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1.
Anal Bioanal Chem ; 412(19): 4681-4690, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32451642

RESUMO

A new straightforward gel permeation chromatography (GPC) method was developed to calculate the drug encapsulation efficiency and loading content of Poly(lactic acid) nanoparticles (PLA NPs) loaded with Salinomycin (Sal), exploiting the capability of this technique to separate a macromolecular/molecular mixture on the basis of the molecular weight of each component. The proposed GPC method allowed Sal detection until 1% of Sal content in PLA NPs, avoiding sample pre-treatments. The method was validated by wave voltammetry (SW) technique, using a slightly modified literature procedure, useful to detect Sal in the concentration range 0.4 ≤ C/µmol/L ≤ 12 (linear concentration range). PLA-based NPs were prepared by nanoprecipitation with either native and functionalized PLA. Specifically, folate-decorated PLA NPs (PLA-FA NPs) were obtained by CuAAC click functionalization of alkyne-grafted PLA with azide-folate. Sal-loaded NPs were characterized physicochemically and morphologically. They exhibited adequate physicochemical properties, good drug encapsulation efficiency (98 ± 0.5% and 99 ± 0.5%), and loading content (8.8 ± 0.1% and 8.9 ± 0.1% for PLA/Sal and PLA-FA/Sal NPs, respectively). The size of empty PLA NPs resulted smaller (90 ± 3.2 nm and 680 ± 15.3 nm, for PLA NPs and PLA-FA NPs respectively) than the correspondent drug-loaded NPs (110 ± 3.8 nm and 875 ± 20.5 nm, respectively). Their biological activity was assessed on osteosarcoma bulk cells MG63, healthy osteoblast cell line (hFOB1.19), and enriched osteosarcoma cancer stem cells (CSCs), showing cell-depending effect. Entrapped Sal maintained its cytotoxic effect on CSCs and MG63 cells, with a potency comparable to the free drug and no evident benefit was detected for folate-decorated PLA NPs respect to native PLA NPs. Graphical abstract.

2.
J Mater Sci Mater Med ; 30(12): 136, 2019 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-31802234

RESUMO

Many medical-related scientific discoveries arise from trial-error patterns where the processes involved must be refined and modified continuously before any product could be able to reach the final costumers. One of the elements affecting negatively these processes is the inaccuracy of two-dimension (2D) standard culture systems, carried over in plastic plates or similar, in replicating complex environments and patterns. Consequently, animal tests are required to validate every in vitro finding, at the expenses of more funds and ethical issues. A possible solution relies in the implementation of three-dimension (3D) culture systems as a fitting gear between the 2D tests and in vivo tests, aiming to reduce the negative in vivo outcomes. These 3D structures are depending from the comprehension of the extracellular matrix (ECM) and the ability to replicate it in vitro. In this article a comparison of efficacies between these two culture systems was taken as subject, human mesenchymal stem cells (hMSCs) was utilized and a hybrid scaffold made by a blend of chitosan, gelatin and biomineralized gelatin was used for the 3D culture system.


Assuntos
Técnicas de Cultura de Células , Células-Tronco Mesenquimais/fisiologia , Osteogênese/fisiologia , Tecidos Suporte , Materiais Biocompatíveis , Diferenciação Celular , Humanos , Teste de Materiais
3.
Front Cell Dev Biol ; 7: 268, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31799249

RESUMO

The regeneration of bone fractures, resulting from trauma, osteoporosis or tumors, is a major problem in our super-aging society. Bone regeneration is one of the main topics of concern in regenerative medicine. In recent years, stem cells have been employed in regenerative medicine with interesting results due to their self-renewal and differentiation capacity. Moreover, stem cells are able to secrete bioactive molecules and regulate the behavior of other cells in different host tissues. Bone regeneration process may improve effectively and rapidly when stem cells are used. To this purpose, stem cells are often employed with biomaterials/scaffolds and growth factors to accelerate bone healing at the fracture site. Briefly, this review will describe bone structure and the osteogenic differentiation of stem cells. In addition, the role of mesenchymal stem cells for bone repair/regrowth in the tissue engineering field and their recent progress in clinical applications will be discussed.

4.
Aging Clin Exp Res ; 2019 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-31595428

RESUMO

The aging of the world population is increasingly claimed as an alarming situation, since an ever-raising number of persons in advanced age but still physically active is expected to suffer from invalidating and degenerative diseases. The impairment of the endogenous healing potential provoked by the aging requires the development of more effective and personalized therapies, based on new biomaterials and devices able to direct the cell fate to stimulate and sustain the regrowth of damaged or diseased tissues. To obtain satisfactory results, also in cases where the cell senescence, typical of the elderly, makes the regeneration process harder and longer, the new solutions have to possess excellent ability to mimic the physiological extracellular environment and thus exert biomimetic stimuli on stem cells. To this purpose, the "biomimetic concept" is today recognized as elective to fabricate bioactive and bioresorbable devices such as hybrid osteochondral scaffolds and bioactive bone cements closely resembling the natural hard tissues and with enhanced regenerative ability. The review will illustrate some recent results related to these new biomimetic materials developed for application in different districts of the musculoskeletal system, namely bony, osteochondral and periodontal regions, and the spine. Further, it will be shown how new bioactive and superparamagnetic calcium phosphate nanoparticles can give enhanced results in cardiac regeneration and cancer therapy. Since tissue regeneration will be a major demand in the incoming decades, the high potential of biomimetic materials and devices is promising to significantly increase the healing rate and improve the clinical outcomes even in aged patients.

5.
Mater Sci Eng C Mater Biol Appl ; 96: 234-247, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30606529

RESUMO

The present work describes biomimetic hybrid microspheres made of collagen type I-like peptide matrix (RCP) mineralised with Fe2+/Fe3+ doping hydroxyapatite (RCPFeHA) by a bio-inspired process. Superparamagnetic RCPFeHA microspheres are obtained by emulsification of the hybrid slurries in the presence of citrate ions, to achieve a biomimetic surface functionalisation improving the bioactivity and the dispersion ability in cell culture medium. A biological in vitro study correlates the osteoblast cells behaviour to calcium and iron ions released by the hybrid microspheres in culture media mimicking physiological or inflammatory environment, evidencing a clear triggering of cell activity and bio-resorption ability. In presence of the microspheres, the osteoblast cells maintain their typical morphology and no cell damage were detected, whereas also showing up-regulation of osteogenic markers. The ability of the hybrid microspheres to undergo bio-resorption and release bioactive ions in response to different environmental stimuli without harmful effects opens new perspectives in bone regeneration, as magnetically active bone substitute with potential ability of drug carrier and smart response in the presence of inflammatory states.


Assuntos
Materiais Biomiméticos/química , Substitutos Ósseos/química , Óxido Ferroso-Férrico/química , Campos Magnéticos , Microesferas , Osteoblastos/metabolismo , Animais , Linhagem Celular , Camundongos , Osteoblastos/citologia
6.
Biomater Sci ; 7(1): 307-321, 2018 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-30468436

RESUMO

The present work investigates heterogeneous gas-solid reactions involved in the biomorphic transformation of natural wood into large 3-D hydroxyapatite (HA) scaffolds recapitulating physico-chemical, morphological and mechanical features typical of natural bone. In particular, we found that the use of a reactive CO2/H2O gas mixture, under supercritical conditions at high pressure, permits to control heterogeneous CaO-CO2 reactions throughout the whole bulk and to direct the nucleation-growth of CaCO3 at a relatively low temperature, thus obtaining a highly reactive 3-D precursor enabling the formation of a large biomorphic HA scaffold preserving fine nanostructure by a hydrothermal process. To the best of our knowledge, the application of heterogeneous chemical reactions in the 3-D state is an original way to generate large HA scaffolds maintaining bio-relevant ionic substitutions, with specific regard to Mg2+, Sr2+ and CO32- ions, conferring a superior ability to guide cell fate. We hypothesize that the original nanostructure of the final 3-D HA scaffold, not achievable by the classic sintering procedure, and the multi-scale hierarchical organization inherited by the original template, account for its high compression strength with damage-tolerant mechanical behaviour. The ability of the new scaffold to induce bone regeneration is attested by the overexpression of genes, early and late markers of the osteogenic differentiation pathway, and by the in vivo osteoinductivity. We hypothesize that the unique association of bioactive chemical composition, nanostructure and multi-scale hierarchy can synergistically act as instructing signals for cells to generate new bone tissue with organized 3-D architecture. These results point to its great applicative potential for the regeneration of large bone defects, which is a still unmet clinical need.


Assuntos
Substitutos Ósseos/química , Durapatita/química , Tecidos Suporte/química , Animais , Regeneração Óssea , Células Cultivadas , Força Compressiva , Teste de Materiais , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos C57BL , Osteogênese , Coelhos , Engenharia Tecidual
7.
Langmuir ; 34(40): 12036-12048, 2018 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-30204449

RESUMO

Nanocrystalline apatites mimicking bone mineral represent a versatile platform for biomedical applications thanks to their similarity to bone apatite and the possibility to (multi)functionalize them so as to provide "à la carte" properties. One relevant domain is in particular oncology, where drug-loaded biomaterials and engineered nanosystems may be used for diagnosis, therapy, or both. In a previous contribution, we investigated the adsorption of doxorubicin onto two nanocrystalline apatite substrates, denoted HA and FeHA (superparamagnetic apatite doped with iron ions), and explored these drug-loaded systems against tumor cells. To widen their applicability in the oncology field, here we examine the interaction between the same two substrates and two other molecules: folic acid (FA), often used as cell targeting agent, and the anticancer drug methotrexate (MTX), an antifolate analogue. In a first stage, we investigated the adsorptive behavior of FA (or MTX) on both substrates, evidencing their specificities. At low concentration, typically under 100 mmol/L, adsorption onto HA was best described using the Sips isotherm model, while the formation of a calcium folate secondary salt was evidenced at high concentration by Raman spectroscopy. Adsorption onto FeHA was instead fitted to the Langmuir model. A larger adsorptive affinity was found for the FeHA substrate compared to HA; accordingly, a faster release was noticed from HA. In vitro tests carried out on human osteosarcoma cell line (SAOS-2) allowed us to evaluate the potential of these compounds in oncology. Finally, in vivo (subcutaneous) implantations in the mouse were run to ascertain the biocompatibility of the two substrates. These results should allow a better understanding of the interactions between FA/MTX and bioinspired nanocrystalline apatites in view of applications in the field of cancer.


Assuntos
Antineoplásicos/farmacologia , Antagonistas do Ácido Fólico/farmacologia , Ácido Fólico/química , Hidroxiapatitas/química , Metotrexato/farmacologia , Adsorção , Animais , Antineoplásicos/química , Materiais Biocompatíveis/química , Materiais Biocompatíveis/toxicidade , Linhagem Celular Tumoral , Liberação Controlada de Fármacos , Antagonistas do Ácido Fólico/química , Humanos , Hidroxiapatitas/toxicidade , Metotrexato/química , Camundongos Endogâmicos C57BL , Nanopartículas/química , Nanopartículas/toxicidade
8.
Int J Biol Macromol ; 106: 739-748, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28827204

RESUMO

This study focuses on the development of novel bone-like scaffolds by bio-inspired, pH-driven, mineralization of type I collagen matrix with magnesium-doped hydroxyapatite nanophase (MgHA/Coll). To this aim, this study evaluates the altered modifications in the obtained composite due to different crosslinkers such as dehydrothermal treatment (DHT), 1,4-butanediol diglycidyl ether (BDDGE) and ribose in terms of morphological, physical-chemical and biological properties. The physical-chemical properties of the composites evaluated by XRD, FTIR, ICP and TGA demonstrated that the chemical mimesis of bone was effectively achieved using the in-lab biomineralization process. Furthermore, the presence of various crosslinkers greatly promoted beneficial enzymatic resistivity and swelling ability. The morphological results revealed highly porous and fibrous micro-architecture with total porosity above 85% with anisotropic pore size within the range of 50-200µm in all the analysed composites. The mechanical behaviour in response to compressive forces demonstrated enhanced compressive modulus in all crosslinked composites, suggesting that mechanical behaviour is largely dependent on the type of crosslinker used. The biomimetic compositional and morphological features of the composites elicited strong cell-material interaction. Therefore, the results showed that by activating specific crosslinking mechanisms, hybrid composites can be designed and tailored to develop tissue-specific biomimetic biomaterials for hard tissue engineering.


Assuntos
Colágeno/química , Reagentes para Ligações Cruzadas/química , Durapatita/química , Medicina Regenerativa , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/uso terapêutico , Biomimética , Butileno Glicóis/química , Colágeno/uso terapêutico , Durapatita/uso terapêutico , Porosidade , Engenharia Tecidual/métodos , Tecidos Suporte/química
9.
Biomed Mater ; 12(5): 055002, 2017 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-28573980

RESUMO

In this study, ribose was proposed as a promising, non-toxic, low-cost crosslinker to enhance the structural integrity and stiffness of type I collagen matrices. The main objective was to determine the optimal conditions of glycation by ribose to fabricate 3D porous collagen scaffolds and to verify their effectiveness for use as scaffolds for cartilage tissue engineering, by physicochemical and biological characterization. Two different crosslinking strategies were investigated including variation in the amount of ribose and the time of reaction: pre-crosslinking (PRE) and post-crosslinking (POST). All ribose-glycated collagen scaffolds demonstrated good swelling properties and interconnected porous microstructure suitable for cell growth and colonization. The POST samples were superior to PRE, in terms of porosity, degree of crosslinking, fluid uptake ability, and resistance to enzymatic digestion. Moreover, the mechanical properties of the scaffolds were significantly improved upon glycation when compared to non-crosslinked collagen, manifesting the best performance for POST matrices crosslinked for 5 d and in the highest amount of sugar. In vitro studies analyzing cell-material interactions revealed scaffold cytocompatibility with higher cell viability and cell proliferation as well as higher glycosaminoglycan secretion for POST scaffolds with respect to PRE. This report demonstrated the feasibility of developing 3D collagen scaffolds by ribose glycation and highlighted the POST-crosslinking strategy as being more favorable than the PRE-crosslinking to achieve scaffolds suitable for cartilage regeneration.


Assuntos
Colágeno/química , Ribose/química , Engenharia Tecidual , Tecidos Suporte/química , Animais , Materiais Biocompatíveis/química , Cartilagem/metabolismo , Cartilagem/patologia , Proliferação de Células , Sobrevivência Celular , Colágeno Tipo I/química , Colagenases/química , Reagentes para Ligações Cruzadas/química , Glicosaminoglicanos/química , Concentração de Íons de Hidrogênio , Teste de Materiais , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Eletrônica de Varredura , Porosidade , Pressão , Regeneração , Espectroscopia de Infravermelho com Transformada de Fourier , Estresse Mecânico , Temperatura
10.
Tissue Eng Part A ; 23(23-24): 1423-1435, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28637399

RESUMO

The need of synthetic bone grafts that recreate from macro- to nanoscale level the biochemical and biophysical cues of bone extracellular matrix has been a major driving force for the development of new generation of biomaterials. In this study, synthetic bone substitutes have been synthesized via biomimetic mineralization of a recombinant collagen type I-derived peptide (RCP), enriched in tri-amino acid sequence arginine-glycine-aspartate (RGD). Three-dimensional (3D) isotropic porous scaffolds of three different compositions are developed by freeze-drying: non-mineralized (RCP, as a control), mineralized (Ap/RCP), and mineralized scaffolds in the presence of magnesium (MgAp/RCP) that closely imitate bone composition. The effect of mineral phase on scaffold pore size, porosity, and permeability, as well as on their in vitro kinetic degradation, is evaluated. The ultimate goal is to investigate how chemical (i.e., surface chemistry and ion release from scaffold) together with physical signals (i.e., surface nanotopography) conferred via biomimetic mineralization can persuade and guide mesenchymal stem cell (MSC) interaction and fate. The three scaffold compositions showed optimum pore size and porosity for osteoconduction, without significant differences between them. The degradation tests confirmed that MgAp/RCP scaffolds presented higher reactivity under physiological condition compared to Ap/RCP ones. The in vitro study revealed an enhanced cell growth and proliferation on MgAp/RCP scaffolds at day 7, 14, and 21. Furthermore, MgAp/RCP scaffolds potentially promoted cell migration through the inner areas reaching the bottom of the scaffold after 14 days. MSCs cultured on MgAp/RCP scaffolds displayed higher gene and protein expressions of osteogenic markers when comparing them with the results of those MSCs grown on RCP or Ap/RCP scaffolds. This work highlights that mineralization of recombinant collagen mimicking bone mineral composition and morphology is a versatile approach to design smart scaffold interface in a 3D model guiding MSC fate.


Assuntos
Materiais Biomiméticos , Osso e Ossos/química , Calcificação Fisiológica , Diferenciação Celular/efeitos dos fármacos , Colágeno , Células-Tronco Mesenquimais/metabolismo , Tecidos Suporte/química , Animais , Materiais Biomiméticos/química , Materiais Biomiméticos/farmacologia , Colágeno/química , Colágeno/farmacologia , Células-Tronco Mesenquimais/citologia , Camundongos , Proteínas Recombinantes/química , Proteínas Recombinantes/farmacologia
11.
Mater Sci Eng C Mater Biol Appl ; 77: 594-605, 2017 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-28532070

RESUMO

This study explores for the first time the application of ribose as a highly biocompatible agent for the crosslinking of hybrid mineralized constructs, obtained by bio-inspired mineralization of self-assembling Type I collagen matrix with magnesium-doped-hydroxyapatite nanophase, towards a biomimetic mineralized 3D scaffolds (MgHA/Coll) with excellent compositional and structural mimicry of bone tissue. To this aim, two different crosslinking mechanisms in terms of pre-ribose glycation (before freeze drying) and post-ribose glycation (after freeze drying) were investigated. The obtained results explicate that with controlled freeze-drying, highly anisotropic porous structures with opportune macro-micro porosity are obtained. The physical-chemical features of the scaffolds characterized by XRD, FTIR, ICP and TGA demonstrated structural mimicry analogous to the native bone. The influence of ribose greatly assisted in decreasing solubility and increased enzymatic resistivity of the scaffolds. In addition, enhanced mechanical behaviour in response to compressive forces was achieved. Preliminary cell culture experiments reported good cytocompatibility with extensive cell adhesion, proliferation and colonization. Overall, scaffolds developed by pre-ribose glycation process are preferred, as the related crosslinking technique is more facile and robust to obtain functional scaffolds. As a proof of concept, we have demonstrated that ribose crosslinking is cost-effective, safe and functionally effective. This study also offers new insights and opportunities in developing promising scaffolds for bone tissue engineering.


Assuntos
Biomimética , Proliferação de Células , Colágeno , Durapatita , Porosidade , Ribose , Engenharia Tecidual , Tecidos Suporte
12.
Inorg Chem ; 56(8): 4447-4459, 2017 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-28379709

RESUMO

Doping of biocompatible nanomaterials with magnetic phases is currently one of the most promising strategies for the development of advanced magnetic biomaterials. However, especially in the case of iron-doped magnetic hydroxyapatites, it is not clear if the magnetic features come merely from the magnetic phases/ions used as dopants or from complex mechanisms involving interactions at the nanoscale. Here, we report an extensive chemical-physical and magnetic investigation of three hydroxyapatite nanocrystals doped with different iron species and containing small or no amounts of maghemite as a secondary phase. The association of several investigation techniques such as X-ray absorption spectroscopy, Mössbauer, magnetometry, and TEM allowed us to determine that the unusual magnetic properties of Fe2+/3+-doped hydroxyapatites (FeHA) occur by a synergy of two different phenomena: i.e., (i) interacting superparamagnetism due to the interplay between iron-doped apatite and iron oxide nanoparticles as well as to the occurrence of dipolar interactions and (ii) interacting paramagnetism due to Fe3+ ions present in the superficial hydrated layer of the apatite nanophase and, to a lesser extent, paramagnetism due to isolated Fe3+ ions in the apatite lattice. We also show that a major player in the activation of the above phenomena is the oxidation of Fe2+ into Fe3+, as induced by the synthesis process, and their consequent specific positioning in the FeHA structure.


Assuntos
Hidroxiapatitas/química , Ferro/química , Fenômenos Magnéticos , Nanopartículas/química , Tamanho da Partícula , Propriedades de Superfície
13.
PLoS One ; 12(2): e0172100, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28196118

RESUMO

Strontium-substituted apatitic bone cements enriched with sodium alginate were developed as a potential modulator of bone cells fate. The biological impact of the bone cement were investigated in vitro through the study of the effect of the nanostructured apatitic composition and the doping of strontium on mesenchymal stem cells, pre-osteoblasts and osteoclasts behaviours. Up to 14 days of culture the bone cells viability, proliferation, morphology and gene expression profiles were evaluated. The results showed that different concentrations of strontium were able to evoke a cell-specific response, in fact an inductive effect on mesenchymal stem cells differentiation and pre-osteoblasts proliferation and an inhibitory effect on osteoclasts activity were observed. Moreover, the apatitic structure of the cements provided a biomimetic environment suitable for bone cells growth. Therefore, the combination of biological features of this bone cement makes it as promising biomaterials for tissue regeneration.


Assuntos
Cimentos para Ossos/farmacologia , Proliferação de Células/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Estrôncio/farmacologia , Animais , Cimentos para Ossos/química , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Teste de Materiais , Células-Tronco Mesenquimais/citologia , Camundongos , Osteoblastos/citologia , Estrôncio/química
14.
Int J Biol Macromol ; 95: 1199-1209, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27836656

RESUMO

The present study aims to investigate the physical-chemical and biological features exhibited by porous scaffolds for regeneration of cartilaginous tissues obtained through stabilization of 3D gelatin hydrogels by physical (DHT), chemical (BDDGE) and natural (Genipin) cross-linking approaches. The study aimed at comparatively assessing the porous microstructure and the long-term resistance of the scaffolds upon degradation in wet physiological conditions (37°C, pH=7.4). The degree of cross-linking increases as function of incorporation of cross-linkers which was maximum up to 73% for BDDGE. The infrared spectroscopy and thermal analysis confirmed the gelatin structure was preserved during the cross-linking treatments. Mechanical properties of the scaffolds were analysed by static and dynamic compression test, which showed different viscoelastic behaviour upon various cross-linking strategies. The biological performance of the scaffolds investigated using human chondrocytes showed good cell adhesion, viability and proliferation, as well as extensive 3D scaffold colonization. Besides, the analysis of gene expression related to the formation of new chondral tissue reported increasing ability with time in the formation of new extra-cellular matrix. In conclusion, out of three different cross-linking methods, the gelatin scaffolds subjected to dehydrothermal treatment (DHT) represented to be the most favourable 3D scaffold for cartilage regeneration.


Assuntos
Butileno Glicóis/química , Reagentes para Ligações Cruzadas/química , Gelatina/química , Iridoides/química , Engenharia Tecidual/métodos , Tecidos Suporte , Animais , Adesão Celular/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Condrócitos/citologia , Condrócitos/efeitos dos fármacos , Elasticidade , Gelatina/isolamento & purificação , Gelatina/farmacologia , Temperatura Alta , Humanos , Concentração de Íons de Hidrogênio , Porosidade , Pele/química , Suínos , Viscosidade
15.
Mol Med Rep ; 14(5): 4248-4254, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27748851

RESUMO

Skeletal unloading leads to hypoxia in the bone microenvironment, resulting in imbalanced bone remodeling that favors bone resorption. Osteocytes, the mechanosensors of bone, have been demonstrated to orchestrate bone homeostasis. Hypoxic osteocytes either undergo apoptosis or actively stimulate osteoclasts to remove bone matrix during hypoxia. Oxygen­regulated protein 150 (ORP150) is an endoplasmic reticulum­associated chaperone that has been observed to serve an important role in the cellular adaptation to hypoxia and in preventing cellular apoptosis in various tissue types. The current study hypotheses that expression of ORP150 would be increased in osteocytes under hypoxic conditions (1% O2). The MLO­Y4 osteocyte cell line was cultured under normoxic or hypoxic conditions for up to 72 h. It was demonstrated that 1% O2 significantly induced hypoxia after 16 h and up to 72 h, significantly reduced cell number at 8 and 48 h, induced cell death at 8, 24 and 48 h and induced apoptosis at 16, 24 and 48 h. Significant differences in ORP150 mRNA were observed at 72 h, however no differences were observed in the protein expression levels. The relative increase in ORP150 mRNA observed in hypoxia, compared with normoxia, may support its cytoprotective role in oxygen­deprived conditions.


Assuntos
Remodelação Óssea/genética , Reabsorção Óssea/genética , Proteínas de Choque Térmico HSP70/genética , Proteômica , Animais , Apoptose/genética , Reabsorção Óssea/metabolismo , Reabsorção Óssea/patologia , Hipóxia Celular/genética , Microambiente Celular/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Choque Térmico HSP70/biossíntese , Elevação dos Membros Posteriores , Camundongos , Osteoclastos/metabolismo , Osteócitos/metabolismo , Osteócitos/patologia , Oxigênio/metabolismo
16.
Artigo em Inglês | MEDLINE | ID: mdl-27446909

RESUMO

The development of bio-devices for complete regeneration of ligament and tendon tissues is presently one of the biggest challenges in tissue engineering. Such device must simultaneously possess optimal mechanical performance, suitable porous structure, and biocompatible microenvironment. This study proposes a novel collagen-BDDGE-elastin (CBE)-based device for tendon tissue engineering, by the combination of two different modules: (i) a load-bearing, non-porous, "core scaffold" developed by braiding CBE membranes fabricated via an evaporative process and (ii) a hollow, highly porous, "shell scaffold" obtained by uniaxial freezing followed by freeze-drying of CBE suspension, designed to function as a physical guide and reservoir of cells to promote the regenerative process. Both core and shell materials demonstrated good cytocompatibility in vitro, and notably, the porous shell architecture directed cell alignment and population within the sample. Finally, a prototype of the core module was implanted in a rat tendon lesion model, and histological analysis demonstrated its safety, biocompatibility, and ability to induce tendon regeneration. Overall, our results indicate that such device may have the potential to support and induce in situ tendon regeneration.

17.
J Struct Biol ; 196(2): 138-146, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27374321

RESUMO

Understanding the mineralization mechanism of synthetic protein has recently aroused great interest especially in the development of advanced materials for bone regeneration. Herein, we propose the synthesis of composite materials through the mineralization of a recombinant collagen type I derived protein (RCP) enriched with RGD sequences in the presence of magnesium ions (Mg) to closer mimic bone composition. The role of both RCP and Mg ions in controlling the precipitation of the mineral phase is in depth evaluated. TEM and X-ray powder diffraction reveal the crystallization of nanocrystalline apatite (Ap) in all the evaluated conditions. However, Raman spectra point out also the precipitation of amorphous calcium phosphate (ACP). This amorphous phase is more evident when RCP and Mg are at work, indicating the synergistic role of both in stabilizing the amorphous precursor. In addition, hybrid matrices are prepared to tentatively address their effectiveness as scaffolds for bone tissue engineering. SEM and AFM imaging show an homogeneous mineral distribution on the RCP matrix mineralized in presence of Mg, which provides a surface roughness similar to that found in bone. Preliminary in vitro tests with pre-osteoblast cell line show good cell-material interaction on the matrices prepared in the presence of Mg. To the best of our knowledge this work represents the first attempt to mineralize recombinant collagen type I derived protein proving the simultaneous effect of the organic phase (RCP) and Mg on ACP stabilization. This study opens the possibility to engineer, through biomineralization process, advanced hybrid matrices for bone regeneration.


Assuntos
Regeneração Óssea , Calcificação Fisiológica , Engenharia Tecidual/métodos , Animais , Apatitas , Biomimética/métodos , Linhagem Celular , Colágeno Tipo I/metabolismo , Magnésio , Camundongos , Minerais , Engenharia de Proteínas
18.
Artigo em Inglês | MEDLINE | ID: mdl-27376060

RESUMO

Tooth loss is a common result of a variety of oral diseases due to physiological causes, trauma, genetic disorders, and aging and can lead to physical and mental suffering that markedly lowers the individual's quality of life. Tooth is a complex organ that is composed of mineralized tissues and soft connective tissues. Dentin is the most voluminous tissue of the tooth and its formation (dentinogenesis) is a highly regulated process displaying several similarities with osteogenesis. In this study, gelatin, thermally denatured collagen, was used as a promising low-cost material to develop scaffolds for hard tissue engineering. We synthetized dentin-like scaffolds using gelatin biomineralized with magnesium-doped hydroxyapatite and blended it with alginate. With a controlled freeze-drying process and alginate cross-linking, it is possible to obtain scaffolds with microscopic aligned channels suitable for tissue engineering. 3D cell culture with mesenchymal stem cells showed the promising properties of the new scaffolds for tooth regeneration. In detail, the chemical-physical features of the scaffolds, mimicking those of natural tissue, facilitate the cell adhesion, and the porosity is suitable for long-term cell colonization and fine cell-material interactions.

19.
Nanomedicine (Lond) ; 11(16): 2119-30, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27463861

RESUMO

AIM: Synthesis of superparamagnetic hybrid nanobeads (MHNs) made of iron-substituted hydroxyapatite nanophase mineralizing a self-assembling alginate (Alg) matrix to be used as drug carriers, with ability of remote activation by magnetic signaling. MATERIALS & METHODS: Iron-doped apatite was heterogeneously nucleated on the self-assembling Alg matrix by a bioinspired mineralization process and MHNs are formed by a subsequent emulsification by oil-in-water technique. RESULTS: The obtained MHNs exhibited biomimetic composition, adequate swelling properties in physiological-like environment and superparamagnetic properties. The assembling of Alg induced the egg-like rearrangement of the mineralized composite that was then stabilized through cross-linking reaction with calcium ions. CONCLUSION: The new MHNs can be considered as a promising biocompatible and bio-resorbable drug delivery system with magnetic properties, thus opening to smart applications in nanomedicine.


Assuntos
Alginatos/química , Portadores de Fármacos/química , Durapatita/química , Ferro/química , Imãs/química , Nanopartículas/química , Cálcio/química , Sistemas de Liberação de Medicamentos , Ácido Glucurônico/química , Ácidos Hexurônicos/química , Nanopartículas/ultraestrutura
20.
J Biomed Nanotechnol ; 12(5): 909-21, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-27305814

RESUMO

Superparamagnetic nanoparticles offer several opportunities in nanomedicine and magnetic cell targeting. They are considered to be an extremely promising approach for the translation of cell-based therapies from the laboratory to clinical studies. In fact, after injection, the magnetic labeled cells could be driven by a static magnetic field and localized to the target site where they can perform their specific role. In this study, innovative iron-doped hydroxyapatite nanoparticles (FeHA NPs) were tested with mesenchymal stem cells (MSCs) as tools for cell therapy. Results showed that FeHA NPs could represent higher cell viability in'respect to commercial superparamagnetic iron oxide nanoparticles (SPION) at four different concentrations ranging from 10 µg/ml up to 200 µg/ml and would also upregulate an early marker involved in commitment and differentiation of MSCs. Moreover, FeHA NPs were uptaken without negatively affecting the cell behavior and their ultrastructure. Thus obtained magnetic cells were easily guided by application of a static magnetic field. This work demonstrates the promising opportunities of FeHA NPs in MSCs labeling due to the unique features of fast degradation and very low iron content of FeHA NPs compared to SPIONs. Likewise, due to the intrinsic properties of FeHA NPs, this approach could be simply transferred to different cell types as an effective magnetic carrier of drugs, growth factors, miRNA, etc., offering favorable prospects in nanomedicine.


Assuntos
Durapatita/farmacologia , Ferro/farmacologia , Magnetismo/métodos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Nanopartículas/química , Coloração e Rotulagem , Animais , Proteína Morfogenética Óssea 2/genética , Proteína Morfogenética Óssea 2/metabolismo , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Endocitose/efeitos dos fármacos , Imunofluorescência , Perfilação da Expressão Gênica , Campos Magnéticos , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/ultraestrutura , Nanopartículas/ultraestrutura , Osteogênese/efeitos dos fármacos , Osteogênese/genética , Tamanho da Partícula , Coelhos , Difração de Raios X
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