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Cardiac microthrombi are postulated to underlie cardiac injury in critical COVID-19. To determine pathogenic mechanism(s) of cardiac injury in fatal COVID-19, we conducted a single-center prospective cohort study of 69 consecutive COVID-19 decedents. Microthrombi was the most commonly detected acute cardiac histopathologic feature (n=48, 70%). We tested associations of cardiac microthrombi with biomarkers of inflammation, cardiac injury, and fibrinolysis and with in-hospital antiplatelet therapy, therapeutic anticoagulation, and corticosteroid treatment, while adjusting for multiple clinical factors, including COVID-19 therapies. Higher peak ESR and CRP during hospitalization were independently associated with higher odds of microthrombi (ESR, Pnon-linearity0.015, Passociation=0.008; CRP per 20mg/L increase, OR 1.17, 95%CI 1.00-1.36). Using single nuclei RNA-sequence analysis, we discovered an enrichment of prothrombotic, anti-fibrinolytic, and extracellular matrix signaling amongst cardiac fibroblasts in microthrombi-positive COVID-19 hearts, compared with microthrombi-negative COVID-19 hearts and non-COVID-19 donor hearts. Our cumulative findings identify these specific transcriptomic changes in cardiac fibroblasts as salient features of COVID-19-associated cardiac microthrombi.

Introduction:Approximately 50% of all heart failure (HF) diagnoses can be classified as HF with preserved ejection fraction (HFpEF), which has no FDA approved therapies. HFpEF is more prevalent in females compared to males, but the underlying mechanisms for the development as a sex-based disorder are unknown. We previously described how slow progressive pressure overload (PO) in male felines recapitulates the HFpEF phenotype but have not investigated the female phenotype.Hypothesis:Females will develop a less severe HFpEF phenotype compared to males under the same pathological stress.Methods & Results:Male (m) and female (f) domestic short felines (age 2mo) underwent aortic constriction (m: n=11; f: n=10) using a customized pre-shaped band or a sham procedure (m: n=7; f: n=7). Before surgery (baseline), there was no difference in body weight (BW) between groups and lung compliance was not different. Echocardiography revealed no significant difference in the ratio of left atrium to aortic root (LA/Ao), LA ejection fraction (LA EF), left ventricle (LV) ejection fraction, LV wall-thickness, and E/A ratio. By 4mo post-surgery, both males and females had developed cardiac dysfunction and decreased lung compliance. At this time, females had significantly smaller BWs than males. Despite the difference in BW, LV wall thickness and changes in E/A ratio were similar in both sexes in banded vs. shams. Importantly, LV EF did not change in any group. There was a decrease in LA EF and increased LA/Ao in all banded animals. Invasive hemodynamics at 4mo post-surgery showed no differences between sexes for the systolic pressure gradient generated by aortic banding. Banded males had higher LV end-diastolic pressure vs. banded females (m: 15.0±2.7mmHg; f: 8.1±1.9mmHg). However, there was a trend towards prolongation of tau and lower dp/dtminin banded females, suggestive of abnormal relaxation. There were no differences between banded males and females in heart weight/BW or cardiomyocyte cross-sectional area and both developed fibrosis.Conclusions:Exposure of male and female felines to PO resulted in similar cardiac hypertrophy, fibrosis, and decreased lung compliance. Females had lower LVEDP than males suggesting they may be protected from diastolic dysfunction.

Introduction:Atrial fibrillation (AF) often arises from dysfunction in the left atrium (LA), a tissue with limited annotation of the non-coding genome and its effects on gene expression. As many AF-associated genetic variants reside in these regions, this knowledge gap hinders efforts to understand AF biology.Methods:In order to generate a model of the LA non-coding genome, we profiled 7 histone post-translational modifications (active: H3K4me3, H3K4me2, H3K4me1, H3K27ac, H3K36me3; repressive: H3K27me3, H3K9me3), CTCFbinding, and gene expression in 4 LA samples from individuals without structural heart disease or AF. We used MACS2 to identify peak regions (P < 0.01), applied a Markov model approach to classify regulatory elements, and then annotated with gene expression. In order to better relate this model with disease biology, we then intersected chromatin states with independently derived eQTL, DNA methylation, and HiC long-range chromatin interaction data from primary human LA and left ventricle.Results:Our model identified 21 states, which recapitulate regulatory motifs such as promoters, enhancers, and repressed regions. Genes were actively regulated by proximal chromatin states, and repressive states was associated with a significant reduction in gene expression (P < 2x10-16). Further, chromatin states were differentially methylated, with promoters being less methylated than repressed regions (P < 2x10-16). For enhancer-gene associations, we identified 142 significantly enriched pathways (P < 0.05; FDR). Intersecting AF, PR, and QT SNP loci with cell type specific promoters identified a gene interaction network dominated by CAV1, CDKN1A, CREBP, NKX2-5and SYNPO2L.Discussion:Our profiling of the non-coding genome in human LA provides new insights into the gene expression and chromatin regulation within this AF-associated tissue. Further analyses of these data will be used to inform the relationship between GWAS variants and AF pathology.