Post-Procedure Monocyte Count Levels Predict Major Adverse Cardiovascular Events (MACE) Following Transcatheter Aortic Valve Implantation (TAVI) for Aortic Stenosis.

BACKGROUND: Aortic stenosis has recently been characterised as having an inflammatory aetiology, beyond the traditional degenerative model. Recruitment of monocytes has been associated with inflammation contributing to progression of calcific aortic-valve disease. Prior research has demonstrated that pre-procedure inflammatory biomarkers do not consistently discriminate poorer outcomes in those with aortic stenosis. It remains, however, unclear if postprocedure inflammatory biomarkers, which are influenced by intraprocedural pro-inflammatory insults, can predict major adverse cardiovascular events (MACE) post transcatheter aortic valve implantation (TAVI). METHOD: All patients with postprocedure monocyte levels undergoing transcatheter aortic valve implantation at The Alfred Hospital, Melbourne, Australia (2008-2019) were included. The highest monocyte count from postprocedure days 1 to 3 was used. Patients were divided into "high" or "low" postprocedure monocyte count groups using the Youden Index. The incidence of 30-day MACE a composite of stroke, acute myocardial infarction, and death) was then compared. RESULTS: In total, 472 patients were included (54% men, median age 84 years). Fourteen (14) patients (3%) suffered a 30-day MACE. Those with high postprocedure monocyte count were more likely to: be hypertensive (p=0.049); have a higher Society of Thoracic Surgeons risk score (p=0.032); and, undergo non-transfemoral access (p=0.018). A high (≥0.975) postprocedure monocyte count was significantly associated with 30-day MACE (odds ratio [OR] 1.16 for each 0.1 increase in monocyte, p=0.025). This association remained present on multivariable analysis adjusted for age, sex, Society of Thoracic Surgeons risk score, and self-expanding valve prosthesis type (OR 1.17, p=0.028). CONCLUSIONS: The association between postprocedure monocytosis and 30-day MACE suggests that minimising peri-procedural inflammatory insults may improve outcomes. This inexpensive and readily available biomarker may also aid in tailored risk stratification for patients.

Platelet-targeted thromboprophylaxis with a human serum albumin fusion drug: Preventing thrombosis and reducing cardiac ischemia/reperfusion injurywithout bleeding complications.

Background: Myocardial infarction (MI) as a consequence of atherosclerosis-associated acute thrombosis is a leading cause of death and disability globally. Antiplatelet and anticoagulant drugs are standard therapies in preventing and treating MI. However, all clinically used drugs are associated with bleeding complications, which ultimately limits their use in patients with a high risk of bleeding. We have developed a new recombinant drug, targ-HSA-TAP, that combines targeting and specific inhibition of activated platelets as well as anticoagulation. This drug is designed and tested for a prolonged circulating half-life, enabling unique thromboprophylaxis without bleeding complications. Methods: Targ-HSA-TAP combines a single-chain antibody (scFv) that targets activated glycoprotein IIb/IIIa on activated platelets, human serum albumin (HSA) for prolonged circulation, and tick anticoagulant peptide (TAP) for coagulation FX inhibition. A non-binding scFv is employed as a non-targeting control (non-targ-HSA-TAP). Its efficacy was investigated in vivo using murine models of acute thrombosis and cardiac ischemia-reperfusion (I/R) injury. Results: Our experiments confirmed the targeting specificity of targ-HSA-TAP to activated platelets and demonstrated effective prevention of platelet aggregation and thrombus formation, as well as FXa inhibition in vitro. Thromboprophylactic administration of targ-HSA-TAP subcutaneously in mice prevented occlusion of the carotid artery after ferric chloride injury as compared to non-targ-HSA-TAP and PBS-control treated mice. By comparing the therapeutic outcomes between targ-TAP and targ-HSA-TAP, we demonstrate the significant improvements brought by the HSA fusion in extending the drug's half-life and enhancing its therapeutic window for up to 16 h post-administration. Importantly, tail bleeding time was not prolonged with targ-HSA-TAP in contrast to the clinically used anticoagulant enoxaparin. Furthermore, in a murine model of cardiac I/R injury, mice administered targ-HSA-TAP 10 h before injury demonstrated preserved cardiac function, with significantly higher ejection fraction and fractional shortening, as compared to the non-targ-HSA-TAP and PBS control groups. Advanced strain analysis revealed reduced myocardial deformation and histology confirmed a reduced infarct size in targ-HSA-TAP treated mice compared to control groups. Conclusion: The inclusion of HSA represents a significant advancement in the design of targeted therapeutic agents for thromboprophylaxis. Our activated platelet-targeted targ-HSA-TAP is a highly effective antithrombotic drug with both anticoagulant and antiplatelet effects while retaining normal hemostasis. The long half-life of targ-HSA-TAP provides the unique opportunity to use this antithrombotic drug for more effective, long-lasting and safer anti-thrombotic prophylaxis. In cases where MI occurs, this prophylactic strategy reduces thrombus burden and effectively reduces cardiac I/R injury.

Development of endothelial-targeted CD39 as a therapy for ischemic stroke.

BACKGROUND: Ischemic stroke is characterized by a necrotic lesion in the brain surrounded by an area of dying cells termed the penumbra. Salvaging the penumbra either with thrombolysis or mechanical retrieval is the cornerstone of stroke management. At-risk neuronal cells release extracellular adenosine triphosphate, triggering microglial activation and causing a thromboinflammatory response, culminating in endothelial activation and vascular disruption. This is further aggravated by ischemia-reperfusion injury that follows all reperfusion therapies. The ecto-enzyme CD39 regulates extracellular adenosine triphosphate by hydrolyzing it to adenosine, which has antithrombotic and anti-inflammatory properties and reverses ischemia-reperfusion injury. OBJECTIVES: The objective off the study was to determine the efficacy of our therapeutic, anti-VCAM-CD39 in ischaemic stroke. METHODS: We developed anti-VCAM-CD39 that targets the antithrombotic and anti-inflammatory properties of recombinant CD39 to the activated endothelium of the penumbra by binding to vascular cell adhesion molecule (VCAM)-1. Mice were subjected to 30 minutes of middle cerebral artery occlusion and analyzed at 24 hours. Anti-VCAM-CD39 or control agents (saline, nontargeted CD39, or anti-VCAM-inactive CD39) were given at 3 hours after middle cerebral artery occlusion. RESULTS: Anti-VCAM-CD39 treatment reduced neurologic deficit; magnetic resonance imaging confirmed significantly smaller infarcts together with an increase in cerebrovascular perfusion. Anti-VCAM-CD39 also restored blood-brain barrier integrity and reduced microglial activation. Coadministration of anti-VCAM-CD39 with thrombolytics (tissue plasminogen activator [tPA]) further reduced infarct volumes and attenuated blood-brain barrier permeability with no associated increase in intracranial hemorrhage. CONCLUSION: Anti-VCAM-CD39, uniquely targeted to endothelial cells, could be a new stroke therapy even when administered 3 hours postischemia and may further synergize with thrombolytic therapy to improve stroke outcomes.

A microfluidic model to study the effects of arrhythmic flows on endothelial cells.

Atrial fibrillation (AF) is the most common type of cardiac arrhythmia and an important contributor to morbidity and mortality. Endothelial dysfunction has been postulated to be an important contributing factor in cardiovascular events in patients with AF. However, how vascular endothelial cells respond to arrhythmic flow is not fully understood, mainly due to the limitation of current in vitro systems to mimic arrhythmic flow conditions. To address this limitation, we developed a microfluidic system to study the effect of arrhythmic flow on the mechanobiology of human aortic endothelial cells (HAECs). The system utilises a computer-controlled piezoelectric pump for generating arrhythmic flow with a unique ability to control the variability in both the frequency and amplitude of pulse waves. The flow rate is modulated to reflect physiological or pathophysiological shear stress levels on endothelial cells. This enabled us to systematically dissect the importance of variability in the frequency and amplitude of pulses and shear stress level on endothelial cell mechanobiology. Our results indicated that arrhythmic flow at physiological shear stress level promotes endothelial cell spreading and reduces the plasma membrane-to-cytoplasmic distribution of ß-catenin. In contrast, arrhythmic flow at low and atherogenic shear stress levels does not promote endothelial cell spreading or redistribution of ß-catenin. Interestingly, under both shear stress levels, arrhythmic flow induces inflammation by promoting monocyte adhesion via an increase in ICAM-1 expression. Collectively, our microfluidic system provides opportunities to study the effect of arrhythmic flows on vascular endothelial mechanobiology in a systematic and reproducible manner.

Heparanase promotes the onset and progression of atherosclerosis in apolipoprotein E gene knockout mice.

BACKGROUND AND AIMS: Atherosclerosis is the primary underlying cause of myocardial infarction and stroke, which are the major causes of death globally. Heparanase (Hpse) is a pro-inflammatory extracellular matrix degrading enzyme that has been implicated in atherogenesis. However, to date the precise roles of Hpse in atherosclerosis and its mechanisms of action are not well defined. This study aims to provide new insights into the contribution of Hpse in different stages of atherosclerosis in vivo. METHODS: We generated Hpse gene-deficient mice on the atherosclerosis-prone apolipoprotein E gene knockout (ApoE-/-) background to investigate the impact of Hpse gene deficiency on the initiation and progression of atherosclerosis after 6 and 14 weeks high-fat diet feeding, respectively. Atherosclerotic lesion development, blood serum profiles, lesion composition and aortic immune cell populations were evaluated. RESULTS: Hpse-deficient mice exhibited significantly reduced atherosclerotic lesion burden in the aortic sinus and aorta at both time-points, independent of changes in plasma cholesterol levels. A significant reduction in the necrotic core size and an increase in smooth muscle cell content were also observed in advanced atherosclerotic plaques of Hpse-deficient mice. Additionally, Hpse deficiency reduced circulating and aortic levels of VCAM-1 at the initiation and progression stages of disease and circulating MCP-1 levels in the initiation but not progression stage. Moreover, the aortic levels of total leukocytes and dendritic cells in Hpse-deficient ApoE-/- mice were significantly decreased compared to control ApoE-/-mice at both disease stages. CONCLUSIONS: This study identifies Hpse as a key pro-inflammatory enzyme driving the initiation and progression of atherosclerosis and highlighting the potential of Hpse inhibitors as novel anti-inflammatory treatments for cardiovascular disease.

Differential effects of physiological agonists on the proteome of platelet-derived extracellular vesicles.

Arterial thrombosis manifesting as heart attack and stroke is the leading cause of death worldwide. Platelets are central mediators of thrombosis that can be activated through multiple activation pathways. Platelet-derived extracellular vesicles (pEVs), also known as platelet-derived microparticles, are granular mixtures of membrane structures produced by platelets in response to various activating stimuli. Initial studies have attracted interest on how platelet agonists influence the composition of the pEV proteome. In the current study, we used physiological platelet agonists of varying potencies which reflect the microenvironments that platelets experience during thrombus formation: adenosine diphosphate, collagen, thrombin as well as a combination of thrombin/collagen to induce platelet activation and pEV generation. Proteomic profiling revealed that pEVs have an agonist-dependent altered proteome in comparison to their cells of origin, activated platelets. Furthermore, we found that various protein classes including those related to coagulation and complement (prothrombin, antithrombin, and plasminogen) and platelet activation (fibrinogen) are attributed to platelet EVs following agonist stimulation. This agonist-dependent altered proteome suggests that protein packaging is an active process that appears to occur without de novo protein synthesis. This study provides new information on the influence of physiological agonist stimuli on the biogenesis and proteome landscape of pEVs.

Endothelial Response to the Combined Biomechanics of Vessel Stiffness and Shear Stress Is Regulated via Piezo1.

How endothelial cells sense and respond to dynamic changes in their biophysical surroundings as we age is not fully understood. Vascular stiffness is clearly a contributing factor not only in several cardiovascular diseases but also in physiological processes such as aging and vascular dementia. To address this gap, we utilized a microfluidic model to explore how substrate stiffness in the presence of shear stress affects endothelial morphology, senescence, proliferation, and inflammation. We also studied the role of mechanosensitive ion channel Piezo1 in endothelial responses under the combined effect of shear stress and substrate stiffness. To do so, we cultured endothelial cells inside microfluidic channels covered with fibronectin-coated elastomer with elastic moduli of 40 and 200 kPa, respectively, mimicking the stiffness of the vessel walls in young and aged arteries. The endothelial cells were exposed to atheroprotective and atherogenic shear stress levels of 10 and 2 dyn/cm2, respectively. Our findings show that substrate stiffness affects senescence under atheroprotective flow conditions and cytoskeleton remodeling, senescence, and inflammation under atherogenic flow conditions. Additionally, we found that the expression of Piezo1 plays a crucial role in endothelial adaptation to flow and regulation of inflammation under both atheroprotective and atherogenic shear stress levels. However, Piezo1 contribution to endothelial senescence was limited to the soft substrate and atheroprotective shear stress level. Overall, our study characterizes the response of endothelial cells to the combined effect of shear stress and substrate stiffness and reveals a previously unidentified role of Piezo1 in endothelial response to vessel stiffening, which potentially can be therapeutically targeted to alleviate endothelial dysfunction in aging adults.

Decoding the role of platelets in tumour metastasis: enigmatic accomplices and intricate targets for anticancer treatments.

The canonical role of platelets as central players in cardiovascular disease by way of their fundamental role in mediating thrombosis and haemostasis is well appreciated. However, there is now a large body of experimental evidence demonstrating that platelets are also pivotal in various physiological and pathophysiological processes other than maintaining haemostasis. Foremost amongst these is the emerging data highlighting the key role of platelets in driving cancer growth, metastasis and modulating the tumour microenvironment. As such, there is significant interest in targeting platelets therapeutically for the treatment of cancer. Therefore, the purpose of this review is to provide an overview of how platelets contribute to the cancer landscape and why platelets present as valuable targets for the development of novel cancer diagnosis tools and therapeutics.

Variability in Contemporary Heparin Prescription and Activated Clotting Time Monitoring During Percutaneous Coronary Intervention: Call for Up-To-Date Evidence-Based Guidelines.

BACKGROUND: Unfractionated heparin (UFH) is the preferred anticoagulant agent in percutaneous coronary intervention (PCI) procedures for minimising the risk of thrombotic complications. Because of the narrow therapeutic range of UFH, some society guidelines have advocated the use of the activated clotting time (ACT) test to monitor anticoagulation intensity during PCI to reduce thrombotic and bleeding complications. We aimed to assess the current practice of UFH prescription and its monitoring in Australia and New Zealand (ANZ). METHOD: We conducted an anonymous voluntary cross-sectional survey of interventional cardiologists (ICs) who were members of the Cardiac Society of Australia and New Zealand in 2022. The survey included 10 questions pertaining to the current practice of anticoagulation during PCI. RESULTS: Of 430 ICs surveyed, 148 responded (response rate, 34.4%). Most ICs (84.4%) prescribed 70-100 IU/kg of UFH for PCI. Over half of ICs (58.7%) routinely measured ACT during PCI, whereas only 22.2% routinely measured ACT after PCI to guide additional UFH prescription. Among ICs who prescribed additional UFH, approximately half (48%) aimed for ACT ≥250 seconds. Factors that influenced post-PCI UFH prescription included vascular access site and concomitant antiplatelet or anticoagulant therapy. CONCLUSIONS: The contemporary practice of UFH prescription during PCI and ACT monitoring in ANZ is variable and based on outdated evidence preceding current drug-eluting stents, antiplatelet therapies, and radial-first practice. Current society guideline recommendations lack clarity and agreement, reflecting the quality of the available evidence. Up-to-date clinical trials evaluating UFH prescription and ACT monitoring are needed to optimise clinical outcomes in contemporary PCI procedures.

Graft-Host Interaction and Its Effect on Wound Repair Using Mouse Models.

Autologous skin grafting has been commonly used in clinics for decades to close large wounds, yet the cellular and molecular interactions between the wound bed and the graft that mediates the wound repair are not fully understood. The aim of this study was to better understand the molecular changes in the wound triggered by autologous and synthetic grafting. Defining the wound changes at the molecular level during grafting sets the basis to test other engineered skin grafts by design. In this study, a full-thickness skin graft (SKH-1 hairless) mouse model was established. An autologous full-thickness skin graft (FTSG) or an acellular fully synthetic Biodegradable Temporising Matrix (BTM) was grafted. The wound bed/grafts were analysed at histological, RNA, and protein levels during the inflammation (day 1), proliferation (day 5), and remodelling (day 21) phases of wound repair. The results showed that in this mouse model, similar to others, inflammatory marker levels, including Il-6, Cxcl-1, and Cxcl-5/6, were raised within a day post-wounding. Autologous grafting reduced the expression of these inflammatory markers. This was different from the wounds grafted with synthetic dermal grafts, in which Cxcl-1 and Cxcl-5/6 remained significantly high up to 21 days post-grafting. Autologous skin grafting reduced wound contraction compared to wounds that were left to spontaneously repair. Synthetic grafts contracted significantly more than FTSG by day 21. The observed wound contraction in synthetic grafts was most likely mediated at least partly by myofibroblasts. It is possible that high TGF-ß1 levels in days 1-21 were the driving force behind myofibroblast abundance in synthetic grafts, although no evidence of TGF-ß1-mediated Connective Tissue Growth Factor (CTGF) upregulation was observed.

Bioengineered Vascular Model of Foam Cell Formation.

Foam cell formation is a complex blood vessel pathology, which is characterized by a series of events, including endothelium dysfunction, inflammation, and accumulation of immune cells underneath the blood vessel walls. Novel bioengineered models capable of recapitulating these events are required to better understand the complex pathological processes underlying the development of foam cell formation and, consequently, advanced bioengineered platforms for screening drugs. Here, we generated a microfluidic blood vessel model, incorporating a three-dimensional (3D) extracellular matrix coated with an endothelial layer. This system enables us to perform experiments under a dynamic microenvironment that recapitulates the complexities of the native vascular regions. Using this model, we studied the effectors that regulate monocyte adhesion and migration, as well as foam cell formation inside vessel walls. We found that monocyte adhesion and migration are regulated by both the endothelium and monocytes themselves. Monocytes migrated into the extracellular matrix only when endothelial cells were cultured in the vessel model. In addition, the exposure of an endothelial layer to tumor necrosis factor α (TNF-α) and low shear stress both increased monocyte migration into the subendothelial space toward the matrix. Furthermore, we demonstrated the process of foam cell formation, 3 days after transmigration of peripheral blood mononuclear cells (PBMCs) into the vessel wall. We showed that pre-exposure of PBMCs to high shear rates increases their adhesion and migration through the TNF-α-treated endothelium but does not affect their capacity to form foam cells. The versatility of our model allows for mechanistic studies on foam cell formation under customized pathological conditions.

Prognosis and Personalized In Silico Prediction of Treatment Efficacy in Cardiovascular and Chronic Kidney Disease: A Proof-of-Concept Study.

(1) Background: Kidney and cardiovascular diseases are responsible for a large fraction of population morbidity and mortality. Early, targeted, personalized intervention represents the ideal approach to cope with this challenge. Proteomic/peptidomic changes are largely responsible for the onset and progression of these diseases and should hold information about the optimal means of treatment and prevention. (2) Methods: We investigated the prediction of renal or cardiovascular events using previously defined urinary peptidomic classifiers CKD273, HF2, and CAD160 in a cohort of 5585 subjects, in a retrospective study. (3) Results: We have demonstrated a highly significant prediction of events, with an HR of 2.59, 1.71, and 4.12 for HF, CAD, and CKD, respectively. We applied in silico treatment, implementing on each patient's urinary profile changes to the classifiers corresponding to exactly defined peptide abundance changes, following commonly used interventions (MRA, SGLT2i, DPP4i, ARB, GLP1RA, olive oil, and exercise), as defined in previous studies. Applying the proteomic classifiers after the in silico treatment indicated the individual benefits of specific interventions on a personalized level. (4) Conclusions: The in silico evaluation may provide information on the future impact of specific drugs and interventions on endpoints, opening the door to a precision-based medicine approach. An investigation into the extent of the benefit of this approach in a prospective clinical trial is warranted.

An Activation-Specific Anti-Mac-1 Designed-Ankyrin-Repeat-Protein Attenuates Colitis in Mice.

(1) Background: Inflammatory bowel diseases are complex and multifactorial disorders of unknown etiology. The extravasation of activated leukocytes is a critical step in the pathogenesis of these diseases. Leukocyte integrin Mac-1 (αMß2; CD11b/CD18) is crucial for the extravasation of myeloid cells, and a novel activation-specific anti-Mac-1 Designed Ankyrin Repeat protein (DARPin F7) is a promising therapeutic agent for inflammatory diseases. In its activated conformation, Mac-1 expresses the high-affinity binding site I-domain, which the DARPin F7 selectively targets. In our study, we aimed to explore the therapeutic potential of anti-Mac-1 DARPin F7 in murine dextrane sodium sulfate (DSS)-induced colitis. (2) Methods: C57BL/6J mice received 3% DSS drinking water for five days, followed by normal drinking water for one week. The mice were treated with DARPin F7 or a control substance daily via intraperitoneal injections. Disease activity index (DAI), colon length, myeloperoxidase (MPO) activity measurements, H&E staining, and qRT-PCR were conducted after euthanizing the mice on day 12. (3) Results: Treatment with DARPin F7 resulted in less pronounced colon shortening and significantly lower histological scores. The DARPin F7-treated animals experienced substantially less disease and myeloperoxidase (MPO) activity. Animals that received DARPin F7 treatment suffered less weight loss and recovered from the weight loss more efficiently. Treatment with DARPin F7 also led to significantly reduced mRNA expression of inflammatory cytokines. (4) Conclusion: Anti-Mac-1 treatment markedly reduced disease activity and inflammatory reaction accompanying DSS-induced colitis in mice.

Mortality data from omission of early thromboprophylaxis in critically ill patients highlights the importance of an individualised diagnosis-related approach.

BACKGROUND: Venous thromboembolism (VTE) prophylaxis is effective in reducing VTE events, however, its impact on mortality is unclear. We examined the association between omission of VTE prophylaxis within the first 24 h after intensive care unit (ICU) admission and hospital mortality. METHODS: Retrospective analysis of prospectively collected data from the Australian New Zealand Intensive Care Society Adult Patient Database. Data were obtained for adult admissions between 2009 and 2020. Mixed effects logistic regression models were used to evaluate the association between omission of early VTE prophylaxis and hospital mortality. RESULTS: Of the 1,465,020 ICU admissions, 107,486 (7.3%) did not receive any form of VTE prophylaxis within the first 24 h after ICU admission without documented contraindication. Omission of early VTE prophylaxis was independently associated with 35% increased odds of in-hospital mortality (odds ratios (OR): 1.35; 95% CI: 1.31-1.41). The associations between omission of early VTE prophylaxis and mortality varied by admission diagnosis. In patients diagnosed with stroke (OR: 1.26, 95% CI: 1.05-1.52), cardiac arrest (OR: 1.85, 95% CI: 1.65-2.07) or intracerebral haemorrhage (OR: 1.48, 95% CI: 1.19-1.84), omission of VTE prophylaxis was associated with increased risk of mortality, but not in patients diagnosed with subarachnoid haemorrhage or head injury. CONCLUSIONS: Omission of VTE prophylaxis within the first 24 h after ICU admission was independently associated with increased risk of mortality that varied by admission diagnosis. Consideration of early thromboprophylaxis may be required for patients with stroke, cardiac arrest and intracerebral haemorrhage but not in those with subarachnoid haemorrhage or head injury. The findings highlight the importance of individualised diagnosis-related thromboprophylaxis benefit-harm assessments.

Activated Coagulation FXII: A Unique Target for In Vivo Molecular Imaging.

BACKGROUND: Current clinical imaging of thromboembolic diseases often relies on indirect detection of thrombi, which may delay diagnosis and ultimately the institution of beneficial, potentially lifesaving treatment. Therefore, the development of targeting tools that facilitate the rapid, specific, and direct imaging of thrombi using molecular imaging is highly sought after. One potential molecular target is FXIIa (factor XIIa), which initiates the intrinsic coagulation pathway but also activates the kallikrein-kinin system, thereby initiating coagulation and inflammatory/immune responses. As FXII (factor XII) is dispensable for normal hemostasis, its activated form (FXIIa) represents an ideal molecular target for diagnostic and therapeutic approaches, the latter combining diagnosis/identification of thrombi and effective antithrombotic therapy. METHODS: We conjugated an FXIIa-specific antibody, 3F7, to a near-infrared (NIR) fluorophore and demonstrated binding to FeCl3-induced carotid thrombosis with 3-dimensional fluorescence emission computed tomography/computed tomography and 2-dimensional fluorescence imaging. We further demonstrated ex vivo imaging of thromboplastin-induced pulmonary embolism and detection of FXIIa in human thrombi produced in vitro. RESULTS: We demonstrated imaging of carotid thrombosis by fluorescence emission computed tomography/computed tomography and measured a significant fold increase in signal between healthy and control vessels from mice injected with 3F7-NIR compared with mice injected with nontargeted probe (P=0.002) ex vivo. In a model of pulmonary embolism, we measured increased NIR signal in lungs from mice injected with 3F7-NIR compared with mice injected with nontargeted probe (P=0.0008) and healthy lungs from mice injected with 3F7-NIR (P=0.021). CONCLUSIONS: Overall, we demonstrate that FXIIa targeting is highly suitable for the specific detection of venous and arterial thrombi. This approach will allow direct, specific, and early imaging of thrombosis in preclinical imaging modalities and may facilitate monitoring of antithrombotic treatment in vivo.

Colchicine exerts anti-atherosclerotic and -plaque-stabilizing effects targeting foam cell formation.

Colchicine is a broad-acting anti-inflammatory agent that has attracted interest for repurposing in atherosclerotic cardiovascular disease. Here, we studied its ability at a human equivalent dose of 0.5 mg/day to modify plaque formation and composition in murine atherosclerosis and investigated its actions on macrophage responses to atherogenic stimuli in vitro. In atherosclerosis induced by high-cholesterol diet, Apoe-/- mice treated with colchicine had 50% reduction in aortic oil Red O+ plaque area compared to saline control (p = .001) and lower oil Red O+ staining of aortic sinus lesions (p = .03). In vitro, addition of 10 nM colchicine inhibited foam cell formation from murine and human macrophages after treatment with oxidized LDL (ox-LDL). Mechanistically, colchicine downregulated glycosylation and surface expression of the ox-LDL uptake receptor, CD36, and reduced CD36+ staining in aortic sinus plaques. It also decreased macrophage uptake of cholesterol crystals, resulting in lower intracellular lysosomal activity, inhibition of the NLRP3 inflammasome, and reduced secretion of IL-1ß and IL-18. Colchicine's anti-atherosclerotic actions were accentuated in a mouse model of unstable plaque induced by carotid artery tandem stenosis surgery, where it decreased lesion size by 48% (p = .01), reduced lipid (p = .006) and necrotic core area (p = .007), increased collagen content and cap-to-necrotic core ratio (p = .05), and attenuated plaque neutrophil extracellular traps (p < .001). At low dose, colchicine's effects were not accompanied by the evidence of microtubule depolymerization. Together, these results show that colchicine exerts anti-atherosclerotic and plaque-stabilizing effects at low dose by inhibiting foam cell formation and cholesterol crystal-induced inflammation. This provides a new framework to support its repurposing for atherosclerotic cardiovascular disease.

Myocarditis: causes, mechanisms, and evolving therapies.

INTRODUCTION: Myocarditis is a severe lymphocyte-mediated inflammatory disorder of the heart, mostly caused by viruses and immune checkpoint inhibitors (ICIs). Recently, myocarditis as a rare adverse event of mRNA vaccines for SARS-CoV-2 has caused global attention. The clinical consequences of myocarditis can be very severe, but specific treatment options are lacking or not yet clinically proven. AREAS COVERED: This paper offers a brief overview of the biology of viruses that frequently cause myocarditis, focusing on mechanisms important for viral entry and replication following host infection. Current and new potential therapeutic targets/strategies especially for viral myocarditis are reviewed systematically. In particular, the immune system in myocarditis is dissected with respect to infective viral and non-infective, ICI-induced myocarditis. EXPERT OPINION: Vaccination is an excellent emerging preventative strategy for viral myocarditis, but most vaccines still require further development. Anti-viral treatments that inhibit viral replication need to be considered following viral infection in host myocardium, as lower viral load reduces inflammation severity. Understanding how the immune system continues to damage the heart even after viral clearance will define novel therapeutic targets/strategies. We propose that viral myocarditis can be best treated using a combination of antiviral agents and immunotherapies that control cytotoxic T cell activity.

Prediction of coronary artery disease using urinary proteomics.

AIMS: Coronary artery disease (CAD) is multifactorial, caused by complex pathophysiology, and contributes to a high burden of mortality worldwide. Urinary proteomic analyses may help to identify predictive biomarkers and provide insights into the pathogenesis of CAD. METHODS AND RESULTS: Urinary proteome was analysed in 965 participants using capillary electrophoresis coupled with mass spectrometry. A proteomic classifier was developed in a discovery cohort with 36 individuals with CAD and 36 matched controls using the support vector machine. The classifier was tested in a validation cohort with 115 individuals who progressed to CAD and 778 controls and compared with two previously developed CAD-associated classifiers, CAD238 and ACSP75. The Framingham and SCORE2 risk scores were available in 737 participants. Bioinformatic analysis was performed based on the CAD-associated peptides. The novel proteomic classifier was comprised of 160 urinary peptides, mainly related to collagen turnover, lipid metabolism, and inflammation. In the validation cohort, the classifier provided an area under the receiver operating characteristic curve (AUC) of 0.82 [95% confidence interval (CI): 0.78-0.87] for the CAD prediction in 8 years, superior to CAD238 (AUC: 0.71, 95% CI: 0.66-0.77) and ACSP75 (AUC: 0.53 and 95% CI: 0.47-0.60). On top of CAD238 and ACSP75, the addition of the novel classifier improved the AUC to 0.84 (95% CI: 0.80-0.89). In a multivariable Cox model, a 1-SD increment in the novel classifier was associated with a higher risk of CAD (HR: 1.54, 95% CI: 1.26-1.89, P < 0.0001). The new classifier further improved the risk reclassification of CAD on top of the Framingham or SCORE2 risk scores (net reclassification index: 0.61, 95% CI: 0.25-0.95, P = 0.001; 0.64, 95% CI: 0.28-0.98, P = 0.001, correspondingly). CONCLUSION: A novel urinary proteomic classifier related to collagen metabolism, lipids, and inflammation showed potential for the risk prediction of CAD. Urinary proteome provides an alternative approach to personalized prevention.

A biomarker that can predict coronary artery disease (CAD) is urgently in need. We developed and validated a urinary proteomic classifier for the prediction of CAD. The proteomic classifier involved in atherosclerosis improved the risk reclassification on top of the clinical risk score.

Quantitative proteomic landscape of unstable atherosclerosis identifies molecular signatures and therapeutic targets for plaque stabilization.

Atherosclerotic plaque rupture leading to myocardial infarction is a major global health burden. Applying the tandem stenosis (TS) mouse model, which distinctively exhibits the characteristics of human plaque instability/rupture, we use quantitative proteomics to understand and directly compare unstable and stable atherosclerosis. Our data highlight the disparate natures and define unique protein signatures of unstable and stable atherosclerosis. Key proteins and pathway networks are identified such as the innate immune system, and neutrophil degranulation. The latter includes calprotectin S100A8/A9, which we validate in mouse and human unstable plaques, and we demonstrate the plaque-stabilizing effects of its inhibition. Overall, we provide critical insights into the unique proteomic landscape of unstable atherosclerosis (as distinct from stable atherosclerosis and vascular tissue). We further establish the TS model as a reliable preclinical tool for the discovery and testing of plaque-stabilizing drugs. Finally, we provide a knowledge resource defining unstable atherosclerosis that will facilitate the identification and validation of long-sought-after therapeutic targets and drugs for plaque stabilization.

Improving the rate of inferior vena cava filter retrieval through multidisciplinary engagement.

Background: The placement of retrievable inferior vena cava (IVC) filters occurs commonly, but retrieval rates remain low. Consequently, there is an unmet clinical need to ensure appropriate follow-up and retrieval of these devices. Objectives: To determine the association between an IVC filter surveillance team with filter retrievals or a documented filter plan, time to retrieval, and incidence of filter complications or recurrent venous thromboembolism. Methods: Ambidirectional cohort study evaluating consecutive IVC filter insertions before and after the implementation of a multidisciplinary surveillance team (MDST). We report an odds ratio (OR) with 95% CIs, adjusted by age, sex, weight, and malignancy status. Results: Overall, 453 patients were included, with 272 individuals in the pre-MDST cohort and 181 individuals in the post-MDST cohort. The MDST was associated with a higher composite primary outcome of IVC filter retrieval or a documented filter plan from 79.4% in the pre-MDST cohort to 96.1% in the post-MDST cohort (OR, 6.44; 95% CI, 3.06-15.84). Compared with the pre-MDST cohort, IVC filter retrieval rates were higher in the post-MDST cohort (52.6%-73.5%, respectively; (OR, 2.50; 95% CI, 1.67-3.78). The MDST was associated with a shorter median time-to-filter retrieval (187-150 days, hazard ratio, 1.78; 95% CI, 1.39-2.29), but there was no significant difference when comparing symptomatic or clinically significant IVC filter complications, recurrent venous thromboembolism, or mortality. Conclusion: Our study demonstrates the importance of a structured program to ensure timely IVC filter retrieval and ultimately improve patient care.