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  1. Mitochondrial morphology provides unique insights into their integrity and function. Among fluorescence microscopy techniques, 3D super-resolution microscopy uniquely enables the analysis of mitochondrial morphological features individually. However, there is a lack of tools to extract morphological parameters from super-resolution images of mitochondria. We report a quantitative method to extract mitochondrial morphological metrics, including volume, aspect ratio, and local protein density, from 3D single-molecule localization microscopy images, with single-mitochondrion sensitivity. We validated our approach using simulated ground-truth SMLM images of mitochondria. We further tested our morphological analysis on mitochondria that have been altered functionally and morphologically in controlled manners. This work sets the stage to quantitatively analyze mitochondrial morphological alterations associated with disease progression on an individual basis.

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  2. We developed a multiscale optical imaging workflow, integrating and correlating visible-light optical coherence tomography, confocal laser scanning microscopy, and single-molecule localization microscopy to investigate mouse cornea damage from thein-vivotissue level to the nanoscopic single-molecule level. We used electron microscopy to validate the imaged nanoscopic structures. We imaged wild-type mice and mice with acute ocular hypertension and examined the effects of Rho-kinase inhibitor application. We defined four types of intercellular tight junction structures as healthy, compact, partially-distorted, and fully-distorted types by labeling the zonula occludens-1 protein in the corneal endothelial cell layer. We correlated the statistics of the four types of tight junction structures with cornea thickness and intraocular pressure. We found that the population of fully-distorted tight junctions correlated well with the level of corneal edema, and applying Rho-kinase inhibitor reduced the population of fully-distorted tight junctions under acute ocular hypertension. Together, these data point to the utility of multiscale optical imaging in revealing fundamental biology relevant to disease and therapeutics.

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  3. Abstract

    Single-molecule localization microscopy (SMLM) breaks the optical diffraction limit by numerically localizing sparse fluorescence emitters to achieve super-resolution imaging. Spectroscopic SMLM or sSMLM further allows simultaneous spectroscopy and super-resolution imaging of fluorescence molecules. Hence, sSMLM can extract spectral features with single-molecule sensitivity, higher precision, and higher multiplexity than traditional multicolor microscopy modalities. These new capabilities enabled advanced multiplexed and functional cellular imaging applications. While sSMLM suffers from reduced spatial precision compared to conventional SMLM due to splitting photons to form spatial and spectral images, several methods have been reported to mitigate these weaknesses through innovative optical design and image processing techniques. This review summarizes the recent progress in sSMLM, its applications, and our perspective on future work.

    Graphical Abstract

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  4. Transposable elements (TEs) and the silencing machinery of their hosts are engaged in a germline arms-race dynamic that shapes TE accumulation and, therefore, genome size. In animal species with extremely large genomes (>10 Gb), TE accumulation has been pushed to the extreme, prompting the question of whether TE silencing also deviates from typical conditions. To address this question, we characterize TE silencing via two pathways—the piRNA pathway and KRAB-ZFP transcriptional repression—in the male and female gonads of Ranodon sibiricus , a salamander species with a ∼21 Gb genome. We quantify 1) genomic TE diversity, 2) TE expression, and 3) small RNA expression and find a significant relationship between the expression of piRNAs and TEs they target for silencing in both ovaries and testes. We also quantified TE silencing pathway gene expression in R. sibiricus and 14 other vertebrates with genome sizes ranging from 1 to 130 Gb and find no association between pathway expression and genome size. Taken together, our results reveal that the gigantic R. sibiricus genome includes at least 19 putatively active TE superfamilies, all of which are targeted by the piRNA pathway in proportion to their expression levels, suggesting comprehensive piRNA-mediated silencing. Testes have higher TE expression than ovaries, suggesting that they may contribute more to the species’ high genomic TE load. We posit that apparently conflicting interpretations of TE silencing and genomic gigantism in the literature, as well as the absence of a correlation between TE silencing pathway gene expression and genome size, can be reconciled by considering whether the TE community or the host is currently “on the attack” in the arms race dynamic. 
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    Free, publicly-accessible full text available February 24, 2024
  5. Abstract

    Magneto‐elastic materials facilitate features such as shape programmability, adaptive stiffness, and tunable strength, which are critical for advances in structural and robotic materials. Magneto‐elastic networks are commonly fabricated by employing hard magnets embedded in soft matrices to constitute a monolithic body. These architected network materials have excellent mechanical properties but damage incurred in extreme loading scenarios are permanent. To overcome this limitation, we present a novel design for elastic bars with permanent fixed dipole magnets at their ends and demonstrate their ability to self‐assemble into magneto‐elastic networks under random vibrations. The magneto‐elastic unit configuration, most notably the orientation of end dipoles, is shown to dictate the self‐assembled network topology, which can range from quasi‐ordered triangular lattices to stacks or strings of particles. Network mechanics are probed with uniaxial tensile tests and design criteria for forming stable lightweight 2D networks are established. It is shown that these magneto‐elastic networks rearrange and break gracefully at their magnetic nodes under large excitations and yet recover their original structure at moderate random excitations. This work paves the way for structural materials that can be self‐assembled and repaired on‐the‐fly with random vibrations, and broadens the applications of magneto‐elastic soft materials.

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  6. Abstract

    This decade has witnessed the tremendous progress in miniaturizing optical imaging systems. Despite the advancements in 3D printing optical lenses at increasingly smaller dimensions, challenges remain in precisely manufacturing the dimensionally compatible optomechanical components and assembling them into a functional imaging system. To tackle this issue, the use of 3D printing to enable digitalized optomechanical component manufacturing, part‐count‐reduction design, and the inclusion of passive alignment features is reported here, all for the ease of system assembly. The key optomechanical components of a penny‐sized accommodating optical microscope are 3D printed in 50 min at a significantly reduced unit cost near $4. By actuating a built‐in voice‐coil motor, its accommodating capability is validated to focus on specimens located at different distances, and a focus‐stacking function is further utilized to greatly extend depth of field. The microscope can be readily customized and rapidly manufactured to respond to task‐specific needs in form factor and optical characteristics.

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  7. Abstract

    One of the most challenging aspects of developing high-energy lithium-based batteries is the structural and (electro)chemical stability of Ni-rich active cathode materials at thermally-abused and prolonged cell cycling conditions. Here, we report in situ physicochemical characterizations to improve the fundamental understanding of the degradation mechanism of charged polycrystalline Ni-rich cathodes at elevated temperatures (e.g., ≥ 40 °C). Using multiple microscopy, scattering, thermal, and electrochemical probes, we decouple the major contributors for the thermal instability from intertwined factors. Our research work demonstrates that the grain microstructures play an essential role in the thermal stability of polycrystalline lithium-based positive battery electrodes. We also show that the oxygen release, a crucial process during battery thermal runaway, can be regulated by engineering grain arrangements. Furthermore, the grain arrangements can also modulate the macroscopic crystallographic transformation pattern and oxygen diffusion length in layered oxide cathode materials.

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  8. By manipulating the spectral dispersion of detected photons, spectroscopic single-molecule localization microscopy (sSMLM) permits concurrent high-throughput single-molecular spectroscopic analysis and imaging. Despite its promising potential, using discrete optical components and managing the delicate balance between spectral dispersion and spatial localization compromise its performance, including non-uniform spectral dispersion, high transmission loss of grating, high optical alignment demands, and reduced precision. We designed a dual-wedge prism (DWP)-based monolithic imaging spectrometer to overcome these challenges. We optimized the DWP for spectrally dispersing focused beam without deviation and with minimal wavefront error. We integrated all components into a compact assembly, minimizing total transmission loss and significantly reducing optical alignment requirements. We show the feasibility of DWP using ray-tracing and numerical simulations. We validated our numerical simulations by experimentally imaging individual nanospheres and confirmed that DWP-sSMLM achieved much improved spatial and spectral precisions of grating-based sSMLM. We also demonstrated DWP-sSMLM in 3D multi-color imaging of cells. 
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