RESUMO
Dental enamel is a principal component of teeth1, and has evolved to bear large chewing forces, resist mechanical fatigue and withstand wear over decades2. Functional impairment and loss of dental enamel, caused by developmental defects or tooth decay (caries), affect health and quality of life, with associated costs to society3. Although the past decade has seen progress in our understanding of enamel formation (amelogenesis) and the functional properties of mature enamel, attempts to repair lesions in this material or to synthesize it in vitro have had limited success4-6. This is partly due to the highly hierarchical structure of enamel and additional complexities arising from chemical gradients7-9. Here we show, using atomic-scale quantitative imaging and correlative spectroscopies, that the nanoscale crystallites of hydroxylapatite (Ca5(PO4)3(OH)), which are the fundamental building blocks of enamel, comprise two nanometric layers enriched in magnesium flanking a core rich in sodium, fluoride and carbonate ions; this sandwich core is surrounded by a shell with lower concentration of substitutional defects. A mechanical model based on density functional theory calculations and X-ray diffraction data predicts that residual stresses arise because of the chemical gradients, in agreement with preferential dissolution of the crystallite core in acidic media. Furthermore, stresses may affect the mechanical resilience of enamel. The two additional layers of hierarchy suggest a possible new model for biological control over crystal growth during amelogenesis, and hint at implications for the preservation of biomarkers during tooth development.
Assuntos
Amelogênese , Esmalte Dentário/química , Ácidos/química , Cálcio/química , Carbonatos/química , Cristalização , Teoria da Densidade Funcional , Esmalte Dentário/ultraestrutura , Durapatita/química , Fluoretos/química , Humanos , Magnésio/química , Microscopia Eletrônica de Transmissão e Varredura , Sódio/química , Tomografia , Difração de Raios XRESUMO
DNA-based biomaterials have been proposed for tissue engineering approaches due to their predictable assembly into complex morphologies and ease of functionalization. For bone tissue regeneration, the ability to bind Ca2+ and promote hydroxyapatite (HAP) growth along the DNA backbone combined with their degradation and release of extracellular phosphate, a known promoter of osteogenic differentiation, make DNA-based biomaterials unlike other currently used materials. However, their use as biodegradable scaffolds for bone repair remains scarce. Here, we describe the design and synthesis of DNA hydrogels, gels composed of DNA that swell in water, their interactions in vitro with the osteogenic cell lines MC3T3-E1 and mouse calvarial osteoblast, and their promotion of new bone formation in rat calvarial wounds. We found that DNA hydrogels can be readily synthesized at room temperature, and they promote HAP growth in vitro, as characterized by Fourier transform infrared spectroscopy, X-ray diffraction, scanning electron microscopy, atomic force microscopy, and transmission electron microscopy. Osteogenic cells remain viable when seeded on DNA hydrogels in vitro, as characterized by fluorescence microscopy. In vivo, DNA hydrogels promote the formation of new bone in rat calvarial critical size defects, as characterized by micro-computed tomography and histology. This study uses DNA hydrogels as a potential therapeutic biomaterial for regenerating lost bone.
Assuntos
Hidrogéis , Osteogênese , Camundongos , Ratos , Animais , Hidrogéis/química , Microtomografia por Raio-X , Regeneração Óssea , Materiais Biocompatíveis/farmacologia , Materiais Biocompatíveis/química , Durapatita/farmacologia , Durapatita/química , Engenharia Tecidual , Alicerces Teciduais/químicaRESUMO
Citrate is a critical metabolic substrate and key regulator of energy metabolism in mammalian cells. It has been known for decades that the skeleton contains most (>85%) of the body's citrate, but the question of why and how this metabolite should be partitioned in bone has received singularly little attention. Here, we show that osteoblasts use a specialized metabolic pathway to regulate uptake, endogenous production, and the deposition of citrate into bone. Osteoblasts express high levels of the membranous Na+-dependent citrate transporter solute carrier family 13 member 5 (Slc13a5) gene. Inhibition or genetic disruption of Slc13a5 reduced osteogenic citrate uptake and disrupted mineral nodule formation. Bones from mice lacking Slc13a5 globally, or selectively in osteoblasts, showed equivalent reductions in cortical thickness, with similarly compromised mechanical strength. Surprisingly, citrate content in mineral from Slc13a5-/- osteoblasts was increased fourfold relative to controls, suggesting the engagement of compensatory mechanisms to augment endogenous citrate production. Indeed, through the coordinated functioning of the apical membrane citrate transporter SLC13A5 and a mitochondrial zinc transporter protein (ZIP1; encoded by Slc39a1), a mediator of citrate efflux from the tricarboxylic acid cycle, SLC13A5 mediates citrate entry from blood and its activity exerts homeostatic control of cytoplasmic citrate. Intriguingly, Slc13a5-deficient mice also exhibited defective tooth enamel and dentin formation, a clinical feature, which we show is recapitulated in primary teeth from children with SLC13A5 mutations. Together, our results reveal the components of an osteoblast metabolic pathway, which affects bone strength by regulating citrate deposition into mineral hydroxyapatite.
Assuntos
Ácido Cítrico , Simportadores , Animais , Camundongos , Ácido Cítrico/metabolismo , Simportadores/metabolismo , Durapatita/metabolismo , Citratos , Ciclo do Ácido Cítrico , Osteoblastos/metabolismo , Mamíferos/metabolismo , Transportadores de Ácidos Dicarboxílicos/metabolismoRESUMO
Natural killer (NK) cells have become a powerful candidate for adoptive tumor immunotherapy, while their therapeutic efficacy in solid tumors remains unsatisfactory. Here, we developed a hybrid module with an injectable hydrogel and hydroxyapatite (HAp) nanobelts for the controlled delivery of NK cells to enhance the therapy of solid tumors. Surface-functionalized HAp nanobelts modified with agonistic antibodies against NKG2D and 4-1BB and cytokines IL-2 and IL-21 support survival and dynamic activation. Thus, the HAp-modified chitosan (CS) thermos-sensitive hydrogel not only improved the retention of NK cells for more than 20 days in vivo but also increased NK cell function by more than one-fold. The unique architecture of this biomaterial complex protects NK cells from the hostile tumor environment and improves antitumor efficacy. The generation of a transient inflammatory niche for NK cells through a biocompatible hydrogel reservoir may be a conversion pathway to prevent cancer recurrence of resectable tumors.
Assuntos
Hidrogéis , Células Matadoras Naturais , Células Matadoras Naturais/imunologia , Animais , Camundongos , Hidrogéis/química , Humanos , Neoplasias/terapia , Neoplasias/imunologia , Imunoterapia/métodos , Durapatita/química , Linhagem Celular Tumoral , Quitosana/química , Subfamília K de Receptores Semelhantes a Lectina de Células NK , Interleucinas/imunologia , Interleucina-2/imunologiaRESUMO
The low sensitivity of nuclear magnetic resonance (NMR) is a major bottleneck for studying biomolecular structures of complex biomolecular assemblies. Cryogenically cooled probe technology overcomes the sensitivity limitations enabling NMR applications to challenging biomolecular systems. Here we describe solid-state NMR studies of the human blood protein vitronectin (Vn) bound to hydroxyapatite (HAP), the mineralized form of calcium phosphate, using a CryoProbe designed for magic angle spinning (MAS) experiments. Vn is a major blood protein that regulates many different physiological and pathological processes. The high sensitivity of the CryoProbe enabled us to acquire three-dimensional solid-state NMR spectra for sequential assignment and characterization of site-specific water-protein interactions that provide initial insights into the organization of the Vn-HAP complex. Vn associates with HAP in various pathological settings, including macular degeneration eyes and Alzheimer's disease brains. The ability to probe these assemblies at atomic detail paves the way for understanding their formation.
Assuntos
Durapatita , Vitronectina , Humanos , Espectroscopia de Ressonância Magnética/métodos , Imageamento por Ressonância Magnética , Ressonância Magnética Nuclear Biomolecular/métodosRESUMO
BACKGROUND: Differentiation of pluripotent stem cells into desired lineages is the key aspect of regenerative medicine and cell-based therapy. Although RNA interference (RNAi) technology is exploited extensively for this, methods for long term silencing of the target genes leading to differentiation remain a challenge. Sustained knockdown of the target gene by RNAi is often inefficient as a result of low delivery efficiencies, protocol induced toxicity and safety concerns related to viral vectors. Earlier, we established octa-arginine functionalized hydroxyapatite nano vehicles (R8HNPs) for delivery of small interfering RNA (siRNA) against a pluripotency marker gene in mouse embryonic stem cells. Although we demonstrated excellent knockdown efficiency of the target gene, sustained gene silencing leading to differentiation was yet to be achieved. METHODS: To establish a sustained non-viral gene silencing protocol using R8HNP, we investigated various methods of siRNA delivery: double delivery of adherent cells (Adh-D), suspension delivery followed by adherent delivery (Susp + Adh), single delivery in suspension (Susp-S) and multiple deliveries in suspension (Susp-R). Sustained knockdown of a pluripotent marker gene followed by differentiation was analysed by reverse transcriptase-PCR, fluoresence-activated cell sorting and immunofluorescence techniques. Impact on cell viability as a result of repeated exposure of the R8HNP was also tested. RESULTS: Amongst the protocols tested, the most efficient knockdown of the target gene for a prolonged period of time was obtained by repeated suspension delivery of the R8HNP-siRNA conjugate. The long-term silencing of a pluripotency marker gene resulted in differentiation of R1 ESCs predominantly towards the extra embryonic and ectodermal lineages. Cells displayed excellent tolerance to repeated exposures of R8HNPs. CONCLUSIONS: The results demonstrate that R8HNPs are promising, biocompatible, non-viral alternatives for prolonged gene silencing and obtaining differentiated cells for therapeutics.
Assuntos
Diferenciação Celular , Durapatita , Células-Tronco Embrionárias Murinas , RNA Interferente Pequeno , Animais , Camundongos , Durapatita/química , Células-Tronco Embrionárias Murinas/metabolismo , Células-Tronco Embrionárias Murinas/efeitos dos fármacos , RNA Interferente Pequeno/genética , Inativação Gênica , Materiais Biocompatíveis/química , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Nanopartículas/química , Transdução Genética , Interferência de RNA , Técnicas de Silenciamento de GenesRESUMO
Accumulation of reactive oxygen species (ROS) in periodontitis exacerbates the destruction of alveolar bone. Therefore, scavenging ROS to reshape the periodontal microenvironment, alleviate the inflammatory response and promote endogenous stem cell osteogenic differentiation may be an effective strategy for treating bone resorption in periodontitis. In this study, sericin-hydroxyapatite nanoparticles (Se-nHA NPs) are synthesized using a biomimetic mineralization method. Se-nHA NPs and proanthocyanidins (PC) are then encapsulated in sericin/sodium alginate (Se/SA) using an electrostatic injection technique to prepare Se-nHA/PC microspheres. Microspheres are effective in scavenging ROS, inhibiting the polarization of macrophages toward the M1 type, and inducing the polarization of macrophages toward the M2 type. In normal or macrophage-conditioned media, the Se-nHA/PC microspheres effectively promoted the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs). Furthermore, the Se-nHA/PC microspheres demonstrated anti-inflammatory effects in a periodontitis rat model by scavenging ROS and suppressing pro-inflammatory cytokines. The Se-nHA/PC microspheres are also distinguished by their capacity to decrease alveolar bone loss, reduce osteoclast activity, and boost osteogenic factor expression. Therefore, the biomimetic Se-nHA/PC composite microspheres have efficient ROS-scavenging, anti-inflammatory, and osteogenic abilities and can be used as a multifunctional filling material for inflammatory periodontal tissue regeneration.
Assuntos
Periodontite , Proantocianidinas , Sericinas , Humanos , Animais , Ratos , Osteogênese , Biomimética , Microesferas , Espécies Reativas de Oxigênio , Regeneração Óssea , Periodontite/terapia , Durapatita , Anti-InflamatóriosRESUMO
BACKGROUND: In recent years, natural bone extracellular matrix (ECM)-inspired materials have found widespread application as scaffolds for bone tissue engineering. However, the challenge of creating scaffolds that mimic natural bone ECM's mechanical strength and hierarchical nano-micro-macro structures remains. The purposes of this study were to introduce an innovative bone ECM-inspired scaffold that integrates a 3D-printed framework with hydroxyapatite (HAp) mineralized graphene oxide-collagen (GO-Col) microscaffolds and find its application in the repair of mandibular bone defects. METHODS: Initially, a 3D-printed polycaprolactone (PCL) scaffold was designed with cubic disks and square pores to mimic the macrostructure of bone ECM. Subsequently, we developed multi-layer mineralized GO-Col-HAp microscaffolds (MLM GCH) to simulate natural bone ECM's nano- and microstructural features. Systematic in vitro and in vivo experiments were introduced to evaluate the ECM-inspired structure of the scaffold and to explore its effect on cell proliferation and its ability to repair rat bone defects. RESULTS: The resultant MLM GCH/PCL composite scaffolds exhibited robust mechanical strength and ample assembly space. Moreover, the ECM-inspired MLM GCH microscaffolds displayed favorable attributes such as water absorption and retention and demonstrated promising cell adsorption, proliferation, and osteogenic differentiation in vitro. The MLM GCH/PCL composite scaffolds exhibited successful bone regeneration within mandibular bone defects in vivo. CONCLUSIONS: This study presents a well-conceived strategy for fabricating ECM-inspired scaffolds by integrating 3D-printed PCL frameworks with multilayer mineralized porous microscaffolds, enhancing cell proliferation, osteogenic differentiation, and bone regeneration. This construction approach holds the potential for extension to various other biomaterial types.
Assuntos
Durapatita , Grafite , Osteogênese , Ratos , Animais , Durapatita/análise , Durapatita/metabolismo , Durapatita/farmacologia , Alicerces Teciduais/química , Regeneração Óssea , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Engenharia Tecidual , Poliésteres/química , Mandíbula , Impressão TridimensionalRESUMO
BACKGROUND: Biological-derived hydroxyapatite is widely used as a bone substitute for addressing bone defects, but its limited osteoconductive properties necessitate further improvement. The osteo-immunomodulatory properties hold crucial promise in maintaining bone homeostasis, and precise modulation of macrophage polarization is essential in this process. Metabolism serves as a guiding force for immunity, and fluoride modification represents a promising strategy for modulating the osteoimmunological environment by regulating immunometabolism. In this context, we synthesized fluorinated porcine hydroxyapatite (FPHA), and has demonstrated its enhanced biological properties and osteogenic capacity. However, it remains unknown whether and how FPHA affects the immune microenvironment of the bone defects. METHODS: FPHA was synthesized and its composition and structural properties were confirmed. Macrophages were cultured with FPHA extract to investigate the effects of FPHA on their polarization and the related osteo-immune microenvironment. Furthermore, total RNA of these macrophages was extracted, and RNA-seq analysis was performed to explore the underlying mechanisms associated with the observed changes in macrophages. The metabolic states were evaluated with a Seahorse analyzer. Additionally, immunohistochemical staining was performed to evaluate the macrophages response after implantation of the novel bone substitutes in critical size calvarial defects in SD rats. RESULTS: The incorporation of fluoride ions in FPHA was validated. FPHA promoted macrophage proliferation and enhanced the expression of M2 markers while suppressing the expression of M1 markers. Additionally, FPHA inhibited the expression of inflammatory factors and upregulated the expression of osteogenic factors, thereby enhancing the osteogenic differentiation capacity of the rBMSCs. RNA-seq analysis suggested that the polarization-regulating function of FPHA may be related to changes in cellular metabolism. Further experiments confirmed that FPHA enhanced mitochondrial function and promoted the metabolic shift of macrophages from glycolysis to oxidative phosphorylation. Moreover, in vivo experiments validated the above results in the calvarial defect model in SD rats. CONCLUSION: In summary, our study reveals that FPHA induces a metabolic shift in macrophages from glycolysis to oxidative phosphorylation. This shift leads to an increased tendency toward M2 polarization in macrophages, consequently creating a favorable osteo-immune microenvironment. These findings provide valuable insights into the impact of incorporating an appropriate concentration of fluoride on immunometabolism and macrophage mitochondrial function, which have important implications for the development of fluoride-modified immunometabolism-based bone regenerative biomaterials and the clinical application of FPHA or other fluoride-containing materials.
Assuntos
Durapatita , Glicólise , Macrófagos , Fosforilação Oxidativa , Ratos Sprague-Dawley , Animais , Durapatita/química , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Fosforilação Oxidativa/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Ratos , Suínos , Proliferação de Células/efeitos dos fármacos , Masculino , Osteogênese/efeitos dos fármacos , Crânio/patologia , Crânio/efeitos dos fármacos , Camundongos , Microambiente Celular/efeitos dos fármacos , Células RAW 264.7 , Osso e Ossos/metabolismo , Osso e Ossos/efeitos dos fármacosRESUMO
Hydrogels from natural polysaccharides are of great interest for tissue engineering. This study aims (1) to prepare hydroxyapatite-loaded macroporous calcium alginate hydrogels by novel one-step technique using internal gelation in water-frozen solutions; (2) to evaluate their physicochemical properties; (3) to estimate their ability to support cell growth and proliferation in vitro. The structure of the hydrogel samples in a swollen state was studied by confocal laser scanning microscopy and was shown to represent a system of interconnected macropores with sizes of tens micron. The swelling behavior of the hydrogels, their mechanical properties (Young's moduli) in function of a hydroxyapatite content (5-30 mass%) were studied. All hydrogel samples loaded with hydroxyapatite were found to support growth and proliferation of mouse fibroblasts (L929) at long-term cultivation for 7 days. The obtained macroporous composite Ca-Alg-HA hydrogels could be promising for tissue engineering.
Assuntos
Alginatos , Durapatita , Hidrogéis , Alginatos/química , Hidrogéis/química , Hidrogéis/síntese química , Durapatita/química , Camundongos , Animais , Porosidade , Fibroblastos/efeitos dos fármacos , Fibroblastos/citologia , Engenharia Tecidual/métodos , Proliferação de Células/efeitos dos fármacos , Ácido Glucurônico/química , Ácidos Hexurônicos/química , Linhagem Celular , Materiais Biocompatíveis/químicaRESUMO
Injectable, tissue mimetic, bioactive, and biodegradable hydrogels offer less invasive regeneration and repair of tissues. The monitoring swelling and in vitro degradation capacities of hydrogels are highly important for drug delivery and tissue regeneration processes. Bioactivity of bone tissue engineered constructs in terms of mineralized apatite formation capacity is also pivotal. We have previously reported in situ forming chitosan-based injectable hydrogels integrated with hydroxyapatite and heparin for bone regeneration, promoting angiogenesis. These hydrogels were functionalized by glycerol and pH to improve their mechano-structural properties. In the present study, functionalized hybrid hydrogels were investigated for their swelling, in vitro degradation, and bioactivity performances. Hydrogels have degraded gradually in phosphate-buffered saline (PBS) with and without lysozyme enzyme. The percentage weight loss of hydrogels and their morphological and chemical properties, and pH of media were analyzed. The swelling ratio of hydrogels (55%-68%(wt), 6 h of equilibrium) indicated a high degree of cross-linking, can be suitable for controlled drug release. Hydrogels have gradually degraded reaching to 60%-70% (wt%) in 42 days in the presence and absence of lysozyme, respectively. Simulated body fluid (SBF)-treated hydrogels containing hydroxyapatite-induced needle-like carbonated-apatite mineralization was further enhanced by heparin content significantly.
Assuntos
Regeneração Óssea , Quitosana , Hidrogéis , Quitosana/química , Hidrogéis/química , Hidrogéis/farmacologia , Regeneração Óssea/efeitos dos fármacos , Sistemas de Liberação de Medicamentos , Durapatita/química , Durapatita/farmacologia , Muramidase/química , Muramidase/farmacologia , Concentração de Íons de Hidrogênio , Materiais Biocompatíveis/química , Heparina/química , Heparina/farmacologiaRESUMO
We investigated the importance of the carboxy group density in bone affinity during the development of peptide-based bone-seeking radiopharmaceuticals and carriers. Oligo-γ-carboxy glutamic acid peptides [(Gla)n] with higher carboxy group density than oligo-glutamic acid peptides [(Glu)n] and oligo-aspartic acid peptides [(Asp)n] were chosen. Using the radiogallium chelator N,N'-bis-[2-hydroxy-5-(carboxyethyl)benzyl]ethylenediamine-N,N'-diacetic acid (HBED-CC), we synthesized [67Ga]Ga-HBED-CC-(Gla)n (n = 1, 2, 5, 8, 11, or 14) with high yields. Hydroxyapatite-binding assays, biodistribution, and SPECT imaging showed higher affinity and bone accumulation for [67Ga]Ga-HBED-CC-(Gla)n compared to [67Ga]Ga-HBED-CC-(Glu)n. Notably, [67Ga]Ga-HBED-CC-(Gla)8 and [67Ga]Ga-HBED-CC-(Gla)11 exhibited superior bone accumulation and rapid blood clearance. SPECT/CT imaging with [67Ga]Ga-HBED-CC-(Gla)8 exclusively visualized the bone tissue. These findings support the potential use of [67Ga]Ga-HBED-CC-(Gla)n as excellent bone-imaging PET probes, suggesting (Gla)n peptides are superior bone-seeking carriers.
Assuntos
Osso e Ossos , Radioisótopos de Gálio , Compostos Radiofarmacêuticos , Tomografia Computadorizada de Emissão de Fóton Único , Animais , Radioisótopos de Gálio/farmacocinética , Radioisótopos de Gálio/química , Compostos Radiofarmacêuticos/farmacocinética , Camundongos , Distribuição Tecidual , Tomografia Computadorizada de Emissão de Fóton Único/métodos , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/metabolismo , Peptídeos/química , Durapatita/química , Masculino , Ácido Glutâmico/metabolismo , FemininoRESUMO
Nonfouling surfaces are crucial in applications such as biosensors, medical implants, marine coatings, and drug delivery vehicles. However, their long-term coating stability and robust surface binding strength in physiological media remain challenging. Herein, a phosphonate-grafted, PEGylated copolymer on the hydroxyapatite (HA) surface is proposed to significantly improve the adsorption stability and thus enhance the biofunction durability accordingly. The phosphoryl (-PO3) grafted branch is employed in the functional polymer to facilitate attaching to the HA substrate. In addition, the polymer integrates the nonfouling polymer brushes of poly(ethylene glycol) (PEG) with the cell-adhesive moiety of cyclic Arg-Gly-Asp-d-Phe-Cys peptides (cRGD). A systematic study on the as-synthesized PEGylated graft copolymer indicates a synergistic binding mechanism of the NH2 and PO3 groups to HA, achieving a high surface coverage with desirable adsorption stability. The cRGD/PEGylated copolymers of optimized grafting architecture are proven to effectively adsorb to HA surfaces as a self-assembled copolymer monolayer, showing stability with minimal desorption even in a complex, physiological medium and effectively preventing nonspecific protein adsorption as examined with X-ray photoelectron spectroscopy (XPS) and a quartz crystal microbalance with dissipation (QCM-D). Direct adhesion assays further confirm that the enhanced coating stability and biofunction durability of the phosphonate-grafted, cRGD-PEGylated copolymer can considerably promote osteoblast attachment on HA surfaces, meanwhile preventing microbial adhesion. This research has resulted in a solution of self-assembly polymer structure optimization that exhibits stable nonfouling characteristics.
Assuntos
Durapatita , Polímeros , Adsorção , Polímeros/química , Polietilenoglicóis/química , Proteínas , Propriedades de SuperfícieRESUMO
Nanocrystalline apatites have been intensively studied for decades, not only for their well-known mimesis of bone apatite but also for applicative purposes, whether as biomaterials for skeletal repair or more recently for a variety of nanomedical applications enabled by their peculiar surface characteristics. Particularly, ion-doped apatites are of great interest because the incorporation of foreign ions in the composition of apatite (nano)crystals alters the bulk and surface properties, modifying their ability to interact with the external environment. This is clearly seen in the physiology of bone tissue, whose mineral phase, a low crystallinity apatitic phase, can dynamically exchange ions with cells, thus driving bone metabolism. Taking bone mineral as a model, the present work describes the development of Mg-doped hydroxyapatite nanoparticles, exploiting hydrothermal synthesis to achieve extents of Mg2+ doping hardly achieved before and using citrate to develop stable apatite colloidal dispersions. Morphological and physicochemical analyses, associated with in-depth investigation of ions populating the apatitic lattice and the nonapatitic surface layer, concurred to demonstrate the cooperative presence of Mg2+ and citrate ions, affecting the dynamic ion retention/release mechanisms. Achieving high Mg2+ doping rates and understanding how Mg doping translates into surface activation of apatite-based nanoparticles is expected to foster the design of novel smart and tunable devices, to adsorb and release ionic species and cargo molecules, with potential innovations in the biomedical field or even beyond, as in catalysis or for environmental remediation.
Assuntos
Magnésio , Nanopartículas , Nanopartículas/química , Magnésio/química , Apatitas/química , Durapatita/química , Propriedades de Superfície , Osso e Ossos/química , Tamanho da PartículaRESUMO
Osteocalcin is the most abundant noncollagenous bone protein and the functions in bone remineralization as well as in inhibition of bone growth have remained unclear. In this contribution, we explain the dual role of osteocalcin in the nucleation of new calcium phosphate during bone remodeling and in the inhibition of hydroxyapatite crystal growth at the molecular scale. The mechanism was derived using pH-resolved all-atom models for the protein, phosphate species, and hydroxyapatite, along with molecular dynamics simulations and experimental and clinical observations. Osteocalcin binds to (hkl) hydroxyapatite surfaces through multiple residues, identified in this work, and the fingerprint of binding residues varies as a function of the (hkl) crystal facet and pH value. On balance, the affinity of osteocalcin to hydroxyapatite slows down crystal growth. The unique tricalcium γ-carboxylglutamic acid (Gla) domain hereby rarely adsorbs to hydroxyapatite surfaces and faces instead toward the solution. The Gla domain enables prenucleation of calcium phosphate for new bone formation at a slightly acidic pH of 5. The growth of prenucleation clusters of calcium phosphate continues upon increase in pH value from 5 to 7 and is much less favorable, or not observed, on the native osteocalcin structure at and above neutral pH values of 7. The results provide mechanistic insight into the early stages of bone remodeling from the molecular scale, help inform mutations of osteocalcin to modify binding to apatites, support drug design, and guide toward potential cures for osteoporosis and hyperosteogeny.
Assuntos
Osso e Ossos , Durapatita , Osteocalcina/genética , Osteocalcina/química , Osteocalcina/metabolismo , Osso e Ossos/metabolismo , Fosfatos de Cálcio/farmacologiaRESUMO
Breast cancer is a common malignant tumor arising in normal mammary epithelial tissues. Nearly 75% of the patients with advanced mammary cancer develop bone metastases, resulting in secondary tumor growth, osteolytic bone degradation, and poor prognosis. The bone matrix comprises a highly hierarchical architecture and is composed of a nonmineral organic part, a predominantly type-I collagen, and a mineral inorganic part composed of hydroxyapatite (HA) nanocrystals (Ca10(PO4)6(OH)2). Although there has been extensive research indicating that the material properties of bone minerals affect metastatic breast cancer, it remains unclear how the microenvironment of the bone matrix, such as the roughness, which changes as a result of osteolytic bone remodeling, affects this disease. In this study, we created HA coatings in situ on polyelectrolyte multilayers (PEMs) by incubating PEMs in a mixture of phosphate and calcium ions. The HA films with distinctive roughness were successfully collected by controlling the incubation time, which served as the simulated microenvironment of the bone matrix. MDA-MB231 breast cancer cells were cultured on HA films, and an optimal roughness was observed in the adhesion, proliferation, and expression of two cytokines closely related to bone metastasis. This study contributed to the understanding of the effect of the microenvironment of the bone matrix, such as the roughness, on the metastasis behavior of breast cancer.
Assuntos
Neoplasias Ósseas , Neoplasias da Mama , Durapatita , Durapatita/química , Humanos , Neoplasias da Mama/patologia , Neoplasias Ósseas/secundário , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Feminino , Microambiente Tumoral/efeitos dos fármacos , Propriedades de Superfície , Proliferação de Células/efeitos dos fármacos , Adesão Celular/efeitos dos fármacosRESUMO
Severe bone defects resulting from trauma and diseases remain a persistent clinical challenge. In this study, a hierarchical biomimetic microporous hydrogel composite scaffold was constructed by mimicking the hierarchical structure of bone. Initially, gelatin methacrylamide (GelMA) and methacrylic anhydride silk fibroin (SilMA) were synthesized, and GelMA/SilMA inks with suitable rheological and mechanical properties were prepared. Biomimetic micropores were then generated by using an aqueous two-phase emulsification method. Subsequently, biomimetic microporous GelMA/SilMA was mixed with hydroxyapatite (HAp) to prepare biomimetic microporous GelMA/SilMA/HAp ink. Hierarchical biomimetic microporous GelMA/SilMA/HAp (M-GSH) scaffolds were then fabricated through digital light processing (DLP) 3D printing. Finally, in vitro experiments were conducted to investigate cell adhesion, proliferation, and inward migration as well as osteogenic differentiation and vascular regeneration effects. In vivo experiments indicated that the biomimetic microporous scaffold significantly promoted tissue integration and bone regeneration after 12 weeks of implantation, achieving 42.39% bone volume fraction regeneration. In summary, this hierarchical biomimetic microporous scaffold provides a promising strategy for the repair and treatment of bone defects.
Assuntos
Acrilamidas , Durapatita , Alicerces Teciduais , Durapatita/química , Alicerces Teciduais/química , Gelatina/química , Osteogênese , Biomimética , Regeneração Óssea , Impressão Tridimensional , Engenharia TecidualRESUMO
Bone defects show a slow rate of osteoconduction and imperfect reconstruction, and the current treatment strategies to treat bone defects suffer from limitations like immunogenicity, lack of cell adhesion, and the absence of osteogenic activity. In this context, bioactive supramolecular peptides and peptide gels offer unique opportunities to develop biomaterials that can play a dominant role in the biomineralization of bone tissues and promote bone formation. In this article, we have demonstrated the potential of six tetrapeptides for specific binding to hydroxyapatite (HAp), a major inorganic component of the bone, and their effect on the growth and osteogenic differentiation of mesenchymal stem cells (MSCs). We adopted a simplistic approach of rationally designing amphiphilic peptides by incorporating amino acids, Ser, pSer, Pro, Hyp, Asp, and Glu, which are present in either collagenous or noncollagenous proteins and render properties like antioxidant, calcification, and mineralization. A total of six tetrapeptides, Trp-Trp-His-Ser (WWHS), Trp-Trp-His-pSer (WWHJ), Trp-Trp-His-Pro (WWHP), Trp-Trp-His-Hyp (WWHO), Trp-Trp-His-Asp (WWHD), and Trp-Trp-His-Glu (WWHE), were synthesized. Four peptides were found to self-assemble into nanofibrillar gels resembling the extracellular matrix (ECM), and the remaining two peptides (WWHJ, WWHP) self-assembled into nanorods. The peptides showed excellent cell adhesion, encapsulation, proliferation, and migration and induced the differentiation of mesenchymal stem cells (MSCs), as evident from the enhanced mineralization, resulting from the upregulation of osteogenic markers, RUNX 2, COL I, OPN, and OCN, alkaline phosphatase (ALP) production, and calcium deposition. The peptides also induced the downregulation of inflammatory markers, TNF-α and iNOS, and the upregulation of the anti-inflammatory marker, IL-10, resulting in M2 macrophage polarization. RANKL and TRAP genes were downregulated in a coculture system of MC3T3-E1 and RAW 264.7 cells, implying that peptides promote osteogenesis and inhibit osteoclastogenesis. The peptide-based biomaterials developed in this work can enhance bone regeneration capacity and show strong potential as scaffolds for bone tissue engineering.
Assuntos
Células-Tronco Mesenquimais , Osteogênese , Aminoácidos/metabolismo , Regeneração Óssea , Materiais Biocompatíveis/farmacologia , Materiais Biocompatíveis/metabolismo , Diferenciação Celular , Durapatita/química , Peptídeos/farmacologia , Peptídeos/metabolismo , Géis/farmacologia , Células CultivadasRESUMO
The effective regeneration of large bone defects via bone tissue engineering is challenging due to the difficulty in creating an osteogenic microenvironment. Inspired by the fibrillar architecture of the natural extracellular matrix, we developed a nanoscale bioengineering strategy to produce bone fibril-like composite scaffolds with enhanced osteogenic capability. To activate the surface for biofunctionalization, self-adaptive ridge-like nanolamellae were constructed on poly(ε-caprolactone) (PCL) electrospinning scaffolds via surface-directed epitaxial crystallization. This unique nanotopography with a markedly increased specific surface area offered abundant nucleation sites for Ca2+ recruitment, leading to a 5-fold greater deposition weight of hydroxyapatite than that of the pristine PCL scaffold under stimulated physiological conditions. Bone marrow mesenchymal stem cells (BMSCs) cultured on bone fibril-like scaffolds exhibited enhanced adhesion, proliferation, and osteogenic differentiation in vitro. In a rat calvarial defect model, the bone fibril-like scaffold significantly accelerated bone regeneration, as evidenced by micro-CT, histological histological and immunofluorescence staining. This work provides the way for recapitulating the osteogenic microenvironment in tissue-engineered scaffolds for bone repair.
Assuntos
Regeneração Óssea , Células-Tronco Mesenquimais , Osteogênese , Poliésteres , Engenharia Tecidual , Alicerces Teciduais , Animais , Alicerces Teciduais/química , Ratos , Regeneração Óssea/efeitos dos fármacos , Células-Tronco Mesenquimais/citologia , Osteogênese/efeitos dos fármacos , Osteogênese/fisiologia , Engenharia Tecidual/métodos , Poliésteres/química , Diferenciação Celular , Ratos Sprague-Dawley , Materiais Biomiméticos/química , Materiais Biomiméticos/farmacologia , Células Cultivadas , Proliferação de Células , Crânio/lesões , Crânio/patologia , Durapatita/química , Durapatita/farmacologiaRESUMO
Few studies tried to explore the mitigation effect and underlying mechanisms of biochar and their complex for negative allelopathy from invasive plants, which may provide a new way in the invasive plant management. Herein, an invasive plant (Solidago canadensis)-derived biochar (IBC) and its composite with hydroxyapatite (HAP/IBC) were synthesized by high temperature pyrolysis, and characterized by scanning electron microscopy, energy dispersion spectrometer, X-ray diffraction, Fourier transform infrared spectroscopy, and X-ray photoelectron spectroscopy. Then, both the batch adsorption and pot experiments were conducted to compare the removal effects of kaempferol-3-O-ß-D-glucoside (C21 H20 O11 , kaempf), an allelochemical from S. canadensis, on IBC and HAP/IBC, respectively. HAP/IBC showed a stronger affinity for kaempf than IBC due to its higher specific surface area, more functional groups (P-O, P-O-P, PO4 3- ), stronger crystallization [Ca3 (PO4 )2 ]. The maximum kaempf adsorption capacity on HAP/IBC was six times higher than on IBC (10.482 mg/g > 1.709 mg/g) via π-π interactions, functional groups, and metal complexation. The kaempf adsorption process could be fitted best by both pseudo-second-order kinetic and Langmuir isotherm models. Furthermore, HAP/IBC addition into soils could enhance and even recover the germination rate and/or seedling growth of tomato inhibited by negative allelopathy from the invasive S. canadensis. These results indicate that the composite of HAP/IBC could more effectively mitigate the allelopathy from S. canadensis than IBC, which may be a potential efficient approach to control the invasive plant and improve invaded soils.