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Adult zebrafish are able to heal large-sized cutaneous wounds in hours with little to no scarring. This rapid re-epithelialization is crucial for preventing infection and jumpstarting the subsequent regeneration of damaged tissues. Despite significant progress in understanding this process, it remains unclear how vast numbers of epithelial cells are orchestrated on an organismic scale to ensure the timely closure of millimeter-sized wounds. Here, we report an unexpected role of adult zebrafish appendages (fins) in accelerating the re-epithelialization process. Through whole-body monitoring of single-cell dynamics in live animals, we found that fin-resident epithelial cells (FECs) are highly mobile and migrate to cover wounds in nearby body regions. Upon injury, FECs readily undergo organ-level mobilization, allowing for coverage of body surfaces of up to 4.78 mm2 in less than 8 h. Intriguingly, long-term fate-tracking experiments revealed that the migratory FECs are not short-lived at the wound site; instead, the cells can persist on the body surface for more than a year. Our experiments on "fin-less" and "fin-gaining" individuals demonstrated that the fin structures are not only capable of promoting rapid re-epithelialization but are also necessary for the process. We further found that fin-enriched extracellular matrix laminins promote the active migration of FECs by facilitating lamellipodia formation. These findings lead us to conclude that appendage structures in regenerative vertebrates, such as fins, may possess a previously unrecognized function beyond serving as locomotor organs. The appendages may also act as a massive reservoir of healing cells, which speed up wound closure and tissue repair.
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Células Epiteliales , Cicatrización de Heridas , Pez Cebra , Animales , Pez Cebra/fisiología , Células Epiteliales/fisiología , Cicatrización de Heridas/fisiología , Repitelización/fisiología , Movimiento Celular , Aletas de Animales/fisiología , Aletas de Animales/lesionesRESUMEN
Electric fields (EF) play an essential role in cancer cell migration. Numerous cancer cell types exhibit electrotaxis under direct current electric fields (dcEF) of physiological electric field strength (EFs). This study investigated the effects of doxycycline on the electrotactic responses of U87 cells. After EF stimulation, U87 cells migrated toward the cathode, whereas doxycycline-treated U87 cells exhibited enhanced cell mobility but hindered cathodal migration. We further investigated the expression of the metastasis-correlated proteins matrix metallopeptidase-2 (MMP-2) and MMP-9 in U87 cells. The levels of MMP-2 in the cells were not altered under EF or doxycycline stimulation. In contrast, the EF stimulation greatly enhanced the levels of MMP-9 and then repressed in doxycycline-cotreated cells, accompanied by reduced cathodal migration. Our results demonstrated that an antibiotic at a non-toxic concentration could suppress the enhanced cell migration accelerated by EF of physiological strength. This finding may be applied as an anti-metastatic treatment for cancers.
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Relying on the biological responses and activity of living cells, bioluminescent whole-cell biosensors generate an optical signal in response to the presence of target compounds. The miniaturization of low-light detectors and their integration with microfluidics have allowed the realization of portable devices for sensitive imaging and quantification of these signals. This review thus focuses on bioluminescence-based whole-cell biosensors, integrated with handheld optical detectors, with an emphasis on the use of the low-light imaging capability of modern smartphones. We highlight state-of-the-art miniaturization techniques applied to the incorporation of living cells into diverse hardware platforms, and to the sensitive recording of the biologically generated photons. The integration of sample introduction, host-cell activity maintenance, luminescence acquisition, data processing/presentation, and portability are also discussed.
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Técnicas Biosensibles , Microfluídica , Microfluídica/métodos , Técnicas Biosensibles/métodos , LuminiscenciaRESUMEN
The applications of red-light photobiomodulation (PBM) to enhance neurite growth have been proposed for many years. However, the detailed mechanisms require further studies. In the present work we used a focused red-light spot to illuminate the junction of the longest neurite and the soma of a neuroblastoma cell (N2a), and demonstrated enhanced neurite growth at 620 nm and 760 nm with adequate illumination energy fluences. In contrast, 680 nm light showed no effect on neurite growth. The neurite growth was accompanied with the increase of intracellular reactive oxygen species (ROS). Using Trolox to reduce the ROS level, this red light-induced neurite growth was hindered. Suppressing the activities of cytochrome c oxidase (CCO) by using either a small-molecule inhibitor or siRNA abrogated the red light-induced neurite growth. These results suggest that red light-induced ROS production through the activation of CCO could be beneficial for neurite growth.
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Complejo IV de Transporte de Electrones , Neuritas , Especies Reactivas de Oxígeno/metabolismo , Complejo IV de Transporte de Electrones/metabolismo , Neuritas/fisiología , Luz , Neuronas/metabolismoRESUMEN
COVID-19 has shown that a highly specific and rapid diagnostic system is a necessity. A spectral imaging-based surface plasmon resonance (SPRi) platform with an integrated microfluidic biosensor to detect oligonucleotide sequences has been proposed to be a promising alternative for infectious diseases due to its safe and straightforward use. Approaches to reduce the DNA probe loading onto gold nanoparticles with various types of polyethylene glycol (PEG) were explored. Here, we demonstrated the stability of functionalised gold nanoparticles with unmodified PEG whilst lowering the probe loading density. The system was evaluated by performing the detection of a mimicking COVID-19 target sequence, single point-mutation sequence and fully mismatch sequence. Highly specific binding of the mimicking COVID-19 target sequence was observed and analysed by the spectral imaging SPR approach. Our work has demonstrated the potential of a controlled probe density using unmodified PEG as an especially promising functionalisation strategy in SPR spectral imaging assays.
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COVID-19 , Nanopartículas del Metal , Oro , Humanos , Límite de Detección , SARS-CoV-2 , Resonancia por Plasmón de SuperficieRESUMEN
Metastasis is a frequent complication of cancer and accounts for more than 60% of patients' mortality. Despite technological advancements, treatment options are still limited. Ion channels participate in the regulation of cell adhesion, whilst the regulation of cell adhesion further controls metastasis formation. However, to develop a new ion channel inhibitor targeting metastasis takes tremendous effort and resources; therefore, drug repurposing is an emerging strategy in oncology. In previous studies, we have developed a metal-based nanoslit surface plasmon resonance (SPR) platform to examine the influence of drugs on the cell adhesion process. In this work, we developed a scanner-based cell adhesion kinetic examination (CAKE) system that is capable of monitoring the cell adhesion process by measuring color changes of SPR biosensors. The system's performance was demonstrated by screening the anti-metastasis ability of compounds from a commercial ion-channel inhibitor library. Out of the 274 compounds from the inhibitor library, zinc pyrithione (ZPT) and terfenadine were demonstrated to influence CL1-5 cell adhesion. The cell responses to the two compounds were then compared with those by traditional cell adhesion assays where similar behavior was observed. Further investigation of the two compounds using wound healing and transwell assays was performed and inhibitions of both cell migration and invasion by the two compounds were also observed. The results indicate that ZPT and terfenadine are potential candidates for anti-metastasis drugs. Our work has demonstrated the label-free drug screening ability of our CAKE system for finding potential drugs for cancer treatment.
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Preparaciones Farmacéuticas , Resonancia por Plasmón de Superficie , Adhesión Celular , Línea Celular Tumoral , Movimiento Celular , HumanosRESUMEN
Physiological electric fields (EF) play vital roles in cell migration, differentiation, division, and death. This paper describes a microfluidic cell culture system that was used for a long-term cell differentiation study using microscopy. The microfluidic system consists of the following major components: an optically transparent electrotactic chip, a transparent indium-tin-oxide (ITO) heater, a culture media-filling pump, an electrical power supply, a high-frequency power amplifier, an EF multiplexer, a programmable X-Y-Z motorized stage, and an inverted phase-contrast microscope equipped with a digital camera. The microfluidic system is beneficial in simplifying the overall experimental setup and, in turn, the reagent and sample consumption. This work involves the differentiation of neural stem and progenitor cells (NPCs) induced by direct current (DC) pulse stimulation. In the stem cell maintenance medium, the mouse NPCs (mNPCs) differentiated into neurons, astrocytes, and oligodendrocytes after the DC pulse stimulation. The results suggest that simple DC pulse treatment could control the fate of mNPCs and could be used to develop therapeutic strategies for nervous system disorders. The system can be used for cell culture in multiple channels, for long-term EF stimulation, for cell morphological observation, and for automatic time-lapse image acquisition. This microfluidic system not only shortens the required experimental time, but also increases the accuracy of control on the microenvironment.
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Diferenciación Celular , Animales , Astrocitos/citología , Técnicas de Cultivo de Célula , Electricidad , Dispositivos Laboratorio en un Chip , Ratones , Células-Madre Neurales/citología , Neuronas/citología , Oligodendroglía/citologíaRESUMEN
High-throughput and dynamic measurement for living cell activities can benefit biological research and drug development. A low-cost metallic nanostructure-based surface plasmon resonance (SPR) imaging platform, comprising multiple aluminum nanoslit arrays and a color image device, is proposed for label-free study of cell and drug interactions. The multiple nanoslit sensing arrays were fabricated using the compression-injection molding process. These sensing chips showed a detectable depth of 600 nm and refractive index resolution of â¼5 × 10-5 refractive index unit (RIU) by using a self-referenced two-color analysis. Two examples of kinetic studies of living cells under various doses of drugs are presented. The focal adhesion kinases inhibitor (FAKi 14) and cell interactions show exponential changes of cellular adhesion and time constants for different concentrations of antiadhesion drugs. The anticancer drug (doxorubicin (DOX))-treated cells show slow increases of SPR signals in the first 2 h due to the nucleus swelling. The DOX-treated cells further process plasma membrane disruption and become floating cells and debris in the medium, resulting in rapid drops of the SPR signals.
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Aluminio/química , Adhesión Celular/efectos de los fármacos , Doxorrubicina/farmacología , Ensayos Analíticos de Alto Rendimiento/métodos , Nanoestructuras/química , Antineoplásicos/farmacología , Línea Celular , Supervivencia Celular , Colorimetría , Glicerol/química , Humanos , Resonancia por Plasmón de Superficie , AguaRESUMEN
We present an integral smartphone-based whole-cell biosensor, LumiCellSense (LCS), which incorporates a 16-well biochip with an oxygen permeable coating, harboring bioluminescent Escherichia coli bioreporter cells, a macro lens, a lens barrel, a metal heater tray, and a temperature controller, enclosed in a light-impermeable case. The luminescence emitted by the bioreporter cells in response to the presence of the target chemicals is imaged by the phone's camera, and a dedicated phone-embedded application, LCS_Logger, is employed to calculate photon emission intensity and plot it in real time on the device's screen. An alert is automatically given when light intensity increases above the baseline, indicating the presence of the target. We demonstrate the efficacy of this system by the detection of residues of an antibiotic, ciprofloxacin (CIP), in whole milk, with a detection threshold of 7.2 ng/mL. This value is below the allowed maximum as defined by European Union regulations.
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Antibacterianos/aislamiento & purificación , Técnicas Biosensibles , Ciprofloxacina/aislamiento & purificación , Leche/química , Animales , Antibacterianos/química , Bovinos , Ciprofloxacina/química , Humanos , Luz , Luminiscencia , Teléfono InteligenteRESUMEN
Adenocarcinoma, large cell carcinoma and squamous cell carcinoma are the most commonly diagnosed subtypes of non-small cell lung cancers (NSCLC). Numerous lung cancer cell types have exhibited electrotaxis under direct current electric fields (dcEF). Physiological electric fields (EF) play key roles in cancer cell migration. In this study, we investigated electrotaxis of NSCLC cells, including human large cell lung carcinoma NCI-H460 and human lung squamous cell carcinoma NCI-H520 cells. Non-cancerous MRC-5 lung fibroblasts were included as a control. After dcEF stimulation, NCI-H460 and NCI-H520 cells, which both exhibit epithelial-like morphology, migrated towards the cathode, while MRC-5 cells, which have fibroblast-like morphology, migrated towards the anode. The effect of doxycycline, a common antibiotic, on electrotaxis of MRC-5, NCI-H460 and NCI-H520 cells was examined. Doxycycline enhanced the tested cells' motility but inhibited electrotaxis in the NSCLC cells without inhibiting non-cancerous MRC-5 cells. Based on our finding, further in-vivo studies could be devised to investigate the metastasis inhibition effect of doxycycline in an organism level.
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Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Movimiento Celular/efectos de los fármacos , Doxiciclina/farmacología , Fenómenos Electrofisiológicos , Neoplasias Pulmonares/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Movimiento Celular/fisiología , Doxiciclina/uso terapéutico , Ensayos de Selección de Medicamentos Antitumorales , Fibroblastos/efectos de los fármacos , Humanos , Pulmón/citología , Pulmón/patología , Neoplasias Pulmonares/patología , Pruebas de ToxicidadRESUMEN
A variety of physiological and pathological processes rely on cell adhesion, which is most often tracked by changes in cellular morphology. We previously reported a novel gold nanoslit-based biosensor that is capable of real-time and label-free monitoring of cell morphological changes and cell viability. However, the preparation of gold biosensors is inefficient, complicated and costly. Recently, nanostructure-based aluminum (Al) sensors have been introduced for biosensing applications. The Al-based sensor has a longer decay length and is capable of analyzing large-sized mass such as cells. Here, we developed two types of double-layer Al nanoslit-based plasmonic biosensors, which were nanofabricated and used to evaluate the correlation between metastatic potency and adhesion of lung cancer and melanoma cell lines. Cell adhesion was determined by Fano resonance signals that were induced by binding of the cells to the nanoslit. The peak and dip of the Fano resonance spectrum respectively reflected long- and short-range cellular changes, allowing us to simultaneously detect and distinguish between focal adhesion and cell spreading. Also, the Al nanoslit-based biosensor chips were used to evaluate the inhibitory effects of drugs on cancer cell spreading. We are the first to report the use of double layer Al nanoslit-based biosensors for detection of cell behavior, and such devices may become powerful tools for anti-metastasis drug screening in the future.
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Aluminio/química , Técnicas Biosensibles/instrumentación , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Quinasa 1 de Adhesión Focal/antagonistas & inhibidores , Neoplasias Pulmonares/metabolismo , Melanoma/metabolismo , Algoritmos , Adhesión Celular , Línea Celular Tumoral , Movimiento Celular , Supervivencia Celular , Diseño de Equipo , Humanos , Nanotecnología , Metástasis de la Neoplasia , Resonancia por Plasmón de SuperficieRESUMEN
Mesenchymal stem cell (MSC) is mechanosensitive and the respond to mechanical force is pattern specific. We previously reported that oscillatory shear stress at 0.5⯱â¯4â¯dyne/cm2 guided MSCs polarity vertical to net flow direction before apolaric stage at 30â¯min resulting in phosphorylation of ß-catenin and inhibition of Wnt signaling. This time, we explored laminar shear stress (LS) at 0.5â¯dyne/cm2 polarized MSCs by guiding F-actin orientation parallel to the flow direction before apolarity at 30â¯min accompanied with activation of Wnt signaling. Time-dependent microarray analysis supported cell-cell junctional complex of MSCs was the major mechanosensor on MSCs to respond 0.5â¯dyne/cm2 LS. Three-dimensional immunofluorescence image confirmed LS promoting ß-catenin nuclear localization during 15â¯min to 1â¯h with a peak at 30â¯min. Functional analysis of proteomic study on MSC with 30â¯min LS stimulation indicated that upregulation of ß-catenin downstream proteins related to cardiovascular development, endothelial cell protection and angiogenesis. Conditioned medium from MSCs with 30â¯min LS stimulation improved the viability of human endothelial cells from oxidative damage. In conclusion, 0.5â¯dyne/cm2 LS on MSCs for 30â¯min guides MSCs lack of polarity and promotes ß-catenin nuclear translocation favoring Wnt activation and paracrine cardiovascular support.
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Núcleo Celular/metabolismo , Células Madre Mesenquimatosas/citología , beta Catenina/metabolismo , Actinas/análisis , Actinas/metabolismo , Polaridad Celular , Células Cultivadas , Células Endoteliales de la Vena Umbilical Humana , Humanos , Células Madre Mesenquimatosas/metabolismo , Mapas de Interacción de Proteínas , Estrés Mecánico , Vía de Señalización Wnt , beta Catenina/análisisRESUMEN
The clinical assessment of short-stranded nucleic acid biomarkers such as miRNAs could potentially provide useful information for monitoring disease progression, prompting definitive treatment decisions. In the past decade, advancements in biosensing technology have led to a shift towards rapid, real-time and label-free detection systems; as such, surface plasmon resonance (SPR) biosensor-based technology has become of high interest. Here, we developed an automated multiplex transmissive surface plasmon resonance (t-SPR) platform with the use of a capped gold nanoslit integrated microfluidic surface plasmon resonance (SPR) biosensor. The automated platform was custom designed to allow the analysis of spectral measurements using wavelength shift (dλ), intensity (dI) and novel area change (dA) for surface binding reactions. A simple and compact nanostructure based biosensor was fabricated with multiplex real-time detection capabilities. The sensitivity and specificity of the microfluidic device was demonstrated through the use of functionalised AuNPs for target molecule isolation and signal enhancement in combination with probes on the CG nanoslit surface. Our work allows for the multiplex detection of miRNA at femtomolar concentrations in complex media such as urine.
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Técnicas Biosensibles , MicroARNs/orina , Resonancia por Plasmón de Superficie , Biomarcadores/orina , Oro , Humanos , Dispositivos Laboratorio en un Chip , Nanopartículas del MetalRESUMEN
We present a rapid and simple approach for sensitive detection of antibiotic residues in food samples based on luminescence induction by live bacterial sensor strains integrated into a CCD-based lens-free optical analyzer (LumiSense). Using ciprofloxacin as a model antibiotic, we demonstrate response times of between 20 and 80 min, and detection thresholds of 8 ng/mL for milk, egg white, and chicken essence, and 64 ng/mL for egg yolk. These values are below the minimal allowed values as defined by European Union regulations. Although not intended to replace traditional analytical equipment and regulation-approved methods, LumiSense and similar systems, sample preparation for which involves only simple mixing, dilution, and homogenization, may nevertheless provide a simple means for high-throughput food sample screening. Graphical abstract Detection of bioluminescence from genetically modified bacteria offers a simple and effective way for monitoring an antibiotic, ciprofloxacin, in milk without prior sample preparation.
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Antibacterianos/análisis , Técnicas Biosensibles , Ciprofloxacina/análisis , Residuos de Medicamentos/análisis , Contaminación de Alimentos , Farmacorresistencia Microbiana , Límite de DetecciónRESUMEN
Microfluidic devices are capable of creating a precise and controllable cellular micro-environment of pH, temperature, salt concentration, and other physical or chemical stimuli. They have been commonly used for in vitro cell studies by providing in vivo like surroundings. Especially, how cells response to chemical gradients, electrical fields, and shear stresses has drawn many interests since these phenomena are important in understanding cellular properties and functions. These microfluidic chips can be made of glass substrates, silicon wafers, polydimethylsiloxane (PDMS) polymers, polymethylmethacrylate (PMMA) substrates, or polyethyleneterephthalate (PET) substrates. Out of these materials, PMMA substrates are cheap and can be easily processed using laser ablation and writing. Although a few microfluidic devices have been designed and fabricated for generating multiple, coexisting chemical and electrical stimuli, none of them was considered efficient enough in reducing experimental repeats, particular for screening purposes. In this report, we describe our design and fabrication of two PMMA-based microfluidic chips for investigating cellular responses, in the production of reactive oxygen species and the migration, under single or coexisting chemical/electrical/shear stress stimuli. The first chip generates five relative concentrations of 0, 1/8, 1/2, 7/8, and 1 in the culture regions, together with a shear stress gradient produced inside each of these areas. The second chip generates the same relative concentrations, but with five different electric field strengths created within each culture area. These devices not only provide cells with a precise, controllable micro-environment but also greatly increase the experimental throughput.
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Dispositivos Laboratorio en un Chip , Técnicas Analíticas Microfluídicas , Electricidad , Humanos , Microfluídica , Estrés MecánicoRESUMEN
We report the differentiation of neural stem and progenitor cells solely induced by direct current (DC) pulses stimulation. Neural stem and progenitor cells in the adult mammalian brain are promising candidates for the development of therapeutic neuroregeneration strategies. The differentiation of neural stem and progenitor cells depends on various in vivo environmental factors, such as nerve growth factor and endogenous EF. In this study, we demonstrated that the morphologic and phenotypic changes of mouse neural stem and progenitor cells (mNPCs) could be induced solely by exposure to square-wave DC pulses (magnitude 300 mV/mm at frequency of 100-Hz). The DC pulse stimulation was conducted for 48 h, and the morphologic changes of mNPCs were monitored continuously. The length of primary processes and the amount of branching significantly increased after stimulation by DC pulses for 48 h. After DC pulse treatment, the mNPCs differentiated into neurons, astrocytes, and oligodendrocytes simultaneously in stem cell maintenance medium. Our results suggest that simple DC pulse treatment could control the fate of NPCs. With further studies, DC pulses may be applied to manipulate NPC differentiation and may be used for the development of therapeutic strategies that employ NPCs to treat nervous system disorders.
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Técnicas de Reprogramación Celular/métodos , Corteza Cerebral/citología , Electricidad , Células-Madre Neurales/citología , Neurogénesis , Animales , Células Cultivadas , Técnicas de Reprogramación Celular/instrumentación , Femenino , Ratones , Ratones Endogámicos ICR , Neuronas/citología , Oligodendroglía/citologíaRESUMEN
Applying uniform electric field (EF) in vitro in the physiological range has been achieved in rectangular shaped microchannels. However, in a circular-shaped device, it is difficult to create uniform EF from two electric potentials due to different electrical resistances originated from the length difference between the diameter of the circle and the length of any parallel chord of the bottom circular chamber where cells are cultured. To address this challenge, we develop a three-dimensional (3D) computer-aided designed (CAD) polymeric insert to create uniform EF in circular shaped multi-well culture plates. A uniform EF with a coefficient of variation (CV) of 1.2% in the 6-well plate can be generated with an effective stimulation area percentage of 69.5%. In particular, NIH/3T3 mouse embryonic fibroblast cells are used to validate the performance of the 3D designed Poly(methyl methacrylate) (PMMA) inserts in a circular-shaped 6-well plate. The CAD based inserts can be easily scaled up (i.e., 100 mm dishes) to further increase effective stimulation area percentages, and also be implemented in commercially available cultureware for a wide variety of EF-related research such as EF-cell interaction and tissue regeneration studies.
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Cell migration is an essential process involved in the development and maintenance of multicellular organisms. Electric fields (EFs) are one of the many physical and chemical factors known to affect cell migration, a phenomenon termed electrotaxis or galvanotaxis. In this paper, a microfluidics chip was developed to study the migration of cells under different electrical and chemical stimuli. This chip is capable of providing four different strengths of EFs in combination with two different chemicals via one simple set of agar salt bridges and Ag/AgCl electrodes. NIH 3T3 fibroblasts were seeded inside this chip to study their migration and reactive oxygen species (ROS) production in response to different EF strengths and the presence of ß-lapachone. We found that both the EF and ß-lapachone level increased the cell migration rate and the production of ROS in an EF-strength-dependent manner. A strong linear correlation between the cell migration rate and the amount of intracellular ROS suggests that ROS are an intermediate product by which EF and ß-lapachone enhance cell migration. Moreover, an anti-oxidant, α-tocopherol, was found to quench the production of ROS, resulting in a decrease in the migration rate.
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Organs are composed of heterotypic cells with patterned architecture that enables intercellular interaction to perform specific functions. In tissue engineering, the ability to pattern heterotypic cells into desired arrangement will allow us to model complex tissues in vitro and to create tissue equivalents for regeneration. This study was aimed at developing a method for fast heterotypic cell patterning with controllable topological manipulation on a glass chip. We found that poly(vinyl alcohol)-coated glass showed a biphasic change in adhesivity to cells in vitro: low adhesivity in the first 24 h and higher adhesivity at later hours due to increased serum protein adsorption. Combining programmable CO2 laser ablation to remove poly(vinyl alcohol) and glass, we were able to create arrays of adhesive microwells of adjustable patterns. We tested whether controllable patterns of epithelial-mesenchymal interaction could be created. When skin dermal papilla cells and fibroblasts were seeded respectively 24 h apart, we were able to pattern these two cells into aggregates of dermal papilla cells in arrays of microwells in a background of fibroblasts sheet. Seeded later, keratinocytes attached to these mesenchymal cells. Keratinocytes contacting dermal papilla cells started to differentiate toward a hair follicle fate, demonstrating patternable epithelial-mesenchymal interaction. This method allows fast adjustable heterotypic cell patterning and surface topology control and can be applied to the investigation of heterotypic cellular interaction and creation of tissue equivalent in vitro.
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Vidrio/química , Láseres de Gas , Cloruro de Polivinilo/química , Adsorción , Animales , Proteínas Sanguíneas/química , Proteínas Sanguíneas/metabolismo , Adhesión Celular/efectos de los fármacos , Células Cultivadas , Materiales Biocompatibles Revestidos/química , Materiales Biocompatibles Revestidos/farmacología , Fibroblastos/citología , Fibroblastos/metabolismo , Folículo Piloso/citología , Folículo Piloso/metabolismo , Queratinocitos/citología , Queratinocitos/metabolismo , Microscopía Confocal , Ratas , Ratas Wistar , Propiedades de Superficie/efectos de la radiaciónRESUMEN
In this work, electrokinetic acceleration of DNA hybridization was investigated by different combinations of frequencies and amplitudes of actuating electric signals. Because the frequencies from low to high can induce different kinds of electrokinetic forces, i.e., electroosmotic to electrothermal forces, this work provides an in-depth investigation of electrokinetic enhanced hybridization. Concentric circular Cr/Au microelectrodes of 350 µm in diameter were fabricated on a glass substrate and probe DNA was immobilized on the electrode surface. Target DNA labeled with fluorescent dyes suspending in solution was then applied to the electrode. Different electrokinetic forces were induced by the application of different electric signals to the circular microelectrodes. Local microfluidic vortexes were generated to increase the collision efficiency between the target DNA suspending in solution and probe DNA immobilized on the electrode surface. DNA hybridization on the electrode surface could be accelerated by the electrokinetic forces. The level of hybridization was represented by the fluorescent signal intensity ratio. Results revealed that such 5-min dynamic hybridization increased 4.5 fold of signal intensity ratio as compared to a 1-h static hybridization. Moreover, dynamic hybridization was found to have better differentiation ability between specific and non-specific target DNA. This study provides a strategy to accelerate DNA hybridization in microsystems.