RESUMO
Atherosclerosis ï¼ASï¼ is the primary reason behind cardiovascular diseases, leading to approximately one-third of global deaths. Developing a novel multi-model probe to detect AS is urgently required. Macrophages are the primary cells from which AS genesis occurs. Utilizing natural macrophage membranes coated on the surface of nanoparticles is an efficient delivery method to target plaque sites. Herein, Fe3 O4 -Cy7 nanoparticles (Fe3 O4 -Cy7 NPs), functionalized using an M2 macrophage membrane and a liposome extruder for Near-infrared fluorescence and Magnetic resonance imaging, are synthesized. These macrophage membrane-coated nanoparticles (Fe3 O4 @M2 NPs) enhance the recognition and uptake using active macrophages. Moreover, they inhibit uptake using inactive macrophages and human coronary artery endothelial cells. The macrophage membrane-coated nanoparticles (Fe3 O4 @M0 NPs, Fe3 O4 @M1 NPs, Fe3 O4 @M2 NPs) can target specific sites depending on the macrophage membrane type and are related to C-C chemofactor receptor type 2 protein content. Moreover, Fe3 O4 @M2 NPs demonstrate excellent biosafety in vivo after injection, showing a significantly higher Fe concentration in the blood than Fe3 O4 -Cy7 NPs. Therefore, Fe3 O4 @M2 NPs effectively retain the physicochemical properties of nanoparticles and depict reduced immunological response in blood circulation. These NPs mainly reveal enhanced targeting imaging capability for atherosclerotic plaque lesions.
Assuntos
Aterosclerose , Nanopartículas , Humanos , Células Endoteliais , Nanopartículas/química , Imageamento por Ressonância Magnética/métodos , Aterosclerose/diagnóstico por imagemRESUMO
Direct injection of cell-laden hydrogels shows high potentials in tissue regeneration for translational therapy. The traditional cell-laden hydrogels are often used as bulk space fillers to tissue defects after injection, likely limiting their structural controllability. On the other hand, patterned cell-laden hydrogel constructs often necessitate invasive surgical procedures. To overcome these problems, herein, we report a unique strategy for encapsulating living human cells in a pore-forming gelatin methacryloyl (GelMA)-based bioink to ultimately produce injectable hierarchically macro-micro-nanoporous cell-laden GelMA hydrogel constructs through three-dimensional (3D) extrusion bioprinting. The hydrogel constructs can be fabricated into various shapes and sizes that are defect-specific. Due to the hierarchically macro-micro-nanoporous structures, the cell-laden hydrogel constructs can readily recover to their original shapes, and sustain high cell viability, proliferation, spreading, and differentiation after compression and injection. Besides, in vivo studies further reveal that the hydrogel constructs can integrate well with the surrounding host tissues. These findings suggest that our unique 3D-bioprinted pore-forming GelMA hydrogel constructs are promising candidates for applications in minimally invasive tissue regeneration and cell therapy.
RESUMO
The purpose of this study was to investigate the cardiac-differentiation potential of induced pluripotent stem cells (iPSCs) generated from human umbilical cord-derived mesenchymal cells. Spontaneous beating colonies were observed at day 7 after the sequential addition of CHIR99021 and IWP-4. The combined use of CHIR99021 and IWP-4 downregulated the expression of pluripotency markers while upregulating cardiac transcription factors and cardiomyocyte-specific markers. The derived cardiomyocytes demonstrated typical sarcomeric structures and action-potential features; most importantly, the derived cells exhibited responsiveness to ß-adrenergic and muscarinic stimulations. The analyses of molecular, structural, and functional properties revealed that the derived cardiomyocytes were similar to cardiomyocytes derived from BJ foreskin fibroblast cells. In summary, our results demonstrate that functional cardiomyocytes can be generated from human umbilical cord-derived cells. The methodology described here has potential as a means for the production of functional cardiomyocytes from discarded human umbilical cord tissue.
RESUMO
The objective of the study was to elucidate the mechanism by which microRNA-34a (miR-34a) influences heart development and participates in the pathogenesis of congenital heart disease (CHD) by targeting NOTCH-1 through the Notch signaling pathway. Forty D7 pregnant mice were recruited for the purposes of the study and served as the CHD (n=20, successfully established as CHD model) and normal (n=20) groups. The positive expression of the NOTCH-1 protein was evaluated by means of immunohistochemistry. Embryonic endocardial cells (ECCs) were assigned into the normal, blank, negative control (NC), miR-34a mimics, miR-34a inhibitors, miR-34a inhibitors+siRNA-NOTCH-1, siRNA-NOTCH-1, miR-34a mimics+NOTCH-1 OE and miR-34a mimics+crispr/cas9 (mutant NOTCH-1) groups. The expressions of miR-34a, NOTCH-1, Jagged1, Hes1, Hey2 and Csx in cardiac tissues and ECCs were determined by both RT-qPCR and western blotting methods. MTT assay and flow cytometry were conducted for cell proliferation and apoptosis measurement. A dual luciferase reporter assay was applied to demonstrate that NOTCH-1 was the target gene of miR-34a. In comparison to the normal group, the expressions of miR-34a, Jagged1, Hes1 and Hey2 displayed up-regulated levels, while the expressions of NOTCH-1 and Csx were down-regulated in the CHD group. Compared with the blank and NC groups, the miR-34a mimics and siRNA-NOTCH-1 groups displayed reduced expressions of NOTCH-1 and Csx as well as a decreased proliferation rate, higher miR-34a, Jagged1, Hes1 and Hey2 expressions and an increased rate of apoptosis; while an reverse trend was observed in the miR-34a inhibitors group. The expressions of MiR-34a recorded increased levels in the miR-34a mimics+NOTCH-1 OE and miR-34a mimics+crispr/cas9 (mutant NOTCH-1) groups, however no changes in the expressions of NOTCH-1, Jagged1, Hes1, Hey2, Csx, as well as cell proliferation and apoptosis were observed when compared to the blank and NC groups. The results of our study demonstrated that miR-34a increases the risk of CHD through its downregulation of NOTCH-1 by modulating the Notch signaling pathway.
Assuntos
Cardiopatias Congênitas/genética , Coração/embriologia , MicroRNAs/metabolismo , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Apoptose/genética , Sequência de Bases , Ciclo Celular , Proliferação de Células , Sobrevivência Celular , Endocárdio/metabolismo , Feminino , Masculino , Camundongos , MicroRNAs/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
We have previously demonstrated that human umbilical cord-derived mesenchymal stem cells (UC-MSCs) can differentiate into cardiomyocyte-like cells. However, no contracting cells were observed during differentiation. In this study, we generated induced pluripotent stem cells (iPSCs) from UC-MSCs using mRNA reprogramming and focused on the differentiation of reprogrammed iPSCs into functional cardiomyocytes. For cardiac differentiation, the spontaneously contracting cell clusters were present on day 8 of differentiation. Immunostaining studies and cardiac-specific gene expression confirmed the cardiomyocyte phenotype of the differentiated cells. Electrophysiology studies indicated that iPSCs derived from UC-MSCs had a capacity for differentiation into nodal-, atrial-, and ventricular-like phenotypes based on action potential characteristics, and the derived cardiomyocytes exhibited responsiveness to ß-adrenergic and muscarinic stimulations. Moreover, the derived cardiomyocytes displayed spontaneous intracellular Ca2+ transients. These results demonstrate that functional cardiomyocytes can be generated from reprogrammed UC-MSCs, and the methodology described here will serve as a useful protocol to obtain functional cardiomyocytes from human mesenchymal stem cells.
Assuntos
Técnicas de Cultura de Células/métodos , Células-Tronco Pluripotentes Induzidas/citologia , Miócitos Cardíacos/citologia , Cordão Umbilical/citologia , Vírus/metabolismo , Cálcio/metabolismo , Sinalização do Cálcio , Diferenciação Celular , Forma Celular , Regulação da Expressão Gênica , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Miócitos Cardíacos/metabolismoRESUMO
Cells release exosomes into extracellular medium. Although the important roles of exosomes in many physiological and pathological processes are being revealed, the mechanism of exosome-cell interaction remains unclear. In this article, employing real-time fluorescence microscopy, the motion of exosomes on the plasma membrane or in the cytoplasm of recipient PC12 cells was observed directly. In addition, several motion modes of exosomes were revealed by single particle tracking (SPT). The changes between motion modes were also detected, presenting the dynamic courses of exosome attachment onto plasma membrane and exosome uptake. Octadecyl rhodamine B chloride (R18) was found to be useful to distinguish endocytosis from fusion during exosome uptake. Colocalization with organelle markers showed exosomes were sorted to acidic vesicles after internalization. The results provide new sight into the exosome-cell interaction mode and the intercellular trafficking of exosomes. This study will help to understand the roles of exosomes at cell level.
Assuntos
Exossomos/fisiologia , Animais , Transporte Biológico Ativo , Sistemas Computacionais , Endocitose , Corantes Fluorescentes , Lisossomos/fisiologia , Microscopia de Fluorescência , Modelos Biológicos , Movimento/fisiologia , Células PC12 , Ratos , RodaminasRESUMO
BACKGROUND: MicroRNAs (miRNAs) regulate many biological processes by post-translational gene silencing. Analysis of miRNA expression profiles is a reliable method for investigating particular biological processes due to the stability of miRNA and the development of advanced sequencing methods. However, this approach is limited by the broad specificity of miRNAs, which may target several mRNAs. RESULT: In this study, we developed a method for comprehensive annotation of miRNA array or deep sequencing data for investigation of cellular biological effects. Using this method, the specific pathways and biological processes involved in Alzheimer's disease were predicted with high correlation in four independent samples. Furthermore, this method was validated for evaluation of cadmium telluride (CdTe) nanomaterial cytotoxicity. As a result, apoptosis pathways were selected as the top pathways associated with CdTe nanoparticle exposure, which is consistent with previous studies. CONCLUSIONS: Our findings contribute to the validation of miRNA microarray or deep sequencing results for early diagnosis of disease and evaluation of the biological safety of new materials and drugs.
Assuntos
MicroRNAs/metabolismo , Transcriptoma , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Compostos de Cádmio/química , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Pontos Quânticos/química , Pontos Quânticos/toxicidade , Telúrio/química , Transcriptoma/efeitos dos fármacosRESUMO
Recently, photodynamic therapy (PDT) has been considered as a new strategy for atherosclerosis treatment. Targeted delivery of photosensitizer could significantly reduce its toxicity and enhance its phototherapeutic efficiency. CD68 is an antibody that can be conjugated to nano-drug delivery systems to actively target plaque sites, owing to its specific binding to CD68 receptors that are highly expressed on the surfaces of macrophage-derived foam cells. Liposomes are very popular nanocarriers due to their ability to encapsulate a wide range of therapeutic compounds including drugs, microRNAs and photosensitizers, and their ability to be surface-modified with targeting moieties leading to the development of nanocarriers with an improved targeted ability. Hence, we designed a Ce6-loaded liposomes using the film dispersion method, followed by the conjugation of CD68 antibody on the liposomal surface through a covalent crosslinking reaction, forming CD68-modified Ce6-loaded liposomes (CD68-Ce6-mediated liposomes). Flow cytometry results indicated that Ce6-containing liposomes were more effective in promoting intracellular uptake after laser irradiation. Furthermore, CD68-modified liposomes significantly strengthened the cellular recognization and thus internalization. Different cell lines have been incubated with the liposomes, and the results showed that CD68-Ce6-mediated liposomes had no significant cytotoxicity to coronary artery endothelial cells (HCAEC) under selected conditions. Interestingly, they promoted autophagy in foam cells through the increase in LC3-â , LC3-â ¡ expression and the decrease in p62 expression, and restrained the migration of mouse aortic vascular smooth muscle cells (MOVAS) in vitro. Moreover, the enhancement of atherosclerotic plaque stability and the reduction in the cholesterol content by CD68-Ce6-mediated liposomes were dependent on transient reactive oxygen species (ROS) generated under laser irradiation. In summary, we demonstrated that CD68-Ce6-mediated liposomes, as a photosensitizer nano-drug delivery system, have an inhibitory effect on MOVAS migration and a promotion of cholesterol efflux in foam cells, and thereby, represent promising nanocarriers for atherosclerosis photodynamic therapy.
Assuntos
Aterosclerose , Nanopartículas , Fotoquimioterapia , Placa Aterosclerótica , Porfirinas , Camundongos , Animais , Fármacos Fotossensibilizantes , Lipossomos , Placa Aterosclerótica/tratamento farmacológico , Células Endoteliais , Fotoquimioterapia/métodos , Aterosclerose/tratamento farmacológico , Porfirinas/farmacologia , Porfirinas/química , Linhagem Celular Tumoral , Nanopartículas/químicaRESUMO
Many tissues have a three-dimensional (3D) anisotropic structure compatible with their physiological functions. Engineering an in vitro 3D tissue having the natural structure and functions is a hotspot in tissue engineering with application for tissue regeneration, drug screening, and disease modeling. Despite various designs that have successfully guided the cellular alignment, only a few of them could precisely control the orientation of each layer in a multilayered construct or achieve adequate cell contact between layers. This study proposed a design of a multilayered 3D cell/scaffold model, that is, the cell-loaded aligned nanofiber film/hydrogel (ANF/Gel) model. The characterizations of the 3D cell-loaded ANF/Gel model in terms of design, construction, morphology, and cell behavior were systematically studied. The ANF was produced by efficiently aligned electrospinning using a self-designed, fast-and-easy collector, which was designed based on the parallel electrodes and modified with a larger gap area up to about 100 cm2. The nanofibers generated by this simple device presented numerous features like high orientation, uniformity in fiber diameter, and thinness. The ANF/Gel-based cell/scaffold model was formed by encapsulating cell-loaded multilayered poly(lactic-co-glycolic acid)-ANFs in hydrogel. Cells within the ANF/Gel model showed high viability and displayed aligned orientation and elongation in accordance with the nanofiber orientation in each film, forming a multilayered tissue having a layer spacing of 60 µm. This study provides a multilayered 3D cell/scaffold model for the in vitro construction of anisotropic engineered tissues, exhibiting potential applications in cardiac tissue engineering.
Assuntos
Nanofibras , Engenharia Tecidual , Hidrogéis/química , Nanofibras/química , Poliésteres/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Engenharia Tecidual/métodos , Alicerces Teciduais/químicaRESUMO
Atherosclerosis, the leading cause of death in the elderly worldwide, is typically characterized by elevated reactive oxygen species (ROS) levels and a chronic inflammatory state at the arterial plaques. Herein, pH-sensitive nanoparticles (HRRAP NPs) co-delivering all-trans retinal (ATR), an antioxidant linked to hyaluronic acid (HA) through a pH-sensitive hydrazone bond, and rapamycin (RAP), an anti-atherosclerotic drug loaded into the nanoparticle core, are developed for targeted combination therapy of atherosclerosis. In this way, HRRAP NPs might simultaneously reduce ROS levels via ATR antioxidant activity and reduce inflammation via the anti-inflammatory effect of RAP. In response to mildly acidic conditions mimicking the lesional inflammation in vitro, HRRAP NPs dissociated and both ATR and RAP were effectively released. The developed HRRAP NPs effectively inhibited pro-inflammatory macrophage proliferation, and displayed dose- and time-dependent specific internalization by different cellular models of atherosclerosis. Also, HRRAP NP combination therapy showed an efficient synergetic anti-atherosclerotic effect in vitro by effectively inhibiting the inflammatory response and oxidative stress in inflammatory cells. More importantly, HR NPs specifically accumulated in the atherosclerotic plaques of apolipoprotein E-deficient (ApoE-/-) mice, by active interaction with HA receptors overexpressed by different cells of the plaque. The treatment with HRRAP NPs remarkably inhibited the progression of atherosclerosis in ApoE-/- mice which resulted in stable plaques with considerably smaller necrotic cores, lower matrix metalloproteinase-9, and decreased proliferation of macrophages and smooth muscle cells (SMCs). Furthermore, HRRAP NPs attenuated RAP adverse effects and exhibited a good safety profile after long-term treatment in mice. Consequently, the developed pH-sensitive HRRAP NP represent a promising nanoplatform for atherosclerosis combination therapy.
Assuntos
Aterosclerose , Nanopartículas , Placa Aterosclerótica , Animais , Apolipoproteínas E , Aterosclerose/tratamento farmacológico , Ácido Hialurônico/química , Concentração de Íons de Hidrogênio , Inflamação/tratamento farmacológico , Camundongos , Camundongos Endogâmicos C57BL , Nanopartículas/química , Placa Aterosclerótica/tratamento farmacológico , Espécies Reativas de Oxigênio , Retinaldeído/uso terapêutico , Sirolimo/farmacologiaRESUMO
The signal-to-noise ratio of a sensor system is determined by the affinity of its active component for the analyte on one hand and its inertness with respect to unrelated stimuli (noise) on the other hand. Nonspecific interactions between the environment and biosensor components (typically constructed from glass, silica, or transition metal oxides) result in nonspecific adsorption onto the latter and constitute a major source of noise. We have previously introduced a polymeric interface for preventing nonspecific adsorption while allowing for high-affinity, specific interactions. It is based on the coassembly of biotinylated and nonbiotinylated poly(l-lysine)-graft-poly(ethylene glycol) from aqueous solutions to negatively charged surfaces, such as Nb(2)O(5). In this study, we investigated by atomic force microscopy the nanoscale organization of this interface for each individual step involved in the preparation of a bioactive interface: polymer adsorption, loading with streptavidin, and binding of biotinylated vesicles.
Assuntos
Complexos Multiproteicos/síntese química , Complexos Multiproteicos/metabolismo , Nióbio/metabolismo , Óxidos/metabolismo , Adsorção , Técnicas Biossensoriais/métodos , Biotinilação , Microscopia de Força Atômica , Polietilenoglicóis/química , Polietilenoglicóis/metabolismo , Polilisina/química , Polilisina/metabolismo , Polímeros/química , Ligação Proteica , Proteínas/química , Razão Sinal-Ruído , Estreptavidina , Propriedades de SuperfícieRESUMO
Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), a biodegradable polyester, has been a good candidate of biomaterial employed in tissue engineering. However, the PHBV film is hydrophobic and has no recognition sites for cell attachment. In this study, PHBV films are activated by ammonia plasma treatment to produce amino groups on the surface, followed by sequential reactions with a heterobifunctional cross-linker containing a segment of poly(ethylene glycol) (PEG) and further with RGD-containing peptides. XPS analyses of modified surfaces after each reaction step reveal that the RGD-containing peptides have been covalently grafted onto PHBV films. The result of cell viability assay indicates that the RGD-modified PHBV films exhibit a distinctly improved cellular compatibility. Moreover, according to the results of serum adsorption tests by optical waveguide lightmode spectroscopy (OWLS) and fibrinogen adsorption tests by enzyme-linked immunosorbent assay (ELISA) on unmodified and modified PHBV surfaces, the introduced PEG chains can significantly decrease the nonspecific adsorption of proteins from serum and fibrinogen from plasma, thus decreasing the risk of thrombus formation and improving the blood compatibility of implanted materials.
Assuntos
Materiais Biocompatíveis/síntese química , Oligopeptídeos/química , Poliésteres/química , Polietilenoglicóis/química , Adsorção , Materiais Biocompatíveis/farmacocinética , Proteínas Sanguíneas/metabolismo , Reagentes de Ligações Cruzadas , Humanos , Ligação Proteica/efeitos dos fármacos , Trombose/induzido quimicamenteRESUMO
The clinical efficacy of Oxaliplatin (L-OHP) is potentially limited by dose-dependent neurotoxicity and high partitioning to erythrocytes in vivo. Long-circulating liposomes could improve the pharmacokinetic profile of L-OHP and thus enhance its therapeutic efficacy and reduce its toxicity. The purpose of this study was to prepare L-OHP long-circulating liposomes (L-OHP PEG lip) by reverse-phase evaporation method (REV) and investigate their pharmacokinetic behavior based on total platinum in rat plasma using atomic absorption spectrometry (AAS). A simple and a sensitive AAS method was developed and validated to determine the total platinum originated from L-OHP liposomes in plasma. Furthermore, long-circulating liposomes were fully characterized in vitro and showed great stability when stored at 4°C for one month. The results showed that the total platinum in plasma of L-OHP long-circulating liposomes displayed a biexponential pharmacokinetic profile with five folds higher bioavailability and longer distribution half-life compared to L-OHP solution. Thus, long-circulating liposomes prolonged L-OHP circulation time and may present a potential candidate for its tumor delivery. Conclusively, the developed AAS method could serve as a reference to investigate the pharmacokinetic behavior of total platinum in biological matrices for other L-OHP delivery systems.
Assuntos
Oxaliplatina/sangue , Oxaliplatina/farmacocinética , Animais , Varredura Diferencial de Calorimetria , Liberação Controlada de Fármacos , Injeções Intravenosas , Cinética , Lipossomos , Concentração Osmolar , Oxaliplatina/administração & dosagem , Tamanho da Partícula , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Espectrofotometria Atômica , TemperaturaRESUMO
The complexity of the nervous system structure and function, and its slow regeneration rate, makes it more difficult to treat compared to other tissues in the human body when an injury occurs. Moreover, the current therapeutic approaches including the use of autografts, allografts, and pharmacological agents have several drawbacks and can not fully restore nervous system injuries. Recently, nanotechnology and tissue engineering approaches have attracted many researchers to guide tissue regeneration in an effective manner. Owing to their remarkable physicochemical and biological properties, two-dimensional (2D) nanomaterials have been extensively studied in the tissue engineering and regenerative medicine field. The great conductivity of these materials makes them a promising candidate for the development of novel scaffolds for neural tissue engineering application. Moreover, the high loading capacity of 2D nanomaterials also has attracted many researchers to utilize them as a drug/gene delivery method to treat various devastating nervous system disorders. This review will first introduce the fundamental physicochemical properties of 2D nanomaterials used in biomedicine and the supporting biological properties of 2D nanomaterials for inducing neuroregeneration, including their biocompatibility on neural cells, the ability to promote the neural differentiation of stem cells, and their immunomodulatory properties which are beneficial for alleviating chronic inflammation at the site of the nervous system injury. It also discusses various types of 2D nanomaterials-based scaffolds for neural tissue engineering applications. Then, the latest progress on the use of 2D nanomaterials for nervous system disorder treatment is summarized. Finally, a discussion of the challenges and prospects of 2D nanomaterials-based applications in neural tissue engineering is provided.
Assuntos
Nanoestruturas , Engenharia Tecidual , Humanos , Nanotecnologia , Sistema Nervoso , Medicina RegenerativaRESUMO
Three-dimensional (3D) cell culture using hydrogel scaffolds can closely resemble the natural extracellular matrix (ECM), which offers appropriate mechanical support for cells and regulates cellular behavior. In this study, a bacterial transpeptidase sortase A (SA) is used to prepare enzymatically cross-linked methacrylated hyaluronic acid (HA) peptides (HAMA-P) hydrogel, which reveals fast gel kinetics under high SA cross-linking concentrations and can be used as an injection hydrogel for tissue repair or extrusive 3D bioprinting. Furthermore, methacrylated gelatin (GelMA) is introduced to build the hybrid hydrogel (HAMA-P-GelMA) with double cross-linking of enzyme-UV, which has shown proper physical properties (mechanical properties, swelling, degradation rate, etc.) of the hydrogel matrix, and displayed desirable effects on cell viability, adhesion, and cell spreading, when compared to GelMA or HAMA-P single-network hydrogels. The HAMA-P-GelMA hybrid hydrogels provide a favorable 3D milieu for cell growth and can be used as a 3D bio-ink or a carrier of stem cells/cytokines for injectable tissue repair and filling.
Assuntos
Hidrogéis , Raios Ultravioleta , Bioimpressão , Técnicas de Cultura de Células , HumanosRESUMO
The development of diversified biomaterials in tissue engineering has been promoted by growing research into carbon-based nanomaterials. Usually, ideal scaffold materials should possess properties similar to the extracellular matrix of natural myocardial tissue. In this study, dopamine-reduced graphene oxide (GO), was prepared and doped into gelatin methacrylate (GelMA) hydrogels, resulting in novel conductive and mechanical properties for controlling cell growth. Cardiomyocytes (CMs) cultured on PDA-rGO-incorporated hydrogels (GelMA-PDA-rGO) had greater cytocompatibility than those cultured on GelMA hydrogels, as evidenced by higher cell survival rates and up-regulation of cardiac-relevant proteins. Finally, electrical stimulation was applied to facilitate the maturation of CMs which was seeded on different hydrogels. The findings revealed that electrical stimulation of conductive hybrid hydrogel scaffolds improved the orientational order parameter of sarcomeres (OOP). In addition, propagation of intercellular pacing signals, which improves the expression of gap junction proteins was noticed, likewise calcium handling capacity was present in conductive hybrid hydrogels compared to those in pure GelMA group. This study has shown that the combination of GelMA-PDA-rGO based conductive hydrogels and electrical stimulation possessed synergistic effects for engineering a more functional and mature myocardium layer as well as further application in drug screening and disease modeling in vitro.
Assuntos
Hidrogéis , Miócitos Cardíacos , Animais , Animais Recém-Nascidos , Estimulação Elétrica , Gelatina , Grafite , Indóis , Polímeros , Ratos , Engenharia Tecidual , Alicerces TeciduaisRESUMO
In vitro model of the human cardiac tissues generated from human induced pluripotent stem cells (hiPSCs) could facilitate drug discovery and patient-specific studies of physiology and disease. However, the immature state of hiPSC-derived cardiomyocytes (hiPSC-CMs) compared to adult myocardium is a key defect that must be overcome to enable the potential applications of hiPSC-CMs in drug testing. For this purpose, we developed a heart-on-a-chip device that contains microfluidic channels for long-term dynamic culture of cells, platinum wire electrodes for electrical stimulation of hiPSC-CMs, and gold electrode arrays as acquisition electrodes for real-time recording electrophysiological signals of cardiac tissues. Human iPSC-CMs cultured on biocompatible hydrogels in the chip chamber can be electrically stimulated to prompt the maturation of cardiomyocytes (CMs) and generate functional cardiac tissues. Drug tests were performed with calcium transient measurements to evaluate drug responsiveness of electrical stimulated and unstimulated cardiac tissues. The results show that only the electrical-stimulated cardiac tissues respond correctly to drug treatment of verapamil and isoprenaline, indicating the reliability of this engineered cardiac tissues for drug testing. The above integrated heart-on-a-chip device provides a promising platform for drug efficacy testing and cardiactoxicity.
Assuntos
Técnicas Biossensoriais , Células-Tronco Pluripotentes Induzidas , Adulto , Diferenciação Celular , Células Cultivadas , Humanos , Dispositivos Lab-On-A-Chip , Miócitos Cardíacos , Reprodutibilidade dos TestesRESUMO
A new preparation procedure was developed for the stable adsorption of either the cytoplasmic or the nuclear face of native (i.e. in physiological buffer without detergent extraction and in the absence of chemical fixatives) Xenopus oocyte nuclear envelopes (NEs) onto silicon (Si) surfaces. This yields optimal structural preservation of the nuclear pore complexes (NPCs) without compromising their functional properties. The functional viability of thus prepared NPCs was documented by time-lapse atomic force microscopy (AFM) of the reversible calcium-mediated opening (i.e. +Ca(2+)) and closing (i.e. -Ca(2+)) of the iris diaphragm-like distal ring topping the NPCs' nuclear baskets. Moreover, site-specific single colloidal gold particle detection was documented by AFM imaging one and the same NPC before and after immuno-gold labeling the sample with a nucleoporin-specific antibody. With this new preparation protocol at hand, we should eventually be able to follow by time-lapse AFM transport of single gold-conjugated cargos across individual NPCs.
Assuntos
Microscopia de Força Atômica/métodos , Poro Nuclear/metabolismo , Animais , Transporte Biológico/fisiologia , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Oócitos/metabolismo , Xenopus laevis/metabolismoRESUMO
Correction for 'High-aspect-ratio water-dispersed gold nanowires incorporated within gelatin methacrylate hydrogels for constructing cardiac tissues in vitro' by Xiao-Pei Li et al., J. Mater. Chem. B, 2020, 8, 7213-7224, DOI: .
RESUMO
The field of cardiac tissue engineering has made significant strides in therapeutic and pharmaceutical applications, highlighted by the development of smart biomaterials. Scaffolds with appropriate properties mimicking the nature of a heart matrix will be highly beneficial for cardiac tissue engineering. In this study, high-aspect-ratio water-dispersed gold nanowires (AuNWs) were synthesized and incorporated into gelatin methacrylate (GelMA) hydrogels, demonstrating enhanced electrical conductivity and mechanical properties of the biomaterial scaffolds. Cardiac cells cultured on GelMA-AuNW hybrid hydrogels exhibited better biological activities such as cell viability and maturation state compared to those cultured on GelMA hydrogels. Moreover, cardiomyocytes showed synchronous beating activity and a faster spontaneous beating rate on GelMA-AuNW hybrid hydrogels. Our strategy of integrating high-aspect-ratio water-dispersed gold nanowires within gelatin methacrylate hydrogels provides a favorable biomaterial scaffold to construct functional cardiac tissue for further applications in cardiac tissue engineering and drug screening.