Biology subjects

Ware, S. A.

Publications and source records attributed to Ware, S. A..

6 recordsLinked to original sources

bioRxiv2026

Whole-organ surface mapping using multiview projection reconstruction

While light-sheet microscopy is emerging as a robust method for volumetric imaging with improved axial resolution, its capability regarding two-dimensional, surface-level mapping is often hindered by limitations in data redundancy and reconstruction efficiency stemming from volumetric registration methods. We demonstrate that a multiview imaging approach in an axially-swept, dithered light-sheet microscope paired with computational image reconstruction of view projections is able to address these trade-offs to enable large-scale mapping of surface structural features, leveraging the advantages of multiview light-sheet in scalable field of view, working distance, and near isotropic resolution across the entire imaging depth. To aid in the acquisition and analysis of two-dimensional surface structures, we present a tailored surface mapping workflow and a Fiji plugin for computational reconstruction, promoting robust and comprehensive visualization of surface features of uncleared volumetric samples. Our strategy, termed projection reconstruction for imaging surface morphology (PRISM), integrates axially swept dithered light-sheet microscopy and post-processing software for multiview imaging. The imaging hardware enables near-isotropic resolution across its entire field of view, while the software implementation leverages rigid and affine transformations to align two-dimensional projections of multiview samples. It is designed to work with the BigStitcher pipeline, leveraging its robust algorithm to provide support for two-dimensional image alignment and stitching. We demonstrate the capability of PRISM in studies of lymphatic network mapping in the epicardial layer of intact mouse hearts, as well as surface profiles of FaDu spheroids labeled with antibody-nanodiamond conjugates. This method allows us to quantify cardiac lymphatic branch numbers, diameters, and lengths of a Prox1-tdTomato mouse cardiac model, as well as cluster number and diameters of epidermal growth factor receptor within a FaDu spheroid labeled with a nanodiamond-antibody conjugate, with a significant reduction of post-processing data size. PRISM leverages multiview image projections to promote studies of cardiac lymphatics in mouse models and surface receptor distributions within spheroid models, enabling efficient surface mapping of large, intact, and uncleared biological samples across a variety of scales.

bioengineering↗

bioRxiv2025

Axially swept dithered light-sheet microscope to reveal cardiac morphology

Understanding cardiac microstructure and vascular networks in their entirety is critical for assessing cardiovascular development, disease progression, and therapeutic interventions. Light-sheet microscopy combined with tissue clearing enables high-resolution volumetric imaging of intact organs but faces limitations in trabeculated myocardium due to trade-offs among light-sheet thickness, effective range, and frame rate. We exploit temporal dynamics that govern illumination-detection interplay to maintain uniform resolution across specimens. Building on this, we implemented high-speed dithered light-sheet (DiLS) illumination, extending the confocal region by over 40% and enhancing the space-bandwidth product while preserving optical sectioning. Integration of DiLS with a sweeping approach establishes the axially swept dithered light-sheet (AS-DiLS), which enhances imaging throughput while preserving axial resolution and enables uniform illumination up to 12.5-millimeter range. AS-DiLS delivers near-isotropic resolution (~2.5 m) for investigating intricate ventricular trabeculae, vasculature, and extracellular matrix, providing a scalable platform for comprehensive cardiovascular morphology and topology assessment from embryos to adults. TeaserVolumetric imaging reveals microstructure and vascular networks in their entirety with near-isotropic resolution.

bioengineering↗

bioRxiv2022

Chronic Glucocorticoid Stress Reveals Increased Energy Expenditure and Accelerated Aging as Cellular Features of Allostatic Load

Stress triggers anticipatory physiological responses that promote survival, a phenomenon termed allostasis. However, the chronic activation of energy-dependent allostatic responses results in allostatic load, a dysregulated state that predicts functional decline, accelerates aging, and increases mortality in humans. The energetic cost and cellular basis for the damaging effects of allostatic load have not been defined. Here, by longitudinally profiling three unrelated primary human fibroblast lines across their lifespan, we find that chronic glucocorticoid exposure increases cellular energy expenditure by [~]60%, along with a metabolic shift from glycolysis to mitochondrial oxidative phosphorylation (OxPhos). This state of stress-induced hypermetabolism is linked to mtDNA instability, non-linearly affects age-related cytokines secretion, and accelerates cellular aging based on DNA methylation clocks, telomere shortening rate, and reduced lifespan. Pharmacologically normalizing OxPhos activity while further increasing energy expenditure exacerbates the accelerated aging phenotype, pointing to total energy expenditure as a potential driver of aging dynamics. Together, our findings define bioenergetic and multi-omic recalibrations of stress adaptation, underscoring increased energy expenditure and accelerated cellular aging as interrelated features of cellular allostatic load.

cell biology↗

bioRxiv2021

A Multi-Omics and Bioenergetics Longitudinal Aging Dataset in Primary Human Fibroblasts with Mitochondrial Perturbations

Aging is a process of progressive change. In order to develop biological models of aging, longitudinal datasets with high temporal resolution are needed. Here we report a multi-omic longitudinal dataset for cultured primary human fibroblasts measured across their replicative lifespans. Fibroblasts were sourced from both healthy donors (n=6) and individuals with lifespan-shortening mitochondrial disease (n=3). The dataset includes cytological, bioenergetic, DNA methylation, gene expression, secreted proteins, mitochondrial DNA copy number and mutations, cell-free DNA, telomere length, and whole-genome sequencing data. This dataset enables the bridging of mechanistic processes of aging as outlined by the "hallmarks of aging", with the descriptive characterization of aging such as epigenetic age clocks. Here we focus on bridging the gap for the hallmark mitochondrial metabolism. Our dataset includes measurement of healthy cells, and cells subjected to over a dozen experimental manipulations targeting oxidative phosphorylation (OxPhos), glycolysis, and glucocorticoid signaling, among others. These experiments provide opportunities to test how cellular energetics affect the biology of cellular aging. All data are publicly available at our webtool: https://columbia-picard.shinyapps.io/shinyapp-Lifespan_Study/

cell biology↗

bioRxiv2021

Extracellular release of mitochondrial DNA is triggered by cigarette smoke and is detected in COPD

Chronic obstructive pulmonary disease (COPD) is characterized by continuous and irreversible inflammation frequently caused by persistent exposure to toxic inhalants such as cigarette smoke (CS). CS may trigger mitochondrial DNA (mtDNA) extrusion into the cytosol, extracellular space, or foster its transfer by extracellular vesicles (EVs). The present study aimed to elucidate whether mtDNA is released upon CS exposure and in COPD. We measured cell-free mtDNA (cf-mtDNA) in the plasma of former smokers affected by COPD, in the serum of mice that developed CS-induced emphysema, and in the extracellular milieu of human bronchial epithelial cells exposed to cigarette smoke extract (CSE). Further, we characterized cells exposed to sublethal and lethal doses of CSE by measuring mitochondrial membrane potential and dynamics, superoxide production and oxidative stress, cell cycle progression, and cytokine expression. Patients with COPD and mice that developed emphysema showed increased levels of cf-mtDNA. In cell culture, exposure to a sublethal dose of CSE decreased mitochondrial membrane potential, increased superoxide production and oxidative damage, dysregulated mitochondrial dynamics, and triggered mtDNA release in extracellular vesicles. The release of mtDNA into the extracellular milieu occurred concomitantly with increased expression of DNase III, DNA-sensing receptors (cGAS, NLRP3), proinflammatory cytokines (IL-1{beta}, IL-6, IL-8, IL-18, CXCL2), and markers of senescence (p16, p21). Exposure to a lethal dose of CSE preferentially induced mtDNA and nuclear DNA release in cell debris. Our findings demonstrate that CS-induced stress triggers mtDNA release and is associated with COPD, supporting cf-mtDNA as a novel signaling response to CS exposure.

cell biology↗

bioRxiv2020

An automated, high throughput methodology optimized for quantitative cell-free mitochondrial and nuclear DNA isolation from plasma

Circulating, cell-free mitochondrial DNA (ccf-mtDNA) and nuclear DNA (ccf-nDNA) are under investigation as biomarkers for various diseases. Optimal ccf-mtDNA isolation parameters, like those outlined for ccf-nDNA, have not been established. Here, we optimized a protocol for both ccf-mtDNA and ccf-nDNA recovery using a magnetic bead-based isolation process on an automated 96-well platform. Using the optimized protocol, our data show 6-fold improved yields of ccf-mtDNA when compared to the starting protocol. Digestion conditions, liquid handling characteristics, and magnetic particle processor programming all contributed to increased recovery and improved reproducibility. To our knowledge, this is the first high-throughput approach optimized for mtDNA and nDNA recovery and serves as an important starting point for clinical studies. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=191 HEIGHT=200 SRC="FIGDIR/small/206987v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@13ab547org.highwire.dtl.DTLVardef@1535020org.highwire.dtl.DTLVardef@b191aaorg.highwire.dtl.DTLVardef@1d23fa4_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗