There is an unmet need to develop low-cost, rapid and highly multiplexed diagnostic technology platforms for quantitatively detecting blood biomarkers to advance clinical diagnostics beyond the single biomarker model. Here we perform nanopore sequencing of DNA-barcoded molecular probes engineered to recognize a panel of analytes. This allows for highly multiplexed and simultaneous quantitative detection of at least 40 targets, such as microRNAs, proteins and neurotransmitters, on the basis of the translocation dynamics of each probe as it passes through a nanopore. Our workflow is built around a commercially available MinION sequencing device, offering a one-hour turnaround time from sample preparation to results. We also demonstrate that the strategy can directly detect cardiovascular disease-associated microRNA from human serum without extraction or amplification. Due to the modularity of barcoded probes, the number and type of targets detected can be significantly expanded.

Fibrosis is a significant global health problem associated with many inflammatory and degenerative diseases affecting multiple organs, individually or simultaneously. Fibrosis develops when extracellular matrix (ECM) remodeling becomes excessive or uncontrolled and is associated with nearly all forms of heart disease. Cardiac fibroblasts and myofibroblasts are the main effectors of ECM deposition and scar formation. The heart is a complex multicellular organ, where the various resident cell types communicate between themselves and with cells of the blood and immune systems. Exosomes, which are small extracellular vesicles, (EVs), contribute to cell-to-cell communication and their pathophysiological relevance and therapeutic potential is emerging. Here, we will critically review the role of endogenous exosomes as possible fibrosis mediators and discuss the possibility of using stem cell-derived and/or engineered exosomes as anti-fibrotic agents.

Aberrant Zn²⁺ homeostasis is associated with dysregulated intracellular Ca²⁺ release, resulting in chronic heart failure. In the failing heart a small population of cardiac ryanodine receptors (RyR2) displays sub-conductance-state gating leading to Ca²⁺ leakage from sarcoplasmic reticulum (SR) stores, which impairs cardiac contractility. Previous evidence suggests contribution of RyR2-independent Ca²⁺ leakage through an uncharacterized mechanism. We sought to examine the role of Zn²⁺ in shaping intracellular Ca²⁺ release in cardiac muscle. Cardiac SR vesicles prepared from sheep or mouse ventricular tissue were incorporated into phospholipid bilayers under voltage-clamp conditions, and the direct action of Zn²⁺ on RyR2 channel function was examined. Under diastolic conditions, the addition of pathophysiological concentrations of Zn²⁺ (≥2 nm) caused dysregulated RyR2-channel openings. Our data also revealed that RyR2 channels are not the only SR Ca²⁺-permeable channels regulated by Zn²⁺. Elevating the cytosolic Zn²⁺ concentration to 1 nm increased the activity of the transmembrane protein mitsugumin 23 (MG23). The current amplitude of the MG23 full-open state was consistent with that previously reported for RyR2 sub-conductance gating, suggesting that in heart failure in which Zn²⁺ levels are elevated, RyR2 channels do not gate in a sub-conductance state, but rather MG23-gating becomes more apparent. We also show that in H9C2 cells exposed to ischemic conditions, intracellular Zn²⁺ levels are elevated, coinciding with increased MG23 expression. In conclusion, these data suggest that dysregulated Zn²⁺ homeostasis alters the function of both RyR2 and MG23 and that both ion channels play a key role in diastolic SR Ca²⁺ leakage.

Heart failure (HF) is a leading cause of morbidity and mortality worldwide, representing a complex clinical syndrome in which the heart’s ability to pump blood efficiently is impaired. HF can be subclassified into heart failure with reduced ejection fraction (HFrEF) and heart failure with preserved ejection fraction (HFpEF), each with distinct pathophysiological mechanisms and varying levels of severity. The progression of HF is significantly driven by cardiac fibrosis, a pathological process in which the extracellular matrix undergoes abnormal and uncontrolled remodelling. Cardiac fibrosis is characterized by excessive matrix protein deposition and the activation of myofibroblasts, increasing the stiffness of the heart, thus disrupting its normal structure and function and promoting lethal arrythmia. MicroRNAs, long non-coding RNAs, and circular RNAs, collectively known as non-coding RNAs (ncRNAs), have recently gained significant attention due to a growing body of evidence suggesting their involvement in cardiac remodelling such as fibrosis. ncRNAs can be found in the peripheral blood, indicating their potential as biomarkers for assessing HF severity. In this review, we critically examine recent advancements and findings related to the use of ncRNAs as biomarkers of HF and discuss their implication in fibrosis development.

We present a novel framework, Opto-SICM, for studies of cellular interactions in live cells with high spatiotemporal resolution. The approach combines scanning ion conductance microscopy, SICM, and cell-type-specific optogenetic interrogation. Light-excitable cardiac fibroblasts (FB) and myofibroblasts (myoFB) were plated together with non-modified cardiomyocytes (CM) and then paced with periodic illumination. Opto-SICM reveals the extent of FB/myoFB-CM cell-cell contacts and the dynamic changes over time not visible by optical microscopy. FB-CM pairs have lower gap junctional expression of connexin-43 and higher contact dynamism compared to myoFB-CM pairs. The responsiveness of CM to pacing via FB/myoFB depends on the dynamics of the contact but not on the area. The non-responding pairs have higher net cell-cell movement at the contact. These findings are relevant to cardiac disease states, where adverse remodeling leads to abnormal electrical excitation of CM. The Opto-SICM framework can be deployed to offer new insights on cellular and subcellular interactions in various cell types, in real-time.