In the modern era of high-quality cross-sectional imaging, pancreatic cysts (PCs) are a common finding. The prevalence of incidental PCs detected on cross-sectional abdominal imaging (such as CT scan) is 3%-14% which increases with age, up to 8% in those 70 years or older. Although PCs can be precursors of future pancreatic adenocarcinoma, imaging modalities such as CT scan, MRI, or endoscopic ultrasound with fine-needle aspiration (EUS-FNA) are suboptimal at risk stratifying the malignant potential of individual cysts. An inaccurate diagnosis could potentially overlook premalignant lesions, which can lead to missed lesions, lead to unnecessary surveillance, or cause significant long-term surgical morbidity from unwarranted removal of benign lesions. Although current guidelines recommend an EUS or MRI for surveillance, they lack the sensitivity to risk stratify and guide management decisions. Needle-based confocal laser endomicroscopy (nCLE) with EUS-FNA can be a superior diagnostic modality for PCs with sensitivity and accuracy exceeding 90%. Despite this, a significant challenge to the widespread use of nCLE is the lack of adequate exposure and training among gastroenterologists for the real-time interpretation of images. Better understanding, training, and familiarization with this novel technique and the imaging characteristics can overcome the limitations of nCLE use, improving clinical care of patients with PCs. Here, we aim to review the types of CLE in luminal and nonluminal gastrointestinal disorders with particular attention to the evaluation of PCs. Furthermore, we discuss the adverse events and safety of CLE.

UNASSIGNED: 4-methacryloyloxyethyl trimellitate anhydride/methyl methacrylate-tri-n-butyl borane (4-META/MMA-TBB) resin is used for indirect restorations. We aimed to evaluate effects of immersion in 4-META/MMA-TBB-activated liquid on the bond strength of root canal dentin.
UNASSIGNED: We used freshly extracted single-rooted human teeth. After decoronation, each root was vertically sectioned into halves; their dentin walls were polished and flattened. The control group underwent dentin treatment with Green Activator. The immersion group was treated with Green Activator and Teeth Primer and immersed in 4-META/MMA-TBB-activated liquid. After bonding the resin blocks with Super-Bond, microtensile bond strength (μTBS) tests were performed (n = 6), and fracture surfaces were analyzed. Before surface treatment, dentin was immersed in a sodium fluorescein solution for 3 h, and resin blocks were bonded with Super-Bond with rhodamine B as in the bond strength test. The bonded cross section was observed using confocal laser scanning microscopy (CLSM).
UNASSIGNED: μTBS was significantly higher in the immersion group than in the control group (61.5 [51.3-66.7] vs. 33.0 [20.4-57.8] MPa; P < 0.05). Fracture mode analysis showed that, compared with the control group, the immersion group had a significantly lower rate of adhesive failure at the dentin interface and a significantly higher rate of cohesive failure in Super-Bond (P < 0.01). CLSM showed a water droplet-like accumulation of fluorescein dye above the hybrid layer in the control group, not in the immersion group.
UNASSIGNED: Immersion in a 4-META/MMA-TBB-activated liquid inhibited water exudation from the root canal dentin and improved the bond strength.

A 14-year-old girl presented with a facial-pigmented lesion suspicious of melanoma clinically and dermoscopically. In vivo, reflectance confocal microscopy (RCM) findings excluded melanoma by revealing typical epidermal honeycomb and cobblestone patterns. Well-defined follicular contours were seen at the dermal-epidermal junction; there were no elongated, \"medusa head-like\" follicular protrusions or folliculotropism, which are classical findings seen in lentigo maligna. With this report, we aim to demonstrate the significance of utilizing RCM technology in difficult to diagnose lentiginous pigmented lesions.

FACT is a developed technique for clearing tissues that does not use acrylamide. Since the removal of lipids is crucial for transparency and efficient antibody staining throughout the tissue, especially for microscopy and imaging, it is a harmful process that can cause the loss of important biological molecules such as proteins. The FACT technique overcomes this by chemically bonding the membrane and intracellular proteins with the extracellular matrix, creating a massive 3D hydrogel matrix and providing structural support to fortify the tissue during processing. Compared to other acrylamide-based techniques, the FACT technique requires less labor and harmful chemicals and is therefore considered a more suitable option. In this study, we describe the complete FACT protocol for antibody staining and imaging of whole-cleared tissues while preserving structure and improving image quality. The protocol includes tissue perfusion, fixation, clearing, antibody staining, refractive index matching (RIM) (), microscopy, and imaging. The timing for each step varies depending on the size, weight, and type of tissue, as well as the type of immunostaining. We provide an example of the FACT protocol using mouse brain tissue, which demonstrates its suitability for molecular interrogation analysis of large tissues. The FACT technique has been successfully performed on different types of tissues, making it a favorable choice for a variety of applications.

Correlative light-electron microscopy (CLEM) has evolved in the last decades, especially after significant developments in sample preparation, imaging acquisition, software, spatial resolution, and equipment, including confocal, live-cell, super-resolution, and electron microscopy (scanning, transmission, focused ion beam, and cryo-electron microscopy). However, the recent evolution of different laser-related techniques, such as mass spectrometry imaging (MSI) and laser capture microdissection, could further expand spatial imaging capabilities into high-resolution OMIC approaches such as proteomic, lipidomics, small molecule, and drug discovery. Here, we will describe a protocol to integrate the detection of rare viral reservoirs with imaging mass spectrometry.

Probe-based confocal laser endoscopy (pCLE) has emerged as a powerful tool for disease diagnosis, yet it faces challenges such as the formation of hexagonal patterns in images due to the inherent characteristics of fiber bundles. Recent advancements in deep learning offer promise in image denoising, but the acquisition of clean-noisy image pairs for training networks across all potential scenarios can be prohibitively costly. Few studies have explored training denoising networks on such pairs. Here, we propose an innovative self-supervised denoising method. Our approach integrates noise prediction networks, image quality assessment networks, and denoising networks in a collaborative, jointly trained manner. Compared to prior self-supervised denoising methods, our approach yields superior results on pCLE images and fluorescence microscopy images. In summary, our novel self-supervised denoising technique enhances image quality in pCLE diagnosis by leveraging the synergy of noise prediction, image quality assessment, and denoising networks, surpassing previous methods on both pCLE and fluorescence microscopy images.

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OBJECTIVE: Defining how the in vivo immune status of peripheral tissues is shaped by the external environment has remained a technical challenge. We recently developed Functional in vivo confocal microscopy (Fun-IVCM) for dynamic, longitudinal imaging of corneal immune cells in living humans. This study investigated the effect of seasonal-driven environmental factors on the morphodynamic features of human corneal immune cell subsets.
METHODS: Longitudinal, observational clinical study.
METHODS: Sixteen healthy participants (aged 18-40 years) attended 2 visits in distinct seasons in Melbourne, Australia (Visit 1, November-December 2021 [spring-summer]; Visit 2, April-June 2022 [autumn-winter]).
METHODS: Environmental data were collected over each period. Participants underwent ocular surface examinations and corneal Fun-IVCM (Heidelberg Engineering). Corneal scans were acquired at 5.5 ± 1.5-minute intervals for up to 5 time points. Time-lapse Fun-IVCM videos were created to analyze corneal immune cells, comprising epithelial T cells and dendritic cells (DCs), and stromal macrophages. Tear cytokines were analyzed using a multiplex bead-based immunoassay.
METHODS: Difference in the density, morphology, and dynamic parameters of corneal immune cell subsets over the study periods.
RESULTS: Visit 1 was characterized by higher temperature, lower humidity, and higher air particulate and pollen levels compared with Visit 2. Clinical ocular surface parameters and the density of immune cell subsets were similar across visits. At Visit 1 , corneal epithelial DCs were larger, with a lower dendrite probing speed (0.38 ± 0.21 vs. 0.68 ± 0.33 μm/min; P < 0.001) relative to Visit 2; stromal macrophages were more circular and had less dynamic activity (Visit 1, 7.2 ± 1.9 vs. Visit 2, 10.3 ± 3.7 dancing index; P < 0.001). Corneal T cell morphodynamics were unchanged across periods. Basal tear levels of interleukin 2 and CXCL10 were relatively lower during spring-summer.
CONCLUSIONS: This study identifies that the in vivo morphodynamics of innate corneal immune cells (DCs, macrophages) are modified by environmental factors, but such effects are not evident for adaptive immune cells (T cells). The cornea is a potential in vivo window to investigate season-dependent environmental influences on the human immune system.
BACKGROUND: Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Plant small RNAs are 21-24 nucleotide, noncoding RNAs that function as regulators in plant growth and development. Colorimetric detection of plant small RNAs was made possible with the introduction of locked nucleic acid probes. However, fluorescent detection of plant small RNAs has been challenging due to the high autofluorescence from plant tissue. Here we report a fluorescent in situ detection method for plant small RNAs. This method can be applied to most plant samples and tissue types and also can be adapted for single-molecule detection of small RNAs with super-resolution microscopy.

Actinic keratosis (AK) is the most common pre-malignant cutaneous lesion of the skin, often associated with field cancerization. Daylight photodynamic therapy (DL-PDT) is used as treatment, showing good histological results. Reflectance confocal microscopy (RCM) may be useful as a non-invasive, real-time approach to monitor treatment, however, there is a lack of data on the correlation between RCM and histopathological findings in AK patients treated with DL-PDT. To correlate histological and RCM findings and evaluate the efficacy of DL-PDT in patients with AK and field cancerization treated with DL-PDT. Patients with field cancerization and a minimum of six AK lesions on the face were included in the study. A single session combining methyl aminolevulinate followed by two-hour daylight exposure of the face was performed. RCM and biopsy were performed before and after three months of the intervention to compare efficacy between patients using the Wilcoxon test, and concordance of the findings based on the different methods was analysed using the Kappa test. Twenty-four patients completed the study. An improvement in photodamage and a decrease in the number of AK lesions (45.3% reduction) was observed. Regression in atypia and dysplasia was observed via histopathology and RCM, however, there was poor agreement between the methods. No changes were observed after treatment for inflammation, fibroplasia and acantholysis. Concordance between histological and RCM findings was poor, suggesting that RCM cannot replace the histopathological examination, however, it may be used as an adjuvant test for follow-up of patients. Despite this, DL-PDT proved to be an effective method for treating AK.