Search

In approaches to analyze material composition or in processing tasks using ultra-short laser ablation, it is of particular interest how ablated materials are distributed across the solid angle in front of the interaction region. We found that with our sub-10-fs laser in the regime from 10 14 W / c m 2 to 10 17 W / c m 2 , the solid angle of the ablation cone decreases significantly along with the laser intensity in a vacuum environment. For this observation, we used ion-beam analysis to investigate the distribution of tungsten collected on silicon catcher plates arranged across the solid angle of the laser-ablation cone. Moreover, we used other post-mortem tools, such as scanning electron microscopy and confocal laser scanning microscopy, to determine the ablation threshold of F t h = 468 mJ / c m 2 of our tungsten samples. Here, clearly, two laser intensity-dependent ablation regimes can be observed in the detection of a crater depth and a ablation cone angle. [ABSTRACT FROM AUTHOR]

Mitochondrial abnormalities underscore a variety of neurologic injuries and diseases and are well-studied in adult populations. Clinical studies identify critical roles of mitochondria in a wide range of developmental brain injuries, but models that capture mitochondrial abnormalities in systems representative of the neonatal brain environment are lacking. Here, we develop an organotypic whole-hemisphere (OWH) brain slice model of mitochondrial dysfunction in the neonatal brain. We extended the utility of complex I inhibitor rotenone (ROT), canonically used in models of adult neurodegenerative diseases, to inflict mitochondrial damage in OWH slices from term-equivalent rats. We quantified whole-slice health over 6 days of exposure for a range of doses represented in ROT literature. We identified 50 nM ROT as a suitable exposure level for OWH slices to inflict injury without compromising viability. At the selected exposure level, we confirmed exposure- and time-dependent mitochondrial responses showing differences in mitochondrial fluorescence and nuclear localization using MitoTracker imaging in live OWH slices and dysregulated mitochondrial markers via RT-qPCR screening. We leveraged the regional structures present in OWH slices to quantify cell density and cell death in the cortex and the midbrain regions, observing higher susceptibilities to damage in the midbrain as a function of exposure and culture time. We supplemented these findings with analysis of microglia and mature neurons showing time-, region-, and exposure-dependent differences in microglial responses. We demonstrated changes in tissue microstructure as a function of region, culture time, and exposure level using live-video epifluorescence microscopy of extracellularly diffusing nanoparticle probes in live OWH slices. Our results highlight severity-, time-, and region-dependent responses and establish a complimentary model system of mitochondrial abnormalities for high-throughput or live-tissue experimental needs. [ABSTRACT FROM AUTHOR]

Background: One of the most severe consequences of ageing is cognitive decline, which is associated with dysfunction of the brain microvasculature. Thus, repairing the brain vasculature could result in healthier brain function. Methods: To better understand the potential beneficial effect of endothelial progenitor cells (EPCs) in vascular repair, we studied the adhesion and integration of EPCs using the early embryonic mouse aorta–gonad–mesonephros – MAgEC 10.5 endothelial cell line. The EPC interaction with brain microvasculature was monitored ex vivo and in vivo using epifluorescence, laser confocal and two-photon microscopy in healthy young and old animals. The effects of senolysis, EPC activation and ischaemia (two-vessel occlusion model) were analysed in BALB/c and FVB/Ant: TgCAG-yfp_sb #27 mice. Results: MAgEC 10.5 cells rapidly adhered to brain microvasculature and some differentiated into mature endothelial cells (ECs). MAgEC 10.5-derived endothelial cells integrated into microvessels, established tight junctions and co-formed vessel lumens with pre-existing ECs within five days. Adhesion and integration were much weaker in aged mice, but were increased by depleting senescent cells using abt-263 or dasatinib plus quercetin. Furthermore, MAgEC 10.5 cell adhesion to and integration into brain vessels were increased by ischaemia and by pre-activating EPCs with TNFα. Conclusions: Combining progenitor cell therapy with senolytic therapy and the prior activation of EPCs are promising for improving EPC adhesion to and integration into the cerebral vasculature and could help rejuvenate the ageing brain. [ABSTRACT FROM AUTHOR]

Lentigo maligna (LM) is a growing problem worldwide and the main type of melanoma in situ in some Caucasian populations. It presents as a spectrum from atypical intraepidermal melanocytic proliferation (AIMP) to invasive lentigo maligna melanoma (LMM). Accurate diagnosis and staging are crucial for determining appropriate management strategies. To assess the role of in vivo reflectance confocal microscopy (RCM) in differentiating early and advanced stages of lentigo maligna. A systematic search was conducted on Medline, PubMed, Scopus, Web of Science, Proquest Central, Embase, Cochrane and Google Scholar. References of included and excluded studies were reviewed for additional sources. Studies involving RCM for AIMP, LM and LMM diagnosis published until 1 December 2023, were selected for analysis. Publications on other non‐invasive imaging techniques, other skin diseases or those addressing only RCM surgical margins and therapeutic response without reporting RCM features were excluded. Each article was reviewed by two experts, with disagreements resolved by a third expert. Data for each RCM feature were pooled, and odds ratios (ORs) and 95% confidence intervals (CIs) were calculated. Features were stratified for architecture, cellular morphology and distribution. Twenty‐seven articles involving 303 lesions were included. Features favouring LM over AIMP included (1) Architecture: the presence of non‐edged papillae (OR 4.50, 95% CI 1.92–10.52) and (2) Cellular distribution: widespread pagetoid cells and junctional atypical cells (OR 25.06, 95% CI 4.19–148.45) and junctional nests (OR 18.06, 95% CI 3.04–106.32). Features favouring LMM over LM included (1) Architecture: epidermal disarray (OR 5.03, 95% CI 1.90–13.22), junctional disarray (OR 4.19, 95% CI 1.54–11.33) and destroyed collagen fibres (OR 3.91, 95% CI 1.80–8.44) and (2) cellular distribution: widespread pagetoid and junctional atypical cells (OR 2.71, 95% CI 1.27–5.75) and junctional nests (OR 4.48, 95% CI 2.12–9.43). This study identifies the RCM features associated with the LM/LMM spectrum. [ABSTRACT FROM AUTHOR]

Background and Aims Methods Results Interpretation Small‐fibre neuropathy (SFN) is associated with glucose dysregulation, including impaired glucose tolerance (IGT) and type 2 diabetes (T2D). Corneal confocal microscopy (CCM) offers a non‐invasive tool to assess corneal nerve damage and dendritic cell density (DCD). In this study, we investigated corneal DCD in patients with SFN and glucose dysregulation, defined as IGT or T2D.We enrolled 38 patients with SFN + glucose dysregulation, 51 with SFN + non‐glucose dysregulation and 20 healthy controls. All participants underwent neurological examination, neurophysiology and CCM.Individuals with SFN and glucose dysregulation had higher DCD compared with healthy controls (p = .01), and mature DCD was higher in IGT SFN patients than in T2D patients.Higher DCD in IGT compared with controls and patients with established T2D may suggest that DCD is a biomarker of early neuropathy. [ABSTRACT FROM AUTHOR]

The article discusses the use of Line-field confocal optical coherence tomography (LC-OCT) in diagnosing alopecia areata (AA) and correlating it with histopathological and dermoscopic findings. The study examined 20 patients with severe AA, revealing specific characteristics such as hyperreflective amorphous material in dilated hair follicles. LC-OCT was found to be a valuable tool for diagnosing and monitoring AA, providing real-time in vivo images with nearly histological resolution. The research contributes to understanding the pathophysiology of AA and may aid in early diagnosis and treatment decisions. [Extracted from the article]

Improving light‐trapping capabilities through surface microstructuring of transparent conductive oxides is a promising approach to enhance solar cell efficiency. This study focuses on treating fluorine‐doped tin oxide (FTO) thin films using four‐beam direct laser interference patterning (DLIP) to create dot‐like periodic surface microstructures. The surface analysis using scanning electron microscopy and confocal microscopy reveals the presence of a periodic square grid of microcraters with a spatial period of ≈700 nm and an average depth ranging between 4 and 18 nm. These structures enhance the dispersion of incoming light up to 1000% in the visible and NIR spectra. When integrated into metal halide perovskite solar cells, FTO films patterned using low fluence conditions lead to a notable increase in the power conversion efficiencies (PCEs) compared to those made using untreated FTO. Importantly, preliminary stability tests on devices based on patterned FTO substrates show significantly improved stability compared to those fabricated using reference unpatterned substrates. These findings demonstrate that a DLIP treatment of FTO substrates is a promising technique that can substantially enhance the efficiency and stability of perovskite photovoltaic devices. [ABSTRACT FROM AUTHOR]

In addressing the critical limitations associated with platelet storage, the study investigates an innovative solution aimed at preventing the unwanted activation of platelets within storage bags. This activation is a key factor that currently restricts the shelf life of platelet products to only 72 h, leading to extreme waste production and high costs. Here, highly effective surfaces are identified for minimizing surface‐induced platelet activation. Using thermal nanoimprint lithography (T‐NIL), a new method is demonstrated for patterning reproducible agarose micro/nanostructures (a natural hydrogel with anti‐platelet adhesive properties) including dots, chains, pills, and squares. The agarose (3%) structured surfaces displayed outstanding flexibility and hydrophilic behavior that prevented platelet adhesion as confirmed by confocal microscopy. Importantly, pill‐shaped structures effectively maintained their ability to prevent platelet adhesion, even after a long cell‐contacting duration. Atomic force microscopy indicated that the effectiveness of dampening platelet adherence is determined by the shape, size, height, and aspect ratio of the structures. A model is provided to explain how the different shapes affect wettability and thereby hinder platelet adherence. The developed anti‐adhesive agarose structured surfaces show promise to revolutionize platelet storage, provide vital insights into biomaterials research, and demonstrate the potential of tailored agarose surfaces in biomedical applications. [ABSTRACT FROM AUTHOR]

Cnidarians are the first phylum of metazoans to exhibit a functional mucus layer. In evolution, mucous cells perform the classical secretory function of mucus and act as sentinels for external insults, particularly in mammals. However, this topic has not been much analysed in the lower metazoans. This study aims to investigate the role of mucous cells in Actinia equina (Anthozoa, Hexacorallia) (Linnaeus, 1758) by using confocal microscopy and bioinformatics. The results show colocalization of Toll‐like receptor 2 (TLR2) and Mucin2 (MUC2) on these cells, suggesting a crosstalk between defence response and mucus secretion in cnidarians. Furthermore, the presence of TLR2 on the surface of mucous cells suggests their involvement in host defence responses, as occurs in vertebrates. Moreover, through in silico analysis, we confirmed the presence of orthologous genes for the tested antibodies, providing additional evidence to support our results from an evolutionary perspective. These data suggest that mucous cells play a defensive role throughout evolution, helping us understand how these cells have developed and adapted across phylogenesis. This information also provides new insights into the defensive strategies of invertebrates and clarifies their evolutionary history. [ABSTRACT FROM AUTHOR]

Hemp fiber is one of the most common reinforcements for polymer matrix composites and it is widely used in various applications due to its environmentally friendliness, favorable physical and mechanical properties. In spite of the numerous merits of the hemp fiber reinforced polymer composites, improvement of poor interfacial properties between the hemp fiber and polymer matrix is limited. The main objective of this work is to investigate the effect of increased fiber surface area and alkaline surface treatment on the mechanical performance of hemp fiber reinforced epoxy composites. An elevated temperature aqueous debundling treatment was employed to generate a large number of individual fibers from fiber bundles and was associated with a threefold increase in specific fiber surface area. The fiber dimension and surface properties were investigated through optical microscopy, confocal microscopy, and scanning electron microscopy. The effects of different debundling treatment times and additional alkaline treatment on the mechanical properties were studied. It was found that composites reinforced by the debundled hemp fiber with further alkaline treatment exhibited the best tensile and flexural properties in comparison to other composites reinforced with debundled hemp fiber without further alkaline treatment or with untreated hemp fiber. Highlights: Provided a new physical method for separating hemp fiber bundle into individual hemp fibers using boiling water and blending.Debundled fibers resulted in a 3.5 times increase in fiber surface area for better bonding with epoxy matrix.Debundling process combined with chemical (alkaline) treatment to further improve the compatibility between individual fiber and epoxy.The effects of the debundling only, alkaline treatment only, and combined debundling + alkaline treatment on composite properties were studied. [ABSTRACT FROM AUTHOR]

Droplet digital PCR (ddPCR) is recognized as a high-precision method for nucleic acid quantification, extensively utilized in biomedical research and clinical diagnostics. This technique employs microfluidic technology to partition the nucleic acid-containing reaction mixture into discrete droplets for amplification, achieving absolute quantification by identifying and enumerating the number of fluorescent droplets. The accuracy of droplet quantification is pivotal to the success of the assay. However, current image-processing tools are operationally complex, and commercial instruments are costly. Moreover, the designed algorithms exhibit a need for enhanced accuracy and are often restricted to use by trained personnel with specific microscopy equipment. In response to these challenges, we introduce an automated device (A-MMD), capable of detecting fluorescent droplets in ddPCR images captured by multiple microscopes. The device integrates three distinct algorithms tailored for the image processing of Laser Scanning Confocal Microscopy (LSCM), inverted microscopy, and self-assembled microscopy. Experimental validation using λ DNA demonstrated a 100.00% identification rate for positive droplets across all three image types, and the average identification rates for total droplets being 99.27% for LSCM, 98.96% for inverted microscopy, and 99.08% for self-assembled microscopy. Furthermore, the A-MMD is equipped with a user-friendly interface (UI) that streamlines the operational process, enabling non-specialists to efficiently perform droplet detection tasks. Our device not only has good environmental adaptability and identification accuracy, but also significantly reduces costs and operational complexity. It offers an economical, efficient, and user-friendly solution for ddPCR image analysis, thereby further propelling the advancement and application of nucleic acid detection technology. [ABSTRACT FROM AUTHOR]

Researchers have shown significant interest in three-dimensional DNA building blocks due to their potential applications in biomedicine and biosensing. This study focuses on the synthesis of an HgII ion-stabilized DNA capsule with T-HgII-T pairs for the purpose of detecting melamine (MA). MA reacts with HgII to form a MA-HgII-MA complex, which causes HgII to leave the capsule shell, ultimately leading to capsule collapse and release of fluorescent cargo as output signal. Density functional theory (DFT) calculations and X-ray absorption spectroscopy (XAS) were used to demonstrate the ability of MA to extract HgII from the T-HgII-T adducts. The DNA capsules were characterized using TEM, SEM, DLS, zeta-potential, and melting curve analysis, which indicated the successful construction of the HgII-intercalated DNA shell. The MA-triggered destruction of the DNA capsules was visualized by confocal microscopy, and the dynamics of decapsulation were evaluated through fluorescent cargo release. The HgII-stabilized DNA capsules enable MA detection with a detection limit of 0.037 µM and are insensitive to potential interfering ions and amino acids. The tests conducted using MA spiked milk solution resulted in recoveries ranging from 109 to 113% (0.1 µM) and 94.5 to 96% (0.5 µM). These results suggest that the system is promising for highly accurate and reproducible monitoring of MA adulteration. [ABSTRACT FROM AUTHOR]

The reflectance confocal microscopy (RCM)–optical coherence tomography (OCT) device has shown utility in detecting and assessing the depth of basal cell carcinoma (BCC) in vivo but is challenging for novices to interpret. Artificial intelligence (AI) applied to RCM-OCT could aid readers. We trained AI models, using OCT rasters of biopsy-confirmed BCC, to detect BCC, create three-dimensional rendering and automatically measure tumour depth. Trained AI models were applied to a separate test set containing rasters of BCC, benign lesions, and healthy skin. Blinded reader analysis and tumour depth correlation with histopathology were conducted. BCC detection improved from viewing OCT rasters only (sensitivity 73.3%, specificity 45.5%) to viewing rasters with AI-generated BCC rendering (sensitivity 86.7%, specificity 48.5%). A Pearson correlation r 2 = 0.59 (P = 0.02) was achieved for the tumour depth measurement between AI and histological measured depths. Thus, addition of AI to the RCM-OCT device may expand its utility widely. [ABSTRACT FROM AUTHOR]

Confocal Laser Scanning Microscopy (CLSM) is an advanced imaging technique used in various fields. It offers several advantages over conventional microscopy due to its high resolution and ability to collect three-dimensional data using optical sectioning. While the resolution in confocal microscopy is comparatively lower than that of electron microscopy, it has effectively bridged the gap between these two imaging techniques. In the medical field, CLSM has a wide range of applications, including the diagnosis and treatment of corneal diseases, the study of biofilms and the diagnosis of various types of cancers, as well as the viral pathogenic cycle, including that of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). CLSM is also beneficial in food technology. The present review synthesises the current knowledge on CLSM, encompassing its applications, technological advancements and limitations. A comprehensive search of relevant databases identified studies and articles published by September 2023. The findings highlight CLSM's versatility, with applications ranging from cell biology to materials science, while also addressing technical challenges and future prospects. [ABSTRACT FROM AUTHOR]

The structure of aspirated coronary thrombus in ST-segment elevation myocardial infarction (STEMI) is still being studied. Our aims were to characterize coronary thrombi of different ages, focusing on the appearance of activated protein C (APC/PC) and its relation to the elements of neutrophil extracellular traps (NETs), and the factors closely related to fibrin as factor XIII (FXIII) and α2 plasmin inhibitor (α2-PI). The thrombi of n = 24 male patients with atherosclerotic coronary plaque rupture related to native coronary artery occlusion were selected for histopathology analysis. Thrombus age was distinguished as fresh, lytic, and organized, and then analyzed by immunofluorescent staining and confocal microscopy. FXIII was present at a high level and showed a high degree of co-localization with fibrin in all stages of thrombus evolution. The amount of α2-PI was low in the fresh thrombi, which increased significantly to the lytic phase. It was evenly distributed and consistently associated with fibrin. APC/PC appeared in the fresh thrombus and remained constant during its evolution. The presence of NET marker and CD66b was most dominant in the lytic phase. APC/PC co-localization with the elements of NET formation shows its role in NET degradation. These observations suggest the importance of searching for further targeted therapeutic strategies in STEMI patients. [ABSTRACT FROM AUTHOR]

The eye, and the cornea in particular, is a common site of chemotherapy induced toxicity, and ocular side effects of both traditional and novel agents have been reported. Corneal confocal microscopy (CCM) is an in vivo technique that allows for the study of all the corneal layers in an easy, non-invasive and reproducible way via the direct visualization of corneal cell morphologies as well as of sub-basal nerve plexus. Thus, it represents a useful way to identify and monitor chemotherapy induced corneal alterations. This work aims to review the use of CCM in identifying corneal toxicity secondary to chemotherapy treatment, as regards both corneal nerves alterations in the setting of chemotherapy induced peripheral neuropathy (CIPN) and other corneal structure changes, particularly involving the corneal epithelium. [ABSTRACT FROM AUTHOR]

Traditionally, firearm and toolmark examiners manually evaluate the similarity of features on two bullets using comparison microscopy. Advances in microscopy have made it possible to collect 3D topographic data, and several automated comparison algorithms have been introduced for the comparison of bullet striae using these data. In this study, open‐source approaches for cross‐correlation, congruent matching profile segments, consecutive matching striations, and a random forest model were evaluated. A statistical characterization of these automated approaches was performed using four datasets of consecutively manufactured firearms to provide a challenging comparison scenario. Each automated approach was applied to all samples in a pairwise fashion, and classification performance was compared. Based on these findings, a Bayesian network was empirically learned and constructed to leverage the strengths of each individual approach, model the relationship between the automated results, and combine them into a posterior probability for the given comparison. The network was evaluated similarly to the automated approaches, and the results were compared. The developed Bayesian network classified 99.6% of the samples correctly, and the resultant probability distributions were significantly separated more so than the automated approaches when used in isolation. [ABSTRACT FROM AUTHOR]

Objectives: To test the performance of ex vivo fluorescence confocal microscopy (FCM; Vivascope 2500M‐G4), as compared to intra‐operative frozen section (IFS) analysis, to evaluate surgical margins during robot‐assisted radical prostatectomy (RARP), with final pathology as the reference standard. Methods: Overall, 54 margins in 45 patients treated with RARP were analysed with: (1) ex vivo FCM; (2) IFS analysis; and (3) final pathology. FCM margins were evaluated by two different pathologists (experienced [M.I.: 10 years] vs highly experienced [G.R.: >30 years]) as strongly negative, probably negative, doubtful, probably positive, or strongly positive. First, inter‐observer agreement (Cohen's κ) between pathologists was tested. Second, we reported the sensitivity, specificity, positive predictive (PPV) and negative predictive value (NPV) of ex vivo FCM. Finally, agreement between ex vivo FCM and IFS analysis (Cohen's κ) was reported. For all analyses, four combinations of FCM results were evaluated. Results: At ex vivo FCM, the inter‐observer agreement between pathologists ranged from moderate (κ = 0.74) to almost perfect (κ = 0.90), according to the four categories of results. Indeed, at ex vivo FCM, the highly experienced pathologist reached the best balance between sensitivity (70.5%) specificity (91.8%), PPV (80.0%) and NPV (87.1%). Conversely, on IFS analysis, the sensitivity, specificity, PPV and NPV were, respectively, 88.2% vs 100% vs 100% vs 94.8%. The agreement between the ex vivo FCM and IFS analyses ranged from moderate (κ = 0.62) to strong (κ = 0.86), according to the four categories of results. Conclusion: Evaluation of prostate margins at ex vivo FCM appears to be feasible and reliable. The agreement between readers encourages its widespread use in daily practice. Nevertheless, as of today, the performance of FCM seems to be sub‐par when compared to the established standard of care (IFS analysis). [ABSTRACT FROM AUTHOR]

Intraoperative frozen section (IFS) is used with the intention to improve functional and oncological outcomes for patients undergoing radical prostatectomy (RP). High resource requirements of IFS techniques such as NeuroSAFE may preclude widespread adoption, even if there are benefits to patients. Recent advances in fresh‐tissue microscopic digital imaging technologies may offer an attractive alternative, and there is a growing body of evidence regarding these technologies. In this narrative review, we discuss some of the familiar limitations of IFS and compare these to the attractive counterpoints of modern digital imaging technologies such as the speed and ease of image generation, the locality of equipment within (or near) the operating room, the ability to maintain tissue integrity, and digital transfer of images. Confocal laser microscopy (CLM) is the modality most frequently reported in the literature for margin assessment during RP. We discuss several imitations and obstacles to widespread dissemination of digital imaging technologies. Among these, we consider how the 'en‐face' margin perspective will challenge urologists and pathologists to understand afresh the meaning of positive margin significance. As a part of this, discussions on how to describe, categorize, react to, and evaluate these technologies are needed to improve patient outcomes. Limitations of this review include its narrative structure and that the evidence base in this field is relatively immature but developing at pace. [ABSTRACT FROM AUTHOR]

Pilostyles, an endoparasitic genus within the Apodanthaceae family, grows inside host stems with flowers and fruits being the only external manifestations. Previous studies of P. berteroi growing on Adesmia trijuga provided limited details of the endophyte and omitted the origin of flowers and sinker structure. This study, using classical methods of optical microscopy applied to the analysis with scanning electron microscopy and confocal laser scanning microscopy, expands the understanding of the P. berteroi/A. trijuga complex. We find that P. berteroi develops isophasically with its host, forming endophytic patches between the host's secondary phloem cells. The parasitized Adesmia stem's cambium primarily produces xylem parenchyma, with limited vessel production and halting fiber formation. The radial polarization of endophytic patches led to the formation of floral meristems. Flowers develop endogenously and emerge by the breakthrough of the host stem. Flowers are connected to the host cambium via chimeric sinkers, combining P. berteroi parenchyma and tracheoids with Adesmia vessels. Unlike previous studies that show uniformity among Pilostyles species, our analysis reveals new insights into the structural interaction between P. berteroi and A. trijuga. [ABSTRACT FROM AUTHOR]

Introduction and Objectives: Prostate apoptosis response protein-4 (PAR-4) is considered a tumor suppressor. However, the role of PAR-4 in hepatocellular carcinoma (HCC) has rarely been reported. The study explores the role of PAR-4 in the malignant behaviors of HCC cells. Materials and Methods: TCGA database was applied to analyze the expression of PAR-4 in HCC. Evaluated PAR-4 relationship with clinical parameters and prognosis by tissue microarray; expression of STAT3, p-STAT3, Src and Ras was detected by Western blotting or laser confocal microscopy. Cell scratch and flow cytometry assays were used to observe IL-6 regulation of the malignant behaviors of HCC cells. The tumorigenic potential of HCC cells in vivo was evaluated in a nude mouse tumor model. Results: Analysis indicated that the expression of PAR-4 in HCC tissues was significantly higher than that in normal liver tissues; and PAR-4 interacted with STAT3. KEGG analysis showed that PAR-4 plays a role in the Janus kinase (JAK)/STAT signaling pathway. The positive expression rate of PAR-4 in HCC tissues was significantly higher than that in adjacent tissues. Positive correlation between IL-6 and PAR-4 expression in the HCC tissues. Exogenous IL-6 significantly promoted the proliferation and migration of HCC cells and up-regulated the expression of PAR-4 and p-STAT3 in HCC cells. Interference of the expression of PAR-4 could reduce the malignant behaviors of HCC cells and inhibit tumorigenesis in a nude mouse tumor model. Conclusions: PAR-4 expression is positively correlated with HCC; PAR-4 promotes malignant behavior of HCC cells mediated by the IL-6/STAT3 signaling pathway. [ABSTRACT FROM AUTHOR]

Lipid nanoparticles (LNPs) are the most clinically advanced RNA delivery technology, but their efficiency is limited by low RNA release after endosome disruption. To improve RNA release, an acid‐responsive polymer is synthesized with which to formulate LNPs for RNA encapsulation and release. Specifically, three acid‐responsive poly(lactic acid)‐block‐poly(carboxybetaine) zwitterionic derivatives are designed and synthesized that are cationic and complexed with RNA at pH 7.4, but are neutral following cleavage at endosomal pH, thereby having lower affinity to RNA. The polymers are formulated into each of the clinically approved Onpattro, Moderna, or Pfizer LNP formulations to produce hybrid polymer‐lipid nanoparticles (PLNPs). With the PLNPs, the IC50 values of multiple small interfering RNAs (siRNAs) decreased up to 5.4‐fold compared to parent LNPs in several cell lines. Moreover, messenger RNA (mRNA) transfection increased up to two fold. The acid‐responsive polymers in PLNPs accounted for the enhanced RNA transfection as this phenomenon is lost with acid‐inert polymers. Confocal microscopy confirmed that cytosolic RNA concentration increased using the acid‐responsive polymers; conversely, uptake and endosomal escape are identical to existing LNPs. This confirmed that enhanced RNA transfection is due to increased RNA dissociation from its carrier. The novel polymer represents a versatile strategy to increase RNA transfection from LNPs. [ABSTRACT FROM AUTHOR]

A common problem in confocal microscopy is the decrease in intensity of excitation light and emission signal from fluorophores as they travel through 3D specimens, resulting in decreased signal detected as a function of depth. Here, we report a visualization program compatible with widely used fluorophores in cell biology to facilitate image interpretation of differential protein disposition in 3D specimens. Glioblastoma cell clusters were fluorescently labeled for mitochondrial complex I (COXI), P2X7 receptor (P2X7R), β-Actin, Ki-67, and DAPI. Each cell cluster was imaged using a laser scanning confocal microscope. We observed up to ~70% loss in fluorescence signal across the depth in Z-stacks. This progressive underrepresentation of fluorescence intensity as the focal plane deepens hinders an accurate representation of signal location within a 3D structure. To address these challenges, we developed ProDiVis: a program that adjusts apparent fluorescent signals by normalizing one fluorescent signal to a reference signal at each focal plane. ProDiVis serves as a free and accessible, unbiased visualization tool to use in conjunction with fluorescence microscopy images and imaging software. [ABSTRACT FROM AUTHOR]

Biopsy-based histology has been the foundation of disease diagnosis and management for over a century. A long-sought goal in dermatology is the development of an imaging modality with sufficient resolution and compositional detail to noninvasively interrogate skin histology in vivo. Here, we describe a system that achieves this goal using cross-sectionally scanned, multimodal microscopy (cross-modal). Cross-modal combines multiphoton and reflectance confocal microscopy into one compact system with coordinated three-axis scanning that preserves optical resolution in cross-section. A custom pinhole-free mechanism employing finite-infinite conjugates further simplifies and stabilizes confocal alignment. Evaluated in participants ages 9–81 and Fitzpatrick skin types (FST) 1–5, cross-modal images revealed histological details analogous to those obtained from traditional biopsied tissue. We observed dermal elastosis in sun-damaged skin, elevated melanin in pigmented skin, basaloid nests in basal cell carcinoma, and elongated rete ridges in seborrheic keratosis, supporting cross-modal's potential to deliver histological insights noninvasively. [ABSTRACT FROM AUTHOR]

The correction notice in the Scientific Reports journal addresses errors in the original article regarding the assembly of figures and images. Specifically, Figure 4B and supplementary Figures S4A, S4C, S4B, and S4D contain overlapping images due to errors during preparation and assembly. The corrected figures show the effect of temperature on exosome uptake by cancer cells and the impact of different uptake inhibitors on labeled exosomes. The authors of the article are Sherif E. Emam, Hidenori Ando, Amr S. Abu Lila, Taro Shimizu, Keiichiro Okuhira, Yu Ishima, Mahmoud A. Mahdy, Fakhr-eldin S. Ghazy, Ikuko Sagawa, and Tatsuhiro Ishida. [Extracted from the article]

Background: Diabetic peripheral neuropathy (DPN) is common and can go unnoticed until it is firmly developed. This study aims to establish a transformer-based deep learning algorithm (DLA) to classify corneal confocal microscopy (CCM) images, identifying DPN in diabetic patients. Methods: Our classification model differs from traditional convolutional neural networks (CNNs) using a Swin transformer network with a hierarchical architecture backbone. Participants included those with (DPN+, n = 57) or without (DPN-, n = 37) DPN as determined by the updated Toronto consensus criteria. The CCM image dataset (consisting of 570 DPN+ and 370 DPN- images, with five images selected from each participant's left and right eyes) was randomly divided into training, validation, and test subsets at a 7:1: 2 ratio, considering individual participants. The effectiveness of the algorithm was assessed using diagnostic accuracy measures, such as sensitivity, specificity, and accuracy, in conjunction with Grad-CAM visualization techniques to interpret the model's decisions. Results: In the DPN + group (n = 12), the transformer model successfully predicted all participants, while in the DPN- group (n = 7), one participant was misclassified as DPN+, with an area under the curve (AUC) of 0.9405 (95% CI 0.8166, 1.0000). Among the DPN + images (n = 120), 117 were correctly classified, and among the DPN- images (n = 70), 49 were correctly classified, with an AUC of 0.8996 (95% CI 0.8502, 0.9491). For single-image predictions, the transformer model achieved a superior AUC relative to the ResNet50 model (0.8761, 95% CI 0.8155, 0.9366), the Inception_v3 model (0.8802, 95% CI 0.8231, 0.9374), and the DenseNet121 model (0.8965, 95% CI 0.8438, 0.9491). Conclusion: Transformer-based networks outperform CNN-based networks in rapid binary DPN classification. Transformer-based DLAs have clinical DPN screening potential. [ABSTRACT FROM AUTHOR]

Senescence, caused by cell‐cycle arrest, is a hallmark of aging. Senescence has also been described in embryogenesis, wound healing, and acute injuries. Sepsis is characterized by a dysregulated host response to infection, leading to organ dysfunction and mortality. Most of the pathophysiology of human sepsis is recapitulated in the mouse model of polymicrobial sepsis, developed by cecal ligation and puncture (CLP). In this report, we demonstrate a rapid onset of cellular senescence in the liver of mice subjected to CLP‐induced sepsis, characterized by the upregulation of p21, p53, and other senescence markers, including SA‐βgal. Using RNAscope, confocal microscopy, and flow cytometry, we further confirm the emergence of p21‐expressing senescence phenotype in the liver 24 h after sepsis induction. Senescence was observed in several cell types in the liver, including hepatocytes, endothelial cells, and macrophages. We determined the landscape of senescence phenotype in murine sepsis by single‐cell sequencing, which further ascertained that this cell fate is not confined to any particular cell type but displays a heterogeneous distribution. Furthermore, we observed a significant reduction in mortality following sepsis when mice were treated with senolytics, a combination of dasatinib and quercetin, before the CLP surgery. Our experiments unequivocally demonstrated a rapid development of cellular senescence with sepsis and, for the first time, described the senescence landscape in the sepsis liver and the possible role of senescent cells in the worsening outcome following sepsis. [ABSTRACT FROM AUTHOR]

PurposeMethodsResultsConclusionsTo compare corneal endothelial changes in patients with Fuchs Uveitis Syndrome (FUS) undergoing phacoemulsification surgery using confocal and specular microscopy.We included 14 patients with unilateral FUS and cataracts who underwent phacoemulsification surgery in a Mexican referral center for inflammatory eye diseases. Preoperative confocal and specular microscopies were conducted, establishing baseline images for subsequent analyses. Surgery on the FUS eye was performed by a single surgeon and an intraocular lens was implanted in all cases. Both specular and confocal microscopy were repeated 6 months after FUS eye surgery and compared with baseline images. We used Image J to do a manual segmentation of KP and determine their density for further analysis hence developing a new tool for confocal microscopy image analysis.All patients underwent uneventful phacoemulsification surgery in the FUS eye. There was no significant change in endothelial cell density (ECD) from 2257 (±508.2) cells/mm2 preoperatively to 2214 (±535.1) cells/mm2 postoperatively (p = 0.809). Confocal microscopy revealed a decrease in Keratic Precipitate Density (KPD) from a median of 1413 (±2809.7) KPs/mm2 preoperatively to a median of 685.5 (1527.9) KPs/mm2 postoperatively (p = 0.036).Phacoemulsification surgery in patients with FUS produces no significant loss of endothelial cells and morphological changes that can be detected by confocal and specular microscopy. We found a reduction in KPD 6 months after surgery on confocal microscopy. Additionally, our manual segmentation technique for KPs utilizing Image J offers a novel and practical approach for confocal microscopy image analysis. [ABSTRACT FROM AUTHOR]

Background: High-risk strains of HPV are known to cause cervical cancer. Multiple clinical studies have emphasized that smoking and drinking are critical risk factors for cervical cancer and its high-grade precursors. In this study, we investigated if smoking and/or drinking augment the molecular mechanisms of cervical carcinogenesis and defined a potential therapeutic approach for their attenuation. Methods: The impact of benzo[a]pyrene (B[a]P) and/or ethanol (EtOH) exposure on cervical cancer cells was assessed by measuring changes in their cell migration and invasion characteristics. Expression of HPV16 E6/E7, NF-κB, cytokines, and inflammation mediators was determined using qRT-PCR, immunoblotting, ELISA, luciferase reporter assay, and confocal microscopy. Herein, we used curcumin (Cur), and PLGA nanoparticle formulation of curcumin (PLGA-Cur) and determined effectiveness of free Cur and PLGA-Cur formulation on smoking and drinking activated NF-κB/IL-6 mediated inflammatory signaling pathways using in vitro cervical cancer models. Results: Treatments with B[a]P and/or EtOH altered the expression of HPV16 E6/E7 oncogenes and EMT markers in cervical cancer cells; it also enhanced migration and invasion. In addition, B[a]P and/or EtOH exposure promoted inflammation pathways through TNF-α and NF-κB signaling, leading to IL-6 upregulation and activation of VEGF. The molecular effects caused by B[a]P and/or EtOH exposure were effectively attenuated by curcumin (Cur)/PLGA-Cur treatment. Conclusions: These data suggest a molecular link between smoking, drinking, and HPV infectivity in cervical carcinogenesis. In addition, attenuation of these effects by treatment with Cur/PLGA-Cur treatment, implies the role of curcumin in cervical cancer prevention and treatment. [ABSTRACT FROM AUTHOR]

Background: There is a need for therapies that can mitigate bone marrow dysfunction and organ toxicity that occur following myeloablative injury and reduced intensity conditioning regimens used in patients undergoing bone marrow transplantation (BMT). The pathogenesis of adverse effects from BMT conditioning has been linked to injury to the vascular endothelium, bone marrow (BM), and other organs. Objective: To evaluate the impact of the thrombopoietin mimetic drug JNJ-26366821 (TPOm) on BM vascular recovery in mice undergoing myeloablative radiation conditioning followed by BMT. Study design: TPOm (doses: 0 µg, 300 µg, 1000 µg per Kg body weight) was administered on Days 0 and 7 after BMT, in mice receiving a total body irradiation (TBI) conditioning regimen (5.5 Gy x 2) before congenic BMT. BM donner cell engraftment was analyzed using flow cytometry on Days 7, 14, and 30 post-BMT. The morphological and biophysical properties of the BM vasculature were evaluated by intravital multiphoton microscopy (MPM) and immunofluorescence confocal imaging. Herein, morphological properties involve microvascular density (MVD), vessel diameter, and vascular area, while biophysical properties include transfer rate (Ktrans) of contrast within the BM vascular niche, as well as the fractional volume (vec) of extracellular extravascular tissue (EES). Results: No significant difference in donor chimerism was observed at days 7, 14, and 30 post-BMT, between TPOm and PBS-treated mice. TPOm intervention improved BM vasculature regeneration in transplanted mice. The MVD, Ktrans, and BM vasculature as well as vascular endothelial growth factor receptor-2 (VEGFR2) in the BM, showed a dose dependent improvement inmice treated with TPOm. On day 14 post-BMT, the group receiving 1000 µg/Kg TPOm showed significant shifts (p-value < 0.05) in MVD, Ktrans, and VEGFR2 expression from their corresponding control types (TPOm dose 0 µg) towards levels comparable to healthy controls. Conclusion: TPOm intervention augments BM vascular structure and function, which may be important for hematopoietic recovery and bone marrow function in radiation conditioned hematopoietic stem cell transplant patients, in addition to enhancing platelet recovery. day 14 post-BMT, the group receiving 1000 µg/Kg TPOm showed significant shifts (p-value < 0.05) in MVD, Ktrans, and VEGFR2 expression from their corresponding control types (TPOm dose 0 µg) towards levels comparable to healthy controls. Conclusion: TPOm intervention augments BM vascular structure and function, which may be important for hematopoietic recovery and bone marrow function in radiation conditioned hematopoietic stem cell transplant patients, in addition to enhancing platelet recovery. [ABSTRACT FROM AUTHOR]

Background and objective: The primary cilium is a small protrusion found on most mammalian cells. It acts as a cellular antenna, being involved in various cell signaling pathways. The length of the primary cilium affects its function. To study the impact of physical or chemical stimuli on cilia, their lengths must be determined easily and reproducibly. Methods: We have developed and evaluated an open-source R package called detectCilia to detect and measure primary cilia automatically. As a case study to demonstrate the capability of our tool, we compared the influence of 4 different cell culture media compositions on the lengths of primary cilia in human chondrocytes. These media compositions include (1) insulin-transferrin-selenium (ITS); (2) ITS and dexamethasone (Dexa); (3) ITS, Dexa, insulin-like growth factor 1 (IGF-1), and transforming growth factor beta 1 (TGF-β1); and (4) fetal bovine serum (FBS). Results: The assessment of detectCilia included a comparison with 2 similar tools: ACDC (Automated Cilia Detection in Cells) and CiliaQ. Several differences and advantages of our package make it a valuable addition to these tools. In the case study, we have observed variations in the ciliary lengths associated with using different media compositions. Conclusions: We conclude that detectCilia can automatically and reproducibly detect and measure primary cilia in confocal microscopy images with low false-positive rates without requiring extensive user interaction. [ABSTRACT FROM AUTHOR]

The glycocalyx is a complex layer of carbohydrate and protein molecules that surrounds the cell membrane of many types of mammalian cells. It serves several important functions, including cell adhesion and communication, and maintain cell shape and stability, especially in the case of erythrocytes. Alteration of glycocalyx composition represents a cardiovascular health threatening. For example, in diabetes mellitus glycocalyx of erythrocytes and of endothelial cells is known to be impaired, a potential source of blood occlusion in microcirculation, which may lead to blindness, and renal failure of patients. The impact of glycocalyx impairment on erythrocyte aggregation remains a largely unexplored research area. We conduct here in vitro-experiments in microfluidic devices in order to investigate erythrocytes aggregation incubated with amylase, an enzyme that partially breaks down glycocalyx molecules. It is found that incubation of erythrocytes by amylase leads to a dramatic increase of their aggregation and stability and alters the aggregates morphologies. Confocal microscopy analysis reveals a significant degradation of the glycocalyx layer, correlated with enhanced erythrocytes aggregation. An increased erythrocyte aggregation in vivo should affect oxygen and other metabolites delivery to organs and tissues. This study brings new elements about elucidation of microscopic origins of erythrocyte aggregation and their potential impact on cardiovascular pathologies. [ABSTRACT FROM AUTHOR]

Ex vivo fusion confocal microscopy (EVFCM) enables the rapid examination of breast tissue and has the potential to reduce the surgical margins and the necessity for further surgeries. Traditional methods, such as frozen section analysis, are limited by the distortion of tissue and artefacts, leading to false negatives and the need for additional surgeries. This study on observational diagnostic accuracy evaluated the ability of EVFCM to detect breast cancer. A total of 36 breast tissue samples, comprising 20 non-neoplastic and 16 neoplastic cases, were analysed using EVFCM and compared to the results obtained from routine histopathology. A Mohs surgeon experienced in EVFCM (evaluator A) and two breast pathologists unfamiliar with EVFCM (evaluators B and C) performed blinded analyses. EVFCM showed high concordance with the histopathology and the detection of neoplasia, with significant kappa values (p < 0.001). Evaluator A achieved 100% sensitivity and specificity. Evaluators B and C achieved a sensitivity of >87%, a specificity of >94%, positive predictive values of >95%, and negative predictive values of 81% and 94%, respectively. EVFCM therefore offers a promising technique for the assessment of margins in breast-conserving surgery. Its widespread adoption could significantly reduce re-excisions, lower healthcare costs, and improve cosmetic and oncological outcomes. [ABSTRACT FROM AUTHOR]

Salmonid rickettsial septicemia (SRS), caused by Piscirickettsia salmonis, has been the most severe health concern for the Chilean salmon industry. The efforts to control P. salmonis infections have focused on using antibiotics and vaccines. However, infected salmonids exhibit limited responses to the treatments. Here, we developed a poly (D, L-lactide-glycolic acid) (PLGA)-nanosystem functionalized with Atlantic salmon IgM (PLGA-IgM) to specifically deliver florfenicol into infected cells. Polymeric nanoparticles (NPs) were prepared via the double emulsion solvent-evaporation method in the presence of florfenicol. Later, the PLGA-NPs were functionalized with Atlantic salmon IgM through carbodiimide chemistry. The nanosystem showed an average size of ~380–410 nm and a negative surface charge. Further, florfenicol encapsulation efficiency was close to 10%. We evaluated the internalization of the nanosystem and its impact on bacterial load in SHK-1 cells by using confocal microscopy and qPCR. The results suggest that stimulation with the nanosystem elicits a decrease in the bacterial load of P. salmonis when it infects Atlantic salmon macrophages. Overall, the IgM-functionalized PLGA-based nanosystem represents an alternative to the administration of antibiotics in salmon farming, complementing the delivery of antibiotics with the stimulation of the immune response of infected macrophages. [ABSTRACT FROM AUTHOR]

Parenchymal distribution of nebulized drug in healthy and diseased lungs has not, as evident from a literature review, been well characterized. We use a vibrating mesh nebulizer to deliver fluorescein solution in vivo to healthy or intratracheal‐lipopolysaccharide (LPS)‐instilled anesthetized rats in dorsal recumbency, or ex vivo to the lungs of LPS‐instilled rats. Following in vivo nebulization (healthy/LPS‐instilled), we quantify fluorescein intensity distribution by confocal microscopy in standard locations on the surface of freshly isolated lungs. Following LPS instillation (in vivo/ex vivo nebulization), we quantify fluorescein intensity in visibly injured locations. In standard locations, there is uniform, low‐intensity basal fluorescein deposition. Focal regions receive high deposition that is, in upper (cranial), middle, and lower (caudal) locations, 6.4 ± 4.9, 3.3 ± 3.0, and 2.3 ± 2.8 times greater, respectively, than average basal intensity. Following LPS instillation, deposition in moderately injured regions can be high or low; deposition in severely injured regions is low. Further, actively phagocytic cells are observed in healthy and LPS‐instilled lungs. And LPS particularly impairs mechanics and activates phagocytic cells in the male sex. We conclude that a low level of nebulized drug can be distributed across the parenchyma excepting to severely injured regions. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is a pathogen associated with severe respiratory infections. The ability of S. aureus to internalize into lung epithelial cells complicates the treatment of respiratory infections caused by this bacterium. In the intracellular environment, S. aureus can avoid elimination by the immune system and the action of circulating antibiotics. Consequently, interfering with S. aureus internalization may represent a promising adjunctive therapeutic strategy to enhance the efficacy of conventional treatments. Here, we investigated the host-pathogen molecular interactions involved in S. aureus internalization into human lung epithelial cells. Lipid raft-mediated endocytosis was identified as the main entry mechanism. Thus, bacterial internalization was significantly reduced after the disruption of lipid rafts with methyl-β-cyclodextrin. Confocal microscopy confirmed the colocalization of S. aureus with lipid raft markers such as ganglioside GM1 and caveolin-1. Adhesion of S. aureus to α5β1 integrin on lung epithelial cells via fibronectin-binding proteins (FnBPs) was a prerequisite for bacterial internalization. A mutant S. aureus strain deficient in the expression of alpha-hemolysin (Hla) was significantly impaired in its capacity to enter lung epithelial cells despite retaining its capacity to adhere. This suggests a direct involvement of Hla in the bacterial internalization process. Among the receptors for Hla located in lipid rafts, caveolin-1 was essential for S. aureus internalization, whereas ADAM10 was dispensable for this process. In conclusion, this study supports a significant role of lipid rafts in S. aureus internalization into human lung epithelial cells and highlights the interaction between bacterial Hla and host caveolin-1 as crucial for the internalization process. [ABSTRACT FROM AUTHOR]

Objective Methods Results Interpretation Clinical Trial Registration Compare the diagnostic characteristics of intraepidermal nerve fiber density (IENFD) and confocal corneal microscopy (CCM) for distal symmetric polyneuropathy (DSP) and small fiber neuropathy (SFN).Participants with obesity were recruited from bariatric surgery clinics and testing was performed prior to surgery. DSP and SFN were determined using the Toronto consensus definitions of probable neuropathy. IENFD was assessed from 3 mm punch biopsies of the distal leg and proximal thigh. CCM was performed on both eyes with manual and automated counting. The Michigan Neuropathy Screening Instrument questionnaire (MNSIq) was also completed. Diagnostic capability was determined using areas under the receiver operating characteristics curve (AUC) from logistic regression.We enrolled 140 participants (mean [standard deviation [SD]] age: 50.3 years [7.1], 77.1% female, BMI: 44.4 kg/m2 [6.7]). In this population, 22.9% had DSP and 14.3% had SFN. Distal leg IENFD had the largest AUC (95% confidence interval) for DSP (0.78, 0.68–0.89) and SFN (0.85, 0.75–0.96). Proximal thigh IENFD (DSP: AUC: 0.59, 0.48–0.69, SFN: AUC: 0.59, 0.46–0.73) and CCM metrics (DSP: AUC range: 0.55–0.60, SFN: AUC range: 0.45–0.62) had poorer diagnostic capability than distal leg IENFD for DSP/SFN (P < 0.05). MNSIq had similar diagnostic capability to distal leg IENFD for both DSP/SFN (DSP: AUC: 0.76, 0.68–0.85, SFN: AUC: 0.81, 0.73–0.88). More participants (52%) preferred skin biopsies to CCM.Distal leg IENFD was the best quantitative measure of DSP/SFN. CCM had poor diagnostic characteristics and fewer patients preferred this test to IENFD. The MNSIq had similar diagnostic characteristics to distal leg IENFD, indicating its value as a diagnostic tool in the clinical setting.clinicaltrials.gov: NCT03617185. [ABSTRACT FROM AUTHOR]

ABSTRACT Background Methods Results Conclusion Early diagnosis of melanoma and prompt effective therapy optimizes prognosis. Reflectance confocal microscopy (RCM) facilitates diagnosis by providing immediate 3D single cell resolution down into the papillary dermis.Consecutive cases were examined using a Vivascope 1500 confocal microscope at a single referral medical practice in Sydney, Australia 2019–2023. Melanoma clinical surface diameters were recorded by 0.1 mm increments up to 6.0 mm. The RCM features recorded were: pagetoid single cells or nests, pleomorphic cell shape, atypical dendritic cells, non‐edged papillae, variation in melanocyte size and confluent sheets of cells. All cases required diagnostic agreement by two dermatohistopathologists using hematoxylin and eosin staining followed by SOX 10 and/or PRAME stains if required.Total cases were 68: 38 males (mean age 57) and 30 females (mean age 64). Melanoma in situ (n = 65) compared to invasive melanoma (n = 3), all males, invasion depth (0.4–0.5 mm). Most frequent RCM features found in 50% or more of cases within all diameter increments were: pagetoid single cells n = 64/68 (94%), pleomorphic cell shape n = 63/68 (93%), epidermal disarray n = 58/68 (85%), and atypical dendritic cells n = 45/68 (66%). Non‐edged dermal papillae were n = 42/68 (62%).Melanoma RCM features were found throughout the diameter ranges. Confocal examination may facilitate early melanoma recognition in these ranges. [ABSTRACT FROM AUTHOR]

Background: Oocytes, the largest cells in mammals, harbor numerous mitochondria within their cytoplasm. These highly dynamic organelles are crucial for providing energy resources and serving as central regulators during oogenesis. Mitochondrial dynamics ensure proper energy distribution for various cellular processes involved in oocyte maturation. Previous studies have used alterations in mitochondrial distribution as a biomarker to assess the oocyte health. However, there are discrepancies between studies regarding mitochondrial distribution profiles in healthy oocytes. Consequently, a comprehensive mitochondrial distribution profile in oocytes during maturation has not been fully characterized. Additionally, there is a lack of objective, quantitative methods to evaluate alterations in mitochondrial distribution profiles in oocytes. Methods: This study aims to provide an in-depth overview of mitochondrial distribution profiles in mouse oocytes at different maturation stages: germinal vesicle (GV) stage, metaphase I (MI), and mature metaphase II (MII). Freshly collected mouse GV, MI and MII oocytes were stained with MitoTracker Red. Confocal microscopy was used to obtain images of mitochondrial distribution profiles in these oocytes. Using the Imaris software, we reconstructed three-dimensional (3D) surface renderings of each oocyte and quantitatively illustrated the mitochondrial distribution profiles. Results: At the GV stage, mitochondria in oocytes were evenly distributed throughout the ooplasm. As oocytes progressed to MI and MII stages, mitochondria aggregated and formed clusters, the mean size of mitochondrial clusters and the proportions of clustered mitochondria increased along with the maturation of oocytes. Conclusions: Our findings reveal that mitochondria in mouse oocytes are highly dynamic, undergoing significant reorganizations during oocyte maturation. We for the first time provided comprehensive mitochondrial distribution profiles in mouse oocytes at the GV, MI and MII stages. These mitochondrial distribution profiles were further quantitatively evaluated. Our methods provide an objective and standardized approach for evaluating alterations in mitochondrial dynamics, which can be used as biomarkers to monitor oocyte conditions during maturation. [ABSTRACT FROM AUTHOR]

pH-induced reactive transport in porous environments is a critical factor in Earth sciences, influencing a range of natural and anthropogenic processes, such as mineral dissolution and precipitation, adsorption and desorption, microbial reactions, and redox transformations. These processes, pivotal to carbon capture and storage (CCS) applications to groundwater remediation, are determined by pH transport. However, the uncertainty in these macroscopic processes' stems from pore-scale heterogeneities and the high diffusion value of the ions and protons forming the pH range. While practical for field-scale applications, traditional macroscopic models often fail to accurately predict experimental and field results in reactive systems due to their inability to capture the details of the pore-scale pH range. This study investigates the interplay between transverse mixing and pH-driven reactions in porous media. It focuses on how porous structure and flow rate affect mixing and chemical reaction dynamics. Utilizing confocal microscopy, the research visualizes fluorescently labeled fluids, revealing variations in mixing patterns from diffusive in homogeneous to shear-driven in heterogeneous media. However, pH-driven reactions show a different pattern, with a faster reaction rate, suggesting quicker pH equilibration between co-flowing fluids than predicted by transverse dispersion or diffusion. The study highlights the unique characteristics of pH change in water, which significantly influences reactive transport in porous media. [ABSTRACT FROM AUTHOR]

The communication between intestinal Glucagon like peptide 1 (GLP-1)-producing cells and the peripheral nervous system has garnered renewed interest considering the availability of anti-obesity and anti-diabetic approaches targeting GLP-1 signaling. While it is well-established that intestinal GLP-1 cells can exert influence through paracrine mechanisms, recent evidence suggests the possible existence of synaptic-like connections between GLP-1 cells and peripheral neurons, including those of the vagus nerve. In this study, using a reporter Phox2b-Cre-Tomato mouse model and super-resolution confocal microscopy, we demonstrated that vagal axons made apparent contacts with less than 0.5% of GLP-1 cells. Moreover, immunohistochemistry combined with super-resolution confocal microscopy revealed abundant post-synaptic density 95 (PSD-95) immunoreactivity within the enteric plexus of the lower intestines of C57/BL6 mice, with virtually none in its mucosa. Lastly, utilizing RNAScope in situ hybridization in the lower intestines of mice, we observed that GLP-1 cells expressed generic markers of secretory cells such as Snap25 and Nefm, but neither synaptic markers such as Syn1 and Nrxn2, nor glutamatergic markers such as Slc17a7. Through theoretical considerations and a critical review of the literature, we concluded that intestinal GLP-1 cells primarily communicate with vagal neurons through paracrine mechanisms, rather than synaptic-like contacts. [ABSTRACT FROM AUTHOR]

Porous organic cobaltocenium‐containing particles are scarce in literature but highly interesting for their electrochemical properties and reusability in, for example, catalysis or magnetic systems. In this work, we present a scalable one‐pot strategy to introduce tailorable amounts of cobaltocenium on a porous substrate, adjusting the electrochemical switching capability. For this purpose, 3‐(triethoxysilyl)propan‐1‐amine (APTES) and ethynyl cobaltocenium hexafluorophosphate is used as functionalization agents for in‐situ catalyst‐free hydroamination, followed by silane condensation at the particles' surface. Functionalized particles are characterized by attenuated total reflection infrared spectroscopy (ATR‐IR), thermogravimetric analysis (TGA), laser scanning confocal microscopy (LSCM), scanning electron microscopy (SEM), energy dispersive X‐ray spectroscopy (EDS), inductively coupled plasma mass spectrometry (ICP‐MS), powder X‐ray diffraction (PXRD) and cyclic voltammetry (CV) showing excellent control over the degree of functionalization, i. e., the added cobaltocenium reagents. The electrochemical stability and good addressability while preserving the porous structure are shown. By utilizing higher amounts of APTES, the overall cobaltocenium amount can be reduced in favor of additional amine groups, strongly affecting the electrochemical behavior, making this functionalization strategy a good platform for metallopolymer immobilization and tailored functionalization. Additionally, thermal treatment of the synthesized metallopolymer microparticles paves the way to magnetic properties with tailorable microporous architectures for end‐of‐life and upcycling aspects. [ABSTRACT FROM AUTHOR]

Purpose: The primary aim of this study was to develop an open-source Python-based software for the automated analysis of dynamic cell behaviors in microphysiological models using non-confocal microscopy. This research seeks to address the existing gap in accessible tools for high-throughput analysis of endothelial tube formation and cell invasion in vitro, facilitating the rapid assessment of drug sensitivity. Methods: Our approach involved annotating over 1000 2 mm Z-stacks of cancer and endothelial cell co-culture model and training machine learning models to automatically calculate cell coverage, cancer invasion depth, and microvessel dynamics. Specifically, cell coverage area was computed using focus stacking and Gaussian mixture models to generate thresholded Z-projections. Cancer invasion depth was determined using a ResNet-50 binary classification model, identifying which Z-planes contained invaded cells and measuring the total invasion depth. Lastly, microvessel dynamics were assessed through a U-Net Xception-style segmentation model for vessel prediction, the DisPerSE algorithm to extract an embedded graph, then graph analysis to quantify microvessel length and connectivity. To further validate our software, we reanalyzed an image set from a high-throughput drug screen involving a chemotherapy agent on a 3D cervical and endothelial co-culture model. Lastly, we applied this software to two naive image datasets from coculture lumen and microvascular fragment models. Results: The software accurately measured cell coverage, cancer invasion, and microvessel length, yielding drug sensitivity IC50 values with a 95% confidence level compared to manual calculations. This approach significantly reduced the image processing time from weeks down to h. Furthermore, the software was able to calculate cell coverage, microvessel length, and invasion depth from two additional microphysiological models that were imaged with confocal microscopy, highlighting the versatility of the software. Conclusions: Our free and open source software offers an automated solution for quantifying 3D cell behavior in microphysiological models assessed using non-confocal microscopy, providing the broader Cellular and Molecular Bioengineering community with an alternative to standard confocal microscopy paired with proprietary software.This software can be found in our GitHub repository: https://github.com/fogg-lab/tissue-model-analysis-tools. [ABSTRACT FROM AUTHOR]

The label‐free imaging of inorganic nanoparticles (NPs) using confocal laser scanning microscopy (CLSM) provides a powerful and versatile tool for studying interactions between NPs and biological systems. Without the need for exogenous labels or markers, it simply benefits from the differential scattering of visible photons between biomaterials and inorganic NPs. Validation experiments conducted on fixed and living cells in real‐time, as well as mouse tissue sections following parenteral administration of NPs. Additionally, by incorporating reporter fluorophores and utilizing both reflectance and fluorescence imaging modalities, the method enables high‐resolution multiplex imaging of cellular structures and NPs. Different sizes and concentrations of Au NPs are tested as for Ag, Fe3O4, and CeO2 NPs, all with biological interest. Overall, the comprehensive study of NP imaging by confocal microscopy in reflectance mode provides valuable insights and tools for researchers interested in monitoring the nano‐bio interactions. [ABSTRACT FROM AUTHOR]

Nanoporous gold electrodes are of great interest in electroanalytical chemistry, because of their unusual activity and large surface area. The electrochemical activity can be further improved by coating with molecular catalysts such as the tetraruthenated cobalt‐tetrapyridylporphyrazines investigated in this work. The plasmonic enhancement of the scattered light at the nanoholes and borders modifies the electrode's optical characteristics, improving the transmission through the surface‐enhanced Raman scattering (SERS) effect. When monitored by hyperspectral dark‐field and confocal Raman microscopy, this effect allows probing of the porphyrazine species at the plasmonic nanholes, improving the understanding of the chemically modified gold electrodes. [ABSTRACT FROM AUTHOR]

Background: Acral melanocytic lesions often pose a diagnostic and therapeutic challenge for many clinicians. Reflectance confocal microscopy (RCM) is an imaging technique widely used for the assessment of skin cancers. The aim of this review is to explore the applicability of RCM for the diagnosis of nevi and melanoma on the acral sites. Methods: Study selection was conducted based on the application of RCM for acral melanocytic lesions. All types of articles (original articles, short reports, and single case reports) were included in the analysis following PRISMA updated guidelines. Results: The search retrieved 18 papers according to the selection criteria; after removing duplicate records and additional articles by one or more of the exclusion criteria, a total of seven studies were carefully evaluated. Conclusions: RCM seems a valuable and useful additional tool for the diagnosis of acral melanocytic lesions, and its use may decrease the need for invasive procedures to some extent. Visualization of deeper layers may be achieved through mechanical removal of the superficial stratum corneum. [ABSTRACT FROM AUTHOR]

Although there have been many theoretical studies on the enhancement effect of nanofiller networks and their interaction with elastomer molecular chains on the mechanical properties of elastomers, its mechanism description is still not completely clear. One of the main obstacles is the lack of quantitative characterization techniques and corresponding theoretical models for the three-dimensional morphology of complex nanofiller networks. In this paper, the precipitated silica-filled silicone rubber was studied by fluorescence labeling combined with laser scanning confocal microscopy, and the real three-dimensional images of dispersion and aggregation structure of filled rubber systems were obtained. The microstructure evolution of nano-particle aggregates caused by the increase in the filler volume fraction was quantitatively described, and the reinforcement mechanism of elastomers with a distribution of aggregates and filler networks composed of nanoparticles was studied. Furthermore, a nano-composite reinforcement model based on volume fraction, particle shape, interaction, and filler dispersion has been proposed. [ABSTRACT FROM AUTHOR]

γ-Secretase plays a pivotal role in the central nervous system. Our recent development of genetically encoded Förster resonance energy transfer (FRET)-based biosensors has enabled the spatiotemporal recording of γ-secretase activity on a cell-by-cell basis in live neurons in culture. Nevertheless, how γ-secretase activity is regulated in vivo remains unclear. Here, we employ the near- infrared (NIR) C99 720-670 biosensor and NIR confocal microscopy to quantitatively record γ-secretase activity in individual neurons in living mouse brains. Intriguingly, we uncovered that γ-secretase activity may influence the activity of γ-secretase in neighboring neurons, suggesting a potential 'cell non- autonomous' regulation of γ-secretase in mouse brains. Given that γ-secretase plays critical roles in important biological events and various diseases, our new assay in vivo would become a new platform that enables dissecting the essential roles of γ-secretase in normal health and diseases. [ABSTRACT FROM AUTHOR]