Mitochondrial sulfur dioxide (SO2) plays important roles in physiological and pathological activities. Unfortunately, it is lack of a reliable tool to precisely visualize the mitochondrial SO2 and elaborate its complicated functions in various cytoactivities. Here we report a mitochondrial-immobilized fluorescent probe PM-Cl consisting of coumarin and benzyl chloride modified benzothiazole, which enables selective visualization of mitochondrial SO2via chemical immobilization. The spectral results demonstrated that probe PM-Cl could respond to SO2 with high selectivity and sensitivity. Co-localization and the fluorescence of cytolysis extraction verified the excellent mitochondrial targeting and anchoring abilities. Due to the chemical immobilization, probe PM-Cl could firmly retain into mitochondria after stimulation of carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and H2O2. Significantly, a series of fluorescence images are indicative of capability for detecting the fluctuations of SO2 in mitochondria during ferroptosis. Furthermore, PM-Cl also could visualize SO2 in myocardium and muscle tissues after the stimulation of CCCP. Taken together, probe PM-Cl is a very potential molecular tool for precisely detecting mitochondrial SO2 to explore its complex functions in physiological and pathological activities.

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive and lethal malignancy with extremely poor patient survival rates. A key reason for the poor prognosis is the lack of effective diagnostic tools to detect the disease at curable, premetastatic stages. Tumor surgical resection is PDAC\'s first-line treatment, however distinguishing between cancerous and healthy tissue with current imaging tools remains a challenge. In this work, we report a DOTA-based fluorescent probe targeting plectin-1 for imaging PDAC with high specificity. To enable heterogeneous functionalization of the DOTA-core with multiple targeting peptide units and the fluorophore, a novel, fully clickable synthetic route that proceeds in one pot was developed. Extensive validation of the probe set the stage for PDAC detection in mice and human tissue. Altogether, these findings may pave the way for improved clinical understanding and early detection of PDAC progression as well as more accurate resection criteria.

Imaging mass spectrometry (IMS) enables highly multiplexed, untargeted tissue mapping for a broad range of molecular classes, facilitating in situ biological discovery. Yet, challenges persist in molecular specificity, which is the ability to discern one molecule from another, and spatial specificity, which is the ability to link untargeted imaging data to specific tissue features. Instrumental developments have dramatically improved IMS spatial resolution, allowing molecular observations to be more readily associated with distinct tissue features across spatial scales, ranging from larger anatomical regions to single cells. High-performance mass analyzers and systems integrating ion mobility technologies are also becoming more prevalent, further improving molecular coverage and the ability to discern chemical identity. This review provides an overview of recent advancements in high-specificity IMS that are providing critical biological context to untargeted molecular imaging, enabling integrated analyses, and addressing advanced biomedical research applications.

Hypertrophic cardiomyopathy (HCM) is a common hereditable cardiomyopathy that affects between 1:200 to 1:500 of the general population. The role of cardiovascular magnetic resonance (CMR) imaging in the management of HCM has expanded over the past 2 decades to become a key informant of risk in this patient population, delivering unique insights into tissue health and its influence on future outcomes. Numerous mature CMR-based techniques are clinically available for the interrogation of tissue health in patients with HCM, inclusive of contrast and noncontrast methods. Late gadolinium enhancement imaging remains a cornerstone technique for the identification and quantification of myocardial fibrosis with large cumulative evidence supporting value for the prediction of arrhythmic outcomes. T1 mapping delivers improved fidelity for fibrosis quantification through direct estimations of extracellular volume fraction but also offers potential for noncontrast surrogate assessments of tissue health. Water-sensitive imaging, inclusive of T2-weighted dark blood imaging and T2 mapping, have also shown preliminary potential for assisting in risk discrimination. Finally, emerging techniques, inclusive of innovative multiparametric methods, are expanding the utility of CMR to assist in the delivery of comprehensive tissue characterization toward the delivery of personalized HCM care. In this narrative review we summarize the contemporary landscape of CMR techniques aimed at characterizing tissue health in patients with HCM. The value of these respective techniques to identify patients at elevated risk of future cardiovascular outcomes are highlighted.

In optical imaging, optical clearing agents are commonly used to enhance the structural details of a sample. The current study investigates how to use it to improve the data obtained by an optical coherence tomography angiography system. A natural edible oil with no chemical base has been used for optical clearing. In-vivo testing on mice and humans yielded excellent optical clearing. Using computational techniques, the improvement in angiography signal caused by the optical clearing agent is investigated qualitatively and quantitatively. Compared to the control group, applying the edible oil-based optical clearing agent demonstrated improved vessel percentage and refined vascular signal intensity along depth.

Dopaminergic (DAergic) neurodegeneration in the substantia nigra pars compacta of the human brain is the pathological feature associated with Parkinson\'s disease (PD). Drosophila also exhibits mobility defects and diminished levels of brain dopamine on exposure to neurotoxicants mimicking PD. Our laboratory demonstrated in a Drosophila model of sporadic PD that there is no decrease in DAergic neuronal number; instead, there is a significant reduction in tyrosine hydroxylase (TH) fluorescence intensity (FI). Here, we present a sensitive assay based on the quantification of FI of the secondary antibody (ab). As the FI is directly proportional to the amount of TH synthesis, its reduction under PD conditions denotes the decrease in the TH synthesis, suggesting DAergic neuronal dysfunction. Therefore, FI quantification is a refined and sensitive method to understand the early stages of DAergic neurodegeneration. FI quantification is performed using the ZEN 2012 SP2 single-user software; a license must be acquired to utilize the imaging system to interactively control image acquisition, image processing, and analysis. This method will be of good use to biologists, as it can also be used with little modification to characterize the extent of degeneration and changes in the level of degeneration in response to drugs in different cell types. Unlike the expensive and cumbersome confocal microscopy, the present method will be an affordable option for fund-constrained neurobiology laboratories. Key features • Allows characterizing the incipient DAergic and other catecholaminergic neurodegeneration, even in the absence of loss of neuronal cell body. • Great alternative for the fund-constrained neurobiology laboratories in developing countries to utilize this method in different cell types and their response to drugs/nutraceuticals.

Sarcoidosis embodies a complex inflammatory disorder spanning multiple systems, with its origin remaining elusive. It manifests as the infiltration of inflammatory cells that coalesce into distinctive noncaseous granulomas within afflicted organs. Unraveling this disease necessitates the utilization of cellular or tissue-based imaging methods to both visualize and characterize the biochemistry of these sarcoid granulomas. Although hematoxylin and eosin stain, standard in routine use alongside cytological stains have found utility in diagnosis within clinical contexts, special stains such as Masson\'s trichrome, reticulin, methenamine silver, and Ziehl-Neelsen provide additional varied perspectives of sarcoid granuloma imaging. Immunohistochemistry aids in pinpointing specific proteins and gene expressions further characterizing these granulomas. Finally, recent advances in spatial transcriptomics promise to divulge profound insights into their spatial orientation and three-dimensional (3-D) molecular mapping. This review focuses on a range of preexisting imaging methods employed for visualizing sarcoid granulomas at the cellular level while also exploring the potential of the latest cutting-edge approaches like spatial transcriptomics and matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI), with the overarching goal of shedding light on the trajectory of sarcoidosis research.

Using high-fidelity, permeable, lipophilic, and bright fluorophores for imaging lipid droplets (LDs) in tissues holds immense potential in diagnosing conditions such as diabetic or alcoholic fatty liver disease. In this work, we utilized linear and Λ-shaped polarity-sensitive fluorescent probes for imaging LDs in both cellular and tissue environments, specifically in rats with diabetic and alcoholic fatty liver disease. The fluorescent probes possess several key characteristics, including high permeability, lipophilicity, and brightness, which make them well-suited for efficient LD imaging. Notably, the probes exhibit a substantial Stokes shift, with 143 nm for DCS and 201 nm for DCN with selective targeting of the lipid droplets. Our experimental investigations successfully differentiated morphological variations between diseased and normal tissues in three distinct tissue types: liver, adipose, and small intestine. They could help provide pointers for improved detection and understanding of LD-related pathologies.

Inflammation is a highly dynamic process with immune cells that continuously interact with each other and parenchymal components as they migrate through tissue. The dynamic cellular responses and interaction patterns are a function of the complex tissue environment that cannot be fully reconstructed ex vivo, making it necessary to assess cell dynamics and changing spatial patterning in vivo. These dynamics often play out deep within tissues, requiring the optical focus to be placed far below the surface of an opaque organ. With the emergence of commercially available two-photon excitation lasers that can be combined with existing imaging systems, new avenues for imaging deep tissues over long periods of time have become available. We discuss a selected subset of studies illustrating how two-photon microscopy (2PM) has helped to relate the dynamics of immune cells to their in situ function and to understand the molecular patterns that govern their behavior in vivo. We also review some key practical aspects of 2PM methods and point out issues that can confound the results, so that readers can better evaluate the reliability of conclusions drawn using this technology.

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