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CD40 signalling is involved in chronic inflammation, which is increasingly recognized as an important factor in initiation and progression of various cardiovascular diseases. Inhibition of the CD40-TRAF6 pathway was previously shown to be effective in preventing atherosclerosis and obesity in mice (Seijkens, JACC 2018, Chatzigeorgiou, PNAS 2014). In the present study we investigated the effect of CD40-TRAF6 inhibitor, compound 6877002 on left ventricular (LV) and coronary microvascular function in an adult swine model of familial hypercholesterolemia (FH). For this purpose, 10 adult male FH swine (3 years old, homozygous LDLR R84C mutation) received an atherogenic diet (10% lard and 0.75% cholesterol) for 9 months. During the last 3 months, 5 swine were subjected to treatment with the 6877002 compound (0.16 mg/kg i.v), while 5 animals received the vehicle i.v. At the end of treatment, swine were anesthetized and LV performance was studied under closed-chest conditions by pressure-volume loops constructed at baseline and during cardiac preload reduction. Subsequently, small coronary arteries (±250 μm diameter) were harvested from the LV and endothelium-dependent response to bradykinin in the presence and absence of endothelial nitric oxide (eNOS) blockade with LNAME, as well as endothelium-independent responses to the nitric oxide donor SNAP were studied in vitro. 6877002-treated animals showed a lower LV end-diastolic pressure (0±2 mmHg) as compared to vehicle-treated animals (6±1 mmHg, p≤0.05), and trends towards a reduced LV stiffness, reflected in a lower slope of the end diastolic pressure-volume relationship (sEDPVR: 0.16±0.02 mmHg/ml vs 0.20±0.01 mmHg/ml, p=0.07), and a higher ejection fraction (EF 46±2% vs 40±2%, p=0.08). Data from isolated small coronary arteries indicated that endothelium-independent vasorelaxation in response to SNAP was similar in both 6877002- and vehicle-treated animals, indicating unaltered vascular smooth smooth mucle cell responsiveness to NO. However, 6877002 treatment improved endothelium-dependent vasorelaxation to bradykinin (p≤0.05), as compared to vessels isolated from vehicle-treated animals. Moreover, eNOS blockade with LNAME blunted the endothelium-dependent vasorelaxation to bradykinin only in vessels from 6877002-treated animals, but not in those from the vehicle-treated swine, suggesting that NO bioavailability was completely lost in the vehicle-treated FH-animals, which was at least partially restored by treatment with 6877002 Conclusion: – Initation of CD40-TRAF6 pathway inhibition in adult FH swine receiving a high fat diet reduced LV end-diastolic pressure and enhanced LV passive compliance. The improved LV performance was accompanied by an amelioration of coronary microvascular endothelial dysfunction, characterized by the partial restoration of NO bioavailability. Funding: Grant 2020B008 RECONNEXT This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

The translation of AI-generated brain metastases (BM) segmentation into clinical practice relies heavily on diverse, high-quality annotated medical imaging datasets. The BraTS-METS 2023 challenge has gained momentum for testing and benchmarking algorithms using rigorously annotated internationally compiled real-world datasets. This study presents the results of the segmentation challenge and characterizes the challenging cases that impacted the performance of the winning algorithms. Untreated brain metastases on standard anatomic MRI sequences (T1, T2, FLAIR, T1PG) from eight contributed international datasets were annotated in stepwise method: published UNET algorithms, student, neuroradiologist, final approver neuroradiologist. Segmentations were ranked based on lesion-wise Dice and Hausdorff distance (HD95) scores. False positives (FP) and false negatives (FN) were rigorously penalized, receiving a score of 0 for Dice and a fixed penalty of 374 for HD95. The mean scores for the teams were calculated. Eight datasets comprising 1303 studies were annotated, with 402 studies (3076 lesions) released on Synapse as publicly available datasets to challenge competitors. Additionally, 31 studies (139 lesions) were held out for validation, and 59 studies (218 lesions) were used for testing. Segmentation accuracy was measured as rank across subjects, with the winning team achieving a LesionWise mean score of 7.9. The Dice score for the winning team was 0.65 ± 0.25. Common errors among the leading teams included false negatives for small lesions and misregistration of masks in space. The Dice scores and lesion detection rates of all algorithms diminished with decreasing tumor size, particularly for tumors smaller than 100 mm3. In conclusion, algorithms for BM segmentation require further refinement to balance high sensitivity in lesion detection with the minimization of false positives and negatives. The BraTS-METS 2023 challenge successfully curated well- annotated, diverse datasets and identified common errors, facilitating the translation of BM segmentation across varied environments and providing the tools for future development of personalized volumetric reports to patients undergoing BM treatment.

Introduction: Atherosclerosis, the underlying cause of cardiovascular disease (CVD), is a lipid-driven chronic inflammatory disease. We have identified the co-stimulatory CD40-CD40L immune checkpoint dyad as a key driver of inflammation during atherosclerosis. CD40 and CD40L exert cell-type divergent functions via different signaling pathways. Hypothesis: We hypothesize that CD40 on endothelial cell (ECs) induces endothelial-to-mesenchymal transition (EndMT)-mediated vascular inflammation in atherosclerosis. Methods & Results: Using single cell RNAseq, we found that CD40 (TNFRSF5) is expressed on PDE3A + GJA5 + and KDR + COL15A1 + ECs of advanced human carotid atherosclerotic plaques. CD40 is highly expressed in the endothelial lining of human and mouse atherosclerotic plaques, and colocalizes with the EndMT markers fibronectin and collagen and adhesion molecule ICAM-1. CD40L activation of human aortic endothelial cells (HAECs) results in expression of the EndMT markers fibronectin, vimentin, sm22α, FSP1 and snail. Activation of CD40 in EC induces filamin A dependent translocation of CD40 into lipid rafts, increases TNF-receptor associated factor (TRAFs) -2, -3 and 6 levels, induces Akt signaling and increases expression of VCAM1 and CCL2. Conclusions: The endothelial cell has been more and more regarded as a cell type that plays a major role in driving inflammation. Our data suggest that the CD40-CD40L axis plays a significant role in EC-mediated inflammation in atherosclerosis by driving EndMT.

SUMMARY Bloodstream infections (BSIs) are a significant cause of mortality and morbidity. Rapid identification of pathogens and detection of a few resistance markers from positive blood cultures are now possible through the increased availability of commercial rapid diagnostic tests, including nucleic acid amplification tests and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. This document describes the clinical utility of rapid diagnostics performed on positive blood cultures and provides evidence-based laboratory medicine guidelines for using rapid tests to diagnose BSIs in hospitalized adult and pediatric patients. This guideline was developed for use by medical (a.k.a. clinical) microbiologists, medical laboratory professionals, infectious disease clinicians, pharmacists, hospital administrators, healthcare providers, and other stakeholders associated with BSIs. A panel of experts, including medical microbiologists and experts in systematic literature review, was assembled to formulate the Population–Intervention–Comparison–Outcome (PICO) question, review the literature, and provide recommendations for using rapid tests to diagnose BSI and improve patient outcomes. A comprehensive literature search of four electronic bibliographic databases (MEDLINE, Embase, CINAHL, and Cochrane) was conducted to identify studies with measurable outcomes. The panel followed a systematic process, which included a standardized methodology for rating the certainty of the evidence and strength of recommendations using the GRADE (Grading of Recommendations, Assessment, Development, and Evaluation) approach. The panel evaluated the literature to answer the question: Does using rapid diagnostic tests improve clinical outcomes in adult and pediatric patients hospitalized with a BSI? Peer-reviewed literature was available to address three outcomes, including time to targeted therapy, mortality, and length of hospital stay. In general, the quality of the evidence was low to moderate due to the paucity of controlled, randomized clinical trial studies. However, eight recommendations were made based on evidence derived from the systematic review of the published literature. To answer the PICO question, the expert committee recommended using rapid diagnostic tests combined with active communication to decrease the time to targeted therapy and length of stay (strong recommendation). While the strength of the evidence for the impact on mortality is low, the panel supports using rapid tests to impact these outcomes. A summary of the recommendations is listed in the Executive Summary, which includes a detailed description of the background, methods, evidence summary, and rationale that supports each recommendation in the full text.

AbstractSecure messaging platforms were designed to optimize healthcare communication by providing asynchronous, bidirectional, Health Insurance Portability and Accountability Act‐compliant text messaging options. Proponents argue that these platforms streamline workflows, foster coordination across multidisciplinary teams, and improve patient safety. However, critics highlight their potential to disrupt workflows through overwhelming message volumes and task interruptions, increased risk for miscommunication, and possible detrimental impacts on patient care. While secure messaging shows promise for modernizing hospital communication, it requires institutional guidelines and thoughtful use to mitigate inefficiencies and risks.

10510 Background: The impact of germline pathogenic or likely pathogenic variants (PVs) in cancer predisposition genes on overall survival (OS) after breast cancer diagnosis is not well defined. In particular, unbiased OS estimates from population-based studies accounting for clinical subtypes of breast cancer (based on ER, PR, and HER2 status) and for genes other than BRCA1/2 are lacking. Methods: The study included 25,168 prospectively followed women with locoregional invasive breast cancer within the population-based CARRIERS study. Germline sequencing using a custom multigene amplicon-based panel was performed to identify PVs. OS was compared between germline PV carriers in ATM, BRCA1, BRCA2, CHEK2 and PALB2 and non-carriers (negative for germline PVs in 13 known breast cancer predisposition genes) for each clinical subtype of breast cancer from the time of breast cancer diagnosis to death or last follow up in a multivariable Cox proportional hazard regression analysis adjusting for age at diagnosis, race/ethnicity, histology, TNM stage, type of surgery, use of adjuvant radiation, chemotherapy and endocrine agents. Results: Among 25,168 women with breast cancer, germline PVs in one of the five breast cancer predisposition genes were detected in 5.8% of the women [ ATM : 0.8%, BRCA1 : 1.3%, BRCA2: 1.6%, CHEK2 : 1.5%, and PALB2 : 0.6%]. Among women with ER+ breast cancer, compared to non-carriers, a significantly worse OS was noted for BRCA1 (Hazard Ratio (HR): 1.8, 95% Confidence Interval (CI): 1.1 – 3.0, p=0.02) and BRCA2 (HR: 1.8, 95%CI: 1.4 – 2.4, p<0.001), but not for ATM (HR: 1.2, 95%CI: 0.8 – 1.7, p=0.40) or CHEK2 (HR:1.0, 95%CI: 0.8 – 1.3, p=1.0) PV carriers. Worse OS was noted in PALB2 PV carriers with ER+ breast cancer (HR:1.5, 95%CI: 0.9 – 2.2, p=0.09), although it did not reach statistical significance. Similar HR estimates for OS as with ER+ breast cancer were observed in women with ER+/HER2- breast cancer for BRCA1 , BRCA2 and PALB2 PV carriers, but only BRCA2 results were statistically significant. Among women with ER-negative breast cancer, compared to non-carriers, a significant difference in OS was not observed for PV carriers in ATM (HR:0.8, 95%CI: 0.4 – 1.8, p=0.63), BRCA1 (HR:1.2, 95%CI: 0.8 – 1.7, p=0.35), BRCA2 (HR: 0.8, 95%CI: 0.5 – 1.2, p=0.29), CHEK2 (HR: 0.7, 95%CI: 0.3 – 1.4), or PALB2 (HR: 0.9, 95%CI: 0.5 -1.7, p=0.78). Conclusions: The suggestive differences in OS by ER status of the tumor in BRCA1 and BRCA2 PV carriers warrant further investigation of underlying tumor biology and assessment of endocrine sensitivity of breast cancer in germline PV carriers.

Low-grade endometrial stromal sarcomas (LG-ESS) are the second most common malignant uterine mesenchymal tumors, but in contrast to the more common leiomyosarcomas, they are often characterized by a prolonged and relatively indolent course. However, a subset of patients experience significant morbidity or die of disease, and it is difficult to predict which tumors will behave aggressively, with most published studies limited in either the number of tumors or the depth of pathologic parameters evaluated. Thus, we studied the clinicopathologic features of LG-ESS in 102 patients ranging from 21 to 74 (median: 47) years. All were treated with hysterectomy and staged according to both the FIGO 2018 system (stage IA=22, IB=36, I-not otherwise specified=5, II=16, III=13, IV=10) and the FIGO 1988 system (stage I=62, II=1, III=17, IV=22). Tumors measured 1.2-49 (median: 7) cm. Microscopically, 69 involved the endometrium while 33 were centered in the myometrium. Thirteen showed only minimal infiltration of the myometrium while the rest displayed the typical extensive myometrial permeation. The cervical stroma was involved in 18, the uterine serosa in 27, and the parametrium in 22. Conventional morphology resembling proliferative endometrial stroma was seen in 95, fibroblastic appearance in 35, smooth muscle differentiation in 23, sex cord-like differentiation in 21, stromal hyalinization in 21, and myxoid stroma in 9. Less common features included glandular differentiation resembling adenomyosis (n=5), pseudopapillary pattern (n=1), deciduoid appearance (n=2), adipocytic differentiation (n=2), multinucleated cells (n=2), and rhabdomyoblastic differentiation (n=1). Mitoses ranged from <1 to 20 per 10 high-power fields (median=3). Lymphovascular invasion and infarct-type necrosis were present in 64 and 23, respectively. Follow-up was available in all patients ranging from 16 to 358 (median: 79) months. Forty-six received adjuvant treatment as hormonal therapy (n=34), radiation (n=4), radiation and hormonal therapy (n=4), chemotherapy (n=3), or chemotherapy and radiation (n=1). Three patients had persistent unresected tumor following surgery, and an additional 34 had recurrences at intervals of 3 to 272 (median: 79) months, including 2 tumors with minimal infiltration. At last follow-up, 75 patients were alive with no evidence of disease, 14 were alive with disease, and 9 died of disease at intervals of 16 to 167 (median=70) months. Four died of unrelated causes without recurrence. Five-year recurrence-free survival (RFS) and disease-specific survival (DSS) were 80% and 94%, while 10-year RFS and DSS were 51% and 87%, respectively. On statistical analysis, cervical stromal involvement ( P =0.018) and myxoid stroma ( P <0.001) were associated with shorter recurrence-free survival. Tumors lacking a conventional component had worse disease-specific survival ( P =0.048). All other clinical and morphologic features, including stage, were not significantly associated with outcome. On multivariate analysis, only cervical stromal involvement remained an independent predictor of recurrence-free survival ( P =0.047; HR: 16.939) and no factors were independently predictive of disease-specific survival. Our findings highlight the difficulty in predicting outcomes in these tumors, likely due to slow progression and frequent treatment responses even in the recurrent setting. We confirm the potential for recurrence even in tumors initially showing minimal infiltration. Cervical stromal involvement and lack of conventional morphology are potential novel risk factors that should be further evaluated in subsequent studies.

Cardiovascular disease is the global #1 cause of mortality and morbidity. The majority of cardiovascular diseases is caused by atherosclerosis, a lipid-driven, inflammatory disease of the middle- and large-sized arteries. The disease is characterized by the formation of atherosclerotic plaques throughout the arterial tree. Over the years, insights into the pathogenesis of atherosclerosis have shifted from a ‘lipid-driven’ model to a ‘response-to-injury’ perspective, and more recently to a ‘lipid-driven inflammatory disease’ viewpoint. We are now aware that a network of multiple immune cell-types and -subsets of the innate and adaptive immune system inhabit our arteries. Intricate interactions between these immune cellsubsets, non-immune cells, and local environmental substances such as lipids, cell debris and calcium cause a fluidic balance of pro-inflammatory and regulatory responses. A dysregulation of this balance towards a pro-inflammatory milieu drives atherosclerotic disease progression. Although we have acknowledged that atherosclerosis is an inflammatory disease, state-of-theart treatments are still based on lipid lowering, anti-hypertensive and lifestyle-changing strategies. In the past decade, clinical phase I, II and III trials targeting the immune system revealed that patients tolerate immunotherapy, show decreased inflammation and/or had a reduction in cardiovascular endpoints. However, the search for novel immunotherapeutic targets and treatment regimens as well as stratification of patients who would benefit from such treatments to combat atherosclerotic cardiovascular disease is only just beginning. In this review article, we will highlight the newest insights on the different cell subsets and components of the immune system in atherosclerosis and elaborate on current and future immunotherapeutics to treat atherosclerotic cardiovascular disease.