RESUMO
Direct reprogramming is an efficient strategy to produce cardiac lineage cells necessary for cardiac tissue engineering and drug testing for cardiac toxicity. However, functional maturation of reprogrammed cardiomyocytes, which is of great importance for their regenerative potential and drug response, still remains challenging. In this study, we propose a novel electrode platform to promote direct cardiac reprogramming and improve the functionality of reprogrammed cardiac cells. Nonviral cardiac reprogramming was improved via a three-dimensional spheroid culture of chemically induced cardiomyocytes exposed to a small-molecule cocktail. A micropillar electrode array providing biphasic electrical pulses mimicking the heartbeat further enhanced maturation and electrophysiological properties of reprogrammed cardiac spheroids, leading to proper responses and increased sensitivity to drugs. On the basis of our results, we conclude that our device may have a wider application in the generation of functional cardiac cells for regenerative medicine and screening of novel drugs.
Assuntos
Células-Tronco Pluripotentes Induzidas , Preparações Farmacêuticas , Eletrodos , Frequência Cardíaca , Miócitos CardíacosRESUMO
CpG, 5'-C-phosphate-G-3', islands (CGIs) have long been known for their association with enhancers, silencers, and promoters, and for their epigenetic signatures. They are maintained in embryonic stem cells (ESCs) in a poised but inactive state via the formation of bivalent chromatin containing both active and repressive marks. CGIs also occur within coding sequences, where their functional role has remained obscure. Intragenic CGIs (iCGIs) are largely absent from housekeeping genes, but they are found in all genes associated with organ development and cell lineage control. In this paper, we investigated the epigenetic status of iCGIs and found that they too reside in bivalent chromatin in ESCs. Cell type-specific DNA methylation of iCGIs in differentiated cells was linked to the loss of both the H3K4me3 and H3K27me3 marks, and disruption of physical interaction with promoter regions, resulting in transcriptional activation of key regulators of differentiation such as PAXs, HOXs, and WNTs. The differential epigenetic modification of iCGIs appears to be mediated by cell type-specific transcription factors distinct from those bound by promoter, and these transcription factors may be involved in the hypermethylation of iCGIs upon cell differentiation. iCGIs thus play a key role in the cell type-specific regulation of transcription.
Assuntos
Diferenciação Celular/genética , Ilhas de CpG/genética , Metilação de DNA/genética , Epigênese Genética/genética , Linhagem da Célula/genética , Cromatina/genética , Células-Tronco Embrionárias/citologia , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica no Desenvolvimento , Histonas/genética , Humanos , Regiões Promotoras GenéticasRESUMO
While neural cell transplantation represents a promising therapy for neurodegenerative diseases, the formation of functional networks of transplanted cells with host neurons constitutes one of the challenging steps. Here, we introduce a magnetic guidance methodology that controls neurite growth signaling via magnetic nanoparticles (MNPs) conjugated with antibodies targeting the deleted in colorectal cancer (DCC) receptor (DCC-MNPs). Activation of the DCC receptors by clusterization and subsequent axonal growth of the induced neuronal (iN) cells was performed in a spatially controlled manner. In addition to the directionality of the magnetically controlled axon projection, axonal growth of the iN cells assisted the formation of functional connections with pre-existing primary neurons. Our results suggest magnetic guidance as a strategy for improving neuronal connectivity by spatially guiding the axonal projections of transplanted neural cells for synaptic interactions with the host tissue.
Assuntos
Anticorpos/química , Axônios/metabolismo , Reprogramação Celular , Receptor DCC/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Campos Magnéticos , Nanopartículas de Magnetita/química , Receptor DCC/antagonistas & inibidores , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Neuritos/metabolismoRESUMO
Electrically conductive hyaluronic acid (HA) hydrogels incorporated with single-walled carbon nanotubes (CNTs) and/or polypyrrole (PPy) were developed to promote differentiation of human neural stem/progenitor cells (hNSPCs). The CNT and PPy nanocomposites, which do not easily disperse in aqueous phases, dispersed well and were efficiently incorporated into catechol-functionalized HA (HA-CA) hydrogels by the oxidative catechol chemistry used for hydrogel cross-linking. The prepared electroconductive HA hydrogels provided dynamic, electrically conductive three-dimensional (3D) extracellular matrix environments that were biocompatible with hNSPCs. The HA-CA hydrogels containing CNT and/or PPy significantly promoted neuronal differentiation of human fetal neural stem cells (hfNSCs) and human induced pluripotent stem cell-derived neural progenitor cells (hiPSC-NPCs) with improved electrophysiological functionality when compared to differentiation of these cells in a bare HA-CA hydrogel without electroconductive motifs. Calcium channel expression was upregulated, depolarization was activated, and intracellular calcium influx was increased in hNSPCs that were differentiated in 3D electroconductive HA-CA hydrogels; these data suggest a potential mechanism for stem cell neurogenesis. Overall, our bioinspired, electroconductive HA hydrogels provide a promising cell-culture platform and tissue-engineering scaffold to improve neuronal regeneration.
Assuntos
Hidrogéis/química , Células-Tronco Neurais/citologia , Neurogênese , Alicerces Teciduais/química , Catecóis/química , Linhagem Celular , Condutividade Elétrica , Humanos , Ácido Hialurônico/química , Hidrogéis/farmacologia , Nanotubos de Carbono/química , Células-Tronco Neurais/efeitos dos fármacos , Células-Tronco Neurais/metabolismo , Polímeros/química , Pirróis/químicaRESUMO
Using small interfering RNA (siRNA) to regulate gene expression is an emerging strategy for stem cell manipulation to improve stem cell therapy. However, conventional methods of siRNA delivery into stem cells based on solution-mediated transfection are limited due to low transfection efficiency and insufficient duration of cell-siRNA contact during lengthy culturing protocols. To overcome these limitations, a bio-inspired polymer-mediated reverse transfection system is developed consisting of implantable poly(lactic-co-glycolic acid) (PLGA) scaffolds functionalized with siRNA-lipidoid nanoparticle (sLNP) complexes via polydopamine (pDA) coating. Immobilized sLNP complexes are stably maintained without any loss of siRNA on the pDA-coated scaffolds for 2 weeks, likely due to the formation of strong covalent bonds between amine groups of sLNP and catechol group of pDA. siRNA reverse transfection with the pDA-sLNP-PLGA system does not exhibit cytotoxicity and induces efficient silencing of an osteogenesis inhibitor gene in human adipose-derived stem cells (hADSCs), resulting in enhanced osteogenic differentiation of hADSCs. Finally, hADSCs osteogenically committed on the pDA-sLNP-PLGA scaffolds enhanced bone formation in a mouse model of critical-sized bone defect. Therefore, the bio-inspired reverse transfection system can provide an all-in-one platform for genetic modification, differentiation, and transplantation of stem cells, simultaneously enabling both stem cell manipulation and tissue engineering.
Assuntos
Tecido Adiposo/citologia , Diferenciação Celular/fisiologia , Osteogênese/fisiologia , Células-Tronco/citologia , Regeneração Óssea/genética , Regeneração Óssea/fisiologia , Diferenciação Celular/genética , Humanos , Ácido Láctico/química , Osteogênese/genética , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , RNA Interferente Pequeno/química , Engenharia Tecidual/métodos , Alicerces Teciduais/químicaRESUMO
Over the last few decades, stem cell therapies have been highlighted for their potential to heal damaged tissue and aid in tissue reconstruction. However, materials used to deliver and support implanted cells often display limited efficacy, which has resulted in delaying translation of stem cell therapies into the clinic. In our previous work, we developed a mussel-inspired, catechol-functionalized hyaluronic acid (HA-CA) hydrogel that enabled effective cell transplantation due to its improved biocompatibility and strong tissue adhesiveness. The present study was performed to further expand the utility of HA-CA hydrogels for use in stem cell therapies to treat more clinically relevant tissue defect models. Specifically, we utilized HA-CA hydrogels to potentiate stem cell-mediated angiogenesis and osteogenesis in two tissue defect models: critical limb ischemia and critical-sized calvarial bone defect. HA-CA hydrogels were found to be less cytotoxic to human adipose-derived stem cells (hADSCs) in vitro compared to conventional photopolymerized HA hydrogels. HA-CA hydrogels also retained the angiogenic functionality of hADSCs and supported osteogenic differentiation of hADSCs. Because of their superior tissue adhesiveness, HA-CA hydrogels were able to mediate efficient engraftment of hADSCs into the defect regions. When compared to photopolymerized HA hydrogels, HA-CA hydrogels significantly enhanced hADSC-mediated therapeutic angiogenesis (promoted capillary/arteriole formation, improved vascular perfusion, attenuated ischemic muscle degeneration/fibrosis, and reduced limb amputation) and bone reconstruction (mineralized bone formation, enhanced osteogenic marker expression, and collagen deposition). This study proves the feasibility of using bioinspired HA-CA hydrogels as functional biomaterials for improved tissue regeneration in critical tissue defects.
Assuntos
Tecido Adiposo/citologia , Catecóis/química , Ácido Hialurônico/química , Hidrogéis/química , Neovascularização Fisiológica/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Transplante de Células-Tronco , Células-Tronco/citologia , Animais , Proteína Morfogenética Óssea 2/metabolismo , Catecóis/farmacologia , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Traumatismos Craniocerebrais/terapia , Modelos Animais de Doenças , Feminino , Membro Posterior/irrigação sanguínea , Humanos , Ácido Hialurônico/farmacologia , Hidrogéis/farmacologia , Isquemia/terapia , Camundongos Nus , Crânio/efeitos dos fármacos , Crânio/lesões , Alicerces Teciduais/químicaRESUMO
Membrane transporters belonging to the multidrug and toxic compound extrusion family mediate the efflux of unrelated pharmaceuticals from the interior of the cell in organisms ranging from bacteria to human. These proteins are thought to fall into two classes that couple substrate efflux to the influx of either Na(+) or H(+). We studied the energetics of drug extrusion by NorM from Vibrio cholerae in proteoliposomes in which purified NorM protein was functionally reconstituted in an inside-out orientation. We establish that NorM simultaneously couples to the sodium-motive force and proton-motive force, and biochemically identify protein regions and residues that play important roles in Na(+) or H(+) binding. As the positions of protons are not available in current medium and high-resolution crystal structures of multidrug and toxic compound extrusion transporters, our findings add a previously unrecognized parameter to mechanistic models based of these structures.
Assuntos
Antiporters/metabolismo , Proteínas de Bactérias/metabolismo , Prótons , Sódio/metabolismo , Vibrio cholerae/metabolismo , Antiporters/química , Antiporters/genética , Ácido Aspártico/química , Ácido Aspártico/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Sítios de Ligação , Transporte Biológico , Western Blotting , Membrana Celular/metabolismo , DNA/metabolismo , Etídio/metabolismo , Ácido Glutâmico/química , Ácido Glutâmico/metabolismo , Lactococcus lactis/genética , Lactococcus lactis/metabolismo , Modelos Moleculares , Mutação , Ligação Proteica , Estrutura Terciária de Proteína , Proteolipídeos/metabolismo , Força Próton-Motriz , Vibrio cholerae/genéticaRESUMO
This study evaluates the efficacy of a decellularized intestine tissue-derived extracellular matrix (Intestine ECM) as a scaffold for culturing colorectal cancer (CRC) organoids and establishing cellderived xenograft (CDX) models, comparing its performance to traditional Matrigel. Intestine ECM demonstrates comparable support for organoid formation and cellular function, highlighting its potential as a more physiologically relevant and reproducible platform. Our findings suggest that Intestine ECM enhances the mimetic environment for colon epithelium, supporting comparable growth and improved differentiation compared to Matrigel. Moreover, when used as a delivery carrier, Intestine ECM significantly increases the growth rate of CDX models using patient-derived primary colorectal cancer cells. This enhancement demonstrates Intestine ECM's role not only as a scaffold but also as a vital component of the tumor microenvironment, facilitating more robust tumorigenesis. These findings advocate for the broader application of Intestine ECM in cancer model systems, potentially leading to more accurate preclinical evaluations and the development of targeted cancer therapies.
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Inflammatory bowel disease (IBD), including Crohn's disease and ulcerative colitis, is a family of chronic disorders along the gastrointestinal tract. Because of its idiopathic nature, IBD does not have a fundamental cure; current available therapies for IBD are limited to prolonged doses of immunomodulatory agents. While these treatments may reduce inflammation, limited therapeutic efficacy, inconsistency across patients, and adverse side effects from aggressive medications remain as major drawbacks. Recently, excessive production and accumulation of neutrophil extracellular traps (NETs) also known as NETosis have been identified to exacerbate inflammatory responses and induce further tissue damage in IBD. Such discovery invited many researchers to investigate NETs as a potential therapeutic target. DNase-I is a natural agent that can effectively destroy NETs and, therefore, potentially reduce NETs-induced inflammations even without the use of aggressive drugs. However, low stability and rapid clearance of DNase-I remain as major limitations for further therapeutic applications. In this research, polymeric nanozymes were fabricated to increase the delivery and therapeutic efficacy of DNase-I. DNase-I was immobilized on the surface of polymeric nanoparticles to maintain its enzymatic properties while extending its activity in the colon. Delivery of DNase-I using this platform allowed enhanced stability and prolonged activity of DNase-I with minimal toxicity. When administered to animal models of IBD, DNase-I nanozymes successfully alleviated various pathophysiological symptoms of IBD. More importantly, DNase-I nanozyme administration successfully attenuated neutrophil infiltration and NETosis in the colon compared to free DNase-I or mesalamine.
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Thein vitrosimulation of organs resolves the accuracy, ethical, and cost challenges accompanyingin vivoexperiments. Organoids and organs-on-chips have been developed to model thein vitro, real-time biological and physiological features of organs. Numerous studies have deployed these systems to assess thein vitro, real-time responses of an organ to external stimuli. Particularly, organs-on-chips can be most efficiently employed in pharmaceutical drug development to predict the responses of organs before approving such drugs. Furthermore, multi-organ-on-a-chip systems facilitate the close representations of thein vivoenvironment. In this review, we discuss the biosensing technology that facilitates thein situ, real-time measurements of organ responses as readouts on organ-on-a-chip systems, including multi-organ models. Notably, a human-on-a-chip system integrated with automated multi-sensing will be established by further advancing the development of chips, as well as their assessment techniques.
Assuntos
Sistemas Microfisiológicos , Organoides , HumanosRESUMO
Developing bioelectronics that retains their long-term functionalities in the human body during daily activities is a current critical issue. To accomplish this, robust tissue adaptability and biointerfacing of bioelectronics should be achieved. Hydrogels have emerged as promising materials for bioelectronics that can softly adapt to and interface with tissues. However, hydrogels lack toughness, requisite electrical properties, and fabrication methodologies. Additionally, the water-swellable property of hydrogels weakens their mechanical properties. In this work, an intrinsically nonswellable multifunctional hydrogel exhibiting tissue-like moduli ranging from 10 to 100 kPa, toughness (400-873 J m-3 ), stretchability (≈1000% strain), and rapid self-healing ability (within 5 min), is developed. The incorporation of carboxyl- and hydroxyl-functionalized carbon nanotubes (fCNTs) ensures high conductivity of the hydrogel (≈40 S m-1 ), which can be maintained and recovered even after stretching or rupture. After a simple chemical modification, the hydrogel shows tissue-adhesive properties (≈50 kPa) against the target tissues. Moreover, the hydrogel can be 3D printed with a high resolution (≈100 µm) through heat treatment owing to its shear-thinning capacity, endowing it with fabrication versatility. The hydrogel is successfully applied to underwater electromyography (EMG) detection and ex vivo bladder expansion monitoring, demonstrating its potential for practical bioelectronics.
Assuntos
Hidrogéis , Nanotubos de Carbono , Humanos , Hidrogéis/química , Nanotubos de Carbono/química , Condutividade ElétricaRESUMO
Organoids show great potential in clinical translational research owing to their intriguing properties to represent a near physiological model for native tissues. However, the dependency of organoid generation on the use of poorly defined matrices has hampered their clinical application. Current organoid culture systems mostly reply on biochemical signals provided by medium compositions and cell-cell interactions to control growth. Recent studies have highlighted the importance of the extracellular matrix (ECM) composition, cell-ECM interactions, and mechanical signals for organoid expansion and differentiation. Thus, several hydrogel systems prepared using natural or synthetic-based materials have been designed to recreate the stem cell niche in vitro, providing biochemical, biophysical, and mechanical signals. In this review, we discuss how recapitulating multiple aspects of the tissue-specific environment through designing and applying matrices could contribute to accelerating the translation of organoid technology from the laboratory to therapeutic and pharmaceutical applications.
RESUMO
A 3D microenvironment with dynamic cell-biomaterial interactions was developed using a dual-responsive system for in situ microenvironment remodeling and control of cellular function. A visible-light-responsive polymer was utilized to prepare a hydrogel with photodegradation properties, enabling in situ microenvironment remodeling. Additionally, a vascular endothelial growth factor (VEGF) gene activation unit that was responsive to the same wavelength of light was incorporated to support the potential application of the system in regenerative medicine. Following light exposure, the mechanical properties of the photodegradable hydrogel gradually deteriorated, and product analysis confirmed the degradation of the hydrogel, and thereby, 3D microenvironment remodeling. In situ microenvironment remodeling influenced stem cell proliferation and enlargement within the hydrogel. Furthermore, stem cells engineered to express light-activated VEGF and incorporated into the dual-responsive system were applied to wound healing and an ischemic hindlimb model, proving their potential application in regenerative medicine.
Assuntos
Hidrogéis , Fator A de Crescimento do Endotélio Vascular , Animais , Materiais Biocompatíveis/farmacologia , Hidrogéis/metabolismo , Luz , Ativação Transcricional , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Direct cardiac reprogramming has emerged as a promising therapeutic approach for cardiac regeneration. Full chemical reprogramming with small molecules to generate cardiomyocytes may be more amenable than genetic reprogramming for clinical applications as it avoids safety concerns associated with genetic manipulations. However, challenges remain regarding low conversion efficiency and incomplete cardiomyocyte maturation. Furthermore, the therapeutic potential of chemically induced cardiomyocytes (CiCMs) has not been investigated. Here, we report that a three-dimensional microenvironment reconstituted with decellularized heart extracellular matrix can enhance chemical reprogramming and cardiac maturation of fibroblasts to cardiomyocytes. The resultant CiCMs exhibit elevated cardiac marker expression, sarcomeric organization, and improved electrophysiological features and drug responses. We investigated the therapeutic potential of CiCMs reprogrammed in three-dimensional heart extracellular matrix in a rat model of myocardial infarction. Our platform can facilitate the use of CiCMs for regenerative medicine, disease modeling, and drug screening.
Assuntos
Miócitos Cardíacos , Regeneração , Ratos , Animais , Miócitos Cardíacos/metabolismo , Medicina Regenerativa/métodos , Matriz Extracelular , Fibroblastos/metabolismoRESUMO
Pluripotent and direct reprogramming technologies hold great potential for tissue repair and restoration of tissue and organ function. The implementation of induced pluripotent stem cells and directly reprogrammed cells in biomedical research has resulted in a significant leap forward in the highly promising area of regenerative medicine. While these therapeutic strategies are promising, there are several obstacles to overcome prior to the introduction of these therapies into clinical settings. Bioengineering technologies, such as biomaterials, bioprinting, microfluidic devices, and biostimulatory systems, can enhance cell viability, differentiation, and function, in turn the efficacy of cell therapeutics generated via pluripotent and direct reprogramming. Therefore, cellular reprogramming technologies, in combination with tissue-engineering platforms, are poised to overcome current bottlenecks associated with cell-based therapies and create new ways of producing engineered tissue substitutes.
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Skeletal muscle has an inherent capacity for spontaneous regeneration. However, recovery after severe injuries such as volumetric muscle loss (VML) is limited. There is therefore a need to develop interventions to induce functional skeletal muscle restoration. One suggested approach includes tissue-engineered muscle constructs. Tissue-engineering treatments have so far been impeded by the lack of reliable cell sources and the challenges in engineering of suitable tissue scaffolds. To address these challenges, muscle extracellular matrix (MEM) and induced skeletal myogenic progenitor cells (iMPCs) are integrated within thermally drawn fiber based microchannel scaffolds. The microchannel fibers decorated with MEM enhance differentiation and maturation of iMPCs. Furthermore, engraftment of these bioengineered hybrid muscle constructs induce de novo muscle regeneration accompanied with microvessel and neuromuscular junction formation in a VML mouse model, ultimately leading to functional recovery of muscle activity.
Assuntos
Músculo Esquelético/lesões , Músculo Esquelético/fisiologia , Animais , Humanos , Porosidade , Regeneração , Engenharia TecidualRESUMO
Immunomodulation in the local tissue microenvironment is pivotal for the determination of macrophage phenotypes and regulation of functions necessary for pro-healing effects. Herein, we demonstrate that a lymph node extracellular matrix (LNEM) prepared by the decellularization of lymph node tissues can mimic lymph node microenvironments for immunomodulation in two-dimensional (2D) and three-dimensional (3D) formats. The LNEM exhibits strengthened immunomodulatory effects in comparison to conventional collagen-based platforms. A 3D LNEM hydrogel is more effective than the 2D LNEM coating in inducing M2 macrophage polarization. The 3D LNEM induces macrophage elongation and enhances the M2-type marker expression and the secretion of anti-inflammatory cytokines. Additionally, the phagocytic function of macrophages is improved upon exposure to the intricate 3D LNEM environment. We demonstrate the reduced susceptibility of liver organoids to a hepatotoxic drug when co-cultured with macrophages in a 3D LNEM. This effect could be attributed to the enhanced anti-inflammatory functions and indicates its potential as a drug-testing platform that enables drug responses similar to those observed in vivo. Finally, the implantation of an LNEM hydrogel in a mouse volumetric muscle loss model facilitates the recruitment of host macrophages to the site of injury and enhances macrophage polarization toward the M2 phenotype for tissue healing in vivo. Therefore, 3D immune system-mimicking biomaterials could serve as useful platforms for tissue modeling and regenerative medicine development.
Assuntos
Matriz Extracelular/química , Linfonodos/química , Ativação de Macrófagos , Macrófagos/imunologia , Alicerces Teciduais/química , Animais , Materiais Biocompatíveis/química , Matriz Extracelular/imunologia , Imunomodulação , Linfonodos/imunologia , Macrófagos/citologia , SuínosRESUMO
Brain organoids derived from human pluripotent stem cells provide a highly valuable in vitro model to recapitulate human brain development and neurological diseases. However, the current systems for brain organoid culture require further improvement for the reliable production of high-quality organoids. Here, we demonstrate two engineering elements to improve human brain organoid culture, (1) a human brain extracellular matrix to provide brain-specific cues and (2) a microfluidic device with periodic flow to improve the survival and reduce the variability of organoids. A three-dimensional culture modified with brain extracellular matrix significantly enhanced neurogenesis in developing brain organoids from human induced pluripotent stem cells. Cortical layer development, volumetric augmentation, and electrophysiological function of human brain organoids were further improved in a reproducible manner by dynamic culture in microfluidic chamber devices. Our engineering concept of reconstituting brain-mimetic microenvironments facilitates the development of a reliable culture platform for brain organoids, enabling effective modeling and drug development for human brain diseases.
Assuntos
Encéfalo/crescimento & desenvolvimento , Encéfalo/fisiologia , Dispositivos Lab-On-A-Chip , Neurogênese/fisiologia , Organoides/crescimento & desenvolvimento , Organoides/fisiologia , Animais , Encéfalo/citologia , Meios de Cultura , Fenômenos Eletrofisiológicos , Matriz Extracelular/fisiologia , Estudos de Viabilidade , Perfilação da Expressão Gênica , Humanos , Hidrogéis , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Modelos Anatômicos , Modelos Neurológicos , Neurogênese/genética , Neuroglia/citologia , Neuroglia/fisiologia , Técnicas de Cultura de Órgãos/instrumentação , Técnicas de Cultura de Órgãos/métodos , Organoides/citologia , SuínosRESUMO
Myelination by oligodendrocytes (OLs) is a key developmental milestone in terms of the functions of the central nervous system (CNS). Demyelination caused by defects in OLs is a hallmark of several CNS disorders. Although a potential therapeutic strategy involves treatment with the myelin-forming cells, there is no readily available source of these cells. OLs can be differentiated from pluripotent stem cells; however, there is a lack of efficient culture systems that generate functional OLs. Here, we demonstrate biomimetic approaches to promote OL differentiation from human-induced pluripotent stem cells (iPSCs) and to enhance the maturation and myelination capabilities of iPSC-derived OL (iPSC-OL). Functionalization of culture substrates using the brain extracellular matrix (BEM) derived from decellularized human brain tissue enhanced the differentiation of iPSCs into myelin-expressing OLs. Co-culture of iPSC-OL with induced neuronal (iN) cells on BEM substrates, which closely mimics the in vivo brain microenvironment for myelinated neurons, not only enhanced myelination of iPSC-OL but also improved electrophysiological function of iN cells. BEM-functionalized aligned electrospun nanofibrous scaffolds further promoted the maturation of iPSC-OLs, enhanced the production of myelin sheath-like structures by the iPSC-OL, and enhanced the neurogenesis of iN cells. Thus, the biomimetic strategy presented here can generate functional OLs from stem cells and facilitate myelination by providing brain-specific biochemical, biophysical, and structural signals. Our system comprising stem cells and brain tissue from human sources could help in the establishment of human demyelination disease models and the development of regenerative cell therapy for myelin disorders.
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Encéfalo/metabolismo , Matriz Extracelular/química , Bainha de Mielina/fisiologia , Diferenciação Celular , Linhagem Celular , Técnicas de Cocultura , Fenômenos Eletrofisiológicos , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Proteína Básica da Mielina/metabolismo , Nanofibras/química , Neurogênese , Neurônios/citologia , Neurônios/metabolismo , Neurotransmissores/farmacologia , Oligodendroglia/citologia , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/metabolismoRESUMO
Artificial taste devices for tastant sensing and taste information standardization are attracting increasing attention with the exponential growth of the food and beverage industries. Despite recent developments in artificial taste sensors incorporating polymers, lipid membranes, and synthetic vesicles, current devices have limited functionality and sensitivity, and are complex to manufacture. Moreover, such synthetic systems cannot simulate the taste signal transmissions that are critical for complicated taste perception. The current document describes a primary taste cell-based artificial tongue that can mimic taste sensing. To maintain viable and functional taste cells required for in vitro tastant sensing, a tongue extracellular matrix (TEM) prepared by decellularization of tongue tissue was applied to two- and three-dimensional taste cell cultures. The TEM-based system recreates the tongue's microenvironment and significantly improves the functionality of taste cells for sensing tastant molecules by enhancing cellular adhesion and gustatory gene expression compared with conventional collagen-based systems. The TEM-based platform simulates signal transmission from tastant-treated taste cells to adjacent neuronal cells, which was impossible with previous artificial taste sensors. The artificial tongue device may provide highly efficient, functional sensors for tastant detection and in vitro organ models that mimic the tongue allowing elucidation of the mechanisms of taste.