Longer and better lives for patients with atrial fibrillation: the 9th AFNET/EHRA consensus conference.

AIMS: Recent trial data demonstrate beneficial effects of active rhythm management in patients with atrial fibrillation (AF) and support the concept that a low arrhythmia burden is associated with a low risk of AF-related complications. The aim of this document is to summarize the key outcomes of the 9th AFNET/EHRA Consensus Conference of the Atrial Fibrillation NETwork (AFNET) and the European Heart Rhythm Association (EHRA). METHODS AND RESULTS: Eighty-three international experts met in Münster for 2 days in September 2023. Key findings are as follows: (i) Active rhythm management should be part of the default initial treatment for all suitable patients with AF. (ii) Patients with device-detected AF have a low burden of AF and a low risk of stroke. Anticoagulation prevents some strokes and also increases major but non-lethal bleeding. (iii) More research is needed to improve stroke risk prediction in patients with AF, especially in those with a low AF burden. Biomolecules, genetics, and imaging can support this. (iv) The presence of AF should trigger systematic workup and comprehensive treatment of concomitant cardiovascular conditions. (v) Machine learning algorithms have been used to improve detection or likely development of AF. Cooperation between clinicians and data scientists is needed to leverage the potential of data science applications for patients with AF. CONCLUSIONS: Patients with AF and a low arrhythmia burden have a lower risk of stroke and other cardiovascular events than those with a high arrhythmia burden. Combining active rhythm control, anticoagulation, rate control, and therapy of concomitant cardiovascular conditions can improve the lives of patients with AF.

A RIVA-DM Subanalysis Investigating Patients With Nonvalvular Atrial Fibrillation and Type 2 Diabetes Aged Under Versus Over 80 Years.

BACKGROUND: Advanced age and type 2 diabetes (T2D) are common in patients with nonvalvular atrial fibrillation (NVAF). We evaluated the impact of age on the effectiveness and safety of rivaroxaban versus warfarin in this population. METHODS: We analyzed electronic health record data from November 2010, to December 2019 including adults with NVAF and T2D, newly started on rivaroxaban or warfarin. Propensity score-overlap weighted hazard ratios (HRs) for stroke/systemic embolism (SSE), hospitalization for major or clinically relevant nonmajor bleeding (CRNMB), vascular death, major adverse limb events (MALE), major bleeding, and intracranial hemorrhage (ICH) were calculated for older (≥80 years) and younger (<80 years) cohorts. RESULTS: We included 32 078 rivaroxaban and 83 971 warfarin users (6606 rivaroxaban and 25,335 warfarin patients were aged ≥80 years). No significant interaction for rivaroxaban versus warfarin by age was observed for any outcome, including SSE (HR = 1.05 vs 0.95), hospitalization for major or CRNMB (HR = 1.06 vs 0.90), vascular death (HR = 0.92 vs 0.90), MALE (HR = 0.80 vs 0.76), major bleeding or ICH. CONCLUSIONS: The effectiveness and safety of rivaroxaban versus warfarin remained consistent across patient age subgroups.

Impact of non-adherence to direct oral anticoagulants amongst Swedish patients with non-valvular atrial fibrillation: results from a real-world cost-utility analysis.

AIMS: A third of non-valvular atrial fibrillation (NVAF) patients are non-adherent to direct oral anticoagulants (DOACs). Estimates of the economic value of full adherence and the cost of two types of adherence improving interventions are important to healthcare planners and decision-makers. METHODS: A cost-utility analysis estimated the impact of non-adherence over a 20-year horizon, for a patient cohort with a mean age of 77 years, based on data from the Stockholm Healthcare database of NVAF patients with incident stroke between 2011 and 2018. Adherence was defined using a medication possession ratio (MPR) cut-off of 90%; primary outcomes were the number of ischemic strokes and associated incremental cost-utility ratio. RESULTS: Hypothetical comparisons between cohorts of 1,000 patients with varying non-adherence levels and full adherence (MPR >90%) predicted an additional number of strokes ranging from 117 (MPR = 81-90%) to 866 (MPR <60%), and years of life lost ranging from 177 (MPR = 81- 90%) to 1,318 (MPR < 60%; discounted at 3%). Chronic disease co-management intervention occurring during each DOAC prescription renewal and patient education intervention at DOAC initiation will be cost-saving to the health system if its cost is below SEK 143 and SEK 4,655, and cost-effective if below SEK 858 and SEK 28,665, respectively. CONCLUSION: Adherence improving interventions for NVAF patients on DOACs such as chronic disease co-management and patient education can be cost-saving and cost-effective, within a range of costs that appear reasonable to the Swedish healthcare system.

Atrial fibrillation (AF) is the most common type of chronic cardiac arrhythmia and a major risk factor for ischemic stroke (IS). The objective of this study was to compare the costs and health outcomes associated with adherence to direct oral anticoagulant (DOAC) therapy in Sweden. The study also aimed to demonstrate the potential benefits of developing interventions to improve DOAC adherence. DOAC therapy (DOACs; apixaban, dabigatran, edoxaban, and rivaroxaban) has been approved in Europe for the prevention of stroke in adult patients with AF. It has been demonstrated to provide warfarin-similar reductions in stroke risk in NVAF patients, with reductions in mortality and intracranial hemorrhage. However, non-adherence to DOAC medication prevents patients and healthcare systems from fully benefiting from DOAC therapy, resulting in a lower benefit than those seen in randomized controlled trials. DOAC non-adherence is where AF patients deviate from the DOAC treatment regimen as prescribed by health providers. This study suggested that non-adherence to DOAC therapy has a substantial impact on ischemic stroke risk and significant additional healthcare system costs. Patient education and chronic disease co-management (two types of DOAC adherence improving intervention) can be cost-saving and cost-effective within a range of costs that appear reasonable to the Swedish healthcare system. Healthcare policy-makers should prioritize initiatives aimed at improving DOAC adherence in order to improve outcomes in AF.

Urinary miRNA Profiles in Chronic Kidney Injury-Benefits of Extracellular Vesicle Enrichment and miRNAs as Potential Biomarkers for Renal Fibrosis, Glomerular Injury, and Endothelial Dysfunction.

Micro-RNAs (miRNAs) are regulators of gene expression and play an important role in physiological homeostasis and disease. In biofluids, miRNAs can be found in protein complexes or in extracellular vesicles (EVs). Altered urinary miRNAs are reported as potential biomarkers for chronic kidney disease (CKD). In this context, we compared established urinary protein biomarkers for kidney injury with urinary miRNA profiles in obese ZSF1 and hypertensive renin transgenic rats. Additionally, the benefit of urinary EV enrichment was investigated in vivo and the potential association of urinary miRNAs with renal fibrosis in vitro. Kidney damage in both rat models was confirmed by histopathology, proteinuria, and increased levels of urinary protein biomarkers. In total, 290 miRNAs were elevated in obese ZSF1 rats compared with lean controls, whereas 38 miRNAs were altered in obese ZSF1 rats during 14-26 weeks of age. These 38 miRNAs correlated better with disease progression than established urinary protein biomarkers. MiRNAs increased in obese ZSF1 rats were associated with renal inflammation, fibrosis, and glomerular injury. Eight miRNAs were also changed in urinary EVs of renin transgenic rats, including one which might play a role in endothelial dysfunction. EV enrichment increased the number and detection level of several miRNAs implicated in renal fibrosis in vitro and in vivo. Our results show the benefit of EV enrichment for miRNA detection and the potential of total urine and urinary EV-associated miRNAs as biomarkers of altered kidney physiology, renal fibrosis and glomerular injury, and disease progression in hypertension and obesity-induced CKD.

Differential responses of induced pluripotent stem cell-derived cardiomyocytes to anisotropic strain depends on disease status.

Primary dilated cardiomyopathy (DCM) is a non-ischemic heart disease with impaired pumping function of the heart. In this study, we used human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from a healthy volunteer and a primary DCM patient to investigate the impact of DCM on iPSC-CMs׳ responses to different types of anisotropic strain. A bioreactor system was established that generates cardiac-mimetic forces of 150 kPa at 5% anisotropic cyclic strain and 1 Hz frequency. After confirming cardiac induction of the iPSCs, it was determined that fibronectin was favorable to other extracellular matrix protein coatings (gelatin, laminin, vitronectin) in terms of viable cell area and density, and was therefore selected as the coating for further study. When iPSC-CMs were exposed to three strain conditions (no strain, 5% static strain, and 5% cyclic strain), the static strain elicited significant induction of sarcomere components in comparison to other strain conditions. However, this induction occurred only in iPSC-CMs from a healthy volunteer ("control iPSC-CMs"), not in iPSC-CMs from the DCM patient ("DCM iPSC-CMs"). The donor type also significantly influenced gene expressions of cell-cell and cell-matrix interaction markers in response to the strain conditions. Gene expression of connexin-43 (cell-cell interaction) had a higher fold change in healthy versus diseased iPSC-CMs under static and cyclic strain, as opposed to integrins α-5 and α-10 (cell-matrix interaction). In summary, our iPSC-CM-based study to model the effects of different strain conditions suggests that intrinsic, genetic-based differences in the cardiomyocyte responses to strain may influence disease manifestation in vivo.

Patterned polymer matrix promotes stemness and cell-cell interaction of adult stem cells.

BACKGROUND: The interaction of stem cells with their culture substrates is critical in controlling their fate and function. Declining stemness of adult-derived human mesenchymal stem cells (hMSCs) during in vitro expansion on tissue culture polystyrene (TCPS) severely limits their therapeutic efficacy prior to cell transplantation into damaged tissues. Thus, various formats of natural and synthetic materials have been manipulated in attempts to reproduce in vivo matrix environments in which hMSCs reside. RESULTS: We developed a series of patterned polymer matrices for cell culture by hot-pressing poly(ε-caprolactone) (PCL) films in femtosecond laser-ablated nanopore molds, forming nanofibers on flat PCL substrates. hMSCs cultured on these PCL fiber matrices significantly increased expression of critical self-renewal factors, Nanog and OCT4A, as well as markers of cell-cell interaction PECAM and ITGA2. The results suggest the patterned polymer fiber matrix is a promising model to maintain the stemness of adult hMSCs. CONCLUSION: This approach meets the need for scalable, highly repeatable, and tuneable models that mimic extracellular matrix features that signal for maintenance of hMSC stemness.

Phage-display-guided nanocarrier targeting to atheroprone vasculature.

In regions of the circulation where vessels are straight and unbranched, blood flow is laminar and unidirectional. In contrast, at sites of curvature, branch points, and regions distal to stenoses, blood flow becomes disturbed. Atherosclerosis preferentially develops in these regions of disturbed blood flow. Current therapies for atherosclerosis are systemic and may not sufficiently target these atheroprone regions. In this study, we sought to leverage the alterations on the luminal surface of endothelial cells caused by this atheroprone flow for nanocarrier targeting. In vivo phage display was used to discover unique peptides that selectively bind to atheroprone regions in the mouse partial carotid artery ligation model. The peptide GSPREYTSYMPH (PREY) was found to bind 4.5-fold more avidly to the region of disturbed flow and was used to form targeted liposomes. When administered intravenously, PREY-targeted liposomes preferentially accumulated in endothelial cells in the partially occluded carotid artery and other areas of disturbed flow. Proteomic analysis and immunoblotting indicated that fibronectin and Filamin-A were preferentially bound by PREY nanocarriers in vessels with disturbed flow. In additional experiments, PREY nanocarriers were used therapeutically to deliver the nitric oxide synthase cofactor tetrahydrobiopterin (BH4), which we have previously shown to be deficient in regions of disturbed flow. This intervention increased vascular BH4 and reduced vascular superoxide in the partially ligated artery in wild-type mice and reduced plaque burden in the partially ligated left carotid artery of fat fed atheroprone mice (ApoE(-/-)). Targeting atheroprone sites of the circulation with functionalized nanocarriers provides a promising approach for prevention of early atherosclerotic lesion formation.

Tissue sodium storage: evidence for kidney-like extrarenal countercurrent systems?

Recent evidence from chemical analysis of tissue electrolyte and water composition has shown that body Na(+) content in experimental animals is not constant, does not always readily equilibrate with water, and cannot be exclusively controlled by the renal blood purification process. Instead, large amounts of Na(+) are stored in the skin and in skeletal muscle. Quantitative non-invasive detection of Na(+) reservoirs with sodium magnetic resonance imaging ((23)NaMRI) suggests that this mysterious Na(+) storage is not only an animal research curiosity but also exists in humans. In clinical studies, tissue Na(+) storage is closely associated with essential hypertension. In animal experiments, modulation of reservoir tissue Na(+) content leads to predictable blood pressure changes. The available evidence thus suggests that the patho(?)-physiological process of Na(+) storage might be of relevance for human health and disease.

Oligoproline-derived nanocarrier for dual stimuli-responsive gene delivery.

Gene therapy is a promising method for the treatment of vascular disease; however, successful strategies depend on the development of safe and effective delivery technologies with specific targeting to a diseased point of vasculature. Reactive oxygen species (ROS) are overproduced by vascular smooth muscle cells (VSMCs) at critical stages of atherosclerosis progression. Therefore, ROS were exploited as a stimulus for vascular targeted gene delivery in this study. A combination of bio-conjugation methods and controlled reverse addition-fragmentation chain-trasfer (RAFT) polymerization was utilized to synthesize a new ROS-cleavable, pH-responsive mPEG113-b-CP5K-b-PDMAEMA42-b-P(DMAEMA22-co-BMA40-co-PAA24) (PPDDBP) polymer as a nanocarrier for plasmid DNA (pDNA) delivery. The ros degradability of PPDDBP polymers was confirmed by SIN-1-mediated cleavage of CP5K peptide linkers through a shift in GPC chromatogram with an appearance of mPEG shoulder peak and an increase in zeta potential (ζ). The polyplex nanocarrier also demonstrated effective PDNA loading, serum stability, and hemocompatibility, indicating its excellent performance under physiological conditions. The polyplexes demonstrated ideal pH responsiveness for endosomal escape and effective ROS responsiveness for improved targeting in an in vitro model of pathogenic VSMCs in terms of both uptake and expression of reporter gene. These data suggest this novel nanocarrier polyplex system is a promising gene delivery tool for preventing or treating areas of high ROS, such as atherosclerotic lesions.

Femtosecond laser-patterned nanopore arrays for surface-mediated peptide treatment.

The major goal of this study was to create easy-to-use, reusable substrates capable of storing any peptides or bioactive molecules for a desired period of time until cells uptake them without the need for bioactive molecule or peptide-specific techniques. Nanopore arrays of uniform size and distribution were machined into fused silica substrates using femtosecond laser ablation and loaded with peptides by simple adsorption. The nanopore substrates were validated by examining the effect of N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) loaded nanopores on macrophage phagocytosis and intracellular production of reactive oxygen species (ROS) with and without the pro-inflammatory lipopolysaccharide (LPS). Our results demonstrated that nanopores were generated in a uniform array fashion. Ac-SDKP peptides were stably stored in nanopores and internalized by macrophages. Significant reductions in ROS production and phagocytosis in macrophages were observed over control substrates, even in combination with LPS stimulation, indicating that loading Ac-SDKP peptides in pores significantly improved the anti-inflammatory effects. FROM THE CLINICAL EDITOR: This team of scientists intended to create easy-to-use, reusable substrates for storing peptides or bioactive molecules for a desired period of time before cellular uptake occurs, and without the need for bioactive molecule or peptide-specific techniques. They demonstrate the successful generation of nanopores in a uniform array that stably stores Ac-SDKP peptides in the nanopores. When peptides were internalized by macrophages, significant reductions in ROS production and phagocytosis were observed, indicating improved anti-inflammatory effects.

Scaling and systems biology for integrating multiple organs-on-a-chip.

Coupled systems of in vitro microfabricated organs-on-a-chip containing small populations of human cells are being developed to address the formidable pharmacological and physiological gaps between monolayer cell cultures, animal models, and humans that severely limit the speed and efficiency of drug development. These gaps present challenges not only in tissue and microfluidic engineering, but also in systems biology: how does one model, test, and learn about the communication and control of biological systems with individual organs-on-chips that are one-thousandth or one-millionth of the size of adult organs, or even smaller, i.e., organs for a milliHuman (mHu) or microHuman (µHu)? Allometric scaling that describes inter-species variation of organ size and properties provides some guidance, but given the desire to utilize these systems to extend and validate human pharmacokinetic and pharmacodynamic (PK/PD) models in support of drug discovery and development, it is more appropriate to scale each organ functionally to ensure that it makes the suitable physiological contribution to the coupled system. The desire to recapitulate the complex organ-organ interactions that result from factors in the blood and lymph places a severe constraint on the total circulating fluid (~5 mL for a mHu and ~5 µL for a µHu) and hence on the pumps, valves, and analytical instruments required to maintain and study these systems. Scaling arguments also provide guidance on the design of a universal cell-culture medium, typically without red blood cells. This review presents several examples of scaling arguments and discusses steps that should ensure the success of this endeavour.

Neurovascular unit on a chip: implications for translational applications.

The blood-brain barrier (BBB) dynamically controls exchange between the brain and the body, but this interaction cannot be studied directly in the intact human brain or sufficiently represented by animal models. Most existing in vitro BBB models do not include neurons and glia with other BBB elements and do not adequately predict drug efficacy and toxicity. Under the National Institutes of Health Microtissue Initiative, we are developing a three-dimensional, multicompartment, organotypic microphysiological system representative of a neurovascular unit of the brain. The neurovascular unit system will serve as a model to study interactions between the central nervous system neurons and the cerebral spinal fluid (CSF) compartment, all coupled to a realistic blood-surrogate supply and venous return system that also incorporates circulating immune cells and the choroid plexus. Hence all three critical brain barriers will be recapitulated: blood-brain, brain-CSF, and blood-CSF. Primary and stem cell-derived human cells will interact with a variety of agents to produce critical chemical communications across the BBB and between brain regions. Cytomegalovirus, a common herpesvirus, will be used as an initial model of infections regulated by the BBB. This novel technological platform, which combines innovative microfluidics, cell culture, analytical instruments, bioinformatics, control theory, neuroscience, and drug discovery, will replicate chemical communication, molecular trafficking, and inflammation in the brain. The platform will enable targeted and clinically relevant nutritional and pharmacologic interventions for or prevention of such chronic diseases as obesity and acute injury such as stroke, and will uncover potential adverse effects of drugs. If successful, this project will produce clinically useful technologies and reveal new insights into how the brain receives, modifies, and is affected by drugs, other neurotropic agents, and diseases.

Cell interaction study method using novel 3D silica nanoneedle gradient arrays.

Understanding cellular interactions with culture substrate features is important to advance cell biology and regenerative medicine. When surface topographical features are considerably larger in vertical dimension and are spaced at least one cell dimension apart, the features act as 3D physical barriers that can guide cell adhesion, thereby altering cell behavior. In the present study, we investigated competitive interactions of cells with neighboring cells and matrix using a novel nanoneedle gradient array. A gradient array of nanoholes was patterned at the surface of fused silica by single-pulse femtosecond laser machining. A negative replica of the pattern was extracted by nanoimprinting with a thin film of polymer. Silica was deposited on top of the polymer replica to form silica nanoneedles. NIH 3T3 fibroblasts were cultured on silica nanoneedles and their behavior was studied and compared with those cultured on a flat silica surface. The presence of silica nanoneedles was found to enhance the adhesion of fibroblasts while maintaining cell viability. The anisotropy in the arrangement of silica nanoneedles was found to affect the morphology and spreading of fibroblasts. Additionally, variations in nanoneedle spacing regulated cell-matrix and cell-cell interactions, effectively preventing cell aggregation in areas of tightly-packed nanoneedles. This proof-of-concept study provides a reproducible means for controlling competitive cell adhesion events and offers a novel system whose properties can be manipulated to intimately control cell behavior.

Modular polymer design to regulate phenotype and oxidative response of human coronary artery cells for potential stent coating applications.

Polymer properties can be tailored by copolymerizing subunits with specific physico-chemical characteristics. Vascular stent materials require biocompatibility, mechanical strength, and prevention of restenosis. Here we copolymerized poly(ε-caprolactone) (PCL), poly(ethylene glycol) (PEG), and carboxyl-PCL (cPCL) at varying molar ratios and characterized the resulting material properties. We then performed a short-term evaluation of these polymers for their applicability as potential coronary stent coating materials with two primary human coronary artery cell types: smooth muscle cells (HCASMC) and endothelial cells (HCAEC). Changes in proliferation and phenotype were dependent upon intracellular reactive oxygen species (ROS) levels, and 4%PEG-96%PCL-0%cPCL was identified as the most appropriate coating material for this application. After 3days on this substrate HCASMC maintained a healthy contractile phenotype and HCAEC exhibited a physiologically relevant proliferation rate and a balanced redox state. Other test substrates promoted a pathological, synthetic phenotype of HCASMC and/or hyperproliferation of HCAEC. Phenotypic changes of HCASMC appeared to be modulated by the Young's modulus and surface charge of the test substrates, indicating a structure-function relationship that can be exploited for intricate control over vascular cell functions. These data indicate that tailored copolymer properties can direct vascular cell behavior and provide insights for further development of biologically instructive stent coating materials.

Physiologically relevant oxidative degradation of oligo(proline) cross-linked polymeric scaffolds.

Chronic inflammation-mediated oxidative stress is a common mechanism of implant rejection and failure. Therefore, polymer scaffolds that can degrade slowly in response to this environment may provide a viable platform for implant site-specific, sustained release of immunomodulatory agents over a long time period. In this work, proline oligomers of varying lengths (P(n)) were synthesized and exposed to oxidative environments, and their accelerated degradation under oxidative conditions was verified via high performance liquid chromatography and gel permeation chromatography. Next, diblock copolymers of poly(ethylene glycol) (PEG) and poly(ε-caprolactone) (PCL) were carboxylated to form 100 kDa terpolymers of 4%PEG-86%PCL-10%cPCL (cPCL = poly(carboxyl-ε-caprolactone); i% indicates molar ratio). The polymers were then cross-linked with biaminated PEG-P(n)-PEG chains, where P(n) indicates the length of the proline oligomer flanked by PEG chains. Salt-leaching of the polymeric matrices created scaffolds of macroporous and microporous architecture, as observed by scanning electron microscopy. The degradation of scaffolds was accelerated under oxidative conditions, as evidenced by mass loss and differential scanning calorimetry measurements. Immortalized murine bone-marrow-derived macrophages were then seeded on the scaffolds and activated through the addition of γ-interferon and lipopolysaccharide throughout the 9-day study period. This treatment promoted the release of H(2)O(2) by the macrophages and the degradation of proline-containing scaffolds compared to the control scaffolds. The accelerated degradation was evidenced by increased scaffold porosity, as visualized through scanning electron microscopy and X-ray microtomography imaging. The current study provides insight into the development of scaffolds that respond to oxidative environments through gradual degradation for the controlled release of therapeutics targeted to diseases that feature chronic inflammation and oxidative stress.