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Missense variants throughout ACTA2, encoding smooth muscle α-actin (αSMA), predispose to adult onset thoracic aortic disease, but variants disrupting arginine 179 (R179) lead to Smooth Muscle Dysfunction Syndrome (SMDS) characterized by childhood-onset diverse vascular diseases. Our data indicate that αSMA localizes to the nucleus in wildtype (WT) smooth muscle cells (SMCs), enriches in the nucleus with SMC differentiation, and associates with chromatin remodeling complexes and SMC contractile gene promotors, and the ACTA2 p.R179 variant decreases nuclear localization of αSMA. SMCs explanted from a SMC-specific conditional knockin mouse model, Acta2SMC-R179C/+, are less differentiated than WT SMCs, both in vitro and in vivo, and have global changes in chromatin accessibility. Induced pluripotent stem cells from patients with ACTA2 p.R179 variants fail to fully differentiate from neural crest cells to SMCs, and single cell transcriptomic analyses of an ACTA2 p.R179H patient’s aortic tissue shows increased SMC plasticity. Thus, nuclear αSMA participates in SMC differentiation and loss of this nuclear activity occurs with ACTA2 p.R179 pathogenic variants.

Atherosclerotic plaques consist mostly of smooth muscle cells (SMC), and genes that influence SMC phenotype can modulate coronary artery disease (CAD) risk. Allelic variation at 15q22.33 has been identified by genome-wide association studies to modify the risk of CAD and is associated with the expression of

We demonstrated that joint delivery of human-derived endothelial progenitor cell (EPC) and smooth muscle cell (SMC) sheets mimics the native architecture of structurally mature blood vessels and contributes to limiting ventricular remodeling using a confluent SMC-EPC bi-level cell sheet engineered by cell-sheet technology and transplanted into an athymic rodent model of myocardial infarction. Enhanced vasculogenic potential was observed in vitro when EPCs were stimulated with SMC-conditioned culture medium, augmenting angiogenesis in vivo. Increased structurally mature vessel density, myocardial upregulation of biological adhesion, and vasculature developmental genes in the ischemic border zone myocardium showed interaction between cells and the extracellular matrix. Cell fate tracking experiments featuring xenogeneic transplantation showed transplanted EPCs and SMCs to have elements of the newly formed vasculature. Specialized magnetic resonance imaging of the cell-sheet-transplanted rodents suggested prolonged cell retention. The robust angiogenic effect of the transplanted cell sheets induced reverse ventricular remodeling of the ischemic heart. Bioinformatic analyses indicated that these cell sheets promote transcriptome-wide changes in the left ventricular response to acute ischemia, promote productive remodeling, and prevent pathological ventricular dilation. Thus, the human-derived, spatially arranged SMC-EPC bi-level cell sheet is a promising therapy for increasing myocardial viability and limiting adverse ventricular remodeling after myocardial infarction.

Atherosclerotic plaques consist mostly of smooth muscle cells (SMC), and genes that influence SMC biology can modulate coronary artery disease (CAD) risk. Allelic variation at 15q22.33 has been identified by genome-wide association studies to modify the risk of CAD, and is associated with expression of SMAD3 in SMC, but the mechanism by which this gene modifies CAD risk remains poorly understood. SMC-specific deletion of Smad3 in a murine atherosclerosis model resulted in greater plaque burden, more positive remodeling, and increased vascular calcification. Single-cell transcriptomic analyses revealed that loss of Smad3 altered SMC transition cell state toward two fates: a novel SMC phenotype that governs both vascular remodeling and recruitment of inflammatory cells, as well as a chondromyocyte fate. The remodeling population was marked by uniquely high Mmp3 and Cxcl12 expression, and its appearance correlated with higher risk plaque features such as increased positive remodeling and macrophage content. Further, investigation of transcriptional mechanisms by which Smad3 alters SMC cell fate revealed novel roles for Hox and Sox transcription factors whose direct interaction with Smad3 regulate an extensive transcriptional program balancing remodeling and vascular extracellular matrix with significant implications for atherosclerotic and Mendelian aortic aneurysmal diseases. Together, these data suggest that Smad3 expression in SMC inhibits the emergence of specific SMC phenotypic transition cells that mediate adverse plaque features, including positive remodeling, monocyte recruitment, and vascular calcification.

Introduction This quality improvement project (QI) focused on reducing procedural pain. Phase I of this project demonstrated a need to improve our patients' and nurses’ satisfaction with pain control management during hydrotherapy. In this phase (Phase II), we educated the nursing staff on 1) appropriate dosing and timing of opioid administration (Oral and IV) and 2) need for more frequent midazolam administration before hydrotherapy. We also assessed the safety and potential benefits of ketamine for procedural pain control. Methods Patients undergoing wound cleaning procedures were surveyed for up to three hydrotherapy encounters. Nurses were educated on procedural pain control prior to this current phase. A convenience sample of patients underwent ketamine administration. Ketamine was administered per protocol starting at 0.3 or 0.4 mg/kg with intermittent boluses of 0.25 mg/kg as needed. Demographics, opioid (oral morphine equivalents, OME), midazolam, and ketamine doses and time of administration, and adverse events were collected. Patient pain scores (1-10) before and after hydrotherapy and patient and nurse satisfaction scores (1-10) after hydrotherapy were collected. Paired t-tests and one-way ANOVA were performed. P < 0.05 was considered significant. Results During Phase II, 9 patients and 19 encounters were analyzed; of which ketamine was administered for 10 events. Compared to Phase I (28 Patients and 50 encounters), education of the nursing staff resulted in a significant improvement in giving midazolam prior to hydrotherapy (78.9% vs. 10%; p < 0.001) and opioid administration (27.4 OME vs. 20.2 OME, p = 0.043). Timing of opioid administration did not improve when compared to Phase I (Outside the window of efficacy: 42.1% vs. 43.8%, p = 0.903). Both patient (8.7 ± 2.4 vs. 7.7 ± 2.1; p = 0.011) and nurse (8.2 ± 2.1 vs. 7.3 ± 1.9; p = 0.021) satisfaction scores regarding pain control improved. None of the patients who received ketamine experienced adverse effects. Notably, the use of ketamine improved nurse satisfaction scores with patient pain control (9.2 ± 0.8 vs. 7.1 ± 2.6; p = 0.035). While patients on ketamine were more satisfied with pain control, significance was not reached (9.4 ± 1.1 vs. 7.9 ± 3.2; p = 0.315). Conclusions Our data demonstrate that nurse education regarding medication administration was effective, but further education is needed to improve the timing of opioid administration. Ketamine offers additional benefits that deserve additional study.

e18650 Background: Tracking diagnostic discrepancies is a common quality indicator in anatomic pathology. Many cases are re-reviewed as care is transferred from one facility to another. Most published data on these discrepancies is from the perspective of the tertiary receiving facility. Disparities in patient access to pathology expertise and technologies in small community hospitals can affect the accuracy of cancer diagnosis and the quality of care. Mary Lanning Healthcare (MLH) is a regional community hospital with a well-established referral pattern to several neighboring health systems. As part of a quality assurance process, we evaluated the diagnostic concordance rate of our cancer related pathology diagnoses with the referred institutions. Methods: Between 2017 and 2020, a cohort of cancer related cases was identified where the initial diagnosis was at MLH, then as part of coordinated care, a second pathologic interpretation was rendered at another instution. Data regarding specimen type, discrepancy in original diagnosis, nature and severity of discrepancy, disagreement in histologic grade, concordance or lack thereof with third party reviewers, and distribution among reviewing pathologists were collected. Results: A total of 521 cancer related cases (890 specimens) were sent to 16 facilities for second opinion. There were 46 (5%) discrepancies. Majority of them 45 (98%) were minor. Third party review of one major discrepancy came back in agreement with our original diagnosis. The most common discrepancy was interobserver variability in findings without strictly defined criteria. A change in histologic grade was the second most frequent deviation with prostate and breast being the most common sites. Upgrades and downgrades to the original diagnoses were nearly evenly split (19 to 21). A total of 33 pathologists were involved. While one reviewing pathologist generated 30% of all discrepancies, 3 other pathologists accounted for 67%. Conclusions: Accurate pathologic diagnoses significantly impact clinical outcomes. Our data which represents a rural community-based cancer program identified an overall discrepancy rate in the range of 0.1 to 1.1% for second opinion review. The vast majority of differences were minor in nature with no change in patient care and could be attributed to expected interobserver variability.[Table: see text]