Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 10.489
Filtrar
1.
Int J Nanomedicine ; 15: 5825-5838, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32821104

RESUMO

Background and Purpose: The extracellular matrix (ECM) derived from bone marrow mesenchymal stem cells (BMSCs) has been used in regenerative medicine because of its good biological activity; however, its poor mechanical properties limit its application in bone regeneration. The purpose of this study is to construct a three dimensional-printed hydroxyapatite (3D-HA)/BMSC-ECM composite scaffold that not only has biological activity but also sufficient mechanical strength and reasonably distributed spatial structure. Methods: A BMSC-ECM was first extracted and formed into micron-sized particles, and then the ECM particles were modified onto the surface of 3D-HA scaffolds using an innovative linking method to generate composite 3D-HA/BMSC-ECM scaffolds. The 3D-HA scaffolds were used as the control group. The basic properties, biocompatibility and osteogenesis ability of both scaffolds were tested in vitro. Finally, a critical skull defect rat model was created and the osteogenesis effect of the scaffolds was evaluated in vivo. Results: The compressive modulus of the composite scaffolds reached 9.45±0.32 MPa, which was similar to that of the 3D-HA scaffolds (p>0.05). The pore size of the two scaffolds was 305±47 um and 315±34 um (p>0.05), respectively. A CCK-8 assay indicated that the scaffolds did not have cytotoxicity. The composite scaffolds had good cell adhesion ability, with a cell adhesion rate of up to 76.00±6.17% after culturing for 7 hours, while that of the 3D-HA scaffolds was 51.85±4.77% (p<0.01). In addition, the composite scaffold displayed higher alkaline phosphatase (ALP) activity, osteogenesis-related mRNA expression, and calcium nodule formation, thus confirming that the composite scaffolds had good osteogenic activity. The composite scaffolds exhibited good bone repair in vivo and were superior to the 3D-HA scaffolds. Conclusion: We conclude that BMSC-ECM is a good osteogenic material and that the composite scaffolds have good osteogenic ability, which provides a new method and concept for the repair of bone defects.


Assuntos
Durapatita/farmacologia , Matriz Extracelular/metabolismo , Células-Tronco Mesenquimais/citologia , Tecidos Suporte/química , Animais , Regeneração Óssea/efeitos dos fármacos , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/patologia , Adesão Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Matriz Extracelular/efeitos dos fármacos , Hidrodinâmica , Masculino , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/ultraestrutura , Osteogênese/efeitos dos fármacos , Osteogênese/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Cicatrização/efeitos dos fármacos
2.
PLoS One ; 15(8): e0237116, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32857787

RESUMO

Bone metastases are a frequent complication in prostate cancer, and several studies have shown that vitamin D deficiency promotes bone metastases. However, while many studies focus on vitamin D's role in cell metabolism, the effect of chronically low vitamin D levels on bone tissue, i.e. insufficient mineralization of the tissue, has largely been ignored. To investigate, whether poor tissue mineralization promotes cancer cell attachment, we used a fluorescence based adhesion assay and single cell force spectroscopy to quantify the adhesion of two prostate cancer cell lines to well-mineralized and demineralized dentin, serving as biomimetic bone model system. Adhesion rates of bone metastases-derived PC3 cells increased significantly on demineralized dentin. Additionally, on mineralized dentin, PC3 cells adhered mainly via membrane anchored surface receptors, while on demineralized dentin, they adhered via cytoskeleton-anchored transmembrane receptors, pointing to an interaction via exposed collagen fibrils. The adhesion rate of lymph node derived LNCaP cells on the other hand is significantly lower than that of PC3 and not predominately mediated by cytoskeleton-linked receptors. This indicates that poor tissue mineralization facilitates the adhesion of invasive cancer cells by the exposure of collagen and emphasizes the disease modifying effect of sufficient vitamin D for cancer patients.


Assuntos
Calcificação Fisiológica , Adesão Celular , Neoplasias da Próstata/metabolismo , Animais , Materiais Biomiméticos/química , Linhagem Celular Tumoral , Colágeno/metabolismo , Citoesqueleto/metabolismo , Dentina/química , Elefantes , Humanos , Masculino , Receptores de Superfície Celular/metabolismo , Tecidos Suporte/química , Vitamina D/metabolismo
3.
Small ; 16(38): e2003010, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32815251

RESUMO

Currently, mesenchymal stem cells (MSCs)-based therapies for bone regeneration and treatments have gained significant attention in clinical research. Though many chemical and physical cues which influence the osteogenic differentiation of MSCs have been explored, scaffolds combining the benefits of Zn2+ ions and unique nanostructures may become an ideal interface to enhance osteogenic and anti-infective capabilities simultaneously. In this work, motivated by the enormous advantages of Zn-based metal-organic framework-derived nanocarbons, C-ZnO nanocarbons-modified fibrous scaffolds for stem cell-based osteogenic differentiation are constructed. The modified scaffolds show enhanced expression of alkaline phosphatase, bone sialoprotein, vinculin, and a larger cell spreading area. Meanwhile, the caging of ZnO nanoparticles can allow the slow release of Zn2+ ions, which not only activate various signaling pathways to guide osteogenic differentiation but also prevent the potential bacterial infection of implantable scaffolds. Overall, this study may provide new insight for designing stem cell-based nanostructured fibrous scaffolds with simultaneously enhanced osteogenic and anti-infective capabilities.


Assuntos
Carbono/química , Células-Tronco Mesenquimais/citologia , Nanofibras/química , Osteogênese/fisiologia , Tecidos Suporte/química , Óxido de Zinco/química , Fosfatase Alcalina/metabolismo , Diferenciação Celular/fisiologia , Células Cultivadas , Humanos , Sialoproteína de Ligação à Integrina/metabolismo , Teste de Materiais , Células-Tronco Mesenquimais/metabolismo , Microscopia Eletrônica de Varredura , Nanofibras/ultraestrutura , Transdução de Sinais , Engenharia Tecidual , Vinculina/metabolismo
4.
Int J Nanomedicine ; 15: 5027-5042, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32764934

RESUMO

Background: Bactericidal capacity, durable inhibition of biofilm formation, and a three-dimensional (3D) porous structure are the emphases of infected bone defect (IBD) treatment via local scaffold implantation strategy. Purpose: In this study, silver nanoparticle (AgNP)-loaded nano-hydroxyapatite (nHA)@ reduced graphene oxide (RGO) 3D scaffolds (AHRG scaffolds) were designed to alleviate bone infection, inhibit biofilm formation, and promote bone repair through the synergistic effects of AgNPs, RGO, and nHA. Materials and Methods: AHRGs were prepared using a one-step preparation method, to create a 3D porous scaffold to facilitate a uniform distribution of AgNPs and nHA. Methicillin-resistant Staphylococcus aureus (MRSA) was used as a model-resistant bacterium, and the effects of different silver loadings on the antimicrobial activity and cytocompatibility of materials were evaluated. Finally, a rabbit IBD model was used to evaluate the therapeutic effect of the AHRG scaffold in vivo. Results: The results showed successful synthesis of the AHRG scaffold. The ideal 3D porous structure was verified using scanning electron microscopy and transmission electron microscopy, and X-ray photoelectron spectroscopy and selected area electron diffraction measurements revealed uniform distributions of AgNP and nHA. In vitro antibacterial and cytocompatibility indicated that the 4% AHRG scaffolds possessed the most favorable balance of bactericidal properties and cytocompatibility. In vivo evaluation of the IBD model showed promising treatment efficacy of AHRG scaffolds. Conclusion: The as-fabricated AHRG scaffolds effectively eliminated infection and inhibited biofilm formation. IBD repair was facilitated by the bactericidal properties and 3D porous structure of the AHRG scaffold, suggesting its potential in the treatment of IBDs.


Assuntos
Antibacterianos/farmacologia , Doenças Ósseas Infecciosas/terapia , Grafite/química , Nanopartículas Metálicas/química , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Antibacterianos/química , Biofilmes/efeitos dos fármacos , Regeneração Óssea/efeitos dos fármacos , Substitutos Ósseos , Osso e Ossos/efeitos dos fármacos , Modelos Animais de Doenças , Durapatita/química , Feminino , Masculino , Teste de Materiais , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Porosidade , Coelhos , Ratos , Prata/química , Prata/farmacologia , Infecções Estafilocócicas/terapia
5.
Life Sci ; 257: 118038, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32622947

RESUMO

PURPOSE: The importance of regeneration in large bone defects forces the orthopedic surgeons to search for a proper methodology. The present experiment evaluated the capability of polylactic acid/polycaprolactone/hydroxyapatite (PLA/PCL/HA) scaffold loaded with and without mesenchymal stem cells (MSCs) on bone regeneration. METHODS: Fourier transform infrared spectrometry, X-ray diffraction, scanning electron microscopy, and rheology methodologies were used to characterize the scaffold. Forty Wistar rats were randomly divided into the four groups including the untreated defects as the control group and three other groups in which the bone defects were treated with autologous bones (autograft group), the PLA/PCL/HA scaffolds (PLA/PCL/HA group), and the MSCs-seeded scaffolds (MSCs-seeded PLA/PCL/HA group). RESULTS: Based on the qRT-PCR results, significantly higher expression levels of osteocalcin, osteopontin, and CD31 were seen in the cell-seeded scaffold group compared to the control group (P < 0.05). The CT scanning and radiographic images depicted significantly more newly formed bonny tissue in the MSCs-loaded scaffold and autograft groups than the untreated group (P < 0.001). The immunohistochemistry, biomechanical, histopathologic, and histomorphometric evaluations demonstrated significantly improved regeneration in the autograft and MSCs-loaded scaffold groups compared to the non-treated group (P < 0.05). There were significant differences between the scaffold and untreated groups in all in vivo evaluations (P < 0.05). CONCLUSION: The MSCs enhanced bone healing potential of the PLA/PCL/HA scaffold and the MSCs-seeded scaffold was comparable to the autograft as the golden treatment regimen (P > 0.05).


Assuntos
Regeneração Óssea/fisiologia , Transplante de Células-Tronco Mesenquimais/métodos , Engenharia Tecidual/métodos , Animais , Regeneração Óssea/efeitos dos fármacos , Osso e Ossos/metabolismo , Durapatita/química , Masculino , Células-Tronco Mesenquimais/fisiologia , Poliésteres/química , Rádio (Anatomia)/metabolismo , Ratos , Ratos Wistar , Tecidos Suporte/química
6.
Adv Exp Med Biol ; 1250: 49-61, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32601937

RESUMO

A wide variety of hydrogels have been proposed for tissue engineering applications, cell encapsulation, and bioinks for bioprinting applications. Cell-laden hydrogel constructs rely on natural hydrogels such as alginate, agarose, chitosan, collagen, gelatin, fibroin, and hyaluronic acid (HA), as well as on synthetic hydrogels such as poloxamers (Pluronics®) and polyethylene glycol (PEG). Alginate has become more and more important in the last years, thanks to the possibility to prepare alginate hydrogels suitable for cell encapsulation mainly because of the mild and reversible cross-linking conditions. In this paper alginate will be described in detail with respect to its chemistry, cross-linking behavior, biocompatibility, manufacturing capacity, and possible modifications.


Assuntos
Alginatos , Encapsulamento de Células , Hidrogéis , Engenharia Tecidual , Tecidos Suporte , Alginatos/química , Bioimpressão , Hidrogéis/química , Tecidos Suporte/química
7.
Int J Nanomedicine ; 15: 3903-3920, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32606657

RESUMO

Background: Researchers are trying to study the mechanism of neural stem cells (NSCs) differentiation to oligodendrocyte-like cells (OLCs) as well as to enhance the selective differentiation of NSCs to oligodendrocytes. However, the limitation in nerve tissue accessibility to isolate the NSCs as well as their differentiation toward oligodendrocytes is still challenging. Purpose: In the present study, a hybrid polycaprolactone (PCL)-gelatin nanofiber scaffold mimicking the native extracellular matrix and axon morphology to direct the differentiation of bone marrow-derived NSCs to OLCs was introduced. Materials and Methods: In order to achieve a sustained release of T3, this factor was encapsulated within chitosan nanoparticles and chitosan-loaded T3 was incorporated within PCL nanofibers. Polyaniline graphene (PAG) nanocomposite was incorporated within gelatin nanofibers to endow the scaffold with conductive properties, which resemble the conductive behavior of axons. Biodegradation, water contact angle measurements, and scanning electron microscopy (SEM) observations as well as conductivity tests were used to evaluate the properties of the prepared scaffold. The concentration of PAG and T3-loaded chitosan NPs in nanofibers were optimized by examining the proliferation of cultured bone marrow-derived mesenchymal stem cells (BMSCs) on the scaffolds. The differentiation of BMSCs-derived NSCs cultured on the fabricated scaffolds into OLCs was analyzed by evaluating the expression of oligodendrocyte markers using immunofluorescence (ICC), RT-PCR and flowcytometric assays. Results: Incorporating 2% PAG proved to have superior cell support and proliferation while guaranteeing electrical conductivity of 10.8 × 10-5 S/cm. Moreover, the scaffold containing 2% of T3-loaded chitosan NPs was considered to be the most biocompatible samples. Result of ICC, RT-PCR and flow cytometry showed high expression of O4, Olig2, platelet-derived growth factor receptor-alpha (PDGFR-α), O1, myelin/oligodendrocyte glycoprotein (MOG) and myelin basic protein (MBP) high expressed but low expression of glial fibrillary acidic protein (GFAP). Conclusion: Considering surface topography, biocompatibility, electrical conductivity and gene expression, the hybrid PCL/gelatin scaffold with the controlled release of T3 may be considered as a promising candidate to be used as an in vitro model to study patient-derived oligodendrocytes by isolating patient's BMSCs in pathological conditions such as diseases or injuries. Moreover, the resulted oligodendrocytes can be used as a desirable source for transplanting in patients.


Assuntos
Materiais Biomiméticos/farmacologia , Células da Medula Óssea/citologia , Diferenciação Celular , Nanofibras/química , Células-Tronco Neurais/citologia , Oligodendroglia/citologia , Tecidos Suporte/química , Compostos de Anilina/química , Animais , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Condutividade Elétrica , Gelatina/química , Grafite/química , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Nanofibras/ultraestrutura , Células-Tronco Neurais/metabolismo , Oligodendroglia/efeitos dos fármacos , Poliésteres/química , Ratos , Suínos , Tri-Iodotironina/farmacologia
8.
Proc Natl Acad Sci U S A ; 117(27): 15482-15489, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32571918

RESUMO

Bioelectronic scaffolds that support devices while promoting tissue integration could enable tissue hybrids with augmented electronic capabilities. Here, we demonstrate a photo-cross-linkable silk fibroin (PSF) derivative and investigate its structural, electrical, and chemical properties. Lithographically defined PSF films offered tunable thickness and <1-µm spatial resolution and could be released from a relief layer yielding freestanding scaffolds with centimeter-scale uniformity. These constructs were electrically insulating; multielectrode arrays with PSF-passivated interconnects provided stable electrophysiological readouts from HL-1 cardiac model cells, brain slices, and hearts. Compared to SU8, a ubiquitous biomaterial, PSF exhibited superior affinity toward neurons which we attribute to its favorable surface charge and enhanced attachment of poly-d-lysine adhesion factors. This finding is of significant importance in bioelectronics, where tight junctions between devices and cell membranes are necessary for electronic communication. Collectively, our findings are generalizable to a variety of geometries, devices, and tissues, establishing PSF as a promising bioelectronic platform.


Assuntos
Materiais Biocompatíveis/efeitos da radiação , Fontes de Energia Bioelétrica , Fibroínas/efeitos da radiação , Engenharia Tecidual , Tecidos Suporte/química , Animais , Materiais Biocompatíveis/química , Encéfalo , Adesão Celular , Linhagem Celular , Feminino , Fibroínas/química , Coração , Teste de Materiais , Camundongos , Células-Tronco Neurais , Raios Ultravioleta
9.
PLoS One ; 15(6): e0234087, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32511282

RESUMO

BACKGROUND: Ventricular septal perforation and left ventricular aneurysm are examples of potentially fatal complications of myocardial infarction. While various artificial materials are used in the repair of these issues, the possibility of associated infection and calcification is non-negligible. Cell-seeded biodegradable tissue-engineered patches may be a potential solution. This study evaluated the feasibility of a new left ventricular patch rat model to study neotissue formation in biodegradable cardiac patches. METHODS: Human induced pluripotent stem cell-derived cardiac progenitor cells (hiPS-CPCs) were cultured onto biodegradable patches composed of polyglycolic acid and a 50:50 poly (l-lactide-co-ε-caprolactone) copolymer for one week. After culturing, patches were implanted into left ventricular walls of male athymic rats. Unseeded controls were also used (n = 10/group). Heart conditions were followed by echocardiography and patches were subsequently explanted at 1, 2, 6, and 9 months post-implantation for histological evaluation. RESULT: Throughout the study, no patches ruptured demonstrating the ability to withstand the high pressure left ventricular system. One month after transplantation, the seeded patch did not stain positive for human nuclei. However, many new blood vessels formed within patches with significantly greater vessels in the seeded group at the 6 month time point. Echocardiography showed no significant difference in left ventricular contraction rate between the two groups. Calcification was found inside patches after 6 months, but there was no significant difference between groups. CONCLUSION: We have developed a surgical method to implant a bioabsorbable scaffold into the left ventricular environment of rats with a high survival rate. Seeded hiPS-CPCs did not differentiate into cardiomyocytes, but the greater number of new blood vessels in seeded patches suggests the presence of cell seeding early in the remodeling process might provide a prolonged effect on neotissue formation. This experiment will contribute to the development of a treatment model for left ventricular failure using iPS cells in the future.


Assuntos
Células-Tronco Pluripotentes Induzidas/citologia , Miócitos Cardíacos/citologia , Engenharia Tecidual , Implantes Absorvíveis , Animais , Modelos Animais de Doenças , Ecocardiografia , Ventrículos do Coração/metabolismo , Ventrículos do Coração/patologia , Humanos , Masculino , Infarto do Miocárdio/patologia , Infarto do Miocárdio/terapia , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/transplante , Poliésteres/química , Ácido Poliglicólico/química , Ratos , Ratos Nus , Tecidos Suporte/química , Troponina T/metabolismo , Função Ventricular
10.
J Vis Exp ; (159)2020 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-32510497

RESUMO

Electrospinning affords researchers the opportunity to fabricate reproducible micro to nanoscale polymer fibers. The 3D fibrous architecture of electrospun polymers is regarded as a structural imitation of the extracellular matrix (ECM). Hence, electrospun fibers fabricated from biocompatible polymers have been widely investigated by tissue engineering researchers for their potential role as an artificial ECM for guiding tissue growth both in vitro and in vivo. All cells are acutely sensitive to their mechanical environment. This has been demonstrated by the discovery of multiple mechanotransduction pathways intrinsically linked to the cytoskeletal actin filaments. The cytoskeleton acts as a mechanical sensor that can direct the functionality and differentiation of the host cell depending on the stiffness and morphology of its substrate. Electrospun fibers can be tuned both in terms of fiber size and morphology to easily modulate the mechanical environment within a fibrous polymer scaffold. Here, methods for electrospinning polycaprolactone (PCL) for three distinct morphologies at two different fiber diameters are described. The morphological fiber categories consist of randomly oriented fibers, aligned fibers, and porous cryogenically spun fibers, with 1 µm and 5 µm diameters. The methods detailed within this study are proposed as a platform for investigating the effect of electrospun fiber architecture on tissue generation. Understanding these effects will allow researchers to optimize the mechanical properties of electrospun fibers and demonstrate the potential of this technology more thoroughly.


Assuntos
Poliésteres/química , Engenharia Tecidual/métodos , Diferenciação Celular , Células Hep G2 , Humanos , Gases em Plasma/química , Porosidade , Esterilização , Resistência à Tração , Tecidos Suporte/química
11.
Gene ; 757: 144852, 2020 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-32599019

RESUMO

Until now, various methods have been introduced to fabricate 3D scaffolds to provide a suitable substrate for cell growth and proliferation and subsequent use in tissue engineering to repair damaged tissues. The 3D scaffolds can simulate the natural cellular microenvironment well. Herein, the decellularized leaf spinach has been used which not only have no problems associated with artificial scaffolds, but they also do not cost significantly. Decellularized scaffolds surface properties were characterized by the investigation of scaffolds surface roughness, hydrophilicity, mechanical properties, size and shape of porosities and specific surface area. In the next step, osteogenic differentiation potential of bone marrow derived mesenchymal stem cells cultured on the scaffold and culture plate (as a control) was evaluated using alizarin staining and calcium content, alkaline phosphatase activity and bone related genes expression assays. The results indicated that the surface properties and shape of scaffold pores were effective in the stem cells binding, growth and proliferation. This higher biocompatibility due to the ideal surface hydrophilicity as well as high specific surface area due to the presence of a rough grid surface ultimately increased the efficiency of stem cell's bone differentiation. Taken together, it can be concluded that the decellularized spinach leaf scaffold, due to its easy availability, low prices and high efficiency, can be considered as a promising potential candidate for use as a proper substrate for stem cell growth and differentiation in bone tissue engineering.


Assuntos
Diferenciação Celular , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/metabolismo , Osteocalcina/genética , Folhas de Planta/química , Tecidos Suporte/química , Fosfatase Alcalina/genética , Fosfatase Alcalina/metabolismo , Biomineralização , Cálcio/metabolismo , Linhagem Celular , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Humanos , Células-Tronco Mesenquimais/citologia , Osteoblastos/citologia , Osteocalcina/metabolismo , Spinacia oleracea/química
12.
PLoS One ; 15(6): e0234482, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32520967

RESUMO

Three-dimensional polydimethylsiloxane platforms were developed to mimic the extracellular matrix with blood vessels by having scaffolds with micropatterns, porous membrane and trenches. Precisely controlled physical dimensions, layouts, and topography as well as different surface chemical treatments were applied to study their influences on nasopharyngeal carcinoma cell (10-15 µm in diameter) migration in mimicked platforms over 15-hour of time-lapse imaging. By placing the pores at different distance from the edges of the trenches, pores with different trench sidewall exposures and effective sizes were generated. Pores right next to the trench sidewalls showed the highest cell traversing probability, most likely related to the larger surface contact area with cells along the sidewalls. Straight grating oriented perpendicular to trenches below the top layer increased cell traversing probability. Pore shape as well as pore size influenced the cell traversing probability and cells could not traverse through pores that were 6 µm or less in diameter, which is much smaller than the cell size. Trench depth of 15 µm could induce more cells to traverse through the porous membrane, while shallower trenches impeded cell traversing and longer time was needed for cells to traverse because 3 and 6 µm deep trenches were much smaller than cell size which required large cell deformation. Hydrophobic surface coating on the top layer and fibronectin in pores and trenches increased the cell traversing probability and reduced the pore size that cells could traverse from 8 to 6 µm, which indicated that cells could have larger deformation with certain surface coatings.


Assuntos
Materiais Biomiméticos/química , Movimento Celular , Dimetilpolisiloxanos/química , Linhagem Celular Tumoral , Matriz Extracelular/química , Humanos , Interações Hidrofóbicas e Hidrofílicas , Porosidade , Tecidos Suporte/química
13.
Int J Nanomedicine ; 15: 3483-3496, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32523344

RESUMO

Background: The use of polycaprolactone (PCL) for bone defects in a clinical setting is limited due to a lack of bioactivity. Exosomes derived from mesenchymal stem cells (MSCs) have an important immunoregulatory potential and together with S-nitrosoglutathione (GSNO) they possess therapeutic potential for bone regeneration. Materials and Methods: In this study, PCL was modified with GSNO and MSC-derived exosomes and the impact on macrophages and osteogenes is evaluated. Results: MSC-derived exosomes exhibited a cup-shaped morphology and were internalized by macrophages and human bone marrow-derived mesenchymal stromal cells (hBMSCs). The pattern of internalization of scaffold-immobilized exosomes was similar in RAW264.7 cells and hBMSCs after 4h and 24h of co-culture. Assessment of macrophage morphology under inflammatory conditions by scanning electronic microscopy (SEM) and confocal microscopy demonstrated macrophages were significantly elongated and expression of pro-inflammatory genes markedly decreased when co-cultured with PCL/PDA + GSNO + exosome scaffolds. Furthermore, this scaffold modification significantly enhanced osteogenic differentiation of hBMSCs. Discussion: This study demonstrated the possibility of using a GSNO- and exosome-based strategy to adapt barrier membrane scaffolds. PCL/PDA + GSNO + exosome scaffolds may serve as an important barrier membrane for osteogenesis and tissue regeneration.


Assuntos
Exossomos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Osteogênese , S-Nitrosoglutationa/metabolismo , Animais , Diferenciação Celular , Endocitose , Exossomos/ultraestrutura , Humanos , Inflamação/patologia , Macrófagos/patologia , Macrófagos/ultraestrutura , Camundongos , Células RAW 264.7 , Tecidos Suporte/química
14.
PLoS One ; 15(5): e0232670, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32421748

RESUMO

Complex skin wounds have always been a significant health and economic problem worldwide due to their elusive and sometimes poor or non-healing conditions. If not well-treated, such wounds may lead to amputation, infections, cancer, or even death. Thus, there is a need to efficiently generate multifunctional skin grafts that address a wide range of skin conditions, including non-healing wounds, and enable the regeneration of new skin tissue. Here, we propose studying pristine graphene and two of its oxygen-functionalized derivatives-high and low-oxygen graphene films-as potential substrates for skin cell proliferation and differentiation. Using BJ cells (human foreskin-derived fibroblasts) to represent basic skin cells, we show that the changes in surface properties of pristine graphene due to oxygen functionalization do not seem to statistically impact the normal proliferation and maturation of skin cells. Our results indicate that the pristine and oxidized graphenes presented relatively low cytotoxicity to BJ fibroblasts and, in fact, support their growth and bioactivity. Therefore, these graphene films could potentially be integrated into more complex skin regenerative systems to support skin regeneration. Because graphene's surface can be relatively easily functionalized with various chemical groups, this finding presents a major opportunity for the development of various composite materials that can act as active components in regenerative applications such as skin regeneration.


Assuntos
Fibroblastos/citologia , Grafite/química , Tecidos Suporte/química , Linhagem Celular , Proliferação de Células , Sobrevivência Celular , Humanos , Propriedades de Superfície , Engenharia Tecidual
15.
Int J Nanomedicine ; 15: 3039-3056, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32431500

RESUMO

Background: Electrospinning is a widely used technology that can produce scaffolds with high porosity and surface area for bone regeneration. However, the small pore sizes in electrospun scaffolds constrain cell growth and tissue-ingrowth. In this study, novel drug-loading core-shell scaffolds were fabricated via electrospinning and freeze drying to facilitate the repair of tibia bone defects in rabbit models. Materials and Methods: The collagen core scaffolds were freeze-dried containing icariin (ICA)-loaded chitosan microspheres. The shell scaffolds were electrospun using collagen, polycaprolactone and hydroxyapatite materials to form CPH composite scaffolds with the ones containing ICA microspheres named CPHI. The core-shell scaffolds were then cross-linked by genipin. The morphology, microstructure, physical and mechanical properties of the scaffolds were assessed. Rat marrow mesenchymal stem cells from the wistar rat were cultured with the scaffolds. The cell adhesion and proliferation were analysed. Adult rabbit models with tibial plateau defects were used to evaluate the performance of these scaffolds in repairing the bone defects over 4 to 12 weeks. Results: The results reveal that the novel drug-loading core-shell scaffolds were successfully fabricated, which showed good physical and chemical properties and appropriate mechanical properties. Furthermore, excellent cells attachment was observed on the CPHI scaffolds. The results from radiography, micro-computed tomography, histological and immunohistochemical analysis demonstrated that abundant new bones were formed on the CPHI scaffolds. Conclusion: These new core-shell composite scaffolds have great potential for bone tissue engineering applications and may lead to effective bone regeneration and repair.


Assuntos
Regeneração Óssea , Flavonoides/farmacologia , Tíbia/efeitos dos fármacos , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Regeneração Óssea/efeitos dos fármacos , Quitosana/química , Colágeno/química , Durapatita/química , Flavonoides/administração & dosagem , Flavonoides/química , Masculino , Teste de Materiais , Células-Tronco Mesenquimais/citologia , Microesferas , Poliésteres/química , Porosidade , Coelhos , Ratos Wistar , Tíbia/diagnóstico por imagem , Microtomografia por Raio-X
16.
PLoS One ; 15(5): e0232081, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32374763

RESUMO

The reproduction of reliable in vitro models of human skeletal muscle is made harder by the intrinsic 3D structural complexity of this tissue. Here we coupled engineered hydrogel with 3D structural cues and specific mechanical properties to derive human 3D muscle constructs ("myobundles") at the scale of single fibers, by using primary myoblasts or myoblasts derived from embryonic stem cells. To this aim, cell culture was performed in confined, laminin-coated micrometric channels obtained inside a 3D hydrogel characterized by the optimal stiffness for skeletal muscle myogenesis. Primary myoblasts cultured in our 3D culture system were able to undergo myotube differentiation and maturation, as demonstrated by the proper expression and localization of key components of the sarcomere and sarcolemma. Such approach allowed the generation of human myobundles of ~10 mm in length and ~120 µm in diameter, showing spontaneous contraction 7 days after cell seeding. Transcriptome analyses showed higher similarity between 3D myobundles and skeletal signature, compared to that found between 2D myotubes and skeletal muscle, mainly resulting from expression in 3D myobundles of categories of genes involved in skeletal muscle maturation, including extracellular matrix organization. Moreover, imaging analyses confirmed that structured 3D culture system was conducive to differentiation/maturation also when using myoblasts derived from embryonic stem cells. In conclusion, our structured 3D model is a promising tool for modelling human skeletal muscle in healthy and diseases conditions.


Assuntos
Técnicas de Cultura de Células , Fibras Musculares Esqueléticas/citologia , Músculo Esquelético/citologia , Engenharia Tecidual , Tecidos Suporte/química , Animais , Técnicas de Cultura de Células/instrumentação , Técnicas de Cultura de Células/métodos , Diferenciação Celular , Células Cultivadas , Dimetilpolisiloxanos/química , Humanos , Hidrogéis/química , Teste de Materiais , Camundongos , Modelos Biológicos , Conformação Molecular , Desenvolvimento Muscular , Músculo Esquelético/fisiologia , Mioblastos/citologia , Mioblastos/fisiologia , Análise de Célula Única/instrumentação , Análise de Célula Única/métodos , Engenharia Tecidual/instrumentação , Engenharia Tecidual/métodos
17.
Proc Natl Acad Sci U S A ; 117(23): 12817-12825, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32444491

RESUMO

Morphogenesis, tumor formation, and wound healing are regulated by tissue rigidity. Focal adhesion behavior is locally regulated by stiffness; however, how cells globally adapt, detect, and respond to rigidity remains unknown. Here, we studied the interplay between the rheological properties of the cytoskeleton and matrix rigidity. We seeded fibroblasts onto flexible microfabricated pillar arrays with varying stiffness and simultaneously measured the cytoskeleton organization, traction forces, and cell-rigidity responses at both the adhesion and cell scale. Cells adopted a rigidity-dependent phenotype whereby the actin cytoskeleton polarized on stiff substrates but not on soft. We further showed a crucial role of active and passive cross-linkers in rigidity-sensing responses. By reducing myosin II activity or knocking down α-actinin, we found that both promoted cell polarization on soft substrates, whereas α-actinin overexpression prevented polarization on stiff substrates. Atomic force microscopy indentation experiments showed that this polarization response correlated with cell stiffness, whereby cell stiffness decreased when active or passive cross-linking was reduced and softer cells polarized on softer matrices. Theoretical modeling of the actin network as an active gel suggests that adaptation to matrix rigidity is controlled by internal mechanical properties of the cytoskeleton and puts forward a universal scaling between nematic order of the actin cytoskeleton and the substrate-to-cell elastic modulus ratio. Altogether, our study demonstrates the implication of cell-scale mechanosensing through the internal stress within the actomyosin cytoskeleton and its coupling with local rigidity sensing at focal adhesions in the regulation of cell shape changes and polarity.


Assuntos
Citoesqueleto/metabolismo , Módulo de Elasticidade , Mecanotransdução Celular , Tecidos Suporte/química , Actinina/metabolismo , Polaridade Celular , Reagentes para Ligações Cruzadas/química , Citoesqueleto/ultraestrutura , Fibroblastos/metabolismo , Humanos , Modelos Teóricos , Miosinas/metabolismo
18.
Med Sci (Paris) ; 36(4): 382-388, 2020 Apr.
Artigo em Francês | MEDLINE | ID: mdl-32356715

RESUMO

As burden of chronic respiratory diseases is constantly increasing, improving in vitro lung models is essential in order to reproduce as closely as possible the complex pulmonary architecture, responsible for oxygen uptake and carbon dioxide clearance. The study of diseases that affect the respiratory system has benefited from in vitro reconstructions of the respiratory epithelium with inserts in air/liquid interface (2D) or in organoids able to mimic up to the arborescence of the respiratory tree (3D). Recent development in the fields of pluripotent stem cells-derived organoids and genome editing technologies has provided new insights to better understand pulmonary diseases and to find new therapeutic perspectives.


Assuntos
Técnicas de Cultura de Células , Pulmão/citologia , Organoides/citologia , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/fisiologia , Animais , Bioengenharia/métodos , Bioengenharia/tendências , Dióxido de Carbono/farmacologia , Dióxido de Carbono/fisiologia , Técnicas de Cultura de Células/métodos , Técnicas de Cultura de Células/tendências , Células Cultivadas , Edição de Genes/métodos , Edição de Genes/tendências , Humanos , Pulmão/patologia , Pulmão/fisiologia , Modelos Biológicos , Organoides/patologia , Organoides/fisiologia , Oxigênio/farmacologia , Oxigênio/fisiologia , Troca Gasosa Pulmonar/fisiologia , Mucosa Respiratória/citologia , Mucosa Respiratória/efeitos dos fármacos , Tecidos Suporte/química
19.
Pharm Res ; 37(6): 97, 2020 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-32409985

RESUMO

PURPOSE: Subcutaneously or intramuscularly administered biodegradable microsphere formulations have been successfully exploited in the management of chronic conditions for over two decades, yet mechanistic understanding of the impact of formulation attributes on in vivo absorption rate from such systems is still in its infancy. METHODS: Suspension formulation physicochemical attributes may impact particulate deposition in subcutaneous (s.c.) tissue. Hence, the utility of synchrotron X-ray micro-computed tomography (µCT) for assessment of spatial distribution of suspension formulation components (PLG microspheres and vehicle) was evaluated in a porcine s.c. tissue model. Optical imaging of dyed vehicle and subsequent microscopic assessment of microsphere deposition was performed in parallel to compare the two approaches. RESULTS: Our findings demonstrate that synchrotron µCT can be applied to the assessment of microsphere and vehicle distribution in s.c. tissue, and that microspheres can also be visualised in the absence of contrast agent using this approach. The technique was deemed superior to optical imaging of macrotomy for the characterisation of microsphere deposition owing to its non-invasive nature and relatively rapid data acquisition time. CONCLUSIONS: The method outlined in this study provides a proof of concept feasibility for µCT application to determining the vehicle and suspended PLG microspheres fate following s.c. injection. A potential application for our findings is understanding the impact of injection, device and formulation variables on initial and temporal depot geometry in pre-clinical or ex-vivo models that can inform product design. Graphical abstract.


Assuntos
Materiais Biocompatíveis/química , Meios de Contraste/química , Microesferas , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Suspensões/química , Tomografia Computadorizada por Raios X/métodos , Animais , Composição de Medicamentos , Imageamento Tridimensional , Injeções Subcutâneas , Intensificação de Imagem Radiográfica , Suínos , Síncrotrons , Tecidos Suporte/química
20.
PLoS One ; 15(5): e0230354, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32413029

RESUMO

Bone marrow stroma influences metastatic prostate cancer (PCa) progression, latency, and recurrence. At sites of PCa bone metastasis, cancer-associated fibroblasts and tumor-associated macrophages interact to establish a perlecan-rich desmoplastic stroma. As a heparan sulfate proteoglycan, perlecan (HSPG2) stores and stabilizes growth factors, including heparin-binding Wnt3A, a positive regulator of PCa cell growth. Because PCa cells alone do not induce CAF production of perlecan in the desmoplastic stroma, we sought to discover the sources of perlecan and its growth factor-releasing modifiers SULF1, SULF2, and heparanase in PCa cells and xenografts, bone marrow fibroblasts, and macrophages. SULF1, produced primarily by bone marrow fibroblasts, was the main glycosaminoglycanase present, a finding validated with primary tissue specimens of PCa metastases with desmoplastic bone stroma. Expression of both HSPG2 and SULF1 was concentrated in αSMA-rich stroma near PCa tumor nests, where infiltrating pro-tumor TAMs also were present. To decipher SULF1's role in the reactive bone stroma, we created a bone marrow biomimetic hydrogel incorporating perlecan, PCa cells, macrophages, and fibroblastic bone marrow stromal cells. Finding that M2-like macrophages increased levels of SULF1 and HSPG2 produced by fibroblasts, we examined SULF1 function in Wnt3A-mediated PCa tumoroid growth in tricultures. Comparing control or SULF1 knockout fibroblastic cells, we showed that SULF1 reduces Wnt3A-driven growth, cellularity, and cluster number of PCa cells in our 3D model. We conclude that SULF1 can suppress Wnt3A-driven growth signals in the desmoplastic stroma of PCa bone metastases, and SULF1 loss favors PCa progression, even in the presence of pro-tumorigenic TAMs.


Assuntos
Neoplasias Ósseas/metabolismo , Proteoglicanas de Heparan Sulfato/metabolismo , Neoplasias da Próstata/metabolismo , Sulfotransferases/metabolismo , Engenharia Tecidual/métodos , Tecidos Suporte/química , Via de Sinalização Wnt , Neoplasias Ósseas/secundário , Fibroblastos Associados a Câncer/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Humanos , Hidrogéis/química , Macrófagos/metabolismo , Masculino , Neoplasias da Próstata/patologia , Células Estromais/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA