Lung carcinogenesis is causally linked to cigarette smoking, in part by epigenetic changes. We tested whether accumulated epigenetic change in smokers is apparent in bronchial basal cells as cells of origin of squamous cell carcinoma. Using an EM-seq platform covering 53.8 million CpGs (96% of the entire genome) at an average of 7.5 sequencing reads per CpG site at a single base resolution, we evaluated cytology-normal basal cells bronchoscopically brushed from the in situ tobacco smoke-exposed ‘bronchial epithelial field’ and isolated by short-term primary culture from 54 human subjects. We found that mean methylation was globally lower in ever (former and current) smokers versus never smokers (p = 0.0013) across promoters, CpG shores, exons, introns, 3’-UTRs, and intergenic regions, but not in CpG islands. Among 6mers with dinucleotides flanking CpG, those containing CGCG showed no effect from smoking, while those flanked with TT and AA displayed the strongest effects. At the gene level, smoking-related differences in methylation level were observed in CDKL1, ARTN, EDC3, CYP1B1, FAM131A, and MAGI2. Among candidate cancer genes, smoking reduced the methylation level in KRAS, ROS1, CDKN1A, CHRNB4, and CADM1. We conclude that smoking reduces long-term epigenome-wide methylation in bronchial stem cells, is impacted by the flanking sequence, and persists indefinitely beyond smoking cessation.

Endometrial and ovarian cancers are among the most common gynecologic malignancies in the U.S., with 5-year overall survival rates of 81% and 50% respectively. Ovarian cancer often presents late due to asymptomatic early stages and nonspecific symptoms in advanced stages. Few markers exist for early detection, and they lack the sensitivity and specificity needed for high-risk screening. Given the absence of informative protein blood biomarkers, cell-free DNA analysis has become a promising tool for early detection, treatment evaluation, and monitoring. However, previous studies of circulating tumor DNA (ctDNA) in gynecological cancers had limited sensitivity. We adapted MASQ (Multiplex Accurate Sensitive Quantitation), originally developed to measure residual cells in leukemia, to the measurement of ctDNA in solid cancers. MASQ detects patient-specific mutations simultaneously in up to 50 loci with exceptional sensitivity, below one part per million. This study uses MASQ to measure tumor-specific variants in blood (cell-free DNA) from endometrial and ovarian cancer patients, aiming to determine the range of ctDNA signals at presentation and explore correlations with clinicopathological features and patient outcomes. We analyzed samples from 46 gynecologic cancer patients (12 ovarian, 34 endometrial) treated at Northwell Health. Samples were collected in collaboration with gynecologic oncology research coordinators and the Northwell Health Biorepository. Peripheral blood was collected at diagnosis (n=46) and post-surgery (n=14), along with surgical tumor specimens. Whole genome sequencing of tumor and blood identified thousands of somatic tumor variants per patient. For each patient, 30 representative variants were selected for MASQ sequencing. MASQ libraries were generated from cell-free DNA isolated from pre- and post-surgery plasma. Ultrasensitive variant counts across 30 loci were aggregated and analyzed for clinical correlations. ctDNA signal was detected in 35 of 46 cases at presentation, with variant allele frequencies ranging from below 10^-4 to 10^-1. ctDNA signal was higher in ovarian cancer (13/13 detected) than endometrial cancer, and higher in aggressive Type II endometrial cancer (15/16) vs. Type I (5/13). ctDNA levels correlated with tumor size and copy number imbalance, but not with total cell-free DNA. Detection of ctDNA predicted worse overall survival (p=0.023) and was the strongest predictor of survival in multivariate analysis. Post-surgery follow-up in 10 of 14 patients showed significant ctDNA reduction following treatment, highlighting MASQ’s sensitivity in tracking treatment response. This study highlights the potential of sensitive ctDNA detection for early diagnosis and monitoring in gynecologic cancers. However, limited cell-free DNA abundance necessitates even more sensitive methods to enhance clinical utility. Andrea B. Moffitt, Jude Kendall, Joan Alexander, Asya Stepansky, Arisa Kapedani, Zihua Wang, Dan Levy, Kenny Ye, Abba M. Krieger, Marina Frimer, Gary L. Goldberg, Michael Wigler. Personalized detection of circulating cell-free tumor DNA in gynecologic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5890.

BRD4 is a bromodomain-containing transcriptional co-regulator that plays important roles in driving transcription by binding to histone acetyl-lysines at enhancers and promoters while recruiting additional transcriptional cofactors. While the mechanisms by which BRD4 regulates transcription have been explored, the critical acetylations primarily responsible for targeting it to chromatin remain unclear. Through a machine learning approach, we determined that distinct sets of histone acetylations dominate the prediction of chromatin accessibility and BRD4 binding in distinct chromatin contexts (e.g. intergenic enhancers, gene body enhancers and promoters). Using human fibroblasts engineered to predominantly express specific histones with lysine-to-arginine mutations, we demonstrate that one such acetylation, H2BK120ac, is required to recruit BRD4 specifically to intergenic enhancers, while not affecting chromatin accessibility. Loss of H2BK120ac did not affect BRD4 binding to either promoters or gene body enhancers, demonstrating that the rules governing BRD4 recruitment to regulatory regions depends on the specific genomic context. Highlighting the importance of H2BK120ac in directing BRD4 recruitment, we found that expression of the H2BK120R mutant significantly reduces the phenotypes driven by BRD4-NUT, an oncogenic fusion protein that drives NUT midline carcinoma. This work demonstrates the critical nature that genomic context plays in BRD4 recruitment to distinct classes of regulatory elements, and suggests that intergenic and gene body enhancers represent classes of functional distinct elements.

Lung diseases, including lung cancer, are rising causes of global mortality. Despite novel imaging technologies and the development of biomarker assays, the detection of lung cancer remains a significant challenge. However, the lung communicates directly with the external environment and releases aerosolized droplets during normal tidal respiration, which can be collected, stored and analzsed as exhaled breath condensate (EBC). A few studies have suggested that EBC contains extracellular vesicles (EVs) whose microRNA (miRNA) cargos may be useful for evaluating different lung conditions, but the cellular origin of these EVs remains unknown. In this study, we used nanoparticle tracking, transmission electron microscopy, Western blot analyses and super resolution nanoimaging (ONi) to detect and validate the identity of exhaled EVs (exh-EVs). Using our customizable antibody-purification assay, EV-CATCHER, we initially determined that exh-EVs can be selectively enriched from EBC using antibodies against three tetraspanins (CD9, CD63 and CD81). Using ONi we also revealed that some exh-EVs harbour lung-specific proteins expressed in bronchiolar Clara cells (Clara Cell Secretory Protein [CCSP]) and Alveolar Type II cells (Surfactant protein C [SFTPC]). When conducting miRNA next generation sequencing (NGS) of airway samples collected at five different anatomic levels (i.e., mouth rinse, mouth wash, bronchial brush, bronchoalveolar lavage [BAL] and EBC) from 18 subjects, we determined that miRNA profiles of exh-EVs clustered closely to those of BAL EVs but not to those of other airway samples. When comparing the miRNA profiles of EVs purified from matched BAL and EBC samples with our three tetraspanins EV-CATCHER assay, we captured significant miRNA expression differences associated with smoking, asthma and lung tumor status of our subjects, which were also reproducibly detected in EVs selectively purified with our anti-CCSP/SFTPC EV-CATCHER assay from the same samples, but that confirmed their lung tissue origin. Our findings underscore that enriching exh-EV subpopulations from EBC allows non-invasive sampling of EVs produced by lung tissues.

Objectives Soccer heading is adversely associated with neurocognitive performance, but whether greater neck strength or anthropometrics mitigates these outcomes is controversial. Here, we examine the effect of neck strength or anthropometrics on associations of soccer heading with neurocognitive outcomes in a large cohort of adult amateur players. Methods 380 adult amateur league soccer players underwent standardized measurement of neck strength (forward flexion, extension, left lateral flexion, right lateral flexion) and head/neck anthropometric measures (head circumference, neck length, neck circumference and neck volume). Participants were assessed for heading (HeadCount) and cognitive performance (Cogstate) on up to 7 visits over a period of two years. Principal components analysis (PCA) was performed on 8 neck strength and anthropometric measures. We used generalized estimating equations to test the moderation effect of each of the three PCs on 8 previously identified adverse associations of 2-week and 12-month heading estimates with cognitive performance (psychomotor speed, immediate verbal recall, verbal episodic memory, attention, working memory) and of unintentional head impacts on moderate to severe central nervous system symptoms. Results 3 principal components (PC’s) account for 80% of the variance in the PCA. In men, PC1 represents head/neck anthropometric measures, PC2 represents neck strength measures, and PC3 represents the flexor/extensor (F/E) ratio. In women, PC1 represents neck strength, PC2 represents anthropometrics, and PC3 represents the F/E ratio. Of the 48 moderation effects tested, only one showed statistical significance after Bonferroni correction, which was not robust to extensive sensitivity analyses. Conclusion Neither neck strength nor anthropometrics mitigate adverse associations of soccer heading with cognitive performance in adult amateur players.

Rationale and Objectives This study examined the brain effects of mild severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection which are incompletely understood. Our objective was to ascertain within-person changes associated with mild coronavirus disease 2019 (COVID-19) in otherwise healthy adults. Materials and Methods We leveraged existing pre-pandemic baseline neuroimaging and neurocognitive data, and collected follow-up data from uninfected controls and individuals with prior mild COVID-19, during December 2020 and January 2021, when vaccines were not yet available. We compared change during follow-up in patients (n=5) versus controls (n=15). Results We identified a decrease of intracellular volume fraction (ICVF), decrease of isotropic volume fraction (ISO) and decrease of orientation dispersion index (ODI) in multiple inferior frontal regions of interest in COVID-19 patients; this longitudinal change was significantly different from the control group which demonstrated increases in equivalent measures. This pattern suggests injury with neuronal loss and/or inflammation as underlying mechanisms. Neurocognitive studies identified a pattern of cognitive decline (processing speed, executive function, verbal learning, working memory) in patients, that did not reach significance. Conclusion Our pilot data suggests that mild COVID-19 may result in brain pathology and impact neurocognitive function in younger adults in a manner parallel to prior findings in older individuals. Though findings may not generalize to other SARS-CoV-2variants, larger longitudinal studies of mild COVID-19 should be undertaken to understand the potential clinical implications of these findings over the longer term.

Survival following lung transplant is low. With limited donor lung availability, predicting post-transplant survival is key. We investigated the predictive value of pre-transplant CT biomarkers on survival. In this single-center retrospective cohort study of adults in a diverse, underserved, urban lung transplant program (11/8/2017–5/20/2022), chest CTs were analyzed using TeraRecon to assess musculature, fat, and bone. Erector spinae and pectoralis muscle area and attenuation were analyzed. Sarcopenia thresholds were 34.3 (women) and 38.5 (men) Hounsfield Units (HU). Visceral and subcutaneous fat area and HU, and vertebral body HU were measured. Demographics and pre-transplant metrics were recorded. Survival analyses included Kaplan–Meier and Cox proportional hazard. The study cohort comprised 131 patients, 50 women, mean age 60.82 (SD 10.15) years, and mean follow-up 1.78 (SD 1.23) years. Twenty-nine percent were White. Mortality was 32.1

There is growing concern that repetitive subconcussive head impacts, independent of concussion, alter brain structure and function, and may disproportionately affect the developing brain. Animal studies of repetitive subconcussive head impacts are needed to begin to characterize the pathological basis and mechanisms underlying imaging and functional effects of repetitive subconcussive head impacts seen in humans. Since repetitive subconcussive head impacts have been largely unexplored in animals, we aimed to characterize the evolution of imaging, behavioural and pathological effects of repetitive subconcussive head impacts in awake adolescent rodents. Awake male and female Sprague Dawley rats (postnatal Day 35) received 140 closed-head impacts over the course of a week. Impacted and sham-impacted animals were restrained in a plastic cone, and unrestrained control animals were included to account for effects of restraint and normal development. Animals (n = 43) underwent repeated diffusion tensor imaging prior to and over 1 month following the final impact. A separate cohort (n = 53) was assessed behaviourally for fine motor control, emotional-affective behaviour and memory at acute and chronic time points. Histological and immunohistochemical analyses, which were exploratory in nature due to smaller sample sizes, were completed at 1 month following the final impact. All animals tolerated the protocol with no overt changes in behaviour or stigmata of traumatic brain injury, such as alteration of consciousness, intracranial haemorrhage or skull fracture. We detected longitudinal, sex-dependent diffusion tensor imaging changes (fractional anisotropy and axial diffusivity decline) in corpus callosum and external capsule of repetitive subconcussive head impact animals, which diverged from both sham and control. Compared to sham animals, repetitive subconcussive head impact animals exhibited acute but transient mild motor deficits. Repetitive subconcussive head impact animals also exhibited chronic anxiety and spatial memory impairment that differed from the control animals, but these effects were not different from those seen in the sham condition. We observed trends in the data for thinning of the corpus callosum as well as regions with elevated Iba-1 in the corpus callosum and cerebral white matter among repetitive subconcussive head impact animals. While replication with larger study samples is needed, our findings suggest that subconcussive head impacts cause microstructural tissue changes in the developing rat brain, which are detectable with diffusion tensor imaging, with suggestion of correlates in tissue pathology and behaviour. The results point to potential mechanisms underpinning consequences of subconcussive head impacts that have been described in humans. The congruence of our imaging findings with human subconcussive head impacts suggests that neuroimaging could serve as a translational bridge to advance study of injury mechanisms and development of interventions.

Frailty is characterized by increased vulnerability to disability and high risk for mortality in older adults. Identification of factors that contribute to frailty resilience is an important step in the development of effective therapies that protect against frailty. First, a reliable quantification of frailty resilience is needed. We developed a novel measure of frailty resilience, the Frailty Resilience Score (FRS), that integrates frailty genetic risk, age, and sex. Application of FRS to the LonGenity cohort (n = 467, mean age 74.4) demonstrated its validity compared to phenotypic frailty and its utility as a reliable predictor of overall survival. In a multivariable-adjusted analysis, 1-standard deviation increase in FRS predicted a 38% reduction in the hazard of mortality, independent of baseline frailty (p < .001). Additionally, FRS was used to identify a proteomic profile of frailty resilience. FRS was shown to be a reliable measure of frailty resilience that can be applied to biological studies of resilience.

To compare residential geography, sex, socioeconomic status (SES), and race/ethnicity of patients screened at Montefiore’s Lung Cancer Screening Program with those of patients diagnosed with lung cancer, assessing whether screening efforts are appropriately focused. This retrospective cohort study involved patients within a multisite urban medical center undergoing lung cancer screening or diagnosed with lung cancer from January 1, 2015 to December 31, 2019. Inclusion criteria were residence within the Bronx, NY and age between 55 and 80 years. Institutional review board approval was obtained. Data were analyzed using the Wilcoxon two-sample t test and χ2. The cohorts comprised 1568 (50.3%) women and 1551 (49.7%) men (mean age 65.6 ± 6.16). The Southeast Bronx had the most diagnosed lung cancers (29.96%) and screenings (31.22%). Sex did not significantly differ (p = 0.053). Cancer and screening cohorts were from impoverished neighborhoods with mean SES of − 3.11 ± 2.78 and − 3.44 ± 2.80 (p < 0.01). The lower tier SES neighborhoods demonstrated more patients in the screening cohort than cancer cohort (p = 0.01). Both cohorts included a majority of Hispanic patients, although race/ethnicity differed significantly (p = 0.01). Lower SES neighborhoods showed no significant difference in race/ethnicity between cancer and screening cohorts (p = 0.262). Though statistically significant differences were found between cohorts, likely due to sample size, few clinically meaningful differences were found, implying our lung cancer screening program was effective in reaching the desired population. Demographics-based programs should be considered in global efforts to screen vulnerable populations.

Purpose To determine long-term clinical outcomes in patients with suspected acute aortic syndrome (AAS) and evaluate the prognostic value of coronary calcium burden as assessed with CT aortography in this symptomatic population. Materials and Methods A retrospective cohort of all patients who underwent emergency CT aortography from January 2007 through January 2012 for suspected AAS was assembled. A medical record survey tool was used to evaluate subsequent clinical events over 10 years of follow-up. Events included death, aortic dissection, myocardial infarction, cerebrovascular accident, and pulmonary embolism. Coronary calcium scores were computed from original images using a validated simple 12-point ordinal method and categorized into none, low (1-3), moderate (4-6), or high (7-12) groupings. Survival analysis with Kaplan-Meier curves and Cox proportional hazard modeling was performed. Results The study cohort comprised 1658 patients (mean age, 60 years ± 16 [SD]; 944 women), with 595 (35.9%) developing a clinical event over a median follow-up of 6.9 years. Patients with high coronary calcium demonstrated the highest mortality rate (adjusted hazard ratio = 2.36; 95% CI: 1.65, 3.37). Patients with low coronary calcium demonstrated lower mortality, but rates were still almost twice as high compared with patients with no detectable calcium (adjusted hazard ratio = 1.89; 95% CI: 1.41, 2.53). Coronary calcium was a strong predictor of major adverse cardiovascular events (P < .001), which persisted after adjustment for common significant comorbidities. Conclusion Patients with suspected AAS had a high rate of subsequent clinical events, including death. CT aortography-based coronary calcium scores strongly and independently predicted all-cause mortality.Keywords: Acute Aortic Syndrome, Coronary Artery Calcium, CT Aortography, Major Adverse Cardiovascular Events, Mortality Supplemental material is available for this article. © RSNA, 2023See also commentary by Weir-McCall and Shambrook in this issue.

Studying thousands of families, we find siblings concordant for autism share more of their parental genomes than expected by chance, and discordant siblings share less, consistent with a role of transmission in autism incidence. The excess sharing of the father is highly significant (p value of 0.0014), with less significance for the mother (p value of 0.31). To compare parental sharing, we adjust for differences in meiotic recombination to obtain a p value of 0.15 that they are shared equally. These observations are contrary to certain models in which the mother carries a greater load than the father. Nevertheless, we present models in which greater sharing of the father is observed even though the mother carries a greater load. More generally, our observations of sharing establish quantitative constraints that any complete genetic model of autism must satisfy, and our methods may be applicable to other complex disorders.

Objective: At certain institutions and radiology practices, a routine lumbar radiographic exam may include 3 views: AP, lateral, and coned-down lateral of the lumbosacral junction. The purpose of this study is to determine whether the third coned-down-lateral view adds significant diagnostic information regarding pathology at the L4-L5 and L5-S1 levels.Materials and methods: This retrospective study includes patients (n = 74) who had a 3-view radiographic exam of the lumbar spine, as well as a CT or MRI within six months. The AP and lateral views were reviewed by three radiologists, both with and without the use of the third, coned-lateral view. Subsequently, the CT and MRI performed within 6 months was reviewed, and the results compared. The primary outcome was detection of abnormal alignment and disc disease at the L4-L5 and L5-S1 levels.Results: For the combined findings of alignment and disc disease at each L4-L5 and L5-S1, there was disagreement between the 2-view and 3-view exams on 18 (of 296) evaluations. Of these 18, the 2-view and the 3-view exam each made positive findings on 9. By the binomial test, there is no evidence that either the 2-view or the 3-view exam tends to make more findings than the other (p = 1). Compared to CT/MRI, the 2-view exam agrees on 74.7 % of evaluations and the 3-view exam agrees on 75.3 %. There is therefore no evidence that the 3-view exam is more accurate than the 2-view exam.Conclusion: Elimination of the coned-down lateral view could reduce radiation exposure and imaging-related costs while maintaining diagnostic quality.

Introduction The objective of this study is to assess the role of age at first exposure (AFE) to soccer heading as a predictor of known adverse associations of recent and longer-term heading with brain microstructure, cognitive, and behavioral features among adult amateur soccer players. Methods The sample included 276 active amateur soccer players (196 male and 81 female) aged 18–53 years old. AFE to soccer heading was treated as a binary variable, dichotomized at ≤ 10 years vs. >10 years old, based on a recently promulgated US Soccer policy, which bans heading for athletes ages 10 and under. Results We found that soccer players who began heading at age 10 or younger performed better on tests of working memory ( p = 0.03) and verbal learning ( p = 0.02), while accounting for duration of heading exposure, education, sex, and verbal intelligence. No difference in brain microstructure or behavioral measures was observed between the two exposure groups. Discussion The findings indicate that, among adult amateur soccer players, AFE to heading before age 10 compared to later start of heading, is not associated with adverse outcomes, and may be associated with better cognitive performance in young adulthood. Cumulative heading exposure across the lifespan, rather than early life exposure, may drive risk for adverse effects and should be the focus of future longitudinal studies to inform approaches to enhance player safety.

Background: mAbs (biologics) are indicated in patients with poorly controlled moderate-to-severe asthma. The process of prior authorization and administration of a biologic requires exceptional commitment from clinical teams. Objective: Our aim was to evaluate the process of approval and administration of biologics for asthma and determine the most common reasons associated with denials of biologics and delays in administration. Methods: We examined the records of patients with asthma who were prescribed biologics from January 2018 to January 2020 at 2 centers, Montefiore Medical Center (Bronx, NY) and Scripps Clinics (San Diego, Calif). Demographics, insurance information, and details on the approval process were collected. Results: After querying of electronic health records, the records of 352 and 70 patients with moderate-to-severe asthma were included from Montefiore and Scripps, respectively. Most patients at Montefiore (58.2%) were insured under Managed Care Medicaid (MC Medicaid), whereas most patients at Scripps (61.4%) had commercial insurance. The median times from prescription to administration of a biologic were similar: 34 days (interquartile range [IQR] = 18-63 days) and 34 days (IQR = 22.5-56.0 days) (P = .97) for Montefiore and Scripps, respectively. However, the median approval time for Montefiore was 6 days (IQR = 1-20 days) and that for Scripps was 22 days (IQR = 10-36 days) (P < .001). Approval times for prescriptions requiring appeals were significantly longer than for prescriptions approved after the initial submission: 23 days versus 2.5 days and 40.5 days versus 15.5 days (for Montefiore and Scripps, respectively [P < .001 for both]). Conclusions: Lengthy appeals contribute to delays between prescribing and administering a biologic. Site-specific practices and insurance coverage influence approval timing of the biologics for asthma.

For detecting field carcinogenesis non-invasively, early technical development and case-control testing of exhaled breath condensate microRNAs was performed. In design, human lung tissue microRNA-seq discovery was reconciled with TCGA and published tumor-discriminant microRNAs, yielding a panel of 24 upregulated microRNAs. The airway origin of exhaled microRNAs was topographically "fingerprinted", using paired EBC, upper and lower airway donor sample sets. A clinic-based case-control study (166 NSCLC cases, 185 controls) was interrogated with the microRNA panel by qualitative RT-PCR. Data were analyzed by logistic regression (LR), and by random-forest (RF) models. Feasibility testing of exhaled microRNA detection, including optimized whole EBC extraction, and RT and qualitative PCR method evaluation, was performed. For sensitivity in this low template setting, intercalating dye-based URT-PCR was superior to fluorescent probe-based PCR (TaqMan). In application, adjusted logistic regression models identified exhaled miR-21, 33b, 212 as overall case-control discriminant. RF analysis of combined clinical + microRNA models showed modest added discrimination capacity (1.1-2.5%) beyond clinical models alone: all subjects 1.1% (p = 8.7e-04)); former smokers 2.5% (p = 3.6e-05); early stage 1.2% (p = 9.0e-03), yielding combined ROC AUC ranging from 0.74 to 0.83. We conclude that exhaled microRNAs are qualitatively measureable, reflect in part lower airway signatures; and when further refined/quantitated, can potentially help to improve lung cancer risk assessment.

Background: Since many COVID-19 publications lack consensus reviews or controls, interpretive accuracy is unclear; abdominal processes unique or infrequent during the pandemic remain unknown. The incidence and nature of CT findings accounting for abdominal complaints in COVID patients, reader agreement and overcalling will be determined.Methods: A retrospective study was performed on COVID patients with abdominal complaints from 3/15/ 2020-5/31/2020 and 11/1/2020-4/15/2021 including matched controls. Reviewers blinded to initial reads interpreted abdominopelvic CT exams, with discordant cases resolved in consensus. Reader agreement was measured by Cohen's Kappa, differences between cohorts by permutation tests and factors affecting false positive/negative rates by Fisher's Exact Test and logistic regression.Results: 116 first wave (average age 65 years [+/- 15.3], 63 [54%] women) and 194 second wave COVID cases (average age 64 years [+/- 16.3], 103 [53%] women) including 116 wave 1 and 194 wave 2 prepandemic controls were included. Concordance was lower among COVID cases than controls (Cohen's Kappa of 0.58 vs. 0.82 [p < 0.001]) and among wave 1 than wave 2 cases (Cohen's Kappa of 0.45 vs. 0.66 [p = 0.052]). With true positives defined as consensus between the initial reader and study reader, false positive rates were higher among COVID cases than controls (OR = 0.42, p = 0.003) and for initial than study reader (OR = 0.36, p < 0.001), but lower in wave 2 than 1 (OR = 0.5, p = 0.028).Conclusion: Greater reader disagreement occurred during COVID than prepandemic with no reader bias as both initial and study readers called more false positives among COVID cases than controls. More overcalling occurred during COVID with colitis and cystitis most common.

The intestinal epithelium consists of cells derived from continuously cycling Lgr5hi intestinal stem cells (Lgr5hi ISCs) that mature developmentally in an ordered fashion as the cells progress along the crypt-luminal axis. Perturbed function of Lgr5hi ISCs with aging is documented, but the consequent impact on overall mucosal homeostasis has not been defined. Using single-cell RNA sequencing, the progressive maturation of progeny was dissected in the mouse intestine, which revealed that transcriptional reprogramming with aging in Lgr5hi ISCs retarded the maturation of cells in their progression along the crypt-luminal axis. Importantly, treatment with metformin or rapamycin at a late stage of mouse lifespan reversed the effects of aging on the function of Lgr5hi ISCs and subsequent maturation of progenitors. The effects of metformin and rapamycin overlapped in reversing changes of transcriptional profiles but were also complementary, with metformin more efficient than rapamycin in correcting the developmental trajectory. Therefore, our data identify novel effects of aging on stem cells and the maturation of their daughter cells contributing to the decline of epithelial regeneration and the correction by geroprotectors.

Abstract New Western-style diet 1 (NWD1), a purified diet establishing mouse exposure to key nutrients recapitulating levels that increase human risk for intestinal cancer, reproducibly causes mouse sporadic intestinal and colonic tumors reflecting human etiology, incidence, frequency, and lag with developmental age. Complex NWD1 stem cell and lineage reprogramming was deconvolved by bulk and single-cell RNA sequencing, single-cell Assay for Transposase-Accessible Chromatin using sequencing, functional genomics, and imaging. NWD1 extensively, rapidly, and reversibly, reprogrammed Lgr5hi stem cells, epigenetically downregulating Ppargc1a expression, altering mitochondrial structure and function. This suppressed Lgr5hi stem cell functions and developmental maturation of Lgr5hi cell progeny as cells progressed through progenitor cell compartments, recapitulated by Ppargc1a genetic inactivation in Lgr5hi cells in vivo. Mobilized Bmi1+, Ascl2hi cells adapted lineages to the nutritional environment and elevated antigen processing and presentation pathways, especially in mature enterocytes, causing chronic, protumorigenic low-level inflammation. There were multiple parallels between NWD1 remodeling of stem cells and lineages with pathogenic mechanisms in human inflammatory bowel disease, also protumorigenic. Moreover, the shift to alternate stem cells reflects that the balance between Lgr5-positive and -negative stem cells in supporting human colon tumors is determined by environmental influences. Stem cell and lineage plasticity in response to nutrients supports historic concepts of homeostasis as a continual adaptation to environment, with the human mucosa likely in constant flux in response to changing nutrient exposures. Implications: Although oncogenic mutations provide a competitive advantage to intestinal epithelial cells in clonal expansion, the competition is on a playing field dynamically sculpted by the nutritional environment, influencing which cells dominate in mucosal maintenance and tumorigenesis.