inflow Tract Development.

The development of the inflow tract is undoubtedly one of the most complex remodeling events in the formation of the four-chambered heart. It involves the creation of two separate atrial chambers, the formation of an atrial/atrioventricular (AV) septal complex, the incorporation of the caval veins and coronary sinus into the right atrium, and the remodeling events that result in pulmonary venous return draining into the left atrium. In these processes, the atrioventricular mesenchymal complex, consisting of the major atrioventricular (AV) cushions, the mesenchymal cap on the primary atrial septum (pAS), and the dorsal mesenchymal protrusion (DMP), plays a crucial role.

Regulating the bioactivity of non-glycosylated recombinant human bone morphogenetic protein-2 to enhance bone regeneration.

Recombinant human bone morphogenetic protein-2 (rhBMP-2) is the predominant growth factor that effectively induces osteogenic differentiation in orthopedic procedures. However, the bioactivity and stability of rhBMP-2 are intrinsically associated with its sequence, structure, and storage conditions. In this study, we successfully determined the amino acid sequence and protein secondary structure model of non-glycosylated rhBMP-2 expressed by an E. coli expression system through X-ray crystal structure analysis. Furthermore, we observed that acidic storage conditions enhanced the proliferative and osteoinductive activity of rhBMP-2. Although the osteogenic activity of non-glycosylated rhBMP-2 is relatively weaker compared to glycosylated rhBMP-2; however, this discrepancy can be mitigated by incorporating exogenous chaperone molecules. Overall, such information is crucial for rationalizing the design of stabilization methods and enhancing the bioactivity of rhBMP-2, which may also be applicable to other growth factors.

A new mRNA structure prediction based approach to identifying improved signal peptides for bone morphogenetic protein 2.

BACKGROUND: Signal peptide (SP) engineering has proven able to improve production of many proteins yet is a laborious process that still relies on trial and error. mRNA structure around the translational start site is important in translation initiation and has rarely been considered in this context, with recent improvements in in silico mRNA structure potentially rendering it a useful predictive tool for SP selection. Here we attempt to create a method to systematically screen candidate signal peptide sequences in silico based on both their nucleotide and amino acid sequences. Several recently released computational tools were used to predict signal peptide activity (SignalP), localization target (DeepLoc) and predicted mRNA structure (MXFold2). The method was tested with Bone Morphogenetic Protein 2 (BMP2), an osteogenic growth factor used clinically for bone regeneration. It was hoped more effective BMP2 SPs could improve BMP2-based gene therapies and reduce the cost of recombinant BMP2 production. RESULTS: Amino acid sequence analysis indicated 2,611 SPs from the TGF-ß superfamily were predicted to function when attached to BMP2. mRNA structure prediction indicated structures at the translational start site were likely highly variable. The five sequences with the most accessible translational start sites, a codon optimized BMP2 SP variant and the well-established hIL2 SP sequence were taken forward to in vitro testing. The top five candidates showed non-significant improvements in BMP2 secretion in HEK293T cells. All showed reductions in secretion versus the native sequence in C2C12 cells, with several showing large and significant decreases. None of the tested sequences were able to increase alkaline phosphatase activity above background in C2C12s. The codon optimized control sequence and hIL2 SP showed reasonable activity in HEK293T but very poor activity in C2C12. CONCLUSIONS: These results support the use of peptide sequence based in silico tools for basic predictions around signal peptide activity in a synthetic biology context. However, mRNA structure prediction requires improvement before it can produce reliable predictions for this application. The poor activity of the codon optimized BMP2 SP variant in C2C12 emphasizes the importance of codon choice, mRNA structure, and cellular context for SP activity.

Podocyte Pathogenic Bone Morphogenetic Protein-2 Pathway and Immune Cell Behaviors in Primary Membranous Nephropathy.

Primary membranous nephropathy (PMN) is one of the leading causes of end-stage renal disease, and the most frequent cause of massive proteinuria in nondiabetic adults, resulting in fatal complications. However, the underlying pathomechanisms of PMN remain largely unclear. Here, single-cell RNA sequencing is employed to analyze kidney biopsies from eleven PMN patients and seven healthy subjects. Profiling 44 060 cells from patients allowed us to characterize the cellular composition and cell-type-specific gene expression in the PMN kidney. The complement-induced BMP2/pSMAD1/COL4 pathway is identified as the pathogenic pathway in podocytes, bridging two key events, i.e., complement system activation and glomerular basement membrane thickening in PMN. Augmented infiltration and activation of myeloid leukocytes and B lymphocytes are found, profiling delicate crosstalk of immune cells in PMN kidneys. Overall, these results provide valuable insights into the roles of podocytes and immune cells in PMN, and comprehensive resources toward the complete understanding of PMN pathophysiology.

Rhizoma Drynariae-derived nanovesicles reverse osteoporosis by potentiating osteogenic differentiation of human bone marrow mesenchymal stem cells via targeting ERα signaling.

Although various anti-osteoporosis drugs are available, the limitations of these therapies, including drug resistance and collateral responses, require the development of novel anti-osteoporosis agents. Rhizoma Drynariae displays a promising anti-osteoporosis effect, while the effective component and mechanism remain unclear. Here, we revealed the therapeutic potential of Rhizoma Drynariae-derived nanovesicles (RDNVs) for postmenopausal osteoporosis and demonstrated that RDNVs potentiated osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) by targeting estrogen receptor-alpha (ERα). RDNVs, a natural product isolated from fresh Rhizoma Drynariae root juice by differential ultracentrifugation, exhibited potent bone tissue-targeting activity and anti-osteoporosis efficacy in an ovariectomized mouse model. RDNVs, effectively internalized by hBMSCs, enhanced proliferation and ERα expression levels of hBMSC, and promoted osteogenic differentiation and bone formation. Mechanistically, via the ERα signaling pathway, RDNVs facilitated mRNA and protein expression of bone morphogenetic protein 2 and runt-related transcription factor 2 in hBMSCs, which are involved in regulating osteogenic differentiation. Further analysis revealed that naringin, existing in RDNVs, was the active component targeting ERα in the osteogenic effect. Taken together, our study identified that naringin in RDNVs displays exciting bone tissue-targeting activity to reverse osteoporosis by promoting hBMSCs proliferation and osteogenic differentiation through estrogen-like effects.

RUNX1-BMP2 promotes vasculogenic mimicry in laryngeal squamous cell carcinoma via activation of the PI3K-AKT signaling pathway.

BACKGROUND: Laryngeal squamous cell carcinoma (LSCC) is one of the most common malignant tumors of the head and neck. Vasculogenic mimicry (VM) is crucial for tumor growth and metastasis and refers to the formation of fluid channels by invasive tumor cells rather than endothelial cells. However, the regulatory mechanisms underlying VM during the malignant progression of LSCC remain largely unknown. METHODS: Gene expression and clinical data for LSCC were obtained from the TCGA and Gene GEO (GSE27020) databases. A risk prediction model associated with VM was established using LASSO and Cox regression analyses. Based on their risk scores, patients with LSCC were categorized into high- and low-risk groups. The disparities in immune infiltration, tumor mutational burden (TMB), and functional enrichment between these two groups were examined. The core genes in LSCC were identified using the machine learning (SVM-RFE) and WGCNA algorithms. Subsequently, the involvement of bone morphogenetic protein 2 (BMP2) in VM and metastasis was investigated both in vitro and in vivo. To elucidate the downstream signaling pathways regulated by BMP2, western blotting was performed. Additionally, ChIP experiments were employed to identify the key transcription factors responsible for modulating the expression of BMP2. RESULTS: We established a new precise prognostic model for LSCC related to VM based on three genes: BMP2, EPO, and AGPS. The ROC curves from both TCGA and GSE27020 validation cohorts demonstrated precision survival prediction capabilities, with the nomogram showing some net clinical benefit. Multiple algorithm analyses indicated BMP2 as a potential core gene. Further experiments suggested that BMP2 promotes VM and metastasis in LSCC. The malignant progression of LSCC is promoted by BMP2 via the activation of the PI3K-AKT signaling pathway, with the high expression of BMP2 in LSCC resulting from its transcriptional activation by runt-related transcription factor 1 (RUNX1). CONCLUSION: BMP2 predicts poor prognosis in LSCC, promotes LSCC VM and metastasis through the PI3K-AKT signaling pathway, and is transcriptionally regulated by RUNX1. BMP2 may be a novel, precise, diagnostic, and therapeutic biomarker of LSCC.

Effect of recombinant human bone morphogenetic protein-2 and osteoprotegerin-Fc in MC3T3-E1 cells.

Objective: We compared the osteoblastogenesis by serially administrating recombinant human bone morphogenetic protein-2 (rhBMP-2) and osteoprotegerin-immunoglobulin Fc segment complex (OPG-Fc). Methods: The MC3T3-E1 preosteoblast cell line was differentiated for 1, 3, and 7 days with a treatment of OPG-Fc in 10~200 ng/mL concentration and the cell viability was evaluated by Cell Counting Kit-8 analysis. The level of differentiation from MC3T3-E1 cells to osteoblasts was determined by alkaline phosphatase activity. The level of runt domain-containing transcription factor 2 (Runx2) and osteopontin (OPN) manifestation, involved in osteoblast differentiation, was examined by real-time polymerase chain reaction and western blotting. Results: During MC3T3-E1 cell differentiation, the differentiation level was high with 1-day treatment using 100 ng/mL OPG-Fc. The treatment with 50 ng/mL rhBMP-2 for 7 days, followed by 1-day treatment with 100 ng/mL OPG-Fc produced the highest differentiation level, which was approximately 5.3 times that of the control group (p<0.05). The expression of Runx2 mRNA significantly increased, reaching 2.5 times the level of the control group under the condition of 7-day treatment with rhBMP-2 and 1-day treatment with OPG-Fc (p<0.001). The expression of Runx2 protein significantly increased to approximately 5.7 times that of the control group under the condition of 7-day treatment with rhBMP-2, followed by 1-day treatment with OPG-Fc (p<0.01). The expression of OPN protein showed no change from that of the control group under various conditions of rhBMP-2 and OPG-Fc combinations. Conclusion: These results imply that the treating preosteoblasts with rhBMP-2 first and then with OPG-Fc increased osteoblast differentiation efficacy.

Bone Substitute Options for Spine Fusion in Patients With Spine Trauma-Part II: The Role of rhBMP.

In Part II, we focus on an important aspect of spine fusion in patients with spine trauma: the pivotal role of recombinant human bone morphogenetic protein-2 (rhBMP-2). Despite the influx of diverse techniques facilitated by technological advancements in spinal surgery, spinal fusion surgery remains widely used globally. The persistent challenge of spinal pseudarthrosis has driven extensive efforts to achieve clinically favorable fusion outcomes, with particular emphasis on the evolution of bone graft substitutes. Part II of this review aims to build upon the foundation laid out in Part I by providing a comprehensive summary of commonly utilized bone graft substitutes for spinal fusion in patients with spinal trauma. Additionally, it will delve into the latest advancements and insights regarding the application of rhBMP-2, offering an updated perspective on its role in enhancing the success of spinal fusion procedures.

Delivery of Stem Cells and BMP-2 With Functionalized Self-Assembling Peptide Enhances Regeneration of Infarcted Myocardium.

Stem cell transplantation is a promising therapeutic strategy for myocardial infarction (MI). However, engraftment, survival and differentiation of the transplanted stem cells in ischemic and inflammatory microenvironment are poor. We designed a novel self-assembly peptide (SAP) by modifying the peptide RADA16 with cell-adhesive motif and BMP-2 (bone morphogenetic protein-2)-binding motif. Effects of the functionalized SAP on adhesion, survival and differentiation of c-kit+ MSCs (mesenchymal stem cells) were examined. Myocardial regeneration, neovascularization and cardiac function were assessed after transplantation of the SAP loading c-kit+ MSCs and BMP-2 in rat MI models. The SAP could spontaneously assemble into well-ordered nanofibrous scaffolds. The cells adhered to the SAP scaffolds and spread well. The SAP protected the cells in the condition of hypoxia and serum deprivation. Following degradation of the SAP, BMP-2 was released sustainedly and induced c-kit+ MSCs to differentiate into cardiomyocytes. At four weeks after transplantation of the SAP loading c-kit+ MSCs and BMP-2, myocardial regeneration and angiogenesis were enhanced, and cardiac function was improved significantly. The cardiomyocytes differentiated from the engrafted c-kit+ MSCs were increased markedly. The differentiated cells connected with recipient cardiomyocytes to form gap junctions. Collagen volume was decreased dramatically. These results suggest that the functionalized SAP promotes engraftment, survival and differentiation of stem cells effectively. Local sustained release of BMP-2 with SAP is a viable strategy to enhance differentiation of the engrafted stem cells and repair of the infarcted myocardium.