Background:Non-atherosclerotic vascular aging (NAVA) contributes to cardiovascular risk through progressive arterial stiffening and endothelial dysfunction. Colchicine, best known for anti-inflammatory activity, has been proposed to protect vascular structure and function. We evaluated whether chronic colchicine mitigates NAVA in a smooth-muscle-specific ERCC1 knockout (SMC-KO) mouse model of DNA-damage-driven vascular aging. Methods:We performed experiments in the SMC-KO treated with colchicine (0.1mg/kg/day) or vehicle from the age of 10 to 22 weeks. Endothelial function was assessed by acetylcholine-induced vasorelaxation, and vascular structure by pulse-wave velocity (PWV), carotid intima-media thickness (cIMT), and elastin integrity. Results:SMC-KO mice developed increased arterial stiffness and impaired acetylcholine-mediated relaxation. Chronic colchicine significantly (p<0.01) lowered PWV, preserved elastin architecture, and improved endothelium-dependent relaxation, while sodium-nitroprusside responses and systemic cytokine levels remained unchanged. Conclusion:Chronic colchicine treatment preserves endothelial function and vascular elastin structure and reduces arterial stiffness in a DNA-damage-driven model of non-atherosclerotic vascular aging. These findings highlight colchicine's pleiotropic vascular benefits beyond anti-inflammation, supporting its potential repurposing for primary prevention in vascular aging and cardiovascular health or the development of colchicine-mimicking drugs with greater selectivity and improved safety profiles.

Ischaemic heart disease shows important differences between men and women, requiring an understanding of sex and gender dissimilarities to improve outcomes. This Scientific Statement provides an updated review of the current knowledge from risk factors to prognosis. It discusses the unequal impact of certain traditional risk factors between men and women, along with additional factors, such as hormonal changes and treatments (including those for transgender people and cancer), pregnancy-related complications, and autoimmune diseases, which contribute to the sex-specific risk profiles. Moreover, it outlines functional and structural sex differences in the pathophysiology (e.g. coronary atheroma plaques and burden, coronary dissection, vasospasm, and microvascular disease) with women being more prone to microvascular disease and endothelial dysfunction, while paradoxically experiencing less severe myocardial ischaemia at similar levels of coronary stenosis. The document further addresses the evaluation of diagnostic tools, which often have a male-centric bias, resulting in underdiagnosis in women who also tend to receive less guideline-recommended treatment. Additionally, women can have different responses and side effects to various preventive and therapeutic treatments, potentially contributing to the worse prognosis documented in acute coronary syndromes with obstructive coronary artery disease, particularly at a young age. Considering all these sex and gender differences and the low enrolment of women in randomized controlled trials, questions arise regarding the optimal treatment for women. Addressing sex differences requires conducting sex-specific research to close the knowledge gap. Overall, the Scientific Statement highlights all relevant sex- and gender-specific dissimilarities to advance clinical practice and identify directions for future research to improve guideline recommendations for equitable care.

BACKGROUND:Heart failure (HF) progression is closely linked to oxidative stress. 5-Oxoproline (5-OP), a product of glutathione degradation, is normally metabolized by 5-oxoprolinase (OPLAH) but accumulates when the gamma-glutamyl cycle is disrupted. Here, we investigated the clinical characteristics of circulating 5-OP, its proteomic correlates, and the associations to outcome in HF. METHODS:In serum of 823 BIOSTAT-CHF patients, 5-OP was quantified by validated liquid chromatography-mass spectrometry and analyzed for associations with clinical outcomes. Proteomic correlates were identified across 355 OLINK proteins using stability selection with Minimax Concave Penalty regression. Mechanistic context was evaluated in a multi-comorbidity, large-animal cardio-kidney-metabolic (CKM) model with regional OPLAH assessment. RESULTS:Higher 5-OP was associated with worse renal function (eGFR declining across 5-OP tertiles, 67.9 to 60.2 mL/min/1.73 m2; p = 0.0012) and higher all-cause mortality (HR 1.55, 95% CI 1.11-2.17, p = 0.010). Per SD increase in log-5-OP, risk for the 2-year composite endpoint increased (HR 1.27, 95% CI 1.05-1.53), with broadly similar associations across CKD strata (interaction p = 0.63). TGF-α was the most robust proteomic correlate (π = 0.70; empirical permutation p = 0.001). In CKM swine, circulating 5-OP was elevated and renal cortical OPLAH protein, but not cardiac OPLAH, was selectively reduced, consistent with a renal contribution to systemic 5-OP elevation. CONCLUSION:Circulating 5-OP identifies HF patients at higher risk and is robustly associated with TGF-α. In a translational swine model, selective loss of renal cortical OPLAH provides tissue context supporting a renal contribution to systemic 5-OP elevation in cardiorenal syndrome.

Immune checkpoint inhibitor (ICI)-mediated myocarditis is a notorious complication of cancer treatment; however, the role of immune checkpoints in the heart during inflammation remains unknown. We investigated myocardial expression of programmed cell death protein 1 (PD-1) and its ligand (PD-L1) in non-ICI myocarditis. We performed a cross-species single-cell/-nucleus RNA sequencing comparative analysis, including 3 different models of myocarditis in mice and human hearts, with in vitro validation in human cells. This provided compelling evidence that PD-1/PD-L1 signaling in both murine and human non-ICI myocarditis is inherent to the myocardial inflammatory response. Specifically, cardiac endothelial cells and fibroblasts exhibited robust and sustained PD-L1 up-regulation in myocarditis, while also coinciding temporally with peak PD-1 expression on T cells, which presents an important defense mechanism. Therefore, immune checkpoints have global importance in myocarditis and may serve as a therapeutic target.

This study tested the hypothesis that K+ serves as an in vivo signal coupling coronary blood flow with the oxidative requirements of the myocardium. Experiments were performed in swine in which coronary parameters and arterial and coronary venous [K+] were measured under baseline conditions, during exogenous administration of K+ (1–5 mM; n = 4), during increases in myocardial oxygen consumption (MVO2) to dobutamine (n = 7) and exercise (n = 6), alterations in coronary perfusion pressure (CPP; n = 8), and systemic hypoxemia (PaO2 to 30 mmHg; n = 7). Exogenous intracoronary K+ increased blood flow ( 20

Myocardial edema significantly develops during current subnormothermic ex situ heart perfusion (ESHP) procedures, resulting in myocardial function decline during prolonged perfusion. A relatively high coronary blood flow (CBF) during ESHP is thought to be responsible for this high degree of myocardial edema formation. In this study, we present a novel tool to calculate CBF based on individual donor (sex and body weight) and perfusate (hemoglobin concentration, oxygen saturation, partial pressure of oxygen [PO2]) characteristics. The tool continuously evaluates the balance between myocardial oxygen consumption (MVO2) and delivery to facilitate adequate and preventing excess perfusion. Taking this personalized approach, the CBF can potentially be lowered while still providing sufficient oxygen to the donor heart. Furthermore, the tool automatically calculates MVO2, ΔPO2, and coronary vascular resistance during ESHP, which aids in the qualitative assessment of the heart before transplantation.

BACKGROUND:Despite successful recanalization after endovascular thrombectomy, more than half of patients with acute ischemic stroke with large-vessel occlusions experience an unsatisfactory outcome. Incomplete microvascular reperfusion may contribute to it, but its occurrence remains debated, partly due to clinical observations of hyperperfusion after recanalization. This study investigates the relationship between ischemia duration, infarct development, microclot presence, and cerebral perfusion in a swine model of focal cerebral ischemia and reperfusion. METHODS:Twenty-three swine underwent craniectomy and were randomized into 5 groups: 1-hour, 2-hour, or 4-hour occlusion followed by 4-hour recanalization, 8-hour occlusion without recanalization (positive control), or without occlusion (sham). Middle cerebral artery occlusion was induced using aneurysm clips, with 3-dimensional digital subtraction angiography confirming occlusion and recanalization. Three-dimensional digital subtraction angiography was used for quantification of area at risk and tissue perfusion. Infarct size was measured using 2,3,5-triphenyl-2H-tetrazolium chloride staining. The presence of microclots was quantified using CD61+ platelet immunostaining (number/mm2). Plasma markers of coagulation activation were measured before and during middle cerebral artery occlusion and after recanalization. RESULTS:Area at risk-normalized infarct size increased significantly with ischemia duration (r=0.8, P<0.001). Compared with the noninfarcted hemisphere, microclots in the infarcted hemisphere increased in middle cerebral artery occlusion groups (0.07 [0.04-0.13] versus 0.33 [0.16-0.71], P=0.003). Microclot density in area at risk region showed a positive correlation with ischemia duration (r=0.7, P=0.007), while no such correlation was observed in remote region. Simultaneously, higher perfusion levels were observed at both 2 hours (1.27±0.19, P<0.001) and 4 hours (1.23±0.16, P<0.001) following recanalization, irrespective of the duration of ischemia. No activation of coagulation was detected in systemic plasma. CONCLUSIONS:Focal cerebral ischemia and reperfusion result in a substantial presence of microclots, which occurs alongside hyperperfusion in this swine model of recanalized acute ischemic stroke. These platelet microclots extend beyond the ischemic area. The density of microclots within the ischemic region increased with prolonged ischemia.

The coronary microvasculature is principally responsible for matching coronary blood flow to myocardial demand of oxygen and nutrients. Short-term control of coronary blood flow is achieved via alterations in coronary microvascular tone, whereas long-term control of coronary flow also involves remodelling of the coronary microvasculature, including adjustments in vascular structure, diameter and density. In the past 50 years, considerable research efforts have been directed at understanding the functional and structural coronary microvascular adaptations involved in matching myocardial oxygen supply to demand, and how these mechanisms are affected by various diseases. In this review article, we will discuss our current understanding of the mechanisms underlying the regulation of coronary microvascular tone under healthy physiological conditions and in ischaemic heart disease. We will specifically discuss the role of microvascular dysfunction in obstructive and non-obstructive coronary artery disease, as studied in large animal models and confirmed in human studies. Future research should be directed at further unravelling the disease-specific mechanisms of coronary microvascular dysfunction in order to identify therapeutic targets to improve microvascular function in patients with ischaemic heart disease.

Background Endovascular catheters and devices used for thrombectomy in patients who had a stroke can damage the vessel lumen leading to microthrombi. During stroke recanalisation, microthrombi could migrate distally and occlude cerebral microvasculature, potentially limiting the benefit of recanalisation therapy.Objectives To describe vascular injury occurring after endovascular therapy (EVT), with stent retrievers (SR) and direct aspiration (DA), to open up avenues for further improvement of EVT technique.Methods SR and DA were performed according to clinical procedures in extracranial vessels in a swine model of thromboembolic arterial occlusion. Treated vessels were collected at 2 hours or 3 days post-EVT to assess respectively acute injury and early healing (remnant vascular injury) as assessed by Evans-Blue (EB) dye exclusion. The presence of microthrombi was quantified using scanning electron microscopy. Markers of coagulation activation were measured periprocedurally in plasma.Results Both SR and DA induced vascular injury. SR tended to result in larger EB positive areas than DA at 2 hours (99.5 vs 84.5; p=0.072) which reached statistical significance at day 3 (78.6 vs 48.6; p=0.040) post-EVT. Both EVT methods similarly yielded microthrombi in treated areas which were still observed at 3 days post-EVT. In addition, both EVT methods immediately increased systemic plasma levels of complexes of intrinsic-pathway coagulation activation: thrombin, Factor IX and Factor Xa:Antithrombin.Conclusions In this preclinical thromboembolic model, SR thrombectomy and DA lead to acute vascular injury, yield microthrombi and trigger contact activation of the coagulation system. At 3 days after intervention, healing remains incomplete, showing remnant vascular injury in the treated arteries, especially in SR thrombectomy.

Objective Assessing myocardial perfusion in acute myocardial infarction is important for guiding clinicians in choosing appropriate treatment strategies. Echocardiography can be used due to its direct feedback and bedside nature, but it currently faces image quality issues and an inability to differentiate coronary macro- from micro-circulation. We previously developed an imaging scheme using high frame-rate contrast-enhanced ultrasound (HFR CEUS) with higher order singular value decomposition (HOSVD) that provides dynamic perfusion and vascular flow visualization. In this study, we aim to show the ability of this technique to image perfusion deficits and investigate the potential occurrence of false-positive contrast detection. Methods We used a porcine model comprising occlusion and release of the left anterior descending coronary artery. During slow contrast agent infusion, the afore-mentioned imaging scheme was used to capture and process the data offline using HOSVD. Results Fast and slow coronary flow was successfully differentiated, presumably representing the different compartments of the micro-circulation. Low perfusion was seen in the area that was affected, as expected by vascular occlusion. Furthermore, we also imaged coronary flow dynamics before, during and after release of the occlusion, the latter showing hyperemia as expected. A contrast agent destruction test showed that the processed images contained actual contrast signal in the cardiac phases with minimal motion. With larger tissue motion, tissue signal leaked into the contrast-enhanced images. Conclusion Our results demonstrate the feasibility of HFR CEUS with HOSVD as a viable option for assessing myocardial perfusion. Flow dynamics were resolved, which potentially helped to directly evaluate coronary flow deficits.

Biological aging varies across individuals and tissues, influencing chronic diseases, including heart failure (HF). Emerging proteome techniques enable quantification of organ-specific aging acceleration (OAA), but whether OAA relates to HF severity and differs by sex remains unclear. We aim to assess the sex-related association between OAA of heart, artery and kidneys and HF severity, and to investigate relevant cardiometabolic risk factors of organ aging. In 556 participants from the HELPFul cohort, we estimated predicted biological age for heart, artery, and kidneys using plasma proteomics and calculated OAA as the deviation from chronological age. Associations between OAA and HF stage, echocardiographic parameters, and cardiometabolic risk factors were evaluated using regression models. Composite indices, including triglyceride-glucose body mass index (TyG-BMI), c-reactive protein-triglyceride glucose index and triglyceride-to-HDL cholesterol ratio were assessed for associations with advanced OAA. Mean age was 63 ± 9 years; 65

Background This study investigated oxygen handling of human hearts donated after circulatory death (DCD) on normothermic ex-situ heart perfusion (ESHP) and evaluated oxygen handling markers as adjuncts to cardiac viability assessment. Methods This single-center retrospective study included human DCD heart transplantation procedures using ESHP. Lactate concentrations, blood gas, myocardial oxygen consumption (MVO2), delivery (MDO2), and extraction (MEO2), coronary blood flow (CBF), coronary vascular resistance (CVR), and adenosine infusion were reported over time. Correlation between parameters was assessed, and statistical testing compared patients who did and did not require extracorporeal membrane oxygenation (ECMO) support after transplantation. Results Lactate concentrations decreased during ESHP in all transplanted hearts (n = 25) and increased in 1 rejected heart. Arterial partial pressure of oxygen (PO2) was 75.2 ± 2.9 kPa, with an arteriovenous ΔPO2 of 44.8 ± 10.4 kPa. Oxygen saturation was 100% in most arterial and venous samples. Average MVO2 was 2.7 ± 0.6 ml/min/100 g myocardium, MDO2 98.5 ± 20.4 ml/min, and MEO2 8.6 ± 1.8%. Average CVR was 0.025 ± 0.006 mm Hg min/ml/100 g and increased over time. ΔPO2 correlated strongly with MVO2 (R = 0.797, p < 0.001) and lactate trend (R = 0.799, p < 0.001) in transplanted hearts, without differences compared to the rejected heart with increasing lactate. Adenosine infusion on ESHP was significantly higher in patients requiring ECMO post-transplantation vs non-ECMO cases (11.7 (4.5-21.0) vs 2.2 (1.5-6.7) ml/h, p = 0.039). Conclusions Hearts on normothermic ESHP receive excessive MDO2, due to high PO2 and CBF, while the MVO2 is relatively low. Thus, CBF and PO2 can potentially be lowered. Furthermore, ΔPO2 could serve as additional marker of metabolic function under these hyperoxic circumstances. The adenosine infusion rate might predict post-transplantation ECMO requirement.

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): CONTRAST CONSORTIUM (CVON, Dutch Heart Foundation) Background Incomplete microvascular reperfusion (IMR) could be a detrimental factor impairing regained functionality in stroke patients despite early recanalization. However, the presence of this phenomenon remains to be ascertained. Gyrencephalic large-animal models of acute ischemic stroke (AIS) such as swine are gaining attention as a next step in translation of preclinical to clinical stroke research, with brain anatomy and hemostasis more similar to humans. However, the occurrence and relevance of IMR in swine models of AIS is not well described yet, since most models do not include recanalization. Purpose We aimed to study the presence of IMR after brain ischemia in a translational swine model of AIS. Methods Anesthetized female Yorkshire-Landrace pigs (n=23) underwent craniotomy to occlude the right-sided middle cerebral arteries with aneurysm clips. Clips were released after 1 (n=5), 2 (n=5), and 4 (n=5) hours of occlusion to allow recanalization for 4 hours (recanalized AIS, n=15) or left in place for 8 hours (non-recanalized AIS, n=5). Three animals underwent the craniotomy without vessel clipping (Sham, n=3). 3D angiography was used to confirm occlusion and recanalization. Brains were collected after sacrifice, stained with TTC and then processed for histologic analysis. Infarct size was determined by TTC staining and expressed as percentage of the infarcted hemisphere. Histologic sections of the upper quadrant of the left and right hemispheres were immuno-stained with CD61 to visualize platelet aggregates in the microvasculature. Intravascular platelet aggregates (Figure 2A) were quantified using automated software analysis (Orbit) (CD61+ aggregates/mm2). Results All animals survived until the end of the procedure. Figure 1 shows differences in infarct size depending on the duration of occlusion (p=0.006). Figure 2 depicts an overall higher presence of platelet (CD61+) aggregates in the right hemisphere (infarct) compared to the left hemisphere (intact) (Figure 2B). There was a significant difference in platelet aggregates between left and right hemisphere but groupwise comparison showed only a trend (p=0.075) for more aggregates in the right hemisphere, compared to left, after 4 hours occlusion. Linear regression model could not identify whether infarct size and duration of occlusion were independent predictors of the number of platelet aggregates in the right hemisphere (p=0.191). Conclusions IMR occurs after brain ischemia in a swine model of AIS as a result of microvascular platelet aggregates. Further analysis and experiments are needed to elucidate which predictors can explain intravascular platelet aggregation.Infarct SizePlatelet aggregates

Coronary atherosclerosis is caused by plaque build-up, with lipids playing a pivotal role in its progression. However, lipid composition and distribution within coronary atherosclerosis remain unknown. This study aims to characterize lipids and investigate differences in lipid composition across disease stages to aid in the understanding of disease progression. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) was used to visualize lipid distributions in coronary artery sections (n=17) from hypercholesterolemic swine. We performed histology on consecutive sections to classify the artery segments and to investigate colocalization between lipids and histological regions of interest in advanced plaque, including necrotic core and inflammatory cells. Segments were classified as healthy (n=6), mild (n=6), and advanced disease (n=5) artery segments. Multivariate data analysis was employed to find differences in lipid composition between the segment types, and the lipids' spatial distribution was investigated using non-negative matrix factorization (NMF). Through this process, MALDI-MSI detected 473 lipid-related features. NMF clustering described three components in positive ionization mode: triacylglycerides (TAG), phosphatidylcholines (PC), and cholesterol species. In negative ionization mode, two components were identified: one driven by phosphatidylinositol(PI)(38:4), and one driven by ceramide-phosphoethanolamine(36:1). Multivariate data analysis showed the association between advanced disease and specific lipid signatures like PC(O-40:5) and cholesterylester(CE)(18:2). Ether-linked phospholipids and LysoPC species were found to colocalize with necrotic core, and mostly CE, ceramide, and PI species colocalized with inflammatory cells. This study, therefore, uncovers distinct lipid signatures correlated with plaque development and their colocalization with necrotic core and inflammatory cells, enhancing our understanding of coronary atherosclerosis progression.

Abstract Background Over time, left ventricular diastolic dysfunction (LVDD) can progress towards heart failure with preserved ejection fraction (HFpEF). Yet, the identification of those at high risk of progression is challenging, and guidance on follow-up or preventive treatment is lacking. Purpose To study the incidence of HFpEF and changes over time in markers of LVDD severity in women and men with pre-clinical LVDD. In addition, we evaluate whether blood pressure and kidney function affect progression of LVDD. Methods We reinvited 146 participants from the HELPFul study (58% women and 42% men) with pre-clinical LVDD after a median follow-up of 4.3 [IQR: 3.9-4.7] years (Figure 1). The follow-up measurements were similar to baseline and encompassed a structured interview, physical examination, blood draw, electrocardiogram and (exercise) echocardiography. We determined HFpEF incidence and report changes over time in cardiovascular risk factors as well as echocardiographic characteristics and biomarkers of LVDD. Additionally, we studied whether changes in blood pressure and kidney function affected LVDD progression using generalized mixed models. LVDD progression was defined as an increase in plasma NT-proBNP levels and HFA-PEFF major functional and morphological abnormalities. All analyses were performed for women and men combined as well as stratified by sex. Results Fifteen (10%) of the 146 participants developed HF of whom 13 had HFpEF (9 women and 4 men) during follow-up. Over time, systolic and diastolic blood pressure levels remained stable while mean kidney function (eGFR) declined from 89±14 to 81±17 mL/min/1.73m2. Median NT-proBNP plasma levels increased from 71 [IQR: 44, 120] to 100 [IQR: 51, 157] pg/mL. The prevalence of major functional abnormalities according to the HFA-PEFF score increased from 84 to 91%, while the prevalence of morphological abnormalities increased from 25% to 39% (Figure 2). A higher systolic blood pressure in women and a higher diastolic blood pressure in men was associated with an increase in NT-proBNP plasma levels over time. Additionally, lower eGFR levels were related to increased NT-proBNP plasma levels over time in both men and women. There was a significant rise in the prevalence of major functional and morphological abnormalities with time, but blood pressure and kidney function did not affect this change over time. Conclusions A small proportion of women and men with preclinical LVDD developed incident HF over a 5-year follow-up period. High blood pressure and reduced kidney function were associated with increased levels of NT-proBNP over time. This highlights the need to further explore cardiorenal protection as a method to prevent HFpEF.Study design and HF outcomesLongitudinal changes in LVDD markers

Coronary microvascular angina occurs in both men and women, however, the majority of patients are postmenopausal women with multiple cardiovascular risk factors, such as diabetes mellitus (DM), chronic kidney disease (CKD) and dyslipidemia. No treatment is available for coronary microvascular dysfunction, requiring more in depth knowledge on the mechanisms underlying this syndrome in this expanding group of patients. Here we studied microvascular function in vivo and and in vitro in isolated small coronary arteries, in postmenopausal miniswine with multiple cardiovascular risk factors. DM (streptozotocin), hypercholesterolemia (HC, high fat, high sugar diet), CKD (renal artery embolization) and menopause (OVX) were induced in 5 adult female minipigs (1.5-2 years old, ±40kg, DM+HC+CKD+OVX) for 6 months, while 11 healthy female minipigs matched for age and weight, on normal pig chow served as controls (Control). At sacrifice, coronary flow reserve (CFR) in response to intracoronary infusion of adenosine was measured under anesthesia, and endothelium-dependent response to bradykinin in the presence and absence of eNOS blockade with LNAME, were studied in vitro in isolated small coronary arteries. 6 months of sustained hyperglycemia (17.5±1.0 in in DM+HC+CKD+OVX vs 8.2±0.9 mmol/l in Control), hypercholesterolemia (15.4±2.0 vs 1.7±0.1 mmol/l) and renal dysfunction (plasma total protein: 84±3 vs 72±1 g/l) were accompanied by systemic inflammation (TNFalpha: 62±7 vs 47±1 pg/ml, all p<0.05 by t-test). Coronary flow reserve was significantly reduced in animals with multiple risk factors (3.4±0.4 in DM+HC+CKD+OVX) as compared to the healthy animals (4.9±0.3 in Control, p<0.05). In vitro, coronary small arteries from DM+HC+CKD+OVX animals showed impaired endothelium-dependent vasodilation to bradykinin, which was mediated by a loss of nitric oxide. Conclusion: Postmenopausal minipigs with multiple risk factors, displayed severe coronary microvascular dysfunction as evidenced by a significantly reduced coronary flow reserve and endothelial dysfunction. The latter was characterized by a marked loss of nitric oxide. Such perturbations may contribute to reduced myocardial perfusion that is often observed in postmenopausal women with multiple cardiovascular risk factors. Funding: Grant 2020B008 RECONNEXT. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Purpose In this study, we explored the role of apoptosis as a potential biomarker for cardiac failure using functional micro-CT and fluorescence molecular tomography (FMT) imaging techniques in Ercc1 mutant mice. Ercc1 is involved in multiple DNA repair pathways, and its mutations contribute to accelerated aging phenotypes in both humans and mice, due to the accumulation of DNA lesions that impair vital DNA functions. We previously found that systemic mutations and cardiomyocyte-restricted deletion of Ercc1 in mice results in left ventricular (LV) dysfunction at older age. Procedures and Results Here we report that combined functional micro-CT and FMT imaging allowed us to detect apoptosis in systemic Ercc1 mutant mice prior to the development of overt LV dysfunction, suggesting its potential as an early indicator and contributing factor of cardiac impairment. The detection of apoptosis in vivo was feasible as early as 12 weeks of age, even when global LV function appeared normal, underscoring the potential of apoptosis as an early predictor of LV dysfunction, which subsequently manifested at 24 weeks. Conclusions This study highlights the utility of combined functional micro-CT and FMT imaging in assessing cardiac function and detecting apoptosis, providing valuable insights into the potential of apoptosis as an early biomarker for cardiac failure.

Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Dutch CardioVascular Alliance (an initiative with support of the Dutch Heart Foundation) Grant 2020B008 RECONNEXT, and the German Center for Cardiovascular Research (DZHK; 81Z0600207) Heart failure with preserved ejection fraction lacks targeted therapies, due to insufficient understanding of pathogenesis. A multiple comorbidity swine model (MCS) was developed by exposure to three risk factors for six months including diabetes (streptozotocin), chronic kidney disease (renal embolization), and an unhealthy diet (high-fat, high salt diet). An additional risk factor ovariectomy was added on top of the MCS model (MCS+). MCS and MCS+ presented with left ventricular diastolic dysfunction, oxidative stress, coronary microvascular dysfunction and inflammation. Furthermore, myocardial oxygen consumption was increased given the same level of cardiac work (Fig. A), suggesting an impairment in myocardial mitochondrial function. Using a combined-omics approach, the purpose of this study is to unravel alterations in gene expression and proteome abundance regarding mitochondrial function in MCS swine. 15 MCS, 4 MCS+ and 10 healthy control female swine were included in the study. Proteome analysis and single nuclei RNA sequencing, were performed on frozen MCS left ventricle myocardial samples. Cardiac mitochondrial function was measured by an O2k-FlouRespirometer ex vivo in MCS+ swine versus control. Proteome analysis showed a reduced abundance of proteins involved in branched-chain amino acid (BCAA) catabolism (Fig. B). Single nuclei RNA sequencing of a subgroup of the same animals demonstrated a downregulation of these genes specifically in cardiomyocyte subpopulations. BCAA can act as a source of ATP via oxidative phosphorylation and can modulate mitochondrial substrate utilisation, and impair mitochondrial function, resulting in increased mitochondrial production of reactive oxygen species (ROS). Mitochondrial function was measured in fresh myocardial tissue, using pyruvate/malate/glutamate as substrate. Low mitochondrial respiratory sensitivity to ADP was observed 26±2.5 in MCS+ vs 43.6±1.5 pmol/(s*mg tissue) in healthy control, reflecting low oxidative capacity. Moreover, the respiratory rate after uncoupling by FCCP was reduced in MCS+ versus healthy control (73.6±11.7 vs 118.4±15.6 pmol/(s*mg tissue). These findings are consistent with our data in the MCS swine and indicate impaired BCAA in the myocardium of animals with comorbidities, which is associated with increased ROS levels (8-isoprostane 12.9±0.8 pg/mg protein in MCS vs 10.3±0.5 in healthy animals) and impaired myocardial efficiency during exercise (Fig. A). A combined omics approach suggested an alteration in cardiac mitochondria. Respiratory analyses showed impaired mitochondrial bioenergetics and increased oxidative stress, which can contribute to diastolic dysfunction.

Assessing the coronary circulation with contrast-enhanced echocardiography has high clinical relevance. However, it is not being routinely performed in clinical practice because the current clinical tools generally cannot provide adequate image quality. The contrast agent's visibility in the myocardium is generally poor, impaired by motion and nonlinear propagation artifacts. The established multipulse contrast schemes (MPCSs) and the more experimental singular value decomposition (SVD) filter also fall short to solve these issues. Here, we propose a scheme to process amplitude modulation/amplitude-modulated pulse inversion (AM/AMPI) echoes with higher order SVD (HOSVD) instead of conventionally summing the complementary pulses. The echoes from the complementary pulses form a separate dimension in the HOSVD algorithm. Then, removing the ranks in that dimension with dominant coherent signals coming from tissue scattering would provide the contrast detection. We performed both in vitro and in vivo experiments to assess the performance of our proposed method in comparison with the current standard methods. A flow phantom study shows that HOSVD on AM pulsing exceeds the contrast-to-background ratio (CBR) of conventional AM and an SVD filter by 10 and 14 dB, respectively. In vivo porcine heart results also demonstrate that, compared to AM, HOSVD improves CBR in open-chest acquisition (up to 19 dB) and contrast ratio (CR) in closed-chest acquisition (3 dB).

Obesity is a modifiable cardiovascular risk factor, but adipose tissue (AT) depots in humans are anatomically, histologically, and functionally heterogeneous. For example, visceral AT is a pro-atherogenic secretory AT depot, while subcutaneous AT represents a more classical energy storage depot. Perivascular adipose tissue (PVAT) regulates vascular biology via paracrine cross-talk signals. In this position paper, the state-of-the-art knowledge of various AT depots is reviewed providing a consensus definition of PVAT around the coronary arteries, as the AT surrounding the artery up to a distance from its outer wall equal to the luminal diameter of the artery. Special focus is given to the interactions between PVAT and the vascular wall that render PVAT a potential therapeutic target in cardiovascular diseases. This Clinical Consensus Statement also discusses the role of PVAT as a clinically relevant source of diagnostic and prognostic biomarkers of vascular function, which may guide precision medicine in atherosclerosis, hypertension, heart failure, and other cardiovascular diseases. In this article, its role as a 'biosensor' of vascular inflammation is highlighted with description of recent imaging technologies that visualize PVAT in clinical practice, allowing non-invasive quantification of coronary inflammation and the related residual cardiovascular inflammatory risk, guiding deployment of therapeutic interventions. Finally, the current and future clinical applicability of artificial intelligence and machine learning technologies is reviewed that integrate PVAT information into prognostic models to provide clinically meaningful information in primary and secondary prevention.