Publications by Year: 2026

A hexanucleotide repeat expansion in C9orf72 is the most common genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. While repeat RNAs are implicated in disease pathogenesis, their mechanisms of action remain incompletely understood. Here, we show that GGGGCC repeat RNA engages chromatin genome-wide preferentially at promoter regions in patient cells. This interaction obstructs RNA polymerase II and transcription factors with GC-rich motifs, leading to broad transcriptional repression. Biochemical assays, single-molecule imaging, and native bisulfite sequencing analyses demonstrate that GGGGCC repeat RNA intrinsically forms DNA:RNA hybrid G-quadruplexes (HQs) with cognate DNA, providing a structural basis for transcriptional interference. Stabilization of these G-quadruplex structures exacerbates neuronal vulnerability to metabolic stress in patient-derived motor neurons and cortical organoids, whereas restoring key gene dysregulation improves resistance. These findings uncover a previously unrecognized trans-acting mechanism whereby repetitive RNAs form hybrid structures with genomic DNA, disrupt gene regulation, and contribute to neurodegeneration.

Cardiac glycosides (CGs) such as ouabain exert positive inotropic effects by inhibiting the Na+-K+-ATPase. CGs' wide spread use is limited by CGs' narrow therapeutic window. Mis- or overdosing with CGs may cause cardiac arrhythmias, resulting from electrolyte disturbances. To study the ethically challenging topic of CG overdosing, we here optimized the in ovo platform to test whether treatment with the selective ouabain antagonist rostafuroxin prevents CG-mediated electrophysiological derangements and arrhythmia by restoring electrolyte homeostasis. We used incubated chicken eggs (iCEs), a 3 R-compliant model, for which we established electrocardiograms (ECGs). ECGs were recorded under 1) baseline conditions, 2) after treatment with ouabain, and 3) after cotreatment with rostafuroxin. Underlying mechanisms of ouabain and rostafuroxin effects were studied using blood gas analysis and fluorescence microscopy. Isolated murine and human cardiomyocytes served as an independent model to confirm in ovo results. Ouabain treatment resulted in increased heart rate variability (HRV), transient sinus arrest, and atrio-ventricular dyssynchrony, accompanied by plasma hyperkalemia and cardiomyocyte Na+ overload. Cotreatment of ouabain and rostafuroxin led to reduced HRV and ameliorated the frequency and duration of transient sinus arrest, whereas plasma K+ levels remained unchanged. In isolated cardiomyocytes, ouabain treatment induced intracellular Na+ overload, which was abolished by additional rostafuroxin treatment. Our work demonstrates the in ovo platform and corresponding readouts as a suitable tool to study cardiac electrophysiology in a 3 R-compliant manner. We found that rostafuroxin treatment ameliorated ouabain-induced electrophysiological disturbances, suggesting rostafuroxin as a potential therapeutic intervention for ouabain mis- or overdosing.NEW & NOTEWORTHY This study evaluates rostafuroxin, a selective ouabain inhibitor, for its potential to antagonize electrophysiological derangements in ouabain overdosing. Methodologically, the study uses the iCE model, previously introduced as a suitable 3 R-compliant cardiovascular research platform. We developed and validated a comprehensive electrophysiological workflow in iCEs to perform our investigations. Ouabain increased heart rate variability, induced arrhythmia and electrolyte imbalances in iCEs, whereas rostafuroxin largely protected them from these effects.

OBJECTIVE: To provide real-world data to inform benchmarking goals and practical issues that influence optimal antenatal corticosteroid timing and to examine patient factors, such as gestational age at steroid administration and presenting diagnoses, associated with steroid administration in relation to delivery.

METHODS: This is a retrospective cohort study of singleton deliveries between July 1, 2016, and December 31, 2024, at two large academic hospitals with level IV neonatal intensive care units in a single health system. The primary cohort of interest was individuals who delivered between 24 0/7 and 33 6/7 weeks of gestation. The primary outcome of interest was the timing of antenatal corticosteroid administration in relation to delivery, categorized as none, delivery between 6 hours and 7 days after the first dose of antenatal corticosteroid ("optimally timed" per the Society for Maternal-Fetal Medicine's quality metric), and delivery less than 6 hours or more than 7 days after the first dose of antenatal corticosteroid ("suboptimally timed"). As a balancing measure to optimally timed antenatal corticosteroid administration, we also examined those who received antenatal corticosteroids before 34 weeks of gestation and delivered at term (after 37 weeks). We reported the rates of optimal timing and term delivery by their corresponding weeks of gestation and performed multivariable logistic regression modeling to understand patient factors and diagnoses associated with antenatal corticosteroid timing.

RESULTS: Among the 1,694 pregnant patients who delivered before 34 weeks of gestation, 961 (56.7%) had optimally timed antenatal corticosteroid administration, 162 (9.6%) received the first dose of antenatal corticosteroids less than 6 hours before delivery, 320 (18.9%) delivered more than 7 days after antenatal corticosteroid administration, and 251 (14.8%) did not receive antenatal corticosteroids. Of those who received antenatal steroids before 34 weeks of gestation, 747 of 2,879 (25.9%) delivered at term. There was little variation in optimal timing or term delivery by gestational age. Clinical factors associated with optimally timed antenatal corticosteroid administration compared with delivery more than 7 days after administration included pregnancy-related hypertensive disorder (adjusted odds ratio [aOR] 1.88, 95% CI, 1.31-2.69), preterm labor (aOR 2.78, 95% CI, 1.32-5.81), premature rupture of membranes (1.37, 95% CI, 1.33-1.42), anxiety disorder (aOR 079, 95% CI, 0.76-0.83), multiparous with no history of preterm birth (aOR 0.81, 95% CI, 0.77-0.86), placenta previa (aOR 0.76, 95% CI, 0.68-0.84), and placenta accreta (aOR 0.83, 95% CI, 0.81-0.85).

CONCLUSION: Achieving optimal timing of antenatal corticosteroid administration remains challenging. These findings underscore the need for improved prediction of preterm delivery and individualized patient assessment to ensure timely access to antenatal corticosteroids for women at risk of preterm birth while reducing unnecessary exposure.

Estrogen receptor (ER) positive breast cancer is the most prevalent subtype, commonly responsive to endocrine therapies. Immune checkpoint inhibitors (ICIs) have limited efficacy in ER-positive disease, highlighting the need for the development of combination immunotherapies for these patients. We previously established that nitroso-N-methylurea-induced mammary tumors in outbred Sprague-Dawley rats mimic immune evasive mechanisms and the heterogeneity of ICI response observed in patients. We identified a "luminal growing" gene signature in ER-positive tumors, which correlated with tumor growth and immune-related differences. Here, we evaluated targeting candidates from this signature KMT5B/C and IKBKE using inhibitors A-196 and IKBKEi respectively, alongside anti-estrogen (fulvestrant) and a TGFβ blocking antibody (NIS793), both individually and in combination with αPD-L1, within this rat model. Fulvestrant emerged as the most effective treatment, inducing regression of most existing tumors and reducing on-treatment tumor burden when combined with αPD-L1. A-196, while ineffective as a monotherapy, demonstrated enhanced response when combined with αPD-L1. Comprehensive tumor profiling through polychromatic flow cytometry and single-cell RNA sequencing revealed that A-196 induced a luminal-to-basal shift in tumor epithelial cells, enhancing antigen presentation, whereas epithelial-to-mesenchymal transition was linked to fulvestrant resistance. Our findings underscore the value of the rat mammary tumor model for preclinical studies in ER-positive breast cancer and advocate for the further validation and potential clinical development of KMT5B/C inhibitors to enhance the efficacy and broaden the applicability of ICI therapy in cancer patients.

Priming rare subdominant precursor B cells in germinal centers (GCs) is a central goal of vaccination to generate broadly neutralizing antibodies (bnAbs) against HIV. Multivalent immunogen display on protein nanoparticle scaffolds can promote such responses, but it also generates scaffold-specific B cells that could theoretically limit bnAb precursor expansion in GCs. We rationally designed DNA origami-based virus-like particles (DNA-VLPs) displaying a germline-targeting HIV envelope protein immunogen, which elicited no scaffold-specific antibody responses. Compared with a state-of-the-art clinical protein nanoparticle, these DNA-VLPs increased the expansion of epitope-specific GC B cells relative to off-target B cells and enhanced expansion of bnAb-lineage B cells in a humanized mouse model of CD4 binding site priming. Thus, minimizing off-target responses enhances bnAb priming and indicates that DNA-VLPs are a promising vaccine platform.

GGGGCC (G4C2) repeat expansion in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Toxicity is thought to result from the accumulation of either repeat RNAs and/or dipeptide repeat proteins (DPRs) translated from repeat-containing transcripts through repeat-associated non-AUG (RAN) translation. To disentangle RNA from DPR toxicity, we mutated a CUG codon predominantly used to initiate DPR translation from all three reading frames. This mutation disrupted DPR synthesis while preserving the expression of repeat-containing RNAs. Despite the accumulation of RNA foci, behavioral deficits and pathological abnormalities, including p-TDP-43 inclusions, STING activation, motor neuron loss, neuroinflammation, and increased plasma neurofilament concentration, were alleviated in C9ORF72 mice. Base editing of the CUG codon also improved molecular phenotypes and survival in patient induced pluripotent stem cell-derived neurons, which highlights the potential of therapeutically targeting DPR production rather than repeat RNAs.

Deficient extinction learning and memory are hypothesized mechanisms for pathological anxiety that are associated with sleep disturbance. fMRI neural activations to threat conditioning, extinction learning, and extinction recall were measured. Activations were compared, in persons with Generalized Anxiety Disorder (GAD), between those with moderate to severe Insomnia Disorder (ID) and those with absent or sub-threshold ID. Relationships of activations with measures of sleep quality and physiology were examined. Between-group comparisons and whole-sample correlation with sleep parameters were examined in relation to large-scale brain networks using a liberal cluster-determining threshold. Localized activations were then identified using family-wise error correction. Activations to the reinforced stimulus (CS+) that increased from the beginning to end ("across") threat conditioning were more extensive within the GAD+ID group. Increased activations to the CS+ across extinction learning were greater within the GAD-ID than the GAD+ID group, and delayed 24 h in the latter. Greater sleep efficiency was associated with decreased activations across threat conditioning, but with increased activations across extinction learning. Better sleep quality promoted greater engagement of neural substrates of extinction learning. The GAD+ID group failed to engage brain areas supporting extinction learning immediately following threat conditioning, but did so when stimuli were again presented following a delay.

Direct measurement of mitochondrial oxygen tension in vivo provides direct information on tissue metabolism and could facilitate new approaches in disease detection, function monitoring, and treatment efficacy assessment with cellular level data, with far superior sensitivity to oxygen changes than blood saturation measures. Here, a platform system was engineered to quantify fast sampling of oxygen partial pressure (pO2) inside tissue by utilizing the inherent rolling shutter readout of a smartphone CMOS camera detector for measuring the oxygen-sensitive time-resolved delayed fluorescence (DF) signal from Protoporphyrin IX (PpIX) which naturally occur in mitochondria for most tissues. The CMOS rolling shutter readout produces a microsecond-level time difference in the pixels row-by-row detection of light, here utilized as a time-gated shutter to sample the time distributed DF intensity. This novel technique eliminated the necessity of high-speed intensified camera with excitation isolation facility as well as advanced precise time synchronization system as required in the conventional time-resolved fluorescence lifetime measurement platforms for quantifying time-dependent very low intensity DF distribution of PpIX conjugated with prompt fluorescence. Both steady state and dynamic performance of the instrument were validated in tissue phantoms at different PpIX concentrations (0.5-10 μM) for wide range of pO2 detection (0-160 mmHg) with tunable fast response time (1-3.5 s) which is substantially faster than electrode-based systems that measure over 10's of seconds. Finally, it was tested in vivo to assess the impact of dynamic inhaled oxygen concentration variation on skin tissue pO2 under the conditions of normoxia, hyperoxia and hypoxia.

Fear is an evolutionarily adaptive mechanism that enables organisms to detect and respond to potential threats. Over the past century, theories of fear have evolved from Pavlovian and behavioral frameworks to neuroscientific models that emphasize specific neural circuits, neurotransmission, and plasticity. This pictorial review synthesizes key concepts underlying the neuroscience of fear, starting with a historical narrative of fear as conceptualized by conditioning and behaviorism theories, to the roles of different neurobiology structures, networks, to concepts of memory consolidation, reconsolidation, and plasticity. We highlight how pathological fear is implicated in disease, integrating evidence from conditioning theories, neuroscience, and additional considerations (ie. Culture and psychosocial context). Finally, we outline therapeutic approaches that leverage neuroscience to target maladaptive fear circuits. Understanding these mechanisms provides a foundation for advancing personalized treatments for fear-related psychopathology.