Time-lapse imaging is a rich data source offering potential kinetic information of cellular activity and behavior. Tracking and extracting measurements of objects from time-lapse datasets are challenges that result from the complexity and dynamics of each object's motion and intensity or the appearance of new objects in the field of view. A wide range of strategies for proper data sampling, object detection, image analysis, and post-analysis interpretation are available. Theory and methods for single-particle tracking, spot detection, and object linking are discussed in this unit, as well as examples with step-by-step procedures for utilizing semi-automated software and visualization tools for achieving tracking results and interpreting this output.

• Klíčová slova
• digital imaging, image analysis, image segmentation methods, object linking, object tracking, sampling frequency, single-particle tracking,
• MeSH
• časosběrné zobrazování MeSH
• Chlamydomonas cytologie   MeSH
• dánio pruhované MeSH
• fluorescence MeSH
• krevní buňky cytologie   MeSH
• malá interferující RNA metabolismus   MeSH
• regionální krevní průtok MeSH
• rozpoznávání automatizované metody   MeSH
• zobrazování trojrozměrné * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• malá interferující RNA MeSH

Automatic detection and segmentation of biological objects in 2D and 3D image data is central for countless biomedical research questions to be answered. While many existing computational methods are used to reduce manual labeling time, there is still a huge demand for further quality improvements of automated solutions. In the natural image domain, spatial embedding-based instance segmentation methods are known to yield high-quality results, but their utility to biomedical data is largely unexplored. Here we introduce EmbedSeg, an embedding-based instance segmentation method designed to segment instances of desired objects visible in 2D or 3D biomedical image data. We apply our method to four 2D and seven 3D benchmark datasets, showing that we either match or outperform existing state-of-the-art methods. While the 2D datasets and three of the 3D datasets are well known, we have created the required training data for four new 3D datasets, which we make publicly available online. Next to performance, also usability is important for a method to be useful. Hence, EmbedSeg is fully open source (https://github.com/juglab/EmbedSeg), offering (i) tutorial notebooks to train EmbedSeg models and use them to segment object instances in new data, and (ii) a napari plugin that can also be used for training and segmentation without requiring any programming experience. We believe that this renders EmbedSeg accessible to virtually everyone who requires high-quality instance segmentations in 2D or 3D biomedical image data.

• Klíčová slova
• Embeddings, Instance Segmentation, Microscopy,
• MeSH
• algoritmy * MeSH
• lidé MeSH
• mikroskopie * metody   MeSH
• počítačové zpracování obrazu metody   MeSH
• zobrazování trojrozměrné metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Data segmentation and object rendering is required for localization super-resolution microscopy, fluorescent photoactivation localization microscopy (FPALM), and direct stochastic optical reconstruction microscopy (dSTORM). We developed and validated methods for segmenting objects based on Delaunay triangulation in 3D space, followed by facet culling. We applied them to visualize mitochondrial nucleoids, which confine DNA in complexes with mitochondrial (mt) transcription factor A (TFAM) and gene expression machinery proteins, such as mt single-stranded-DNA-binding protein (mtSSB). Eos2-conjugated TFAM visualized nucleoids in HepG2 cells, which was compared with dSTORM 3D-immunocytochemistry of TFAM, mtSSB, or DNA. The localized fluorophores of FPALM/dSTORM data were segmented using Delaunay triangulation into polyhedron models and by principal component analysis (PCA) into general PCA ellipsoids. The PCA ellipsoids were normalized to the smoothed volume of polyhedrons or by the net unsmoothed Delaunay volume and remodeled into rotational ellipsoids to obtain models, termed DVRE. The most frequent size of ellipsoid nucleoid model imaged via TFAM was 35 × 45 × 95 nm; or 35 × 45 × 75 nm for mtDNA cores; and 25 × 45 × 100 nm for nucleoids imaged via mtSSB. Nucleoids encompassed different point density and wide size ranges, speculatively due to different activity stemming from different TFAM/mtDNA stoichiometry/density. Considering twofold lower axial vs. lateral resolution, only bulky DVRE models with an aspect ratio >3 and tilted toward the xy-plane were considered as two proximal nucleoids, suspicious occurring after division following mtDNA replication. The existence of proximal nucleoids in mtDNA-dSTORM 3D images of mtDNA "doubling"-supported possible direct observations of mt nucleoid division after mtDNA replication.

• Klíčová slova
• 3D object segmentation, 3D super-resolution microscopy, Delaunay algorithm, Mitochondrial DNA replication, Nucleoids, Principal component analysis,
• MeSH
• algoritmy * MeSH
• analýza hlavních komponent * MeSH
• buňky Hep G2 MeSH
• DNA vazebné proteiny metabolismus   MeSH
• fluorescenční mikroskopie * MeSH
• konformace nukleové kyseliny MeSH
• lidé MeSH
• mitochondriální DNA chemie   metabolismus   MeSH
• mitochondriální proteiny metabolismus   MeSH
• molekulární modely MeSH
• zobrazování trojrozměrné * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA vazebné proteiny MeSH
• mitochondriální DNA MeSH
• mitochondriální proteiny MeSH
• NABP2 protein, human MeSH Prohlížeč

BACKGROUND AND OBJECTIVE: We focus on three-dimensional higher-order tensorial (HOT) images using Finsler geometry. In biomedical image analysis, these images are widely used, and they are based on the diffusion profiles inside the voxels. The diffusion information is stored in the so-called diffusion tensor D. Our objective is to present new methods revealing the architecture of neural fibers in presence of crossings and high curvatures. After tracking the fibers, we achieve direct 3D image segmentation to analyse the brain's white matter structures. METHODS: To deal with the construction of the underlying fibers, the inverse of the second-order diffusion tensor D, understood as the metric tensor D-1, is commonly used in DTI modality. For crossing and highly curved fibers, higher order tensors are more relevant, but it is challenging to find an analogue of such an inverse in the HOT case. We employ an innovative approach to metrics based on higher order tensors to track the fibers properly. We propose to feed the tracked fibers as the internal initial contours in an efficient version of 3D segmentation. RESULTS: We propose a brand-new approach to the inversion of a diffusion HOT, and an effective way of fiber tracking in the Finsler setting, based on innovative classification of the individual voxels. Thus, we can handle complex structures with high curvatures and crossings, even in the presence of noise. Based on our novel tractography approach, we also introduce a new segmentation method. We feed the detected fibers as the initial position of the contour surfaces to segment the image using a relevant active contour method (i.e., initiating the segmentation from inside the structures). CONCLUSIONS: This is a pilot work, enhancing methods for fiber tracking and segmentation. The implemented algorithms were successfully tested on both synthetic and real data. The new features make our algorithms robust and fast, and they allow distinguishing individual objects in complex structures, even under noise.

• Klíčová slova
• Finsler geometry, HARDI, HOT inversion, Segmentation, Tractography, White matter structure,
• MeSH
• algoritmy MeSH
• bílá hmota * diagnostické zobrazování   MeSH
• mozek diagnostické zobrazování   MeSH
• počítačové zpracování obrazu metody   MeSH
• zobrazování difuzních tenzorů * metody   MeSH
• zobrazování trojrozměrné metody   MeSH
• Publikační typ
• časopisecké články MeSH

Image cytometry still faces the problem of the quality of cell image analysis results. Degradations caused by cell preparation, optics, and electronics considerably affect most 2D and 3D cell image data acquired using optical microscopy. That is why image processing algorithms applied to these data typically offer imprecise and unreliable results. As the ground truth for given image data is not available in most experiments, the outputs of different image analysis methods can be neither verified nor compared to each other. Some papers solve this problem partially with estimates of ground truth by experts in the field (biologists or physicians). However, in many cases, such a ground truth estimate is very subjective and strongly varies between different experts. To overcome these difficulties, we have created a toolbox that can generate 3D digital phantoms of specific cellular components along with their corresponding images degraded by specific optics and electronics. The user can then apply image analysis methods to such simulated image data. The analysis results (such as segmentation or measurement results) can be compared with ground truth derived from input object digital phantoms (or measurements on them). In this way, image analysis methods can be compared with each other and their quality (based on the difference from ground truth) can be computed. We have also evaluated the plausibility of the synthetic images, measured by their similarity to real image data. We have tested several similarity criteria such as visual comparison, intensity histograms, central moments, frequency analysis, entropy, and 3D Haralick features. The results indicate a high degree of similarity between real and simulated image data.

• MeSH
• algoritmy MeSH
• buněčné jadérko ultrastruktura   MeSH
• buněčné jádro ultrastruktura   MeSH
• fantomy radiodiagnostické * MeSH
• fluorescenční mikroskopie metody   MeSH
• granulocyty cytologie   MeSH
• HL-60 buňky MeSH
• lidé MeSH
• mikrosféry MeSH
• obrazová cytometrie metody   MeSH
• zobrazování trojrozměrné metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: The presented study investigates the application of bi-arterial 3D printed models to guide transseptal puncture (TSP) in left atrial appendage closure (LAAC). AIMS: The objectives are to (1) test the feasibility of 3D printing (3DP) for TSP guidance, (2) analyse the distribution of the optimal TSP locations, and (3) define a CT-derived 2D parameter suitable for predicting the optimal TSP locations. METHODS: Preprocedural planning included multiplanar CT reconstruction, 3D segmentation, and 3DP. TSP was preprocedurally simulated in vitro at six defined sites. Based on the position of the sheath, TSP sites were classified as optimal, suboptimal, or nonoptimal. The aim was to target the TSP in the recommended position during the procedure. Procedure progress was assessed post hoc by the operator. RESULTS: Of 68 screened patients, 60 patients in five centers (mean age of 74.68 ± 7.64 years, 71.66% males) were prospectively analyzed (3DP failed in one case, and seven patients did not finally undergo the procedure). In 55 patients (91.66%), TSP was performed in the optimal location as recommended by the 3DP. The optimal locations for TSP were postero-inferior in 45.3%, mid-inferior in 45.3%, and antero-inferior in 37.7%, with a mean number of optimal segments of 1.34 ± 0.51 per patient. When the optimal TSP location was achieved, the procedure was considered difficult in only two (3.6%) patients (but in both due to complicated LAA anatomy). Comparing anterior versus posterior TSP in 2D CCT, two parameters differed significantly: (1) the angle supplementary to the LAA ostium and the interatrial septum angle (160.83° ± 9.42° vs. 146.49° ± 8.67°; p = 0.001), and (2) the angle between the LAA ostium and the mitral annulus (95.02° ± 3.73° vs. 107.38° ± 6.76°; p < 0.001), both in the sagittal plane. CONCLUSIONS: In vitro TSP simulation accurately determined the optimal TSP locations for LAAC and facilitated the procedure. More than one-third of the optimal TSP sites were anterior.

• Klíčová slova
• 3D printing, computed tomography, left atrial appendage closure, transseptal puncture,
• MeSH
• 3D tisk MeSH
• fibrilace síní * terapie   chirurgie   MeSH
• lidé MeSH
• počítačová rentgenová tomografie MeSH
• punkce metody   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• síňové ouško * diagnostické zobrazování   chirurgie   MeSH
• výsledek terapie MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

OBJECTIVES: The diploic channels appear to be more developed in humans than in nonhuman primates, suggesting they may be relevant in evolutionary biology. This study is aimed at providing a segmentation procedure for diploic channels and CT analysis, a quantitative description of their variation in modern humans, and paleoanthropological case-studies. MATERIALS AND METHODS: CT data were used for the 2D and 3D visualization, rendering, and measure, of diploic channels in modern and fossil hominids. We analyzed 20 modern human skulls and three Neanderthals. The effect of different resolution factors was evaluated. A specific protocol was designed to segment the vascular network and localize the main branches, reducing the noise of the cancellous bone. RESULTS: We provide a quantitative description of the frontal, parietal, and occipital diploic networks in modern humans and in three Neanderthals. There is a correlation in the degree of vascularization among the different vault areas. No side differences can be detected. The diploic network is commonly connected with the meningeal artery at the temporal fossa, with the emissary veins at the occipital bone, and with the venous sinuses at the parieto-occipital areas. The channels are more developed in the parietal areas. The three Neanderthals show a vascular development, which is in the lower range of the modern human variation. CONCLUSIONS: Modern humans display a large variation in their morphological patterns, being the parietal area the most vascularized. The pattern of the diploic channels may be relevant in anthropology, medicine, and paleontology, taking into account their possible involvement in thermoregulation.

• Klíčová slova
• cranial anatomy, digital anatomy, diploe, paleoanthropology,
• MeSH
• antropologie fyzická MeSH
• dospělí MeSH
• lebka * anatomie a histologie   krevní zásobení   diagnostické zobrazování   MeSH
• lidé MeSH
• neandertálci anatomie a histologie   MeSH
• počítačová rentgenová tomografie MeSH
• zkameněliny * MeSH
• zvířata MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

INTRODUCTION AND OBJECTIVES: Pathologic intimal thickening (PIT) has been considered a benign plaque phenotype. We report plaque phenotypic changes in a baseline/follow-up intravascular ultrasound-based virtual histology study. METHODS: A total of 61 patients with stable coronary artery disease were analyzed from the HEAVEN trial (89 patients randomized between routine statin therapy vs atorvastatin 80mg and ezetimibe 10mg) with serial intravascular ultrasound imaging of nonculprit vessels. We compared changes in 693 baseline and follow-up 5-mm long segments in a novel risk score, Liverpool Active Plaque Score (LAPS), plaque parameters, and plaque composition. RESULTS: The PIT showed the highest increase of risk score and, with fibrous plaque, also the LAPS. Necrotic core (NC) abutting to the lumen increased in PIT (22 ± 51.7; P = .0001) and in fibrous plaque (17.9 ± 42.6; P = .004) but decreased in thin cap fibroatheroma (TCFA) (⿿15.14 ± 52.2; P = .001). The PIT was the most likely of all nonthin cap fibroatheroma plaque types to transform into TCFA at follow-up (11% of all TCFA found during follow-up and 35.9% of newly-developed TCFA), but showed (together with fibrous plaque) the lowest stability during lipid-lowering therapy (24.7% of PIT remained PIT and 24.5% of fibrous plaque remained fibrous plaque). CONCLUSIONS: Over the 1-year follow-up, PIT was the most dynamic of the plaque phenotypes and was associated with an increase of risk score and LAPS (together with fibrous plaque), NC percentage (together with fibrous plaque) and NC abutting to the lumen, despite a small reduction of plaque volume during lipid-lowering therapy. The PIT was the main source for new TCFA segments.

• Klíčová slova
• Angina estable, Atherosclerotic plaque, Ecografía intravascular, Estudio de seguimiento, Follow-up study, Intravascular ultrasound, Lipid, Lípidos, Placa ateroesclerótica, Stable angina,
• MeSH
• anticholesteremika aplikace a dávkování   MeSH
• aterosklerotický plát krev   diagnóza   farmakoterapie   MeSH
• atorvastatin aplikace a dávkování   MeSH
• časové faktory MeSH
• ezetimib aplikace a dávkování   MeSH
• intervenční ultrasonografie metody   MeSH
• kombinovaná farmakoterapie MeSH
• koronární cévy diagnostické zobrazování   MeSH
• LDL-cholesterol krev   MeSH
• lidé středního věku MeSH
• lidé MeSH
• následné studie MeSH
• nemoci koronárních tepen krev   diagnóza   farmakoterapie   MeSH
• progrese nemoci MeSH
• stupeň závažnosti nemoci MeSH
• uživatelské rozhraní počítače * MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zobrazování trojrozměrné MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• multicentrická studie MeSH
• randomizované kontrolované studie MeSH
• Názvy látek
• anticholesteremika MeSH
• atorvastatin MeSH
• ezetimib MeSH
• LDL-cholesterol MeSH

OBJECTIVES: Prenatal identification of the different dentition morphotypes, which exist in the lower molar region of tabby (Ta) adult mice, and investigation of their origin. The mouse Ta syndrome and its counterpart anhidrotic (hypohidrotic) ectodermal dysplasia (EDA) in human are characterized by absence or hypoplasia of sweat glands, hair and teeth. DESIGN: Analysis of tooth morphogenesis using serial histological sections and 3D computer aided reconstructions of the dental epithelium in the cheek region of the mandible. SETTING AND SAMPLE POPULATION: Institute of Experimental Medicine, Academy of Sciences, Prague. Heads of 75 Ta homozygous and hemizygous mice and 40 wild type (WT) control mice aged from embryonic day (ED) 14.0-20.5 (newborns), harvested during 1995-2001. OUTCOME MEASURE: Prenatal identification of five distinct morphotypes of Ta dentition on the basis of differences in tooth number, size, shape, position and developmental stage and of the morphology of the enamel knot in the most mesial tooth primordium. RESULTS: The mesio-distal length of the dental epithelium was similar in the lower cheek region in Ta and WT mice. In Ta embryos, there was altered the mesio-distal segmentation of the dental epithelium giving rise to the individual tooth primordia. Prenatally, two basic morphotypes I and II and their particular subtypes (Ia, Ib, Ic, and IIa, IIb, respectively) of the developing dentition were identified from day 15.5. The incidence of the distinct morphotypes in the present sample did not differ from postnatal data. The proportion of the morphotype I and II was dependent on mother genotype. CONCLUSION: The different dentition morphotypes in Ta mice originate from a defect in the mesio-distal segmentation of the dental epithelium in mouse embryos. This defect presumably leads to variable positions of tooth boundaries that do not correspond to those of the WT molars. One tooth primordium of Ta mice might be derived from adjacent parts of two molar primordia in WT mice.

• MeSH
• abnormality zubů embryologie   MeSH
• ektodermální dysplazie embryologie   genetika   patologie   MeSH
• epitel embryologie   MeSH
• genetické nemoci vázané na chromozom X embryologie   MeSH
• hypohidróza embryologie   genetika   patologie   MeSH
• lidé MeSH
• mandibula MeSH
• modely nemocí na zvířatech MeSH
• moláry abnormality   embryologie   MeSH
• morfogeneze MeSH
• mutantní kmeny myší MeSH
• myši MeSH
• odontogeneze MeSH
• odontometrie MeSH
• počítačové zpracování obrazu MeSH
• zubní zárodek abnormality   embryologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Resurgent interest in synaptic circuitry and plasticity has emphasized the importance of 3D reconstruction from serial section electron microscopy (3DEM). Three volumes of hippocampal CA1 neuropil from adult rat were imaged at X-Y resolution of ~2 nm on serial sections of ~50-60 nm thickness. These are the first densely reconstructed hippocampal volumes. All axons, dendrites, glia, and synapses were reconstructed in a cube (~10 μm(3)) surrounding a large dendritic spine, a cylinder (~43 μm(3)) surrounding an oblique dendritic segment (3.4 μm long), and a parallelepiped (~178 μm(3)) surrounding an apical dendritic segment (4.9 μm long). The data provide standards for identifying ultrastructural objects in 3DEM, realistic reconstructions for modeling biophysical properties of synaptic transmission, and a test bed for enhancing reconstruction tools. Representative synapses are quantified from varying section planes, and microtubules, polyribosomes, smooth endoplasmic reticulum, and endosomes are identified and reconstructed in a subset of dendrites. The original images, traces, and Reconstruct software and files are freely available and visualized at the Open Connectome Project (Data Citation 1).

• MeSH
• elektronová mikroskopie MeSH
• hipokampus anatomie a histologie   MeSH
• krysa rodu Rattus MeSH
• neuropil * MeSH
• počítačové zpracování obrazu MeSH
• software MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• dataset MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH