scholarly journals Zebrafish Vascular Quantification (ZVQ): a tool for quantification of three-dimensional zebrafish cerebrovascular architecture by automated image analysis

Development ◽  
2022 ◽  
Author(s):  
E. C. Kugler ◽  
J. Frost ◽  
V. Silva ◽  
K. Plant ◽  
K. Chhabria ◽  
...  

Zebrafish transgenic lines and light sheet fluorescence microscopy allow in-depth insights into three-dimensional vascular development in vivo. However, quantification of the zebrafish cerebral vasculature in 3D remains highly challenging. Here, we describe and test an image analysis workflow for 3D quantification of the total or regional zebrafish brain vasculature, called zebrafish vasculature quantification “ZVQ”. It provides the first landmark- or object-based vascular inter-sample registration of the zebrafish cerebral vasculature, producing Population Average Maps allowing rapid assessment of intra- and inter-group vascular anatomy. ZVQ also extracts a range of quantitative vascular parameters from a user-specified Region of Interest including volume, surface area, density, branching points, length, radius, and complexity. Application of ZVQ to thirteen experimental conditions, including embryonic development, pharmacological manipulations and morpholino induced gene knockdown, shows ZVQ is robust, allows extraction of biologically relevant information and quantification of vascular alteration, and can provide novel insights into vascular biology. To allow dissemination, the code for quantification, a graphical user interface, and workflow documentation are provided. Together, ZVQ provides the first open-source quantitative approach to assess the 3D cerebrovascular architecture in zebrafish.

2020 ◽  
Author(s):  
E. C. Kugler ◽  
J. Frost ◽  
V. Silva ◽  
K. Plant ◽  
K. Chhabria ◽  
...  

AbstractZebrafish transgenic lines and light sheet fluorescence microscopy allow in-depth insights into vascular development in vivo and 3D. However, robust quantification of the zebrafish cerebral vasculature in 3D remains a challenge, and would be essential to describe the vascular architecture. Here, we report an image analysis pipeline that allows 3D quantification of the total or regional zebrafish brain vasculature. This is achieved by landmark- or object-based inter-sample registration and extraction of quantitative parameters including vascular volume, surface area, density, branching points, length, radius, and complexity. Application of our analysis pipeline to a range of sixteen genetic or pharmacological manipulations shows that our quantification approach is robust, allows extraction of biologically relevant information, and provides novel insights into vascular biology. To allow dissemination, the code for quantification, a graphical user interface, and workflow documentation are provided. Together, we present the first 3D quantification approach to assess the whole 3D cerebrovascular architecture in zebrafish.


2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Garrett M. Fogo ◽  
Anthony R. Anzell ◽  
Kathleen J. Maheras ◽  
Sarita Raghunayakula ◽  
Joseph M. Wider ◽  
...  

AbstractThe mitochondrial network continually undergoes events of fission and fusion. Under physiologic conditions, the network is in equilibrium and is characterized by the presence of both elongated and punctate mitochondria. However, this balanced, homeostatic mitochondrial profile can change morphologic distribution in response to various stressors. Therefore, it is imperative to develop a method that robustly measures mitochondrial morphology with high accuracy. Here, we developed a semi-automated image analysis pipeline for the quantitation of mitochondrial morphology for both in vitro and in vivo applications. The image analysis pipeline was generated and validated utilizing images of primary cortical neurons from transgenic mice, allowing genetic ablation of key components of mitochondrial dynamics. This analysis pipeline was further extended to evaluate mitochondrial morphology in vivo through immunolabeling of brain sections as well as serial block-face scanning electron microscopy. These data demonstrate a highly specific and sensitive method that accurately classifies distinct physiological and pathological mitochondrial morphologies. Furthermore, this workflow employs the use of readily available, free open-source software designed for high throughput image processing, segmentation, and analysis that is customizable to various biological models.


eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Timothy J Duerr ◽  
Ester Comellas ◽  
Eun Kyung Jeon ◽  
Johanna E Farkas ◽  
Marylou Joetzjer ◽  
...  

Measuring nascent macromolecular synthesis in vivo is key to understanding how cells and tissues progress through development and respond to external cues. Here we perform in vivo injection of alkyne- or azide-modified analogs of thymidine, uridine, methionine, and glucosamine to label nascent synthesis of DNA, RNA, protein, and glycosylation. Three-dimensional volumetric imaging of nascent macromolecule synthesis was performed in axolotl salamander tissue using whole-mount click chemistry-based fluorescent staining followed by light sheet fluorescent microscopy. We also developed an image processing pipeline for segmentation and classification of morphological regions of interest and individual cells, and we apply this pipeline to the regenerating humerus. We demonstrate our approach is sensitive to biological perturbations by measuring changes in DNA synthesis after limb denervation. This method provides a powerful means to quantitatively interrogate macromolecule synthesis in heterogenous tissues at the organ, cellular, and molecular levels of organization.


2019 ◽  
Vol 12 (1) ◽  
pp. 31-37
Author(s):  
Dave R Shukla ◽  
Richard J McLaughlin ◽  
Julia Lee ◽  
Ngoc Tram V Nguyen ◽  
Joaquin Sanchez-Sotelo

Background Preoperative planning software has been developed to measure glenoid version, glenoid inclination, and humeral head subluxation on computed tomography (CT) for shoulder arthroplasty. However, most studies analyzing the effect of glenoid positioning on outcome were done prior to the introduction of planning software. Thus, measurements obtained from the software can only be extrapolated to predict failure provided they are similar to classic measurements. The purpose of this study was to compare measurements obtained using classic manual measuring techniques and measurements generated from automated image analysis software. Methods Ninety-five two-dimensional computed tomography scans of shoulders with primary glenohumeral osteoarthritis were measured for version according to Friedman method, inclination according to Maurer method, and subluxation according to Walch method. DICOM files were loaded into an image analysis software (Blueprint, Wright Medical) and the output was compared with values obtained manually using a paired sample t-test. Results Average manual measurements included 13.8° version, 13.2° inclination, and 56.2% subluxation. Average image analysis software values included 17.4° version (3.5° difference, p < 0.0001), 9.2° inclination (3.9° difference, p < 0.001), and 74.2% for subluxation (18% difference, p < 0.0001). Conclusions Glenoid version and inclination values from the software and manual measurement on two-dimensional computed tomography were relatively similar, within approximately 4°. However, subluxation measurements differed by approximately 20%.


Author(s):  
Samuel A. Mihelic ◽  
William A. Sikora ◽  
Ahmed M. Hassan ◽  
Michael R. Williamson ◽  
Theresa A. Jones ◽  
...  

AbstractRecent advances in two-photon microscopy (2PM) have allowed large scale imaging and analysis of cortical blood vessel networks in living mice. However, extracting a network graph and vector representations for vessels remain bottlenecks in many applications. Vascular vectorization is algorithmically difficult because blood vessels have many shapes and sizes, the samples are often unevenly illuminated, and large image volumes are required to achieve good statistical power. State-of-the-art, three-dimensional, vascular vectorization approaches require a segmented/binary image, relying on manual or supervised-machine annotation. Therefore, voxel-by-voxel image segmentation is biased by the human annotator/trainer. Furthermore, segmented images oftentimes require remedial morphological filtering before skeletonization/vectorization. To address these limitations, we propose a vectorization method to extract vascular objects directly from unsegmented images. The Segmentation-Less, Automated, Vascular Vectorization (SLAVV) source code in MATLAB is openly available on GitHub. This novel method uses simple models of vascular anatomy, efficient linear filtering, and low-complexity vector extraction algorithms to remove the image segmentation requirement, replacing it with manual or automated vector classification. SLAVV is demonstrated on three in vivo 2PM image volumes of microvascular networks (capillaries, arterioles and venules) in the mouse cortex. Vectorization performance is proven robust to the choice of plasma- or endothelial-labeled contrast, and processing costs are shown to scale with input image volume. Fully-automated SLAVV performance is evaluated on various, simulated 2PM images based on the large, [1.4, 0.9, 0.6] mm input image, and performance metrics show greater robustness to image quality than an intensity-based thresholding approach.


Author(s):  
Loh Teng Hern Tan ◽  
Liang Ee Low ◽  
Siah Ying Tang ◽  
Wei Hsum Yap ◽  
Lay Hong Chuah ◽  
...  

Three-dimensional cell culture methods revolutionize the field of anticancer drug discovery, forming an important link-bridge between conventional in vitro and in vivo models and conferring significant clinical and biological relevant data. The current work presents an affordable yet reproducible method of generating homogenous 3D tumor spheroids. Also, a new open source software is adapted to perform an automated image analysis of 3D tumor spheroids and subsequently generate a list of morphological parameters of which could be utilized to determine the response of these spheroids toward treatments. Our data showed that this work could serve as a reliable 3D cell culture platform for preclinical cytotoxicity testing of natural products prior to the expensive and time-consuming animal models


Author(s):  
Betsy V. Hunter ◽  
Yasin Y. Dhaher

Increased frontal-plane hip movement of the affected leg during the swing phase is a commonly observed gait adaptation in stroke patients. Recent evidence suggests that pathologically-induced torque coupling may contribute to asymmetric gait behaviors observed following stroke. This study proposes to use a CPG-controlled three-dimensional (3D) bipedal model to quantify the effects of abnormal torque coupling on frontal plane gait kinematics. Model dynamics have been evaluated using overground data collection observed under comparable in vivo experimental conditions. The CPG controller has demonstrated ability to provide sustained stable gait over an inclined surface in a simplified model. Preliminary results indicate that the proposed framework is feasible to control a 3D model for investigating the effects of torque coupling on the abnormal frontal plane kinematics of pathological gait.


2001 ◽  
Author(s):  
Catalin F. Baicu ◽  
Michael R. Zile

Abstract Pathological processes which cause diastolic congestive heart failure (CHF), such as pressure overload hypertrophy (POH), produce abnormalities in the material properties of cardiac muscle cells (cardiomyocytes) and may selectively alter its elastic stiffness, viscosity, or both. Previous methods used to characterize these cardiomyocyte viscoelastic properties were constrained by specific biological and engineering limitations, which prevented testing in conditions that mimic normal physiology. The current study proposes an uniaxial variable-rate stretching method, in which isolated cardiomyocytes embedded in a three-dimensional gel matrix were subjected to stretch. Physiological Ca++ (2.5 mM) and rapid stretch rates up to 100 μm/sec provided experimental conditions parallel to in vivo physiology. The proposed method identified and individually quantified both cellular stiffness and viscosity, and showed that POH increased both elastic and viscous cardiomyocyte diastolic properties.


2012 ◽  
Vol 2012 ◽  
pp. 1-16 ◽  
Author(s):  
Lindsey McKeen Polizzotti ◽  
Basak Oztan ◽  
Chris S. Bjornsson ◽  
Katherine R. Shubert ◽  
Bülent Yener ◽  
...  

Prognosis of breast cancer is primarily predicted by the histological grading of the tumor, where pathologists manually evaluate microscopic characteristics of the tissue. This labor intensive process suffers from intra- and inter-observer variations; thus, computer-aided systems that accomplish this assessment automatically are in high demand. We address this by developing an image analysis framework for the automated grading of breast cancer inin vitrothree-dimensional breast epithelial acini through the characterization of acinar structure morphology. A set of statistically significant features for the characterization of acini morphology are exploited for the automated grading of six (MCF10 series) cell line cultures mimicking three grades of breast cancer along the metastatic cascade. In addition to capturing both expected and visually differentiable changes, we quantify subtle differences that pose a challenge to assess through microscopic inspection. Our method achieves 89.0% accuracy in grading the acinar structures as nonmalignant, noninvasive carcinoma, and invasive carcinoma grades. We further demonstrate that the proposed methodology can be successfully applied for the grading ofin vivotissue samples albeit with additional constraints. These results indicate that the proposed features can be used to describe the relationship between the acini morphology and cellular function along the metastatic cascade.


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