Costantini, IrenePavone, Francesco SaverioPesce, LucaLaurino, Annunziatina...
7页
查看更多>>摘要：Cover-all mapping of the distribution of neurons in the human brain would have a significant impact on the deep understanding of brain function. Therefore, complete knowledge of the structural organization of different human brain regions at the cellular level would allow understanding their role in the functions of specific neural networks. Recent advances in tissue clearing techniques have allowed important advances towards this goal. These methods use specific chemicals capable of dissolving lipids, making the tissue completely transparent by homogenizing the refractive index. However, labeling and clearing human brain samples is still challenging. Here, we present an approach to perform the cellular mapping of the human cerebral cortex coupling immunostaining with SWITCH/TDE clearing and confocal microscopy. A specific evaluation of the contributions of the autofluorescence signals generated from the tissue fixation is provided as well as an assessment of lipofuscin pigments interference. Our evaluation demonstrates the possibility of obtaining an efficient clearing and labeling process of parts of adult human brain slices, making it an excellent method for morphological classification and antibody validation of neuronal and non-neuronal markers. (C) 2021 The Authors. Published by Elsevier Ltd.
查看更多>>摘要：The optical clearing of the cardiac tissue has always been a challenging goal to obtain successful threedimensional reconstructions of entire hearts. Typically, the developed protocols are targeted at the clearing of the brain; cardiac tissue requires proper arrangements to the original protocols, which are usually tough and time-consuming to figure out. Here, we present the application of three different clearing methodologies on mouse hearts: uDISCO, CLARITY, and SHIELD. For each approach, we describe the required optimizations that we have developed to improve the outcome; in particular, we focus on comparing the features of the tissue after the application of each methodology, especially in terms of tissue preservation, transparency, and staining. We found that the uDISCO protocol induces strong fiber delamination of the cardiac tissue, thus reducing the reliability of structural analyses. The CLARITY protocol confers a high level of transparency to the heart and allows deep penetration of the fluorescent dyes; however, it requires long times for the clearing and the tissue loses its robustness. The SHIELD methodology, indeed, is very promising for tissue maintenance since it preserves its consistency and provides ideal transparency, but further approaches are needed to obtain homogeneous staining of the whole heart. Since the CLARITY procedure, despite the disadvantages in terms of tissue preservation and timings, is actually the most suitable approach to image labeled samples in depth, we optimized and performed the methodology also on human cardiac tissue from control hearts and hearts with hypertrophic cardiomyopathy. (c) 2021 Elsevier Ltd. All rights reserved.
Ashton, Jesse L.Trew, Mark L.Sands, Gregory B.Baddeley, David...
15页
查看更多>>摘要：Recent developments in clearing and microscopy enable 3D imaging with cellular resolution up to the whole organ level. These methods have been used extensively in neurobiology, but their uptake in other fields has been much more limited. Application of this approach to the human heart and effective use of the data acquired present challenges of scale and complexity. Four interlinked issues need to be addressed: 1) efficient clearing and labelling of heart tissue, 2) fast microscopic imaging of human-scale samples, 3) handling and processing of multi-terabyte 3D images, and 4) extraction of structural infor-mation in computationally tractable structure-based models of cardiac function. Preliminary studies show that each of these requirements can be achieved with the appropriate application and develop-ment of existing technologies. (c) 2021 Elsevier Ltd. All rights reserved.
查看更多>>摘要：Light Sheet Fluorescence Microscopy (LSFM) has revolutionized how optical imaging of biological specimens can be performed as this technique allows to produce 3D fluorescence images of entire samples with a high spatiotemporal resolution. In this manuscript, we aim to provide readers with an overview of the field of LSFM on ex vivo samples. Recent advances in LSFM architectures have made the technique widely accessible and have improved its acquisition speed and resolution, among other features. These developments are strongly supported by quantitative analysis of the huge image volumes produced thanks to the boost in computational capacities, the advent of Deep Learning techniques, and by the combination of LSFM with other imaging modalities. Namely, LSFM allows for the characterization of biological structures, disease manifestations and drug effectivity studies. This information can ultimately serve to develop novel diagnostic procedures, treatments and even to model the organs physiology in healthy and pathological conditions. (c) 2021 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Silvestri, LudovicoSancataldo, GiuseppePavone, Francesco SaverioRicci, Pietro...
14页
查看更多>>摘要：In recent years, light-sheet fluorescence microscopy (LSFM) has found a broad application for imaging of diverse biological samples, ranging from sub-cellular structures to whole animals, both in-vivo and exvivo, owing to its many advantages relative to point-scanning methods. By providing the selective illumination of sample single planes, LSFM achieves an intrinsic optical sectioning and direct 2D image acquisition, with low out-of-focus fluorescence background, sample photo-damage and photo-bleaching. On the other hand, such an illumination scheme is prone to light absorption or scattering effects, which lead to uneven illumination and striping artifacts in the images, oriented along the light sheet propagation direction. Several methods have been developed to address this issue, ranging from fully optical solutions to entirely digital post-processing approaches. In this work, we present them, outlining their advantages, performance and limitations. (c) 2021 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
查看更多>>摘要：Compressed sensing (CS) is a signal processing approach that solves ill-posed inverse problems, from under-sampled data with respect to the Nyquist criterium. CS exploits sparsity constraints based on the knowledge of prior information, relative to the structure of the object in the spatial or other domains. It is commonly used in image and video compression as well as in scientific and medical applications, including computed tomography and magnetic resonance imaging. In the field of fluorescence microscopy, it has been demonstrated to be valuable for fast and high-resolution imaging, from singlemolecule localization, super-resolution to light-sheet microscopy. Furthermore, CS has found remarkable applications in the field of mesoscopic imaging, facilitating the study of small animals' organs and entire organisms. This review article illustrates the working principles of CS, its implementations in optical imaging and discusses several relevant uses of CS in the field of fluorescence imaging from superresolution microscopy to mesoscopy. (c) 2021 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
查看更多>>摘要：Over the last ten years, developments in whole-brain microscopy now allow for high-resolution imaging of intact brains of small animals such as mice. These complex images contain a wealth of information, but many neuroscience laboratories do not have all of the computational knowledge and tools needed to process these data. We review recent open source tools for registration of images to atlases, and the segmentation, visualisation and analysis of brain regions and labelled structures such as neurons. Since the field lacks fully integrated analysis pipelines for all types of whole-brain microscopy analysis, we propose a pathway for tool developers to work together to meet this challenge.
Akram, Masood A.Ljungquist, BengtAscoli, Giorgio A.
9页
查看更多>>摘要：Advancements in neuroscience research have led to steadily accelerating data production and sharing. The online community repository of neural reconstructions NeuroMorpho.Org grew from fewer than 1000 digitally traced neurons in 2006 to more than 140,000 cells today, including glia that now constitute 10.1% of the content. Every reconstruction consists of a detailed 3D representation of branch geometry and connectivity in a standardized format, from which a collection of morphometric features is extracted and stored. Moreover, each entry in the database is accompanied by rich metadata annotation describing the animal subject, anatomy, and experimental details. The rapid expansion of this resource in the past decade was accompanied by a parallel rise in the complexity of the available information, creating both opportunities and challenges for knowledge mining. Here, we introduce a new summary reporting functionality, allowing NeuroMorpho.Org users to efficiently download digests of metadata and morphometrics from multiple groups of similar cells for further analysis. We demonstrate the capabilities of the tool for both glia and neurons and present an illustrative statistical analysis of the resulting data.
NSTL
Elsevier