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Scientific Reports Feb 2024Many critical issues arise when training deep neural networks using limited biological datasets. These include overfitting, exploding/vanishing gradients and other...
Many critical issues arise when training deep neural networks using limited biological datasets. These include overfitting, exploding/vanishing gradients and other inefficiencies which are exacerbated by class imbalances and can affect the overall accuracy of a model. There is a need to develop semi-supervised models that can reduce the need for large, balanced, manually annotated datasets so that researchers can easily employ neural networks for experimental analysis. In this work, Iterative Pseudo Balancing (IPB) is introduced to classify stem cell microscopy images while performing on the fly dataset balancing using a student-teacher meta-pseudo-label framework. In addition, multi-scale patches of multi-label images are incorporated into the network training to provide previously inaccessible image features with both local and global information for effective and efficient learning. The combination of these inputs is shown to increase the classification accuracy of the proposed deep neural network by 3[Formula: see text] over baseline, which is determined to be statistically significant. This work represents a novel use of pseudo-labeling for data limited settings, which are common in biological image datasets, and highlights the importance of the exhaustive use of available image features for improving performance of semi-supervised networks. The proposed methods can be used to reduce the need for expensive manual dataset annotation and in turn accelerate the pace of scientific research involving non-invasive cellular imaging.
Topics: Humans; Microscopy; Learning; Neural Networks, Computer; Product Labeling; Stem Cells; Image Processing, Computer-Assisted; Supervised Machine Learning
PubMed: 38396157
DOI: 10.1038/s41598-024-54993-y -
Current Oncology (Toronto, Ont.) Feb 2024Fluorescence-guided oncology promises to improve both the detection and treatment of malignancy. We sought to investigate the temporal distribution of indocyanine green...
UNLABELLED
Fluorescence-guided oncology promises to improve both the detection and treatment of malignancy. We sought to investigate the temporal distribution of indocyanine green (ICG), an exogenous fluorophore in human colorectal cancer. This analysis aims to enhance our understanding of ICG's effectiveness in current tumour detection and inform potential future diagnostic and therapeutic enhancements.
METHODS
Fifty consenting patients undergoing treatment for suspected/confirmed colorectal neoplasia provided near infrared (NIR) video and imagery of transanally recorded and ex vivo resected rectal lesions following intravenous ICG administration (0.25 mg/kg), with a subgroup providing tissue samples for microscopic (including near infrared) analysis. Computer vision techniques detailed macroscopic 'early' (<15 min post ICG administration) and 'late' (>2 h) tissue fluorescence appearances from surgical imagery with digital NIR scanning (Licor, Lincoln, NE, USA) and from microscopic analysis (Nikon, Tokyo, Japan) undertaken by a consultant pathologist detailing tissue-level fluorescence distribution over the same time.
RESULTS
Significant intra-tumoural fluorescence heterogeneity was seen 'early' in malignant versus benign lesions. In all 'early' samples, fluorescence was predominantly within the tissue stroma, with uptake within plasma cells, blood vessels and lymphatics, but not within malignant or healthy glands. At 'late' stage observation, fluorescence was visualised non-uniformly within the intracellular cytoplasm of malignant tissue but not retained in benign glands. Fluorescence also accumulated within any present peritumoural inflammatory tissue.
CONCLUSION
This study demonstrates the time course diffusion patterns of ICG through both benign and malignant tumours in vivo in human patients at both macroscopic and microscopic levels, demonstrating important cellular drivers and features of geolocalisation and how they differ longitudinally after exposure to ICG.
Topics: Humans; Tissue Distribution; Indocyanine Green; Colorectal Neoplasms
PubMed: 38392057
DOI: 10.3390/curroncol31020063 -
Archives of Microbiology Feb 2024The River Nile is the main source of fresh water in Egypt, where its water is used for irrigation, drinking, fisheries, industrial uses, and recreation. For sustainable...
The River Nile is the main source of fresh water in Egypt, where its water is used for irrigation, drinking, fisheries, industrial uses, and recreation. For sustainable utilization of the River Nile and its branches in the Nile Delta region, it is necessary to monitor regular investigation for the biodiversity of protozoan fauna in the Damietta branch and other freshwater canals in Dakahlyia Governorate. Water samples were collected monthly from different water sources, for 1 year, and examined for protozoans, using phase-contrast microscopy and recorded video films, The genus Vannella Bovee 1965 is recorded for the first time in four freshwater localities: Demietta branch of the River Nile, Mansouria Canal, Bouhia Canal, and Bahr El-Saghir Canal. A detailed morphological description with a brief report of their locomotion has been given for four morphologically different Vannella species. The locomotive form of Vannella sp.1 has a long pointed posterior tail and 2 lateral posterior processes. Such a tail was absent in other Vannella species. Vannella sp.2 is unique among other recorded species, where its locomotive form possesses a long posterior rounded tail region and a frontal hyaloplasm provided with a wavy surface that forms several lobes and finger-like processes during locomotion. In addition, the hyaloplasm produces several transverse waves that vary in thickness and density. The floating form of Vannella sp.2 is of a radial type and has comparatively long hyaline pointed and spiral pseudopodia. The process of transformation of locomotive form to floating form in Vannella sp.2 has been followed up using several recorded video films. The locomotive form of Vannella sp.3 is bear-shaped, while that of Vannella sp.4 has variable shapes from semicircular to rectangular and sometimes fan-shaped. During movement in vivo, locomotive cells of all Vannella species, except Vannella sp.1, move in nearly a straight line, but there were variations in their rate of locomotion. Vannella sp.4 recorded the highest rate (6.8 µm/s), followed by Vannella sp.2 (4.5 µm/s), Vannella sp.3 (2.4 µm/s), and finally Vannella sp.1 (1.0 µm/s). Molecular studies and transmission electron microscope examinations are still needed to confirm the precise identity of each Vannella species.
Topics: Egypt; Fresh Water; Amoebozoa; Rivers; Water
PubMed: 38376635
DOI: 10.1007/s00203-024-03837-4 -
Clinical Hemorheology and... 2024This review spans a wide arc from the first observations of the early anatomists to the present day. William Harvey was the first to describe the heart as the centre... (Review)
Review
This review spans a wide arc from the first observations of the early anatomists to the present day. William Harvey was the first to describe the heart as the centre of the large and small circulatory system. He thus replaced the previously valid system of Galenos, It was Marcello Malpighi who first described that the capillary system connects the arteries with the veins. In 1688 Antoni van Leeuwenhoek (1632-1686) confirmed these results with a paper on capillary perfusion in the caudal fin of the glass eel. It was then Hermann Boerhave (1668-1738, Leiden) who was the first to carry out microcirculation tests on patients. He studied the microcirculation in the human bulbar conjunctiva. Even today, microcirculation studies in the conjunctiva bulbi of patients are carried out today. Until 1831, it was never quite clear whether the observations reported belonged mainly to the field of microcirculation, which had not yet been defined. This was done in Great Britain by Marshall Hall (1790-1857). Technical Improvements allowed increasingly sophisticated studies of the morphological structure of the terminal vasculature. According to Gustav Ricker (1870-1948, Vienna), the terminal vasculature comprises the functional unit of the smallest arteries, arterioles, capillaries and venules. In 1921 it was still thought that the blood circulation was the sole response to the pumping action of the heart. Even the classic work by Bayliss on the myogenic hypothesis (later referred to as "blood flow autoregulation") initially received little attention. More strikingly, even the findings of August Krogh, for which he received the Nobel Prize in Medicine in 1920 (for his discovery of the mechanisms of capillary motor regulation), were ignored. During an outstanding autoregulation symposium held in 1963 a broad consensus was reached on active and passive mechanisms, which is more or less valid till today. The mechanisms of regulation of capillary blood flow are now largely understood, although not completely resolved. The development of video systems with recording capability and automated off-line recording of capillary erythrocyte velocities allowed the application of morphological and dynamic studies of cutaneous capillaries in humans. These reopened the field of physiological or pathophysiological questions again for many groups worldwide. Since 1955, many publications on "microcirculation (5423)" and "capillary microscopy (2195)" have been listed in pubmed.
Topics: Humans; Capillaries; Erythrocytes; Microcirculation
PubMed: 38363606
DOI: 10.3233/CH-248001 -
Journal of Biomedical Optics Feb 2024There has been significant interest in the development of miniature photoacoustic imaging probes for a variety of clinical uses, including the assessment of tumors and...
SIGNIFICANCE
There has been significant interest in the development of miniature photoacoustic imaging probes for a variety of clinical uses, including the assessment of tumors and minimally invasive surgical guidance. Most of the previously implemented probes are either side viewing or operate in the optical-resolution microscopy mode in which the imaging depth is limited to . We describe a forward-viewing photoacoustic probe that operates in tomography mode and simultaneously provides white light video images.
AIM
We aim to develop a dual-modality endoscope capable of performing high-resolution PAT imaging and traditional white light videoscopy simultaneously in the forward-viewing configuration.
APPROACH
We used a Fabry-Pérot ultrasound sensor that operates in the 1500 to 1600 nm wavelength range and is transparent in the visible and near infrared region (580 to 1250 nm). The FP sensor was optically scanned using a miniature MEMs mirror located at the proximal end of the endoscope, resulting in a system that is sufficiently compact (10 mm outer diameter) and lightweight for practical endoscopic use.
RESULTS
The imaging performance of the endoscope is evaluated, and dual-mode imaging capability is demonstrated using phantoms and abdominal organs of an mouse including spleen, liver, and kidney.
CONCLUSIONS
The proposed endoscope design offers several advantages including the high acoustic sensitivity and wide detection bandwidth of the FP sensor, dual-mode imaging capability, compact footprint, and an all-optical distal end for improved safety. The dual-mode imaging capability also offers the advantage of correlating the tissue surface morphology with the underlying vascular anatomy. Potential applications include the guidance of laparoscopic surgery and other interventional procedures.
Topics: Mice; Animals; Photoacoustic Techniques; Endoscopes; Ultrasonography; Microscopy; Endoscopy
PubMed: 38361504
DOI: 10.1117/1.JBO.29.2.020502 -
Journal of Visualized Experiments : JoVE Jan 2024Reconstituted cytoskeleton composites have emerged as a valuable model system for studying non-equilibrium soft matter. The faithful capture of the dynamics of these 3D,...
Reconstituted cytoskeleton composites have emerged as a valuable model system for studying non-equilibrium soft matter. The faithful capture of the dynamics of these 3D, dense networks calls for optical sectioning, which is often associated with fluorescence confocal microscopes. However, recent developments in light-sheet fluorescence microscopy (LSFM) have established it as a cost-effective and, at times, superior alternative. To make LSFM accessible to cytoskeleton researchers less familiar with optics, we present a step-by-step beginner's guide to building a versatile light-sheet fluorescence microscope from off-the-shelf components. To enable sample mounting with traditional slide samples, this LSFM follows the single-objective light-sheet (SOLS) design, which utilizes a single objective for both the excitation and emission collection. We describe the function of each component of the SOLS in sufficient detail to allow readers to modify the instrumentation and design it to fit their specific needs. Finally, we demonstrate the use of this custom SOLS instrument by visualizing asters in kinesin-driven microtubule networks.
Topics: Microscopy, Fluorescence; Cytoskeleton; Microtubules
PubMed: 38345245
DOI: 10.3791/65411 -
Journal of Visualized Experiments : JoVE Jan 2024Despite the numerous clearing techniques that emerged in the last decade, processing postmortem human brains remains a challenging task due to its dimensions and...
Despite the numerous clearing techniques that emerged in the last decade, processing postmortem human brains remains a challenging task due to its dimensions and complexity, which make imaging with micrometer resolution particularly difficult. This paper presents a protocol to perform the reconstruction of volumetric portions of the human brain by simultaneously processing tens of sections with the SHORT (SWITCH - H2O2 - Antigen Retrieval - 2,2'-thiodiethanol [TDE]) tissue transformation protocol, which enables clearing, labeling, and sequential imaging of the samples with light-sheet fluorescence microscopy (LSFM). SHORT provides rapid tissue clearing and homogeneous multi-labeling of thick slices with several neuronal markers, enabling the identification of different neuronal subpopulations in both white and grey matter. After clearing, the slices are imaged via LSFM with micrometer resolution and in multiple channels simultaneously for a rapid 3D reconstruction. By combining SHORT with LSFM analysis within a routinely high-throughput protocol, it is possible to obtain the 3D cytoarchitecture reconstruction of large volumetric areas at high resolution in a short time, thus enabling comprehensive structural characterization of the human brain.
Topics: Humans; Hydrogen Peroxide; Microscopy, Fluorescence; Brain; Neurons; Neuroimaging; Imaging, Three-Dimensional; Optical Imaging
PubMed: 38345230
DOI: 10.3791/65960 -
Frontiers in Pediatrics 2024The mutations in the (retinitis pigmentosa GTPase regulator) gene are the most common cause of X-linked retinitis pigmentosa (XLRP), a rare genetic disorder affecting...
BACKGROUND
The mutations in the (retinitis pigmentosa GTPase regulator) gene are the most common cause of X-linked retinitis pigmentosa (XLRP), a rare genetic disorder affecting the photoreceptor cells in the retina. Several reported cases identified this gene as a genetic link between retinitis pigmentosa (RP) and primary ciliary dyskinesia (PCD), characterised by impaired ciliary function predominantly in the respiratory tract. Since different mutations in the same gene can result in various clinical manifestations, it is important to describe a correlation between the gene variant and the observed phenotype.
METHODS
Two young brothers from a non-consanguineous Slovak family with diagnosed retinal dystrophy and recurrent respiratory infections were examined. Suspected PCD was diagnosed based on a PICADAR questionnaire, nasal nitric oxide analysis, transmission electron microscopy, high-speed video microscopy analysis, and genetic testing.
RESULTS
We identified a novel frameshift mutation NM_001034853: c.309_310insA, p.Glu104Argfs*12, resulting in a complex X-linked phenotype combining PCD and RP. In our patients, this mutation was associated with normal ultrastructure of respiratory cilia, reduced ciliary epithelium, more aciliary respiratory epithelium, shorter cilia, and uncoordinated beating with a frequency at a lower limit of normal beating, explaining the clinical manifestation of PCD in our patients.
CONCLUSION
The identified novel pathogenic mutation in the gene expands the spectrum of genetic variants associated with the X-linked PCD phenotype overlapping with RP, highlighting the diversity of mutations contributing to the disorder. The described genotype-phenotype correlation can be useful in clinical practice to recognise a broader spectrum of PCD phenotypes as well as for future research focused on the genetic basis of PCD, gene interactions, the pathways implicated in PCD pathogenesis, and the role of RPGR protein for the proper functioning of cilia in various tissues throughout the body.
PubMed: 38333087
DOI: 10.3389/fped.2024.1339664 -
European Journal of Ophthalmology May 2024Ophthalmic microscopes have been crucial in visualizing surgical fields, but their limitations in enhancing the surgical view through digital image processing have...
BACKGROUND
Ophthalmic microscopes have been crucial in visualizing surgical fields, but their limitations in enhancing the surgical view through digital image processing have prompted the development of digital surgical microscopes. The Beyeonics One microscope, a novel digital microscope, offers ophthalmic surgeons a 3D visualization platform and an augmented reality (AR) surgical headset, potentially improving surgical decision-making and outcomes. While its initial use has been described in cataract and corneal surgeries, its application in vitreoretinal surgery remains relatively unexplored.
METHODS
In this interventional case series, we collected data from the medical records of patients who underwent vitreoretinal surgery using the Beyeonics One 3D visualization platform at the Tel Aviv Medical Center. A total of 36 eyes from 36 subjects were included. Surgical techniques included retinal detachment surgeries and macular surgeries, performed by experienced surgeons. The surgical visualization was facilitated by the Beyeonics One 3D head-mounted display (HMD) platform.
RESULTS
The procedures were uneventful, and none intra- or postoperative complications were reported, and surgeons did not experience any signal delay in the real-time video.
DISCUSSION
The Beyeonics One microscope offers several potential advantages in vitreoretinal surgery, including digital image processing, enhanced depth perception through the 3D HMD platform, and hands-free image control using head gestures. While this study demonstrates the feasibility and safety of the Beyeonics One microscope, addressing limitations related to hazy views and optimizing image quality are crucial for consistent visualization.
Topics: Humans; Vitreoretinal Surgery; Imaging, Three-Dimensional; Female; Male; Middle Aged; Aged; Retinal Detachment; Surgery, Computer-Assisted; Microscopy; Adult; Augmented Reality; Retrospective Studies
PubMed: 38327079
DOI: 10.1177/11206721241229115 -
Journal of Visualized Experiments : JoVE Jan 2024The cell membrane is crucial for cell survival, and ensuring its integrity is essential as the cell experiences injuries throughout its entire life cycle. To prevent...
The cell membrane is crucial for cell survival, and ensuring its integrity is essential as the cell experiences injuries throughout its entire life cycle. To prevent damage to the membrane, cells have developed efficient plasma membrane repair mechanisms. These repair mechanisms can be studied by combining confocal microscopy and nanoscale thermoplasmonics to identify and investigate the role of key proteins, such as annexins, involved in surface repair in living cells and membrane model systems. The puncturing method employs a laser to induce highly localized heating upon nanoparticle irradiation. The use of near-infrared light minimizes phototoxicity in the biological sample, while the majority of the absorption takes place in the near-infrared resonant plasmonic nanoparticle. This thermoplasmonic method has been exploited for potential photothermal and biophysical research to enhance the understanding of intracellular mechanisms and cellular responses through vesicle and cell fusion studies. The approach has shown to be complementary to existing methods for membrane disruption, such as mechanically, chemically, or optically induced injuries, and provides a high level of control by inflicting extremely localized injuries. The extent of the injury is limited to the vicinity of the spherical nanoparticle, and no detrimental damage occurs along the beam path as opposed to pulsed lasers using different wavelengths. Despite certain limitations, such as the formation of nanobubbles, the thermoplasmonic method offers a unique tool for investigating cellular responses in plasma membrane repair in an almost native environment without compromising cell viability. When integrated with confocal microscopy, the puncturing method can provide a mechanistic understanding of membrane dynamics in model membrane systems as well as quantitative information on protein responses to membrane damage, including protein recruitment and their biophysical function. Overall, the application of this method to reduced model systems can enhance our understanding of the intricate plasma membrane repair machinery in living cells.
Topics: Cell Membrane; Nanoparticles; Membranes; Cell Survival; Infrared Rays
PubMed: 38314838
DOI: 10.3791/65776