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1.
Anticancer Res ; 41(9): 4259-4269, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34475045

RESUMO

BACKGROUND: Compared to two-dimensional cultures, three-dimensional (3D) cultures have many advantages in cancer studies. Nevertheless, their implementation is unsatisfactory. This study aimed to develop an anchorage-dependent 3D culture model for colorectal cancer research. MATERIALS AND METHODS: Human HCT116, DLD-1 and SW620 colorectal cell lines were cultured in a gelatin sponge, and its applicability for morphological examination was studied. RESULTS: The resulting specimens were suitable for scanning electron microscopy, transmission electron microscopy, and immunohistochemical examination. HCT116 formed smaller structures and migrated through the pores of the sponge. DLD-1 formed larger structures with tight cell-to-cell adhesion. SW620 also formed large structures but small clustered cells tended to attach to the anchorage more favorably. Immunohistochemical staining demonstrated phosphorylated yes-associated protein (YAP) localized near the attachment site in HCT116 cells. CONCLUSION: Because the gelatin sponge provided suitable anchorage and the cultured cells formed distinguishable 3D structures, this method may be useful for further colorectal cancer research.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Técnicas de Cultura de Células/métodos , Neoplasias Colorretais/patologia , Gelatina/química , Tecidos Suporte/química , Fatores de Transcrição/metabolismo , Adesão Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Neoplasias Colorretais/metabolismo , Células HCT116 , Humanos , Fosforilação
2.
Int J Mol Sci ; 22(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34360970

RESUMO

Anterior cruciate ligament (ACL) ruptures are usually treated with autograft implantation to prevent knee instability. Tissue engineered ACL reconstruction is becoming promising to circumvent autograft limitations. The aim was to evaluate the influence of cyclic stretch on lapine (L) ACL fibroblasts on embroidered scaffolds with respect to adhesion, DNA and sulphated glycosaminoglycan (sGAG) contents, gene expression of ligament-associated extracellular matrix genes, such as type I collagen, decorin, tenascin C, tenomodulin, gap junctional connexin 43 and the transcription factor Mohawk. Control scaffolds and those functionalized by gas phase fluorination and cross-linked collagen foam were either pre-cultured with a suspension or with spheroids of LACL cells before being subjected to cyclic stretch (4%, 0.11 Hz, 3 days). Stretch increased significantly the scaffold area colonized with cells but impaired sGAGs and decorin gene expression (functionalized scaffolds seeded with cell suspension). Stretching increased tenascin C, connexin 43 and Mohawk but decreased decorin gene expression (control scaffolds seeded with cell suspension). Pre-cultivation of functionalized scaffolds with spheroids might be the more suitable method for maintaining ligamentogenesis in 3D scaffolds compared to using a cell suspension due to a significantly higher sGAG content in response to stretching and type I collagen gene expression in functionalized scaffolds.


Assuntos
Ligamento Cruzado Anterior/fisiologia , Esferoides Celulares/citologia , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Ligamento Cruzado Anterior/citologia , Adesão Celular , Proliferação de Células , Células Cultivadas , Conexinas/genética , Conexinas/metabolismo , Decorina/genética , Decorina/metabolismo , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Feminino , Fibroblastos/fisiologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Homeostase , Masculino , Poliésteres/química , Coelhos , Regeneração , Esferoides Celulares/metabolismo , Estresse Mecânico
3.
Int J Mol Sci ; 22(16)2021 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-34445255

RESUMO

One of the most important features of striated cardiac muscle is the excitability that turns on the excitation-contraction coupling cycle, resulting in the heart blood pumping function. The function of the heart pump may be impaired by events such as myocardial infarction, the consequence of coronary artery thrombosis due to blood clots or plaques. This results in the death of billions of cardiomyocytes, the formation of scar tissue, and consequently impaired contractility. A whole heart transplant remains the gold standard so far and the current pharmacological approaches tend to stop further myocardium deterioration, but this is not a long-term solution. Electrically conductive, scaffold-based cardiac tissue engineering provides a promising solution to repair the injured myocardium. The non-conductive component of the scaffold provides a biocompatible microenvironment to the cultured cells while the conductive component improves intercellular coupling as well as electrical signal propagation through the scar tissue when implanted at the infarcted site. The in vivo electrical coupling of the cells leads to a better regeneration of the infarcted myocardium, reducing arrhythmias, QRS/QT intervals, and scar size and promoting cardiac cell maturation. This review presents the emerging applications of intrinsically conductive polymers in cardiac tissue engineering to repair post-ischemic myocardial insult.


Assuntos
Arritmias Cardíacas , Materiais Biocompatíveis , Condutividade Elétrica , Infarto do Miocárdio , Miocárdio/metabolismo , Regeneração/efeitos dos fármacos , Tecidos Suporte/química , Animais , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/fisiopatologia , Arritmias Cardíacas/terapia , Materiais Biocompatíveis/química , Materiais Biocompatíveis/uso terapêutico , Humanos , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/fisiopatologia , Infarto do Miocárdio/terapia , Engenharia Tecidual
4.
Int J Mol Sci ; 22(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34445293

RESUMO

In this research, we synthesize and characterize poly(glycerol sebacate) pre-polymer (pPGS) (1H NMR, FTiR, GPC, and TGA). Nano-hydroxyapatite (HAp) is synthesized using the wet precipitation method. Next, the materials are used to prepare a PGS-based composite with a 25 wt.% addition of HAp. Microporous composites are formed by means of thermally induced phase separation (TIPS) followed by thermal cross-linking (TCL) and salt leaching (SL). The manufactured microporous materials (PGS and PGS/HAp) are then subjected to imaging by means of SEM and µCT for the porous structure characterization. DSC, TGA, and water contact angle measurements are used for further evaluation of the materials. To assess the cytocompatibility and biological potential of PGS-based composites, preosteoblasts and differentiated hFOB 1.19 osteoblasts are employed as in vitro models. Apart from the cytocompatibility, the scaffolds supported cell adhesion and were readily populated by the hFOB1.19 preosteoblasts. HAp-facilitated scaffolds displayed osteoconductive properties, supporting the terminal differentiation of osteoblasts as indicated by the production of alkaline phosphatase, osteocalcin and osteopontin. Notably, the PGS/HAp scaffolds induced the production of significant amounts of osteoclastogenic cytokines: IL-1ß, IL-6 and TNF-α, which induced scaffold remodeling and promoted the reconstruction of bone tissue. Initial biocompatibility tests showed no signs of adverse effects of PGS-based scaffolds toward adult BALB/c mice.


Assuntos
Substitutos Ósseos/síntese química , Decanoatos/química , Durapatita/química , Glicerol/análogos & derivados , Polímeros/química , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Materiais Biocompatíveis/síntese química , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacologia , Regeneração Óssea/efeitos dos fármacos , Substitutos Ósseos/química , Substitutos Ósseos/farmacologia , Substitutos Ósseos/uso terapêutico , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/fisiologia , Células Cultivadas , Feminino , Glicerol/química , Humanos , Invenções , Masculino , Teste de Materiais , Camundongos , Camundongos Endogâmicos BALB C , Osteoblastos/efeitos dos fármacos , Osteoblastos/fisiologia , Osteogênese/efeitos dos fármacos , Polímeros/síntese química , Porosidade , Engenharia Tecidual/tendências
5.
Int J Mol Sci ; 22(16)2021 Aug 23.
Artigo em Inglês | MEDLINE | ID: mdl-34445788

RESUMO

Bone formation and growth are crucial for treating bone fractures. Improving bone-reconstruction methods using autologous bone and synthetic implants can reduce the recovery time. Here, we investigated three treatments using two different materials, a bone-derived decellularized extracellular matrix (bdECM) and ß-tricalcium phosphate (ß-TCP), individually and in combination, as osteogenic promoter between bone and 3D-printed polycaprolactone scaffold (6-mm diameter) in rat calvarial defects (8-mm critical diameter). The materials were tested with a human pre-osteoblast cell line (MG63) to determine the effects of the osteogenic promoter on bone formation in vitro. A polycaprolactone (PCL) scaffold with a porous structure was placed at the center of the in vivo rat calvarial defects. The gap between the defective bone and PCL scaffold was filled with each material. Animals were sacrificed four weeks post-implantation, and skull samples were preserved for analysis. The preserved samples were scanned by micro-computed tomography and analyzed histologically to examine the clinical benefits of the materials. The bdECM-ß-TCP mixture showed faster bone formation and a lower inflammatory response in the rats. Therefore, our results imply that a bdECM-ß-TCP mixture is an ideal osteogenic promoter for treating fractures.


Assuntos
Fosfatos de Cálcio/farmacologia , Matriz Extracelular/efeitos dos fármacos , Fraturas Ósseas/tratamento farmacológico , Hidrogéis/farmacologia , Osteogênese/efeitos dos fármacos , Poliésteres/farmacologia , Tecidos Suporte/química , Animais , Matriz Óssea/efeitos dos fármacos , Regeneração Óssea/efeitos dos fármacos , Células Cultivadas , Humanos , Osteoblastos/efeitos dos fármacos , Impressão Tridimensional , Ratos , Ratos Sprague-Dawley , Engenharia Tecidual/métodos
6.
Int J Mol Sci ; 22(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34445106

RESUMO

Tissue decellularization is typically assessed through absorbance-based DNA quantification after tissue digestion. This method has several disadvantages, namely its destructive nature and inadequacy in experimental situations where tissue is scarce. Here, we present an image processing algorithm for quantitative analysis of DNA content in (de)cellularized tissues as a faster, simpler and more comprehensive alternative. Our method uses local entropy measurements of a phase contrast image to create a mask, which is then applied to corresponding nuclei labelled (UV) images to extract average fluorescence intensities as an estimate of DNA content. The method can be used on native or decellularized tissue to quantify DNA content, thus allowing quantitative assessment of decellularization procedures. We confirm that our new method yields results in line with those obtained using the standard DNA quantification method and that it is successful for both lung and heart tissues. We are also able to accurately obtain a timeline of decreasing DNA content with increased incubation time with a decellularizing agent. Finally, the identified masks can also be applied to additional fluorescence images of immunostained proteins such as collagen or elastin, thus allowing further image-based tissue characterization.


Assuntos
Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Colágeno/metabolismo , DNA/metabolismo , Elastina/metabolismo , Matriz Extracelular/metabolismo , Coração/fisiopatologia , Pulmão/metabolismo
7.
Molecules ; 26(15)2021 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-34361602

RESUMO

Repairs of bone defects caused by osteoporosis have always relied on bone tissue engineering. However, the preparation of composite tissue engineering scaffolds with a three-dimensional (3D) macroporous structure poses huge challenges in achieving osteoconduction and osteoinduction for repairing bone defects caused by osteoporosis. In the current study, a three-dimensional macroporous (150-300 µm) reduced graphene oxide/polypyrrole composite scaffold modified by strontium (Sr) (3D rGO/PPY/Sr) was successfully prepared using the oxygen plasma technology-assisted method, which is simple, safe, and inexpensive. The findings of the MTT assay and AO/EB fluorescence double staining showed that 3D rGO/PPY/Sr has a good biocompatibility and effectively promoted MC3T3-E1 cell proliferation. Furthermore, the ALP assay and alizarin red staining showed that 3D rGO/PPY/Sr increased the expression levels of ALP activity and the formation of calcified nodules. The desirable biocompatibility, osteoconduction, and osteoinduction abilities, assure that the 3D macroporous rGO/PPY/Sr composite scaffold offers promising potential for use in the repair of bone defects caused by osteoporosis in bone tissue engineering.


Assuntos
Materiais Biocompatíveis/química , Osteoporose/terapia , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Linhagem Celular , Grafite/química , Camundongos , Osteoblastos , Polímeros/química , Impressão Tridimensional , Pirróis/química , Estrôncio/química
8.
Int J Mol Sci ; 22(16)2021 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-34445150

RESUMO

Titanium is one of the most frequently used materials in bone regeneration due to its good biocompatibility, excellent mechanical properties, and great osteogenic performance. However, osseointegration with host tissue is often not definite, which may cause implant failure at times. The present study investigates the capacity of the mesenchymal stem cell (MSC)-secretome, formulated as a ready-to-use and freeze-dried medicinal product (the Lyosecretome), to promote the osteoinductive and osteoconductive properties of titanium cages. In vitro tests were conducted using adipose tissue-derived MSCs seeded on titanium cages with or without Lyosecretome. After 14 days, in the presence of Lyosecretome, significant cell proliferation improvement was observed. Scanning electron microscopy revealed the cytocompatibility of titanium cages: the seeded MSCs showed a spread morphology and an initial formation of filopodia. After 7 days, in the presence of Lyosecretome, more frequent and complex cellular processes forming bridges across the porous surface of the scaffold were revealed. Also, after 14 and 28 days of culturing in osteogenic medium, the amount of mineralized matrix detected by alizarin red was significantly higher when Lyosecretome was used. Finally, improved osteogenesis with Lyosecretome was confirmed by confocal analysis after 28 and 56 days of treatment, and demonstrating the production by osteoblast-differentiated MSCs of osteocalcin, a specific bone matrix protein.


Assuntos
Regeneração Óssea , Substitutos Ósseos/química , Células-Tronco Mesenquimais/citologia , Titânio/química , Proliferação de Células , Células Cultivadas , Liofilização , Humanos , Células-Tronco Mesenquimais/metabolismo , Osteogênese , Tecidos Suporte/química
9.
Theranostics ; 11(16): 7948-7969, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34335973

RESUMO

Heart disease is the main cause of death worldwide. Because death of the myocardium is irreversible, it remains a significant clinical challenge to rescue myocardial deficiency. Cardiac tissue engineering (CTE) is a promising strategy for repairing heart defects and offers platforms for studying cardiac tissue. Numerous achievements have been made in CTE in the past decades based on various advanced engineering approaches. 3D bioprinting has attracted much attention due to its ability to integrate multiple cells within printed scaffolds with complex 3D structures, and many advancements in bioprinted CTE have been reported recently. Herein, we review the recent progress in 3D bioprinting for CTE. After a brief overview of CTE with conventional methods, the current 3D printing strategies are discussed. Bioink formulations based on various biomaterials are introduced, and strategies utilizing composite bioinks are further discussed. Moreover, several applications including heart patches, tissue-engineered cardiac muscle, and other bionic structures created via 3D bioprinting are summarized. Finally, we discuss several crucial challenges and present our perspective on 3D bioprinting techniques in the field of CTE.


Assuntos
Bioimpressão/métodos , Miocárdio/metabolismo , Engenharia Tecidual/métodos , Materiais Biocompatíveis/química , Biônica/métodos , Bioimpressão/tendências , Procedimentos Cirúrgicos Cardíacos/métodos , Coração/fisiologia , Cardiopatias/fisiopatologia , Cardiopatias/terapia , Humanos , Impressão Tridimensional/tendências , Tecidos Suporte/química
10.
Int J Mol Sci ; 22(15)2021 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-34360604

RESUMO

The musculoskeletal system is a vital body system that protects internal organs, supports locomotion, and maintains homeostatic function. Unfortunately, musculoskeletal disorders are the leading cause of disability worldwide. Although implant surgeries using autografts, allografts, and xenografts have been conducted, several adverse effects, including donor site morbidity and immunoreaction, exist. To overcome these limitations, various biomedical engineering approaches have been proposed based on an understanding of the complexity of human musculoskeletal tissue. In this review, the leading edge of musculoskeletal tissue engineering using 3D bioprinting technology and musculoskeletal tissue-derived decellularized extracellular matrix bioink is described. In particular, studies on in vivo regeneration and in vitro modeling of musculoskeletal tissue have been focused on. Lastly, the current breakthroughs, limitations, and future perspectives are described.


Assuntos
Matriz Extracelular/química , Desenvolvimento Musculoesquelético , Doenças Musculoesqueléticas/terapia , Impressão Tridimensional/instrumentação , Regeneração , Engenharia Tecidual/métodos , Tecidos Suporte/química , Animais , Bioimpressão/métodos , Humanos
11.
Int J Mol Sci ; 22(15)2021 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-34360672

RESUMO

Modular tissue engineering (MTE) is a novel "bottom-up" approach to create engineered biological tissues from microscale repeating units. Our aim was to obtain microtissue constructs, based on polymer microspheres (MSs) populated with cells, which can be further assembled into larger tissue blocks and used in bone MTE. Poly(L-lactide-co-glycolide) MS of 165 ± 47 µm in diameter were produced by oil-in-water emulsification and treated with 0.1 M NaOH. To improve cell adhesion, MSs were coated with poly-L-lysine (PLL) or human recombinant collagen type I (COL). The presence of oxygenated functionalities and PLL/COL coating on MS was confirmed by X-ray photoelectron spectroscopy (XPS). To assess the influence of medium composition on adhesion, proliferation, and osteogenic differentiation, preosteoblast MC3T3-E1 cells were cultured on MS in minimal essential medium (MEM) and osteogenic differentiation medium (OSG). Moreover, to assess the potential osteoblast-osteoclast cross-talk phenomenon and the influence of signaling molecules released by osteoclasts on osteoblast cell culture, a medium obtained from osteoclast culture (OSC) was also used. To impel the cells to adhere and grow on the MS, anti-adhesive cell culture plates were utilized. The results show that MS coated with PLL and COL significantly favor the adhesion and growth of MC3T3-E1 cells on days 1 and 7, respectively, in all experimental conditions tested. On day 7, three-dimensional MS/cell/extracellular matrix constructs were created owing to auto-assembly. The cells grown in such constructs exhibited high activity of early osteogenic differentiation marker, namely, alkaline phosphatase. Superior cell growth on PLL- and COL-coated MS on day 14 was observed in the OSG medium. Interestingly, deposition of extracellular matrix and its mineralization was particularly enhanced on COL-coated MS in OSG medium on day 14. In our study, we developed a method of spontaneous formation of organoid-like MS-based cell/ECM constructs with a few millimeters in size. Such constructs may be regarded as building blocks in bone MTE.


Assuntos
Osso e Ossos/citologia , Matriz Extracelular/química , Microesferas , Osteoblastos/citologia , Osteogênese , Polímeros/química , Engenharia Tecidual/métodos , Animais , Materiais Biocompatíveis/química , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Camundongos , Tecidos Suporte/química
12.
Int J Mol Sci ; 22(16)2021 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-34445320

RESUMO

Meniscus injuries can be highly debilitating and lead to knee osteoarthritis. Progenitor cells from the meniscus could be a superior cell type for meniscus repair and tissue-engineering. The purpose of this study is to characterize meniscus progenitor cells isolated by differential adhesion to fibronectin (FN-prog). Human osteoarthritic menisci were digested, and FN-prog were selected by differential adhesion to fibronectin. Multilineage differentiation, population doubling time, colony formation, and MSC surface markers were assessed in the FN-prog and the total meniscus population (Men). Colony formation was compared between outer and inner zone meniscus digest. Chondrogenic pellet cultures were performed for redifferentiation. FN-prog demonstrated multipotency. The outer zone FN-prog formed more colonies than the inner zone FN-prog. FN-prog displayed more colony formation and a higher proliferation rate than Men. FN-prog redifferentiated in pellet culture and mostly adhered to the MSC surface marker profile, except for HLA-DR receptor expression. This is the first study that demonstrates differential adhesion to fibronectin for the isolation of a progenitor-like population from the meniscus. The high proliferation rates and ability to form meniscus extracellular matrix upon redifferentiation, together with the broad availability of osteoarthritis meniscus tissue, make FN-prog a promising cell type for clinical translation in meniscus tissue-engineering.


Assuntos
Adesão Celular , Fibronectinas/metabolismo , Menisco/citologia , Células-Tronco Mesenquimais/citologia , Engenharia Tecidual/métodos , Idoso , Idoso de 80 Anos ou mais , Células Cultivadas , Condrócitos/citologia , Condrócitos/metabolismo , Condrócitos/fisiologia , Condrogênese , Humanos , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/fisiologia , Pessoa de Meia-Idade , Tecidos Suporte/química
13.
ACS Appl Mater Interfaces ; 13(33): 39135-39141, 2021 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-34374274

RESUMO

Many physiochemical properties of the extracellular matrix (ECM) of muscle tissues, such as nanometer scale dimension, nanotopography, negative charge, and elasticity, must be carefully reproduced to fabricate scaffold materials mimicking muscle tissues. Hence, we developed a muscle tissue ECM-mimicking scaffold using Mo6S3I6 inorganic molecular wires (IMWs). Composed of bio-essential elements and having a nanofibrous structure with a diameter of ∼1 nm and a negative surface charge with high stability, Mo6S3I6 IMWs are ideal for mimicking natural ECM molecules. Once Mo6S3I6 IMWs were patterned on a polydimethylsiloxane surface with an elasticity of 1877.1 ± 22.2 kPa, that is, comparable to that of muscle tissues, the proliferation and α-tubulin expression of myoblasts enhanced significantly. Additionally, the repetitive one-dimensional patterns of Mo6S3I6 IMWs induced the alignment and stretching of myoblasts with enhanced α-tubulin expression and differentiation into myocytes. This study demonstrates that Mo6S3I6 IMWs are promising for mimicking the ECM of muscle tissues.


Assuntos
Materiais Biomiméticos/química , Dimetilpolisiloxanos/química , Matriz Extracelular/metabolismo , Nanofios/química , Tecidos Suporte/química , Materiais Biomiméticos/metabolismo , Diferenciação Celular , Proliferação de Células/efeitos dos fármacos , Humanos , Iodo/química , Molibdênio/química , Músculos/citologia , Mioblastos/citologia , Mioblastos/metabolismo , Enxofre/química , Propriedades de Superfície , Engenharia Tecidual , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo
14.
Chem Commun (Camb) ; 57(60): 7422-7425, 2021 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-34231564

RESUMO

Bioactive molecules that enhance or induce osteogenic potential of bone precursor cells have shown vital roles in bone tissue engineering. Herein, we report the design and synthesis of a novel diketopiperazine (DT) that induces osteoblastic differentiation of pre-osteoblasts and bone-marrow-derived stem cells in vitro and enhances the osteogenic potential of cryogel matrix. Such functional diketopiperazines can serve as potential scaffolds for bone healing and regeneration.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Criogéis/química , Dicetopiperazinas/farmacologia , Osteogênese/efeitos dos fármacos , Peptídeos Cíclicos/farmacologia , Tecidos Suporte/química , Animais , Proliferação de Células/efeitos dos fármacos , Dicetopiperazinas/síntese química , Dicetopiperazinas/toxicidade , Células-Tronco Mesenquimais/efeitos dos fármacos , Camundongos , Osteoblastos/efeitos dos fármacos , Peptídeos Cíclicos/síntese química , Peptídeos Cíclicos/toxicidade , Taurina/análogos & derivados , Taurina/farmacologia , Taurina/toxicidade , Engenharia Tecidual/métodos
15.
Int J Mol Sci ; 22(13)2021 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-34202759

RESUMO

The use of porous three-dimensional (3D) composite scaffolds has attracted great attention in bone tissue engineering applications because they closely simulate the major features of the natural extracellular matrix (ECM) of bone. This study aimed to prepare biomimetic composite scaffolds via a simple 3D printing of gelatin/hyaluronic acid (HA)/hydroxyapatite (HAp) and subsequent biomineralization for improved bone tissue regeneration. The resulting scaffolds exhibited uniform structure and homogeneous pore distribution. In addition, the microstructures of the composite scaffolds showed an ECM-mimetic structure with a wrinkled internal surface and a porous hierarchical architecture. The results of bioactivity assays proved that the morphological characteristics and biomineralization of the composite scaffolds influenced cell proliferation and osteogenic differentiation. In particular, the biomineralized gelatin/HA/HAp composite scaffolds with double-layer staggered orthogonal (GEHA20-ZZS) and double-layer alternative structure (GEHA20-45S) showed higher bioactivity than other scaffolds. According to these results, biomineralization has a great influence on the biological activity of cells. Hence, the biomineralized composite scaffolds can be used as new bone scaffolds in bone regeneration.


Assuntos
Regeneração Óssea , Durapatita , Gelatina , Ácido Hialurônico , Impressão Tridimensional , Engenharia Tecidual , Tecidos Suporte , Biomineralização , Diferenciação Celular , Fenômenos Químicos , Durapatita/química , Módulo de Elasticidade , Gelatina/química , Análise Espectral , Tecidos Suporte/química , Viscosidade
16.
Int J Mol Sci ; 22(13)2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34209772

RESUMO

Due to the limited number of organ donors, 3D printing of organs is a promising technique. Tissue engineering is increasingly using xenogeneic material for this purpose. This study was aimed at assessing the safety of decellularized porcine pancreas, together with the analysis of the risk of an undesirable immune response. We tested eight variants of the decellularization process. We determined the following impacts: rinsing agents (PBS/NH3·H2O), temperature conditions (4 °C/24 °C), and the grinding method of native material (ground/cut). To assess the quality of the extracellular matrix after the completed decellularization process, analyses of the following were performed: DNA concentration, fat content, microscopic evaluation, proteolysis, material cytotoxicity, and most importantly, the Triton X-100 content. Our analyses showed that we obtained a product with an extremely low detergent content with negligible residual DNA content. The obtained results confirmed the performed histological and immuno-fluorescence staining. Moreover, the TEM microscopic analysis proved that the correct collagen structure was preserved after the decellularization process. Based on the obtained results, we chose the most favorable variant in terms of quality and biology. The method we chose is an effective and safe method that gives a chance for the development of transplant and regenerative medicine.


Assuntos
Matriz Extracelular/fisiologia , Pâncreas/ultraestrutura , Engenharia Tecidual/métodos , Tecidos Suporte , Animais , Bioimpressão/métodos , Células Cultivadas , Detergentes/química , Detergentes/farmacologia , Matriz Extracelular/química , Fibroblastos/citologia , Fibroblastos/fisiologia , Teste de Materiais , Camundongos , Octoxinol/química , Octoxinol/farmacologia , Pâncreas/citologia , Pós/química , Impressão Tridimensional , Proteômica , Controle de Qualidade , Suínos , Engenharia Tecidual/normas , Tecidos Suporte/química , Tecidos Suporte/normas
17.
Int J Mol Sci ; 22(13)2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34209789

RESUMO

Near-physiological in vitro thrombogenicity test systems for the evaluation of blood-contacting endothelialized biomaterials requires co-cultivation with platelets (PLT). However, the addition of PLT has led to unphysiological endothelial cell (EC) detachment in such in vitro systems. A possible cause for this phenomenon may be PLT activation triggered by the applied endothelial cell medium, which typically consists of basal medium (BM) and nine different supplements. To verify this hypothesis, the influence of BM and its supplements was systematically analyzed regarding PLT responses. For this, human platelet rich plasma (PRP) was mixed with BM, BM containing one of nine supplements, or with BM containing all supplements together. PLT adherence analysis was carried out in six-channel slides with plasma-treated cyclic olefin copolymer (COC) and poly(tetrafluoro ethylene) (PTFE, as a positive control) substrates as part of the six-channel slides in the absence of EC and under static conditions. PLT activation and aggregation were analyzed using light transmission aggregometry and flow cytometry (CD62P). Medium supplements had no effect on PLT activation and aggregation. In contrast, supplements differentially affected PLT adherence, however, in a polymer- and donor-dependent manner. Thus, the use of standard endothelial growth medium (BM + all supplements) maintains functionality of PLT under EC compatible conditions without masking the differences of PLT adherence on different polymeric substrates. These findings are important prerequisites for the establishment of a near-physiological in vitro thrombogenicity test system assessing polymer-based cardiovascular implant materials in contact with EC and PLT.


Assuntos
Materiais Biocompatíveis/farmacologia , Plaquetas/efeitos dos fármacos , Plaquetas/fisiologia , Meios de Cultura/farmacologia , Adulto , Materiais Biocompatíveis/química , Plaquetas/citologia , Meios de Cultura/química , Endotélio/citologia , Feminino , Humanos , Masculino , Teste de Materiais , Pessoa de Meia-Idade , Ativação Plaquetária/efeitos dos fármacos , Adesividade Plaquetária/efeitos dos fármacos , Agregação Plaquetária/efeitos dos fármacos , Polímeros/farmacologia , Tecidos Suporte/química
18.
Int J Mol Sci ; 22(13)2021 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-34209790

RESUMO

We developed a multi-channel cell chip containing a three-dimensional (3D) scaffold for horizontal co-culture and drug toxicity screening in multi-organ culture (human glioblastoma, cervical cancer, normal liver cells, and normal lung cells). The polydimethylsiloxane (PDMS) multi-channel cell chip (PMCCC) was based on fused deposition modeling (FDM) technology. The architecture of the PMCCC was an open-type cell chip and did not require a pump or syringe. We investigated cell proliferation and cytotoxicity by conducting 3-(4,5-dimethylthiazol-2-yl)-2,5-dphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays and analysis of oleanolic acid (OA)-treated multi-channel cell chips. The results of the MTT and LDH assays showed that OA treatment in the multi-channel cell chip of four cell lines enhanced chemoresistance of cells compared with that in the 2D culture. Furthermore, we demonstrated the feasibility of the application of our multi-channel cell chip in various analysis methods through Annexin V-fluorescein isothiocyanate/propidium iodide staining, which is not used for conventional cell chips. Taken together, the results demonstrated that the PMCCC may be used as a new 3D platform because it enables simultaneous drug screening in multiple cells by single point injection and allows analysis of various biological processes.


Assuntos
Técnicas de Cultura de Células , Avaliação Pré-Clínica de Medicamentos , Técnicas de Cultura de Células/instrumentação , Técnicas de Cultura de Células/métodos , Linhagem Celular Tumoral , Proliferação de Células , Tamanho Celular , Técnicas de Cocultura/instrumentação , Técnicas de Cocultura/métodos , Avaliação Pré-Clínica de Medicamentos/instrumentação , Avaliação Pré-Clínica de Medicamentos/métodos , Células HeLa , Humanos , Dispositivos Lab-On-A-Chip , Teste de Materiais , Tecidos Suporte/química , Testes de Toxicidade/instrumentação , Testes de Toxicidade/métodos
19.
Int J Mol Sci ; 22(14)2021 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-34299345

RESUMO

Reconstruction of the periodontal ligament (PDL) to fulfill functional requirement remains a challenge. This study sought to develop a biomimetic microfibrous system capable of withstanding the functional load to assist PDL regeneration. Collagen-based straight and waveform microfibers to guide PDL cell growth were prepared using an extrusion-based bioprinter, and a laminar flow-based bioreactor was used to generate fluidic shear stress. PDL cells were seeded on the respective microfibers with 0 or 6 dynes/cm2 fluidic shear stress for 1-4 h. The viability, morphology, adhesion pattern, and gene expression levels of PDL cells were assessed. The results revealed that upon bioprinting optimization, collagen-based microfibers were successfully fabricated. The straight microfibers were 189.9 ± 11.44 µm wide and the waveform microfibers were 235.9 ± 11.22 µm wide. Under 6 dynes/cm2 shear stress, PDL cells were successfully seeded, and cytoskeleton expansion, adhesion, and viability were greater. Cyclin D, E-cadherin, and periostin were upregulated on the waveform microfibers. In conclusion, 3D-printed collagen-based waveform microfibers preserved PDL cell viability and exhibited an enhanced tendency to promote healing and regeneration under shear stress. This approach is promising for the development of a guiding scaffold for PDL regeneration.


Assuntos
Colágeno/farmacologia , Osteogênese/efeitos dos fármacos , Ligamento Periodontal/efeitos dos fármacos , Tecidos Suporte/química , Bioimpressão/métodos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Citoesqueleto/efeitos dos fármacos , Humanos , Impressão Tridimensional , Estresse Mecânico
20.
Int J Mol Sci ; 22(14)2021 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-34299344

RESUMO

Bone damage leading to bone loss can arise from a wide range of causes, including those intrinsic to individuals such as infections or diseases with metabolic (diabetes), genetic (osteogenesis imperfecta), and/or age-related (osteoporosis) etiology, or extrinsic ones coming from external insults such as trauma or surgery. Although bone tissue has an intrinsic capacity of self-repair, large bone defects often require anabolic treatments targeting bone formation process and/or bone grafts, aiming to restore bone loss. The current bone surrogates used for clinical purposes are autologous, allogeneic, or xenogeneic bone grafts, which although effective imply a number of limitations: the need to remove bone from another location in the case of autologous transplants and the possibility of an immune rejection when using allogeneic or xenogeneic grafts. To overcome these limitations, cutting edge therapies for skeletal regeneration of bone defects are currently under extensive research with promising results; such as those boosting endogenous bone regeneration, by the stimulation of host cells, or the ones driven exogenously with scaffolds, biomolecules, and mesenchymal stem cells as key players of bone healing process.


Assuntos
Regeneração Óssea/fisiologia , Osso e Ossos/fisiologia , Animais , Rejeição de Enxerto/fisiopatologia , Humanos , Células-Tronco Mesenquimais/fisiologia , Osteogênese/fisiologia , Tecidos Suporte/química , Cicatrização/fisiologia
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