Bone Morphogenetic Protein-2 Peptide Research Articles

3D Niche-Inspired Scaffolds as a Stem Cell Delivery System for the Regeneration of the Osteochondral Interface.

The regeneration of the osteochondral unit represents a challenge due to the distinct cartilage and bone phases. Current strategies focus on the development of multiphasic scaffolds that recapitulate features of this complex unit and promote the differentiation of implanted bone-marrow derived stem cells (BMSCs). In doing so, challenges remain from the loss of stemness during in vitro expansion of the cells and the low control over stem cell activity at the interface with scaffolds in vitro and in vivo. Here, this work scaffolds inspired by the bone marrow niche that can recapitulate the natural healing process after injury. The construct comprises an internal depot of quiescent BMSCs, mimicking the bone marrow cavity, and an electrospun (ESP) capsule that "activates" the cells to migrate into an outer "differentiation-inducing" 3D printed unit functionalized with TGF-β and BMP-2 peptides. In vitro, niche-inspired scaffolds retained a depot of nonproliferative cells capable of migrating and proliferating through the ESP capsule. Invasion of the 3D printed cavity results in location-specific cell differentiation, mineralization, secretion of alkaline phosphatase (ALP) and glycosaminoglycans (GAGs), and genetic upregulation of collagen II and collagen I. In vivo, niche-inspired scaffolds are biocompatible, promoted tissue formation in rat subcutaneous models, and regeneration of the osteochondral unit in rabbit models.

Collagen-Hydroxyapatite Scaffold Platform Incorporating Dimethyloxalylglycine Loaded Polymersomes Decorated with BMP-2 Peptide for Bone Tissue Engineering

Introduction: Recent advances in the field of nanotechnology have led to the development of platforms for healing or repairing damaged tissues. The present study aimed to provide a bone extracellular matrix (ECM) mimetic platform made of hydroxyapatite and collagen loaded with nanopolymersomes encapsulating dimethyloxalylglycine (DMOG) and decorated with BMP-2 peptide in order to accelerate and increase the differentiation of mesenchymal stem cells to bone tissue. Method: In this study, polymersome nanoparticles carrying DMOG and targeted with BMP-2 peptide were prepared. The physicochemical properties of nanoparticles were evaluated. Thereafter, it was loaded into collagen-nanohydroxyapatite scaffolds and the expression levels of osteogenic genes OCN and OPN and angiogenic genes vcam, vwf, and kdr were checked in mesenchymal stem cells. Results: The surface charge and particle size of the synthesized nanoparticles were reported as -6 mv and 141 nm, respectively. The nanoparticles were able to slowly release 35% of the loaded DMOG within 30 days. The results demonstrated that the expression levels of OCN and OPN genes in cells exposed to polymersomes targeted with BMP-2 peptide were 27 and 2 times more than those of non-targeted polymersomes, respectively. In addition, the expression of angiogenic genes, vwf, kdr, and vcam, displayed a significant increase in targeted polymersomes compared to the non-targeted types (11.7-, 12.8-, and 2.17-fold, respectively). Conclusion: It seems that providing an environment similar to the bone ECM by combining an osteomimetic scaffold made of collagen-nanohydroxyapatite, DMOG as an angiogenic factor, and BMP-2 as the osteogenic factor can be effective in accelerating and increasing the differentiation of mesenchymal stem cells into osteoblasts.

Comparison of Osteoconductive Ability of Two Types of Cholesterol-Bearing Pullulan (CHP) Nanogel-Hydrogels Impregnated with BMP-2 and RANKL-Binding Peptide: Bone Histomorphometric Study in a Murine Calvarial Defect Model.

The receptor activator of NF-κB ligand (RANKL)-binding peptide is known to accelerate bone morphogenetic protein (BMP)-2-induced bone formation. Cholesterol-bearing pullulan (CHP)-OA nanogel-crosslinked PEG gel (CHP-OA nanogel-hydrogel) was shown to release the RANKL-binding peptide sustainably; however, an appropriate scaffold for peptide-accelerated bone formation is not determined yet. This study compares the osteoconductivity of CHP-OA hydrogel and another CHP nanogel, CHP-A nanogel-crosslinked PEG gel (CHP-A nanogel-hydrogel), in the bone formation induced by BMP-2 and the peptide. A calvarial defect model was performed in 5-week-old male mice, and scaffolds were placed in the defect. In vivo μCT was performed every week. Radiological and histological analyses after 4 weeks of scaffold placement revealed that the calcified bone area and the bone formation activity at the defect site in the CHP-OA hydrogel were significantly lower than those in the CHP-A hydrogel when the scaffolds were impregnated with both BMP-2 and the RANKL-binding peptide. The amount of induced bone was similar in both CHP-A and CHP-OA hydrogels when impregnated with BMP-2 alone. In conclusion, CHP-A hydrogel could be an appropriate scaffold compared to the CHP-OA hydrogel when the local bone formation was induced by the combination of RANKL-binding peptide and BMP-2, but not by BMP-2 alone.

Engineering immunomodulatory and osteoinductive implant surfaces via mussel adhesion-mediated ion coordination and molecular clicking

Immune response and new tissue formation are important aspects of tissue repair. However, only a single aspect is generally considered in previous biomedical interventions, and the synergistic effect is unclear. Here, a dual-effect coating with immobilized immunomodulatory metal ions (e.g., Zn2+) and osteoinductive growth factors (e.g., BMP-2 peptide) is designed via mussel adhesion-mediated ion coordination and molecular clicking strategy. Compared to the bare TiO2 group, Zn2+ can increase M2 macrophage recruitment by up to 92.5% in vivo and upregulate the expression of M2 cytokine IL-10 by 84.5%; while the dual-effect of Zn2+ and BMP-2 peptide can increase M2 macrophages recruitment by up to 124.7% in vivo and upregulate the expression of M2 cytokine IL-10 by 171%. These benefits eventually significantly enhance bone-implant mechanical fixation (203.3 N) and new bone ingrowth (82.1%) compared to the bare TiO2 (98.6 N and 45.1%, respectively). Taken together, the dual-effect coating can be utilized to synergistically modulate the osteoimmune microenvironment at the bone-implant interface, enhancing bone regeneration for successful implantation.

Chitosan Covalently Functionalized with Peptides Mapped on Vitronectin and BMP-2 for Bone Tissue Engineering.

Worldwide, over 20 million patients suffer from bone disorders annually. Bone scaffolds are designed to integrate into host tissue without causing adverse reactions. Recently, chitosan, an easily available natural polymer, has been considered a suitable scaffold for bone tissue growth as it is a biocompatible, biodegradable, and non-toxic material with antimicrobial activity and osteoinductive capacity. In this work, chitosan was covalently and selectively biofunctionalized with two suitably designed bioactive synthetic peptides: a Vitronectin sequence (HVP) and a BMP-2 peptide (GBMP1a). Nuclear magnetic resonance (NMR), X-ray photoelectron spectroscopy (XPS), and Fourier transform infrared spectroscopy (FT-IR) investigations highlighted the presence of the peptides grafted to chitosan (named Chit-HVP and Chit-GBMP1a). Chit-HVP and Chit-GBMP1a porous scaffolds promoted human osteoblasts adhesion, proliferation, calcium deposition, and gene expression of three crucial osteoblast proteins. In particular, Chit-HVP highly promoted adhesion and proliferation of osteoblasts, while Chit-GBMP1a guided cell differentiation towards osteoblastic phenotype.

Bio-clickable mussel-inspired peptides improve titanium-based material osseointegration synergistically with immunopolarization-regulation

Upon the osteoporotic condition, sluggish osteogenesis, excessive bone resorption, and chronic inflammation make the osseointegration of bioinert titanium (Ti) implants with surrounding bone tissues difficult, often lead to prosthesis loosening, bone collapse, and implant failure. In this study, we firstly designed clickable mussel-inspired peptides (DOPA-N3) and grafted them onto the surfaces of Ti materials through robust catechol-TiO2 coordinative interactions. Then, two dibenzylcyclooctyne (DBCO)-capped bioactive peptides RGD and BMP-2 bioactive domain (BMP-2) were clicked onto the DOPA-N3-coated Ti material surfaces via bio-orthogonal reaction. We characterized the surface morphology and biocompatibility of the Ti substrates and optimized the osteogenic capacity of Ti surfaces through adjusting the ideal ratios of BMP-2/RGD at 3:1. In vitro, the dual-functionalized Ti substrates exhibited excellent promotion on adhesion and osteogenesis of mesenchymal stem cells (MSCs), and conspicuous immunopolarization-regulation to shift macrophages to alternative (M2) phenotypes and inhibit inflammation, as well as enhancement of osseointegration and mechanical stability in osteoporotic rats. In summary, our biomimetic surface modification strategy by bio-orthogonal reaction provided a convenient and feasible method to resolve the bioinertia and clinical complications of Ti-based implants, which was conducive to the long-term success of Ti implants, especially in the osteoporotic or inflammatory conditions.

Effect of a Bone Morphogenetic Protein-2-derived peptide on the expression of tumor marker ZNF217 in osteoblasts and MCF-7 cells

Zinc Finger Protein 217 (ZNF217), a transcription factor and oncogene product, has been found to dysregulate Bone Morphogenetic Protein (BMP) signaling and induce invasion in breast tumors. In this study, the effect of BMP-2 or an active BMP-2 peptide, AISMLYLDEN, on the expression of ZNF217, BMP4 and CDK-inhibitor p21 gene, CDKN1A, was investigated in MCF-7 breast cancer cells. In parallel, the entire protein (BMP-2) as well as the aforementioned peptide were investigated in hDPSCs during osteogenic differentiation. The treatment of MCF-7 cancer cells with different concentrations of peptide AISMLYLDEN showed that the addition of 22.6 ng/ml was more effective in comparison to the other used concentrations. In particular, 48 h after treatment, CDKN1A and BMP4 mRNA levels were substantially increased in contrast to ZNF217 mRNA levels which were decreased. These results are strongly supported by BrdU assay that clearly indicated inhibition of cancer cell proliferation. Taken together, these results open ways for a concurrent use, at appropriate concentrations, of the peptide AISMLYLDEN during conventional therapeutic treatment in breast tumors with a metastatic tendency to the bones. Regarding the effect of the entire protein as well as its peptide on hDPSCs differentiation into osteocytes, the mRNA levels of osteocalcin, an osteogenic marker, showed that the peptide enhanced osteogenesis at a higher degree in comparison to the entire BMP-2 without however altering ZNF217, CDKN1A and BMP4 expression levels, which remained as expected of non-cancer cells.

A Cyclic BMP-2 Peptide Upregulates BMP-2 Protein-Induced Cell Signaling in Myogenic Cells.

In the current study, we designed four cyclic peptide analogues by incorporating two cysteine residues in a BMP-2 linear knuckle epitope in such a way that the active region of the peptide could be either inside or outside the cyclic ring. Bone morphogenetic protein receptor BMPRII was immobilized on the chip surface, and the interaction of the linear and cyclic peptide analogues was studied using surface plasmon resonance (SPR). From the affinity data, the peptides with an active region inside the cyclic ring had a higher binding affinity in comparison to the other peptides. To confirm that our affinity data are in line in vitro, we studied the expression levels of RUNX2 (runt-related transcription factor) and conducted an osteogenic marker alkaline phosphatase (ALP) assay and staining. Based on the affinity data and the in vitro experiments, peptide P-05 could be a suitable candidate for osteogenesis, with higher binding affinity and increased RUNX2 and ALP expression in comparison to the linear peptides.

A Dual Peptide Sustained-Release System Based on Nanohydroxyapatite/Polyamide 66 Scaffold for Synergistic-Enhancing Diabetic Rats' Fracture Healing in Osteogenesis and Angiogenesis.

Diabetes mellitus impairs fracture healing and function of stem cells related to bone regeneration; thus, effective bone tissue engineering therapies can intervene with those dysfunctions. Nanohydroxyapatite/polyamide 66 (n-HA/PA66) scaffold has been used in fracture healing, whereas the low bioactivity limits its further application. Herein, we developed a novel bone morphogenetic protein-2- (BMP-2) and vascular endothelial growth factor- (VEGF) derived peptides-decorated n-HA/PA66 (BVHP66) scaffold for diabetic fracture. The n-HA/PA66 scaffold was functionalized by covalent grafting of BMP-2 and VEGF peptides to construct a dual peptide sustained-release system. The structural characteristics and peptide release profiles of BVHP66 scaffold were tested by scanning electron microscopy, Fourier transform infrared spectroscopy, and fluorescence microscope. Under high glucose (HG) condition, the effect of BVHP66 scaffold on rat bone marrow mesenchymal stem cells’ (rBMSCs) adherent, proliferative, and differentiate capacities and human umbilical vein endothelial cells’ (HUVECs) proliferative and tube formation capacities was assessed. Finally, the BVHP66 scaffold was applied to fracture of diabetic rats, and its effect on osteogenesis and angiogenesis was evaluated. In vitro, the peptide loaded on the BVHP66 scaffold was in a sustained-release mode of 14 days. The BVHP66 scaffold significantly promoted rBMSCs’ and HUVECs’ proliferation and improved osteogenic differentiation of rBMSCs and tube formation of HUVECs in HG environment. In vivo, the BVHP66 scaffold enhanced osteogenesis and angiogenesis, rescuing the poor fracture healing in diabetic rats. Comparing with single peptide modification, the dual peptide-modified scaffold had a synergetic effect on bone regeneration in vivo. Overall, this study reported a novel BVHP66 scaffold with excellent biocompatibility and bioactive property and its application in diabetic fracture.

Preliminary evaluation of BMP-2-derived peptide in repairing a peri-implant critical size defect: A canine model

A synthetic bone morphogenetic protein (BMP)-2-derived peptide has been discovered to promote bone regeneration. The present study investigated the potential of the BMP-2 peptide combined with hydroxyapatite (HAp)/β-tricalcium phosphate (TCP)/collagen (Col) composite in repairing a peri-implant critical size defect. Twenty-four saddle-type alveolar defects (10mm mesiodistally and 4mm apicocoronally) were surgically prepared in edentulous ridges in four male beagle dogs. Following implant placement, the defects with vertically exposed implant fixtures received (a) HAp/TCP/Col composite, (b) HAp/TCP/Col+4mg/mL BMP-2 peptide, (c) HAp/TCP/Col+20mg/mL BMP-2 peptide, or (d) HAp/TCP/Col+0.2mg/mL recombinant human BMP-2 (rhBMP-2). Bone regeneration and mineralization were assessed using radiography, micro-computed tomography (micro-CT), fluorescence labeling, and histologic analyses after healing for 4 or 8 weeks. Implant stability was measured using resonance frequency analysis. The 20mg/mL BMP-2 peptide groups demonstrated a distinguishable advantage in bone regeneration potential over the control groups, as observed on radiographic imaging and histologic examination, although no significant difference was found in implant stability and histomorphometric analysis of mineralization levels. However, the performance of the 20mg/mL BMP-2 peptide groups were inferior to that of the 0.2mg/mL rhBMP-2 groups. The BMP-2 peptide may accelerate peri-implant bone regeneration. The BMP-2 peptide at 20mg/mL still cannot complete bone repair of peri-implant critical size defect. The BMP-2 peptide at 20mg/mL has similar osteoinductive performance to the rhBMP-2 at 0.02mg/mL.

Creation of Bony Microenvironment with Extracellular Matrix Doped-Bioactive Ceramics to Enhance Osteoblast Behavior and Delivery of Aspartic Acid-Modified BMP-2 Peptides.

IntroductionDecellularized matrix from porcine small intestinal submucosa (SIS) endows scaffolds with an ECM-like surface, which enhances stem cell self-renewal, proliferation, and differentiation. Mesoporous bioactive glass (MBG) is extensively recognized as an excellent bio-ceramic for fabricating bone grafts.Materials and MethodsIn the current study, SIS was doped on an MBG scaffold (MBG/SIS) using polyurethane foam templating and polydopamine chemistry method. To mimic the bony environment of a natural bone matrix, an ECM-inspired delivery system was constructed by coupling the BMP2-related peptide P28 to a heparinized MBG/SIS scaffold (MBG/SIS-H-P28). The release of P28 from MBG/SIS-H-P28 and its effects on the proliferation, viability, and osteogenic differentiation of bone marrow stromal stem cells were investigated in vitro and in vivo.ResultsOur research indicated that the novel tissue-derived ECM scaffold MBG/SIS has a hierarchical and interconnected porous architecture, and superior biomechanical properties. MBG/SIS-H-P28 released P28 in a controlled manner, with the long-term release time of 40 d. The results of in vitro experiments showed improvements in cell proliferation, cell viability, alkaline phosphatase activity, and mRNA expression levels of osteogenesis-related genes (Runx-2, OCN, OPN, and ALP) compared to those of MBG/SIS or MBG/SIS-P28 and MBG/SIS-H-P28. The in vivo results demonstrated that MBG/SIS-H-P28 scaffolds evidently increased bone formation in rat calvarial critical-sized defect compared to that in controls.ConclusionMBG/SIS-H-P28 scaffolds show potential as ideal platforms for delivery of P28 and for providing a bony environment for bone regeneration.

Chitosan/PEGDA based scaffolds as bioinspired materials to control in vitro angiogenesis.

In the current work, our purpose was based on the assessment of bioactive chitosan (CS)/Poly(ethylene glycol) diacrylate (PEGDA) based scaffolds ability to stimulate in vitro angiogenesis process. The bioactivation of the scaffolds was accomplished by using organic (BMP-2 peptide) and inorganic (hydroxyapatite nanoparticles) cues. In particular, the properties of the materials in terms of biological response promotion on human umbilical vein endothelial cells (HUVECs) were studied by using in vitro angiogenesis tests based on cell growth and proliferation. Furthermore, our interest was to examine the scaffolds capability to modulate two important steps involved in angiogenesis process: migration and tube formation of cells. Our data underlined that bioactive signals on CS/PEGDA scaffolds surface induce a desirable effect on angiogenic response concerning angiogenic marker expression (CD-31) and endothelial tissue formation (tube formation). Taken together, the results emphasized the concept that bioactive CS/PEGDA scaffolds may be novel implants for stimulating neovascularization of tissue-engineered constructs in regenerative medicine field.

Comparison of liposomal formulations incorporating BMP-2 peptide to induce bone tissue engineering

Objective(s)Fabricating a biomimetic scaffold platform combined with controlled release of bioactive agents is a practical approach for bone tissue engineering. Controlled delivery of peptides and growth factors which play a significant role in osteogenesis is an important issue reducing the associated adverse effects and leading to cost-effectiveness.Materials and MethodsWe developed two liposomal formulations of bone morphogenetic protein-2 (BMP-2) peptide designated as F1 and F2 with controlled release properties. Due to high negative zeta potential of F1 formulation, the surface of the liposomes was decorated with positively charged BMP-2 peptide while the peptide was encapsulated in F2 formulation. Then, we evaluated the hypothesis that whether the electrostatically loaded peptide could act as a ligand and improve the cellular uptake and osteogenic differentiation of mesenchymal stem cells.ResultsBoth formulations were less than 100 nm in size. The release study revealed that both formulations showed a sustained release pattern for 21 days. However, the cumulative releases were 60% and 40% in F1 and F2 formulations, respectively. Flow cytometry analysis indicated that cell internalization of F1 liposomes was more than the other formulation. In the next step, F1 and F2 formulations were attached covalently to our previously developed nanofibrous electrospun scaffold and biocompatibility and osteogenic differentiation of each formulation were studied. The results indicated that the proliferation of the cells seeded on F1 liposcaffold was significantly more than F2 liposcaffold at days 1 and 3. Furthermore, F1 liposcaffold showed superior osteogenic differentiation through measurement of alkaline phosphatase activity which could be due to the higher release pattern of F1 liposomes and their improved cellular uptake.ConclusionOur findings revealed that controlled release BMP-2 decorated liposomal formulations immobilized on nanofibrous electrospun scaffold platform could be a promising candidate for bone regeneration therapeutics and merits further investigation.

Establishment of Collagen: Hydroxyapatite/BMP-2 Mimetic Peptide Composites.

Extensive efforts were undertaken to develop suitable biomaterials for tissue engineering (TE) applications. To facilitate clinical approval processes and ensure the success of TE applications, bioinspired concepts are currently focused on. Working on bone tissue engineering, we describe in the present study a method for biofunctionalization of collagen/hydroxyapatite composites with BMP-2 mimetic peptides. This approach is expected to be fundamentally transferable to other tissue engineering fields. A modified BMP-2 mimetic peptide containing a negatively charged poly-glutamic acid residue (E7 BMP-2 peptide) was used to bind positively charged hydroxyapatite (HA) particles by electrostatic attraction. Binding efficiency was biochemically detected to be on average 85% compared to 30% of BMP-2 peptide without E7 residue. By quartz crystal microbalance (QCM) analysis, we could demonstrate the time-dependent dissociation of the BMP-2 mimetic peptides and the stable binding of the E7 BMP-2 peptides on HA-coated quartz crystals. As shown by immunofluorescence staining, alkaline phosphatase expression is similar to that detected in jaw periosteal cells (JPCs) stimulated with the whole BMP-2 protein. Mineralization potential of JPCs in the presence of BMP-2 mimetic peptides was also shown to be at least similar or significantly higher when low peptide concentrations were used, as compared to JPCs cultured in the presence of recombinant BMP-2 controls. In the following, collagen/hydroxyapatite composite materials were prepared. By proliferation analysis, we detected a decrease in cell viability with increasing HA ratios. Therefore, we chose a collagen/hydroxyapatite ratio of 1:2, similar to the natural composition of bone. The following inclusion of E7 BMP-2 peptides within the composite material resulted in significantly elevated long-term JPC proliferation under osteogenic conditions. We conclude that our advanced approach for fast and cost-effective scaffold preparation and biofunctionalization is suitable for improved and prolonged JPC proliferation. Further studies should prove the functionality of composite scaffolds in vivo.

3D Printed Scaffolds: Decorating 3D Printed Scaffolds with Electrospun Nanofiber Segments for Tissue Engineering (Adv. Biosys. 12/2019)

In article number 1900137, Jingwei Xie and co-workers report a versatile method to modify the surface of 3D printed scaffolds with electrospun short nanofibers. The short nanofiber-modified 3D scaffolds promote cell attachment and proliferation. Further functionalization with BMP2 peptides significantly induces rat bone marrow-derived stem cell differentiation.

Impact of Antifouling PEG Layer on the Performance of Functional Peptides in Regulating Cell Behaviors.

Cell adhesive and other functional peptides (such as RGD, KRSR, YIGSR, VAPG, and BMP-2 peptides) are extensively studied and utilized in tissue engineering scaffolds and biomedical devices to modulate cell functions. Though PEG is frequently used as the antifouling layer, it is unclear how it affects the performance of functional peptides. By analyzing the impact of PEG at short (OEG4), medium (OEG8), and long chain length (PEG2K), we reveal that PEG chain length is critical and a medium-length PEG enables functional peptides to display their optimal and genuine functions in cell adhesion, migration, and differentiation by providing excellent antifouling to minimize background noise of unwanted cell adhesion and high enough surface density of functional peptides. Our result provides new avenues for maximizing the genuine functions of peptides. This study also provides a solution to prevent the heterogeneous and even divergent results caused by inappropriate choice of antifouling PEG and provides a general guidance in identifying new functional peptides.

Bioactivation Routes of Gelatin-Based Scaffolds to Enhance at Nanoscale Level Bone Tissue Regeneration.

The present work is focused on the development of gelatin-based scaffolds crosslinked through carbodiimide reaction and their bioactivation by two different methods: (i) surface modification by inorganic signals represented by hydroxyapatite nanoparticles precipitated on scaffold through biomimetic treatment; (ii) analog of BMP-2 peptide decoration. The results showed the effects of polymer concentration and crosslinking time on the physico-chemical, morphological, and mechanical properties of scaffolds. Furthermore, a comparative study of biological response for both bioactivated structures allowed to evaluate the influence of inorganic and organic cues on cellular behavior in terms of adhesion, proliferation and early osteogenic marker expression. The bioactivation by inorganic cues induced positive cellular response compared to neat scaffolds in terms of increased cell proliferation and early osteogenic differentiation of human mesenchymal stem cell (hMSC), as evidenced by the Alkaline phosphatase (ALP) expression. Similarly BMP-2 peptide decorated scaffolds showed higher values of ALP than biomineralized ones at longer time. The overall results demonstrated that the presence of bioactive signals (either inorganic or organic) at nanoscale level allowed an osteoinductive effect on hMSC in a basal medium, making the modified gelatin scaffolds a promising candidate for bone tissue regeneration.

Enhanced bone regeneration of the silk fibroin electrospun scaffolds through the modification of the graphene oxide functionalized by BMP-2 peptide.

IntroductionBone tissue engineering has become one of the most effective methods to treat bone defects. Silk fibroin (SF) is a natural protein with no physiological activities, which has features such as good biocompatibility and easy processing and causes minimal inflammatory reactions in the body. Scaffolds prepared by electrospinning SF can be used in bone tissue regeneration and repair. Graphene oxide (GO) is rich in functional groups, has good biocompatibility, and promotes osteogenic differentiation of stem cells, while bone morphogenetic protein-2 (BMP-2) polypeptide has an advantage in promoting osteogenesis induction. In this study, we attempted to graft BMP-2 polypeptide onto GO and then bonded the functionalized GO onto SF electrospun scaffolds through electrostatic interactions. The main purpose of this study was to further improve the biocompatibility of SF electrospun scaffolds, which could promote the osteogenic differentiation of bone marrow mesenchymal stem cells and the repair of bone tissue defects.Materials and methodsThe successful synthesis of GO and functionalized GO was confirmed by transmission electron microscope, X-ray photoelectron spectroscopy, and thermogravimetric analysis. Scanning electron microscopy, atomic force microscopy, mechanical test, and degradation experiment confirmed the preparation of SF electrospun scaffolds and the immobilization of GO on the fibers. In vitro experiment was used to verify the biocompatibility of the composite scaffolds, and in vivo experiment was used to prove the repairing ability of the composite scaffolds for bone defects.ResultsWe successfully fabricated the composite scaffolds, which enhanced biocompatibility, not only promoting cell adhesion and proliferation but also greatly enhancing in vitro osteogenic differentiation of bone marrow stromal cells using either an osteogenic or non-osteogenic medium. Furthermore, transplantation of the composite scaffolds significantly promoted in vivo bone formation in critical-sized calvarial bone defects.ConclusionThese findings suggested that the incorporation of BMP-2 polypeptide-functionalized GO into chitosan-coated SF electrospun scaffolds was a viable strategy for fabricating excellent scaffolds that enhance the regeneration of bone defects.

Mineralized nanofiber segments coupled with calcium-binding BMP-2 peptides for alveolar bone regeneration

Bone loss around tooth extraction sites can occur, thus making future placement of dental implants difficult. Alveolar bone regeneration can be guided by the application of a nanofibrous bone graft coupled with osteoinductive proteins/peptides, following tooth loss or tooth extraction. In the present study, we demonstrate the potential of mineralized nanofiber segments coupled with calcium-binding bone morphogenetic protein 2 (BMP-2) mimicking peptides for periodontal bone regeneration. Thin electrospun nanofiber membranes of PLGA-collagen-gelatin (2:1:1 wt ratios) were mineralized in 10× modified simulated body fluid (10× mSBF) and cryocut to segments of 20 µm. For predetermined weights of the mineralized nanofiber segments, it was possible to load various amounts of heptaglutamate E7-domain-conjugated BMP-2 peptide. Mineralized short fiber grafts (2 mg), with and without E7-BMP-2 peptides, were implanted into 2 mm × 2 mm (diameter × depth) critical-sized socket defects created in rat maxillae, following extraction of the first molar teeth. A sustained release profile of E7-BMP-2 from the mineralized nanofiber segments was recorded over 4 weeks. X-ray microcomputed tomography (µ-CT) analysis of peptide-loaded nanofiber graft filled defects revealed ∼3 times greater new bone volume and bone mineral density over 4 weeks in comparison to unfilled control defects. Further, histopathology data confirmed the formation of greater new osseous tissue in the BMP2 peptide-loaded, mineralized nanofiber segment group than that of fibrous connective tissue in the unfilled defect group. Altogether, the mineralized nanofiber segments coupled with E7-BMP-2 peptides may be an effective treatment option for alveolar bone loss and defects. Statement of SignificanceWith the high incidence of dental implants/fixtures for missing teeth, the success of the surgical procedures in restorative dentistry is dictated by the quality and quantity of the supporting alveolar bone. To address the problem of alveolar bone loss and defects due to tumor, periodontitis, or even postextraction remodeling, the present study is the first report on the application of mineralized nanofiber fragments coupled with calcium-binding osteoinductive BMP-2 peptides as a synthetic graft material for oral bone regeneration. The ease of fabrication and application of cryocut mineralized nanofiber fragments as maxillofacial bone defect fillers present a promising alternative to the current dental bone graft formulations. Furthermore, the nanofiber segments may also be utilized for several biomedical applications including hemostasis, soft tissue engineering, and wound healing.

Development of Organic/Inorganic Compatible and Sustainably Bioactive Composites for Effective Bone Regeneration.

In this paper, we demonstrate a strategy of covalently bonding bioactive molecules onto inorganic hydroxyapatite (HAp) to improve the compatibility between organic and inorganic components and endow the bone composites with sustainable bioactivity. Bone morphogenetic protein-2 (BMP-2) peptide covalently immobilized nano-hydroxyapatite (nHAp-BMP-2) is developed to preserve the bioactivity and slow the release of the BMP-2 peptide. Then nHAp-BMP-2 was further incorporated into an ultraviolet-curable mixture of gelatin methacrylamide (GelMA) and four-armed PEG methacrylamide (four-armed PEGMA) to form a Gel/(nHAp-BMP-2) composite. The hydrogen bonding between gelatin and BMP-2 on nHAp-BMP-2 enhanced the compatibility between inorganic and organic components. The Gel/(nHAp-BMP-2) composite exhibited superior biocompatibility caused by gelatin and nHAp-BMP-2, except in a two-dimensional cell culture, the hydrogel was also capable of a three-dimensional cell culture. In addition, the introduction of nHAp-BMP-2 had a positive influence on bone marrow mesenchymal stem cell proliferation, differentiation, and the subsequent calcification on the composite. After treatment of a rat calvarial defect model for 12 weeks, the Gel/(nHAp-BMP-2) group showed the largest new bone volume and the highest ratio of new bone (50.54 ± 13.51 mm3 and 64.38 ± 17.22%, respectively) compared to those of the other groups. These results demonstrate that this way of controlling BMP-2 release is effective and the Gel/(nHAp-BMP-2) composite has great potential in bone regeneration therapy.