The Role of miR-181c in Mechanisms of Diabetes-Impaired Angiogenesis: An Emerging Therapeutic Target for Diabetic Vascular Complications

Diabetes mellitus is estimated to affect up to 700 million people by the year 2045, contributing to an immense health and economic burden. People living with diabetes have a higher risk of developing numerous debilitating vascular complications, leading to an increased need for medical care, a reduced quality of life and increased risk of early death. Current treatments are not satisfactory for many patients who suffer from impaired angiogenesis in response to ischaemia, increasing their risk of ischaemic cardiovascular conditions. These vascular pathologies are characterised by endothelial dysfunction and abnormal angiogenesis, amongst a host of impaired signaling pathways. Therapeutic stimulation of angiogenesis holds promise for the treatment of diabetic vascular complications that stem from impaired ischaemic responses. However, despite significant effort and research, there are no established therapies that directly stimulate angiogenesis to improve ischaemic complications such as ischaemic heart disease and peripheral artery disease, highlighting the immense unmet need. However, despite significant effort and research, there are no established therapies that directly stimulate angiogenesis in a clinical setting, highlighting the immense unmet need. MicroRNAs (miRNAs) are emerging as powerful targets for multifaceted diseases including diabetes and cardiovascular disease. This review highlights the potential role of microRNAs as therapeutic targets for rescuing diabetes-impaired angiogenesis, with a specific focus on miR-181c, which we have previously identified as an important angiogenic regulator. Here we summarise the pathways currently known to be regulated by miR-181c, which include the classical angiogenesis pathways that are dysregulated in diabetes, mitochondrial function and axonal guidance, and describe how these relate both directly and indirectly to angiogenesis. The pleiotropic actions of miR-181c across multiple key angiogenic signaling pathways and critical cellular processes highlight its therapeutic potential as a novel target for treating diabetic vascular complications.Emma L. Solly, Peter J. Psaltis, Christina A. Bursill, and Joanne T. M. Ta

MicroRNAs as therapeutic targets and clinical biomarkers in atherosclerosis

Atherosclerotic cardiovascular disease remains the leading cause of morbidity and mortality worldwide. Atherosclerosis develops over several decades and is mediated by a complex interplay of cellular mechanisms that drive a chronic inflammatory milieu and cell-to-cell interactions between endothelial cells, smooth muscle cells and macrophages that promote plaque development and progression. While there has been significant therapeutic advancement, there remains a gap where novel therapeutic approaches can complement current therapies to provide a holistic approach for treating atherosclerosis to orchestrate the regulation of complex signalling networks across multiple cell types and different stages of disease progression. MicroRNAs (miRNAs) are emerging as important post-transcriptional regulators of a suite of molecular signalling pathways and pathophysiological cellular effects. Furthermore, circulating miRNAs have emerged as a new class of disease biomarkers to better inform clinical diagnosis and provide new avenues for personalised therapies. This review focusses on recent insights into the potential role of miRNAs both as therapeutic targets in the regulation of the most influential processes that govern atherosclerosis and as clinical biomarkers that may be reflective of disease severity, highlighting the potential theranostic (therapeutic and diagnostic) properties of miRNAs in the management of cardiovascular disease.Emma L. Solly, Catherine G. Dimasi, Christina A. Bursill, Peter J. Psaltis, and Joanne T.M. Ta

Inflammation as a therapeutic target in atherosclerosis

Atherosclerotic coronary artery disease (CAD) results from build-up of cholesterol-rich plaques in the walls of the coronary arteries and is a leading cause of death. Inflammation is central to atherosclerosis. Uncontrolled inflammation makes coronary plaques "unstable" and vulnerable to rupture or erosion, leading to thrombosis and myocardial infarction (MI). As multiple inflamed plaques often co-exist in the coronary system, patients are at risk of repeated atherothrombotic cardiovascular events after MI, with rates of 10-12% at one year and 18-20% at three years. This is largely because current therapies for CAD, such as lipid-lowering statins, do not adequately control plaque inflammation. New anti-atherosclerotic agents are therefore needed, especially those that better target inflammation. The recent positive results for the anti-interleukin-1-beta (IL-1β) monoclonal antibody, Canakinumab, in the Canakinumab Anti-inflammatory Thrombosis Outcome Study (CANTOS) clinical trial has provided a major stimulant to the field. It highlights that not only is inflammation important from a pathogenic and risk prediction perspective in CAD, but that reducing inflammation can be beneficial. The challenge is now to find the best strategies to achieve this in real-world practice. This review outlines the role that inflammation plays in atherosclerosis and provides an update on anti-inflammatory therapies currently being investigated to target atherosclerosis.Mau T Nguyen, Sanuja Fernando, Nisha Schwarz, Joanne TM Tan, Christina A Bursill and Peter J Psalti

The mechanisms of restenosis and relevance to next generation stent design

Stents are lifesaving mechanical devices that re-establish essential blood flow to the coronary circulation after significant vessel occlusion due to coronary vessel disease or thrombolytic blockade. Improvements in stent surface engineering over the last 20 years have seen significant reductions in complications arising due to restenosis and thrombosis. However, under certain conditions such as diabetes mellitus (DM), the incidence of stent-mediated complications remains 2–4-fold higher than seen in non-diabetic patients. The stents with the largest market share are designed to target the mechanisms behind neointimal hyperplasia (NIH) through anti-proliferative drugs that prevent the formation of a neointima by halting the cell cycle of vascular smooth muscle cells (VSMCs). Thrombosis is treated through dual anti-platelet therapy (DAPT), which is the continual use of aspirin and a P2Y12 inhibitor for 6–12 months. While the most common stents currently in use are reasonably effective at treating these complications, there is still significant room for improvement. Recently, inflammation and redox stress have been identified as major contributing factors that increase the risk of stent-related complications following percutaneous coronary intervention (PCI). The aim of this review is to examine the mechanisms behind inflammation and redox stress through the lens of PCI and its complications and to establish whether tailored targeting of these key mechanistic pathways offers improved outcomes for patients, particularly those where stent placement remains vulnerable to complications. In summary, our review highlights the most recent and promising research being undertaken in understanding the mechanisms of redox biology and inflammation in the context of stent design. We emphasize the benefits of a targeted mechanistic approach to decrease all-cause mortality, even in patients with diabetes.Jessie Clare, Justin Ganly, Christina A. Bursill, Huseyin Sumer, Peter Kingshott, and Judy B. de Haa

Infrared conductivity of a one-dimensional charge-ordered state: quantum lattice effects

The optical properties of the charge-ordering ($CO$) phase of the one-dimensional (1D) half-filled spinless Holstein model are derived at zero temperature within a well-known variational approach improved including second-order lattice fluctuations. Within the $CO$ phase, the static lattice distortions give rise to the optical interband gap, that broadens as the strength of the electron-phonon ($el-ph$) interaction increases. The lattice fluctuation effects induce a long subgap tail in the infrared conductivity and a wide band above the gap energy. The first term is due to the multi-phonon emission by the charge carriers, the second to the interband transitions accompanied by the multi-phonon scattering. The results show a good agreement with experimental spectra.Comment: 5 figure

Vasculogenic properties of adventitial Sca-1(+)CD45(+) progenitor cells in mice: a potential source of vasa vasorum in atherosclerosis

The cellular origins of vasa vasorum are ill-defined and may involve circulating or local progenitor cells. We previously discovered that murine aortic adventitia contains Sca-1⁺CD45⁺ progenitors that produce macrophages. Here we investigated whether they are also vasculogenic. In aortas of C57BL/6 mice, Sca-1⁺CD45⁺ cells were localised to adventitia and lacked surface expression of endothelial markers (<1% for CD31, CD144, TIE-2). In contrast, they did show expression of CD31, CD144, TIE-2 and VEGFR2 in atherosclerotic ApoE(-/-) aortas. Although Sca-1⁺CD45⁺ cells from C57BL/6 aorta did not express CD31, they formed CD31⁺ colonies in endothelial differentiation media and produced interconnecting vascular-like cords in Matrigel that contained both endothelial cells and a small population of macrophages, which were located at branch points. Transfer of aortic Sca-1⁺CD45⁺ cells generated endothelial cells and neovessels de novo in a hindlimb model of ischaemia and resulted in a 50% increase in perfusion compared to cell-free control. Similarly, their injection into the carotid adventitia of ApoE(-/-) mice produced donor-derived adventitial and peri-adventitial microvessels after atherogenic diet, suggestive of newly formed vasa vasorum. These findings show that beyond its content of macrophage progenitors, adventitial Sca-1⁺CD45⁺ cells are also vasculogenic and may be a source of vasa vasorum during atherogenesis.Deborah Toledo-Flores, Anna Williamson, Nisha Schwarz, Sanuja Fernando, Catherine Dimasi, Tyra A. Witt, Thao M. Nguyen, Amrutesh S . Puranik, Colin D. Chue, Sinny Delacroix, Daniel B. Spoon, Claudine S. Bonder, Christina A. Bursill, Belinda A. Di Bartolo, Stephen J. Nicholls, Robert D. Simari, Peter J. Psalti

Cardiovascular bioimaging of nitric oxide: achievements, challenges, and the future

First published: 19 October 2020Nitric oxide (NO) is a ubiquitous, volatile, cellular signaling molecule that operates across a wide physiological concentration range (pM-µM) in different tissues. It is a highly diffusible messenger and intermediate in various metabolic pathways. NO plays a pivotal role in maintaining optimum cardiovascular function, particularly by regulating vascular tone and blood flow. This review highlights the need for accurate, real-time bioimaging of NO in clinical diagnostic, therapeutic, monitoring, and theranostic applications within the cardiovascular system. We summarize electrochemical, optical, and nanoscale sensors that allow measurement and imaging of NO, both directly and indirectly via surrogate measurements. The physical properties of NO render it difficult to accurately measure in tissues using direct methods. There are also significant limitations associated with the NO metabolites used as surrogates to indirectly estimate NO levels. All these factors added to significant variability in the measurement of NO using available methodology have led to a lack of sensors and imaging techniques of clinical applicability in relevant vascular pathologies such as atherosclerosis and ischemic heart disease. Challenges in applying current methods to biomedical and clinical translational research, including the wide physiological range of NO and limitations due to the characteristics and toxicity of the sensors are discussed, as are potential targets and modifications for future studies. The development of biocompatible nanoscale sensors for use in combination with existing clinical imaging modalities provides a feasible opportunity for bioimaging NO within the cardiovascular system.Achini K. Vidanapathirana, Peter J. Psaltis, Christina A. Bursill, Andrew D. Abell, Stephen J. Nicholls ... et al

Elevated HDL-bound miR-181c-5p level is associated with diabetic vascular complications in Australian Aboriginal people

Published: 02 March 2021Aims/hypothesis: Diabetes is a major burden on Australia’s Indigenous population, with high rates of disease and vascular complications. Diabetic vascular complications are associated with impaired ischaemia-driven angiogenesis. MicroRNAs (miRNAs) are key players in the regulation of angiogenesis. HDL-cholesterol (HDL-c) levels are inversely associated with the risk of developing diabetic complications and HDL can carry miRNAs. HDL-miRNA profiles differ in disease states and may present as biomarkers with the capacity to act as bioactive signalling molecules. Recent studies have demonstrated that HDL becomes dysfunctional in a diabetic environment, losing its vasculo-protective effects and becoming more pro-atherogenic. We sought to determine whether HDL-associated miRNA profiles and HDL functionality were predictive of the severity of diabetic vascular complications in Australia’s Indigenous population. Methods: HDL was isolated from plasma samples from Indigenous participants without diabetes (‘Healthy’), with type 2 diabetes mellitus (‘T2DM’) and with diabetes-associated macrovascular complications (specifically peripheral artery disease, ‘T2DM+Comp’). To assess HDL angiogenic capacity, human coronary artery endothelial cells were treated with PBS, reconstituted HDL (rHDL, positive control) or isolated HDL and then exposed to high-glucose (25 mmol/l) conditions. The expression levels of two anti-angiogenic miRNAs (miR-181c-5p and miR-223-3p) and one pro-angiogenic miRNA (miR-27b-3p) were measured in the HDL fraction, plasma and treated human coronary artery endothelial cells by quantitative real-time PCR. In vitro endothelial tubule formation was assessed using the Matrigel tubulogenesis assay. Results: Strikingly, we found that the levels of the anti-angiogenic miRNA miR-181c-5p were 14-fold higher (1454 ± 1346%) in the HDL from Aboriginal people with diabetic complications compared with both the Healthy (100 ± 121%, p < 0.05) and T2DM (82 ± 77%, p < 0.05) groups. Interestingly, we observed a positive correlation between HDL-associated miR-181c-5p levels and disease severity (p = 0.0020). Under high-glucose conditions, cells treated with rHDL, Healthy HDL and T2DM HDL had increased numbers of tubules (rHDL: 136 ± 8%, p < 0.01; Healthy HDL: 128 ± 6%, p < 0.01; T2DM HDL: 124 ± 5%, p < 0.05) and branch points (rHDL: 138 ± 8%, p < 0.001; Healthy HDL: 128 ± 6%, p < 0.01; T2DM HDL: 127 ± 5%, p < 0.01) concomitant with elevations in mRNA levels of the key hypoxia angiogenic transcription factor HIF1A (rHDL: 140 ± 10%, p < 0.01; Healthy HDL: 136 ± 8%, p < 0.01; T2DM HDL: 133 ± 9%, p < 0.05). However, this increase in angiogenic capacity was not observed in cells treated with T2DM + Comp HDL (tubule numbers: 113 ± 6%, p = 0.32; branch points: 113 ± 5%, p = 0.28; HIF1A: 117 ± 6%, p = 0.43), which could be attributed to the increase in cellular miR-181c-5p levels (T2DM + Comp HDL: 136 ± 7% vs PBS: 100 ± 9%, p < 0.05). Conclusions/interpretation: In conclusion, HDL from Aboriginal people with diabetic complications had reduced angiogenic capacity. This impairment is associated with an increase in the expression of anti-angiogenic miR-181c-5p. These findings provide the rationale for a new way to better inform clinical diagnosis of disease severity with the potential to incorporate targeted, personalised HDL-miRNA intervention therapies to prevent further development of, or to reverse, diabetic vascular complications in Australian Aboriginal people.Kaitlin R. Morrison, Emma L. Solly ... Peter J. Psaltis, Stephen J. Nicholls, Alex Brown, Christina A. Bursill ... et al

A novel ruthenium-based molecular sensor to detect endothelial nitric oxide

Nitric oxide (NO) is a key regulator of endothelial cell and vascular function. The direct measurement of NO is challenging due to its short half-life, and as such surrogate measurements are typically used to approximate its relative concentrations. Here we demonstrate that ruthenium-based [Ru(bpy)2(dabpy)]2+ is a potent sensor for NO in its irreversible, NO-bound active form, [Ru(bpy)2(T-bpy)]2+. Using spectrophotometry we established the sensor's ability to detect and measure soluble NO in a concentration-dependent manner in cell-free media. Endothelial cells cultured with acetylcholine or hydrogen peroxide to induce endogenous NO production showed modest increases of 7.3 ± 7.1% and 36.3 ± 25.0% respectively in fluorescence signal from baseline state, while addition of exogenous NO increased their fluorescence by 5.2-fold. The changes in fluorescence signal were proportionate and comparable against conventional NO assays. Rabbit blood samples immediately exposed to [Ru(bpy)2(dabpy)]2+ displayed 8-fold higher mean fluorescence, relative to blood without sensor. Approximately 14% of the observed signal was NO/NO adduct-specific. Optimal readings were obtained when sensor was added to freshly collected blood, remaining stable during subsequent freeze-thaw cycles. Clinical studies are now required to test the utility of [Ru(bpy)2(dabpy)]2+ as a sensor to detect changes in NO from human blood samples in cardiovascular health and disease.Achini K. Vidanapathirana, Benjamin J. Pullen, Run Zhang, MyNgan Duong, Jarrad M. Goyne, Xiaozhou Zhang, Claudine S. Bonder, Andrew D. Abell, Christina A. Bursill, Stephen J. Nicholls, Peter J. Psalti

Ultrathin monolithic 3D printed optical coherence tomography endoscopy for preclinical and clinical use

Preclinical and clinical diagnostics increasingly rely on techniques to visualize internal organs at high resolution via endoscopes. Miniaturized endoscopic probes are necessary for imaging small luminal or delicate organs without causing trauma to tissue. However, current fabrication methods limit the imaging performance of highly miniaturized probes, restricting their widespread application. To overcome this limitation, we developed a novel ultrathin probe fabrication technique that utilizes 3D microprinting to reliably create side-facing freeform micro-optics (<130 µm diameter) on single-mode fibers. Using this technique, we built a fully functional ultrathin aberration-corrected optical coherence tomography probe. This is the smallest freeform 3D imaging probe yet reported, with a diameter of 0.457 mm, including the catheter sheath. We demonstrated image quality and mechanical flexibility by imaging atherosclerotic human and mouse arteries. The ability to provide microstructural information with the smallest optical coherence tomography catheter opens a gateway for novel minimally invasive applications in disease.Jiawen Li, Simon Thiele, Bryden C. Quirk, Rodney W. Kirk, Johan W. Verjans, Emma Akers ... et al