RESUMEN
Various implementations of mesoscopes provide optical access for calcium imaging across multi-millimeter fields of view in the mammalian brain; however, capturing the activity of the neuronal population within such fields of view near-simultaneously and in a volumetric fashion has remained challenging as approaches for imaging scattering brain tissues typically are based on sequential acquisition. Here we present a modular, mesoscale light-field (MesoLF) imaging hardware and software solution that allows recording from thousands of neurons within volumes of â 4 × 0.2 mm, located at up to 350 µm depth in the mouse cortex, at 18 volumes per second and an effective voxel rate of ~40 megavoxels per second. Using our optical design and computational approach we show recording of ~10,000 neurons across multiple cortical areas in mice using workstation-grade computing resources.
Asunto(s)
Encéfalo , Neuronas , Ratones , Animales , Neuronas/fisiología , Encéfalo/fisiología , Programas Informáticos , Neuroimagen , MamíferosRESUMEN
Fluorescence microscopy has become an indispensable tool for revealing the dynamic regulation of cells and organelles. However, stochastic noise inherently restricts optical interrogation quality and exacerbates observation fidelity when balancing the joint demands of high frame rate, long-term recording and low phototoxicity. Here we propose DeepSeMi, a self-supervised-learning-based denoising framework capable of increasing signal-to-noise ratio by over 12 dB across various conditions. With the introduction of newly designed eccentric blind-spot convolution filters, DeepSeMi effectively denoises images with no loss of spatiotemporal resolution. In combination with confocal microscopy, DeepSeMi allows for recording organelle interactions in four colors at high frame rates across tens of thousands of frames, monitoring migrasomes and retractosomes over a half day, and imaging ultra-phototoxicity-sensitive Dictyostelium cells over thousands of frames. Through comprehensive validations across various samples and instruments, we prove DeepSeMi to be a versatile and biocompatible tool for breaking the shot-noise limit.
Asunto(s)
Dictyostelium , Aumento de la Imagen , Microscopía Confocal/métodos , Relación Señal-Ruido , Microscopía Fluorescente , Procesamiento de Imagen Asistido por Computador/métodosRESUMEN
Widefield microscopy can provide optical access to multi-millimeter fields of view and thousands of neurons in mammalian brains at video rate. However, tissue scattering and background contamination results in signal deterioration, making the extraction of neuronal activity challenging, laborious and time consuming. Here we present our deep-learning-based widefield neuron finder (DeepWonder), which is trained by simulated functional recordings and effectively works on experimental data to achieve high-fidelity neuronal extraction. Equipped with systematic background contribution priors, DeepWonder conducts neuronal inference with an order-of-magnitude-faster speed and improved accuracy compared with alternative approaches. DeepWonder removes background contaminations and is computationally efficient. Specifically, DeepWonder accomplishes 50-fold signal-to-background ratio enhancement when processing terabytes-scale cortex-wide functional recordings, with over 14,000 neurons extracted in 17 h.
Asunto(s)
Encéfalo , Calcio , Animales , Encéfalo/fisiología , Microscopía , Corteza Cerebral , Neuronas/fisiología , MamíferosRESUMEN
Calcium imaging has transformed neuroscience research by providing a methodology for monitoring the activity of neural circuits with single-cell resolution. However, calcium imaging is inherently susceptible to detection noise, especially when imaging with high frame rate or under low excitation dosage. Here we developed DeepCAD, a self-supervised deep-learning method for spatiotemporal enhancement of calcium imaging data that does not require any high signal-to-noise ratio (SNR) observations. DeepCAD suppresses detection noise and improves the SNR more than tenfold, which reinforces the accuracy of neuron extraction and spike inference and facilitates the functional analysis of neural circuits.
Asunto(s)
Potenciales de Acción , Algoritmos , Calcio/metabolismo , Diagnóstico por Imagen/métodos , Procesamiento de Imagen Asistido por Computador/métodos , Neuronas/fisiología , Relación Señal-Ruido , Animales , Femenino , Masculino , Ratones , Ratones Transgénicos , Neuronas/citologíaRESUMEN
Wavefront sensing technique is essential in deep tissue imaging, which guides spatial light modulator to compensate wavefront distortion for better imaging quality. Recently, convolutional neural network (CNN) based sensorless wavefront sensing methods have achieved remarkable speed advantages via single-shot measurement methodology. However, the low efficiency of convolutional filters dealing with circular point-spread-function (PSF) features makes them less accurate. In this paper, we propose a conformal convolutional neural network (CCNN) that boosts the performance by pre-processing circular features into rectangular ones through conformal mapping. The proposed conformal mapping reduces the number of convolutional filters that need to describe a circular feature, thus enables the neural network to recognize PSF features more efficiently. We demonstrate our CCNN could improve the wavefront sensing accuracy over 15% compared to a traditional CNN through simulations and validate the accuracy improvement in experiments. The improved performances make the proposed method promising in high-speed deep tissue imaging.
RESUMEN
BACKGROUND: Hydrogen sulfide poisoning can cause severe myocardial injury, but the damage is subtle and can be easily misdiagnosed. This report presents the dynamic observation of myocardial injury associated with hydrogen sulfide poisoning. CASE REPORT: Two young men presented with symptoms of "lightning-like" death immediately after entering a tank. They were found and rescued in 20 min at a time when they were already in a coma. Case 1 had no spontaneous breathing and pulse, while case 2 had spontaneous breathing and a pulse. Upon transfer to a local hospital, case 1 received continuous cardiopulmonary resuscitation which led to the recovery of his heart rate 3 min after arriving at the hospital. However, the patient remained in a Glasgow coma scale of 3. He was transferred to our hospital where he, unfortunately, died on the seventh day due to multiple organ failure. Case 2 was also transferred to the intensive care unit in our hospital and on the fourth day of hospitalization, the patient presented ST-segment elevation and dynamic changes in markers of myocardial injury. Changes in electrocardiogram and markers of myocardial injury were monitored and examination improved through conventional echocardiography, coronary artery CT, radionuclide myocardial perfusion imaging, and two-dimensional speckle tracking imaging strain. The treatment gradually improved the patient's myocardial injury and was discharged from the hospital. CONCLUSION: Hydrogen sulfide poisoning can cause damage to myocardial function and the damage can be more insidious in nature and with a delayed onset. Recovery from myocardial damage can be very slow.
Asunto(s)
Lesiones Cardíacas/inducido químicamente , Lesiones Cardíacas/patología , Lesiones Cardíacas/terapia , Sulfuro de Hidrógeno/envenenamiento , Miocardio/patología , Traumatismos Ocupacionales/patología , Adulto , Reanimación Cardiopulmonar/métodos , Resultado Fatal , Humanos , Masculino , Resultado del TratamientoRESUMEN
To improve imaging speed, multifocal excitation is widely adopted as a parallel strategy in laser-scanning microscopy. Specifically, axial multifocal microscopy is popular in neuroscience as it enables functional imaging of neurons in multiple depths simultaneously. However, previous phase searching algorithms for axial multi-foci generation generally generate foci of uniform intensities, which cannot compensate the scattering-induced power loss in deep tissue and causes inhomogeneous excitation. Here, we propose a novel adaptive optimization-based phase-searching method (AdaPS) to generate axial multi-foci with arbitrary intensity modulations for scattering-induced loss compensation. By adopting Adaptive Moment Estimation (Adam) as the searching algorithm, our method could escape from unsatisfactory local minima and stably converge to the optimal phase pattern with errors at least an order of magnitude lower. We validate AdaPS through both numerical simulations and experiments and demonstrate that AdaPS could provide uniform multi-depth imaging in scattering phantom and enable high-fidelity multi-depth recordings of neural network dynamics in mouse brain in vivo.
RESUMEN
Compared to point-scanning multiphoton microscopy, line-scanning temporal focusing microscopy (LTFM) is competitive in high imaging speed while maintaining tight axial confinement. However, considering its wide-field detection mode, LTFM suffers from shallow penetration depth as a result of the crosstalk induced by tissue scattering. In contrast to the spatial filtering based on confocal slit detection, here we propose the extended detection LTFM (ED-LTFM), the first wide-field two-photon imaging technique to extract signals from scattered photons and thus effectively extend the imaging depth. By recording a succession of line-shape excited signals in 2D and reconstructing signals under Hessian regularization, we can push the depth limitation of wide-field imaging in scattering tissues. We validate the concept with numerical simulations, and demonstrate the performance of enhanced imaging depth in in vivo imaging of mouse brains.
RESUMEN
While simultaneous phase-contrast and two-photon fluorescence imaging in microscopy can bring abundant biomedical information, it is difficult to retrieve phase information from conventional two-photon microscopes. To realize low-cost, in situ phase-contrast and two-photon fluorescence imaging, we propose Schlieren two-photon microscopy, a method that implements phase-contrast imaging on two-photon microscopes. This method involves spatially modulated fluorescence plates, which are made of two-photon fluorescence dyes or upconversion nanoparticles. We demonstrate that the fluorescence intensity fluctuation reflects the phase gradients of the specimen via theoretical analysis, simulations, and experiments. The proposed method is fully compatible with commercial two-photon microscopes, thus enabling widespread applications in live tissue imaging.
Asunto(s)
Células HeLa/citología , Microscopía de Fluorescencia por Excitación Multifotónica/instrumentación , Microscopía de Contraste de Fase/métodos , Colorantes Fluorescentes , HumanosRESUMEN
This publisher's note amends the page range in Appl. Opt.58, A26 (2019)APOPAI0003-693510.1364/AO.58.000A26.
RESUMEN
Compared with two-photon point-scanning microscopy, line-scanning temporal focusing microscopy breaks the limitation on imaging rate and maintains the axial resolution, which makes it promising for various biomedical studies. However, for deep tissue imaging, it suffers from reduced axial resolution and increased background noise due to sample induced wavefront distortion. Here, we propose a spatio-spectral focal modulation technique to enhance axial resolution and background rejection by simply subtracting an aberrated image, which is induced by a spatial light modulator, from an unaberrated image. The proposed technique could improve the axial resolution by a factor of 1.3 in our implementation, verified by both simulations and experiments. Besides, we show that compared with spatial modulation alone, spatio-spectral modulation induces less peak intensity loss caused by image subtraction. We further demonstrate the performance of our technique on the enhanced axial resolution and background rejection by deep imaging of cleared mouse brains and in vivo imaging of living mouse brains.
RESUMEN
We demonstrate a single-pixel imaging (SPI) method that can achieve pixel resolution beyond the physical limitation of the spatial light modulator (SLM), by adopting sinusoidal amplitude modulation and frequency filtering. Through light field analysis, we observe that the induced intensity with a squared value of the amplitude contains higher frequency components. By filtering out the zero frequency of the sinusoidal amplitude in the Fourier domain, we can separate out the higher frequency components, which enables SPI with higher resolving ability and thus beyond the limitation of the SLM. Further, to address the speed issue in grayscale spatial light modulation, we propose a fast implementation scheme with tens-of-kilohertz refresh rate. Specifically, we use a digital micromirror device (DMD) working at the full frame rate to conduct binarized sinusoidal patterning in the spatial domain and pinhole filtering eliminating the binarization error in the Fourier domain. For experimental validation, we build a single-pixel microscope to retrieve 1200 × 1200-pixel images via a sub-megapixel DMD, and the setup achieves comparable performance to array sensor microscopy and provides additional sectioning ability.
RESUMEN
Multimodal microscopes either use multiple cameras or a single camera to multiplex different modes spatially. The former needs expertise demanding alignment and the latter suffers from limited spatial resolution. Here, we report an alignment-free full-resolution simultaneous fluorescence and phase imaging approach using single-pixel detectors. By combining reference-free interferometry with single-pixel imaging scheme, we employ structured illumination to encode the phase and fluorescence of the sample into two single-pixel detection arms, and then conduct reconstruction computationally from the illumination patterns and recorded correlated measurements. The recovered fluorescence and phase images are inherently aligned thanks to single-pixel imaging scheme. To validate the proposed method, we built a proof-of-concept setup for first imaging the phase of an etched glass with given etching depth and then imaging the phase and fluorescence of the quantum dot sample. This method holds great potential for multispectral fluorescence microscopy with additional single-pixel detectors or a spectrometer. Besides, this cost-efficient multimodal system might find broad applications in biomedical science and material science.
RESUMEN
Hyperspectral imaging is an important tool having been applied in various fields, but still limited in observation of dynamic scenes. In this paper, we propose a snapshot hyperspectral imaging technique which exploits both spectral and spatial sparsity of natural scenes. Under the computational imaging scheme, we conduct spectral dimension reduction and spatial frequency truncation to the hyperspectral data cube and snapshot it in a low cost manner. Specifically, we modulate the spectral variations by several broadband spectral filters, and then map these modulated images into different regions in the Fourier domain. The encoded image compressed in both spectral and spatial are finally collected by a monochrome detector. Correspondingly, the reconstruction is essentially a Fourier domain extraction and spectral dimensional back projection with low computational load. This Fourier-spectral multiplexing in a 2D sensor simplifies both the encoding and decoding process, and makes hyperspectral data captured in a low cost manner. We demonstrate the high performance of our method by quantitative evaluation on simulation data and build a prototype system experimentally for further validation.
RESUMEN
AIM: To observe the effect of human umbilical cord mesenchymal stem cells (hUCMSCs) secretions on the relevant factors in mouse retinal astrocytes, and to investigate the effect of hUCMSCs on the expression of vascular endothelial growth factor-A (VEGF-A) and to observe the therapeutic effect on the mouse model of retinopathy of prematurity (ROP). METHODS: Cultured hUCMSCs and extracted exosomes from them and then retinal astrocytes were divided into control group and hypoxia group. MTT assay, flow cytometry, reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were used to detect related indicators. Possible mechanisms by which hUCMSCs exosomes affect VEGF-A expression in hypoxia-induced mouse retinal astrocytes were explored. At last, the efficacy of exosomes of UCMSCs in a mouse ROP model was explored. Graphpad6 was used to comprehensively process data information. RESULTS: The secretion was successfully extracted from the culture supernatant of hUCMSCs by gradient ultracentrifugation. Reactive oxygen species (ROS) and hypoxia inducible factor-1α (HIF-1α) of mice retinal astrocytes under different hypoxia time and the expression level of VEGF-A protein and VEGF-A mRNA increased, and the ROP cell model was established after 6h of hypoxia. The secretions of medium and high concentrations of hUCMSCs can reduce ROS and HIF-1α, the expression levels of VEGF-A protein and VEGF-A mRNA are statistically significant and concentration dependent. Compared with the ROP cell model group, the expression of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signal pathway related factors in the hUCMSCs exocrine group is significantly decreased. The intravitreal injection of the secretions of medium and high concentrations of hUCMSCs can reduce VEGF-A and HIF-1α in ROP model tissues. HE staining shows that the number of retinal neovascularization in ROP mice decreases with the increase of the dose of hUCMSCs secretion. CONCLUSION: In a hypoxia induced mouse retinal astrocyte model, hUCMSCs exosomes are found to effectively reduce the expression of HIF-1α and VEGF-A, which are positively correlated with the concentration of hUCMSCs exosomes. HUCMSCs exosomes can effectively reduce the number of retinal neovascularization and the expression of HIF-1α and VEGF-A proteins in ROP mice, and are positively correlated with drug dosage. Besides, they can reduce the related factors on the PI3K/AKT/mTOR signaling pathway.
RESUMEN
Low bioavailability of quercetin (Que) reduces its preclinical and clinical benefits. In order to improve Que bioavailability, a novel whey protein isolate (WPI)-zein nanogel was prepared by pH-driven self-assembly and heat-induced gelatinization. The results showed that hydrochloric acid can be substituted by both acetic acid and citric acid during the pH-driven process. After encapsulation, the bioavailability of Que in nanogels (composed of 70 % WPI) induced by different acidifiers increased to 19.89 % (citric acid), 21.65 % (hydrochloric acid) and 24.34 % (acetic acid), respectively. Comparatively, nanogels induced by acetic acid showed higher stability (pH and storage stability), re-dispersibility (75.62 %), Que bioavailability (24.34 %), and antioxidant capacity (36.78 % for DPPH scavenging rates). s improved performance of nanogels. In mechanism, acetic acid significantly balanced different intermolecular forces by weakening "acid-induced denaturation" effect. Moreover, the faster binding of Que and protein as well as higher protein molecular flexibility and randomness (higher ratio of random coil) was also observed in nanogels induced by acetic acid. All of these changes contributed to improve nanogels performances. Overall, WPI-zein nanogels induced by acetic acid might be a safe, efficiency and stable delivery system to improve the bioavailability of hydrophobic active ingredients.
Asunto(s)
Antioxidantes , Disponibilidad Biológica , Nanogeles , Quercetina , Proteína de Suero de Leche , Zeína , Quercetina/química , Quercetina/farmacología , Proteína de Suero de Leche/química , Zeína/química , Antioxidantes/química , Antioxidantes/farmacología , Nanogeles/química , Concentración de Iones de Hidrógeno , Ácido Acético/química , Polietileneimina/química , Polietilenglicoles/química , Estabilidad de Medicamentos , Portadores de Fármacos/químicaRESUMEN
High-performance flexible piezoresistive sensors are becoming increasingly essential in various novel applications such as health monitoring, soft robotics, and human-computer interaction. The evolution of the interfacial contact morphology determines the sensing properties of piezoresistive devices. The introduction of microstructures enriches the interfacial contact morphology and effectively boosts the sensitivity; however, the limited compressibility of conventional microstructures leads to rapid saturation of the sensitivity in the low-pressure range, which hinders their application. Herein, we present a flexible piezoresistive sensor featuring a two-stage micropyramid array structure, which effectively enhances the sensitivity while widening the sensing range. Owing to the synergistic enhancement effect resulting from the sequential contact of micropyramids of various heights, the devices demonstrate remarkable performance, including boosting sensitivity (30.8 kPa-1) over a wide sensing range (up to 200 kPa), a fast response/recovery time (75/50 ms), and an ultralong durability of 15,000 loading-unloading cycles. As a proof of concept, the sensor is applied to detect human physiological and motion signals, further demonstrating a real-time spatial pressure distribution sensing system and a game control system, showing great potential for applications in health monitoring and human-computer interaction.
Asunto(s)
Computadores , Robótica , Humanos , Programas Informáticos , Movimiento (Física) , SensaciónRESUMEN
To improve the techno-functional properties of rapeseed protein (RP), this work tried to regulate the molecular structure of RP via inducing the co-assembly of RP with zein and whey protein (WP). The results showed that WP and zein mainly regulate the folding process of RP through hydrophobic and disulfide bonds, thereby altering the structural conformation and forming stable complex RP (CRP). WP addition not only increased the number of surface charges and hydrophilicity of proteins, but also decreased their sizes, improved the water solubility, as well as the availability of active groups. These changes significantly increased the foaming capacity (from 60 % to 147 %) and in vitro gastric digestion rate (from 10 % to 60 %) of CRP. Besides, WP also contributed to the formation of gels and the regulation of their textural profiles. Comparatively, zein improved the hydrophobicity of CRP and balanced degree of intermolecular forces, which effectively increased the emulsifying activity index of CRP from 22 m2/g to 90 m2/g. Zein decreased the hardness, springiness and water-holding capacity of gel, but increased its gumminess and chewiness. Overall, both WP and zein effectively changed the structural conformation of RP, and improved its techno-functional properties, which provides an effective strategy to modify protein.
Asunto(s)
Brassica rapa , Interacciones Hidrofóbicas e Hidrofílicas , Proteínas de Plantas , Solubilidad , Proteína de Suero de Leche , Zeína , Proteínas de Plantas/química , Zeína/química , Brassica rapa/química , Proteína de Suero de Leche/química , Agua/químicaRESUMEN
Our previous experiments found that rapeseed protein (RP) has applicability in low-moisture textured proteins. The amount of RP added is limited to <20 %, but the addition of 20 % RP still brings some negative effects. Therefore, in order to improve the quality of 20%RP textured protein, this experiment added different proportions of sodium tripolyphosphate (STPP) to improve the quality of the product, and studied the physical-chemical properties and molecular structure changes of the product to explore the possible modification mechanism. The STPP not only improved the expansion characteristics of extrudates, but also increased the brightness of the extrudates, the rehydration rate. In addition, STPP increased the specific mechanical energy during extrusion, decreased the material mass flow rate. Furthermore, STPP decreased the starch digestibility, increased the content of slow-digesting starch and resistant starch. STPP increased the degree of denaturation of extrudate proteins, the proportion of ß-sheets in the secondary structure of proteins, as well as the intermolecular hydrogen bonding interactions. The gelatinization degradation degree of starch molecules also decreased with the addition of STPP. STPP also increased the protein-starch interactions and enhanced the thermal stability of the extrudate. All these indicate that STPP can improve the physical-chemical properties of extrudate.