New Spectroelectrochemical Insights into Manganese and Rhenium Bipyridine Complexes as Catalysts for the Electrochemical Reduction of Carbon Dioxide.

This study aimed to demonstrate the behavior of different complexes using IR spectroelectrochemistry (SEC), a technique that combines IR spectroscopy with electrochemistry. Four different Mn and Re catalysts for electrochemical CO2 reduction were studied in dry acetonitrile. In the case of Mn(apbpy)(CO)3Br (apbpy = 4(4-aminophenyl)-2,2'-bipyridine), SEC suggested that a very slow catalytic reduction of CO2 also occurs in acetonitrile in the absence of proton donors, but at rather negative potentials. In contrast, the corresponding Re(apbpy)(CO)3Br clearly demonstrated slow catalytic conversion at the first reduction potential. Switching to saturated CO2 solutions in a mixture of acetonitrile and 5% water as a proton donor, the SEC of Mn(apbpy)(CO)3Br displayed a faster catalytic behavior.

Development and characterization of anti-fibrotic natural compound similars with improved effectivity.

Cardiac fibroblasts constitute the major cell type of the murine and human heart. Once activated, they contribute to an excessive deposition of extracellular matrix (ECM) leading to cardiac fibrosis and subsequently organ dysfunction. With the exception of the pulmonary drugs, nintedanib and pirfenidone, drugs specifically targeting anti-fibrotic pathways are scarce. We recently performed large library screenings of natural occurring compounds and identified first lead structures with anti-fibrotic properties in vitro and in vivo. In line, we now aimed to improve efficacy of these anti-fibrotic lead structures by combining in vitro validation studies and in silico prediction. Next to this combined approach, we performed large OMICs-multi-panel-based mechanistic studies. Applying human cardiac fibroblasts (HCF), we analysed 26 similars of the initially identified anti-fibrotic lead molecules bufalin and lycorine and determined anti-proliferative activity and potential toxicity in an array of in vitro and ex vivo studies. Of note, even at lower concentrations, certain similars were more effective at inhibiting HCF proliferation than nintedanib and pirfenidone. Additionally, selected similars showed low cytotoxicity on human iPS-derived cardiomyocytes and anti-fibrotic gene regulation in human ex vivo living myocardial slices. Further, array and RNA sequencing studies of coding and non-coding RNAs in treated HCFs revealed strong anti-fibrotic properties, especially with the lycorine similar lyco-s (also known as homoharringtonine), that led to a nearly complete shutdown of ECM production at concentrations 100-fold lower than the previously identified anti-fibrotic compound lycorine without inducing cellular toxicity. We thus identified a new natural compound similar with strong anti-fibrotic properties in human cardiac fibroblasts and human living heart tissue potentially opening new anti-fibrotic treatment strategies.

MicroRNA-449a Inhibits Triple Negative Breast Cancer by Disturbing DNA Repair and Chromatid Separation.

Chromosomal instability (CIN) can be a driver of tumorigenesis but is also a promising therapeutic target for cancer associated with poor prognosis such as triple negative breast cancer (TNBC). The treatment of TNBC cells with defects in DNA repair genes with poly(ADP-ribose) polymerase inhibitor (PARPi) massively increases CIN, resulting in apoptosis. Here, we identified a previously unknown role of microRNA-449a in CIN. The transfection of TNBC cell lines HCC38, HCC1937 and HCC1395 with microRNA-449a mimics led to induced apoptosis, reduced cell proliferation, and reduced expression of genes in homology directed repair (HDR) in microarray analyses. EME1 was identified as a new target gene by immunoprecipitation and luciferase assays. The reduced expression of EME1 led to an increased frequency of ultrafine bridges, 53BP1 foci, and micronuclei. The induced expression of microRNA-449a elevated CIN beyond tolerable levels and induced apoptosis in TNBC cell lines by two different mechanisms: (I) promoting chromatid mis-segregation by targeting endonuclease EME1 and (II) inhibiting HDR by downregulating key players of the HDR network such as E2F3, BIRC5, BRCA2 and RAD51. The ectopic expression of microRNA-449a enhanced the toxic effect of PARPi in cells with pathogenic germline BRCA1 variants. The newly identified role makes microRNA-449a an interesting therapeutic target for TNBC.

Combined high-throughput library screening and next generation RNA sequencing uncover microRNAs controlling human cardiac fibroblast biology.

BACKGROUND: Myocardial fibrosis is a hallmark of the failing heart, contributing to the most common causes of deaths worldwide. Several microRNAs (miRNAs, miRs) controlling cardiac fibrosis were identified in recent years; however, a more global approach to identify miRNAs involved in fibrosis is missing. METHODS AND RESULTS: Functional miRNA mimic library screens were applied in human cardiac fibroblasts (HCFs) to identify annotated miRNAs inducing proliferation. In parallel, miRNA deep sequencing was performed after subjecting HCFs to proliferating and resting stimuli, additionally enabling discovery of novel miRNAs. In-depth in vitro analysis confirmed the pro-fibrotic nature of selected, highly conserved miRNAs miR-20a-5p and miR-132-3p. To determine downstream cellular pathways and their role in the fibrotic response, targets of the annotated miRNA candidates were modulated by synthetic siRNA. We here provide evidence that repression of autophagy and detoxification of reactive oxygen species by miR-20a-5p and miR-132-3p explain some of their pro-fibrotic nature on a mechanistic level. CONCLUSION: We here identified both miR-20a-5p and miR-132-3p as crucial regulators of fibrotic pathways in an in vitro model of human cardiac fibroblast biology.

Natural Compound Library Screening Identifies New Molecules for the Treatment of Cardiac Fibrosis and Diastolic Dysfunction.

BACKGROUND: Myocardial fibrosis is a hallmark of cardiac remodeling and functionally involved in heart failure development, a leading cause of deaths worldwide. Clinically, no therapeutic strategy is available that specifically attenuates maladaptive responses of cardiac fibroblasts, the effector cells of fibrosis in the heart. Therefore, our aim was to develop novel antifibrotic therapeutics based on naturally derived substance library screens for the treatment of cardiac fibrosis. METHODS: Antifibrotic drug candidates were identified by functional screening of 480 chemically diverse natural compounds in primary human cardiac fibroblasts, subsequent validation, and mechanistic in vitro and in vivo studies. Hits were analyzed for dose-dependent inhibition of proliferation of human cardiac fibroblasts, modulation of apoptosis, and extracellular matrix expression. In vitro findings were confirmed in vivo with an angiotensin II-mediated murine model of cardiac fibrosis in both preventive and therapeutic settings, as well as in the Dahl salt-sensitive rat model. To investigate the mechanism underlying the antifibrotic potential of the lead compounds, treatment-dependent changes in the noncoding RNAome in primary human cardiac fibroblasts were analyzed by RNA deep sequencing. RESULTS: High-throughput natural compound library screening identified 15 substances with antiproliferative effects in human cardiac fibroblasts. Using multiple in vitro fibrosis assays and stringent selection algorithms, we identified the steroid bufalin (from Chinese toad venom) and the alkaloid lycorine (from Amaryllidaceae species) to be effective antifibrotic molecules both in vitro and in vivo, leading to improvement in diastolic function in 2 hypertension-dependent rodent models of cardiac fibrosis. Administration at effective doses did not change plasma damage markers or the morphology of kidney and liver, providing the first toxicological safety data. Using next-generation sequencing, we identified the conserved microRNA 671-5p and downstream the antifibrotic selenoprotein P1 as common effectors of the antifibrotic compounds. CONCLUSIONS: We identified the molecules bufalin and lycorine as drug candidates for therapeutic applications in cardiac fibrosis and diastolic dysfunction.

Senescence-induced inflammation: an important player and key therapeutic target in atherosclerosis.

Inflammation is a hallmark and potent driver of pathological vascular remodelling in atherosclerosis. However, current anti-inflammatory therapeutic strategies have shown mixed results. As an alternative perspective on the conundrum of chronic inflammation emerging evidence points towards a small subset of senescent cells as a critical player and central node driving atherosclerosis. Senescent cells belonging to various cell types are a dominant and chronic source of a large array of pro-inflammatory cytokines and various additional plaque destabilizing factors, being involved with various aspects of atherosclerosis pathogenesis. Antagonizing these key agitators of local chronic inflammation and plaque instability may provide a causative and multi-purpose therapeutic strategy to treat atherosclerosis. Anti-senescence treatment options with translational potential are currently in development. However, several questions and challenges remain to be addressed before these novel treatment approaches may enter the clinical setting.

Pleiotropic cardiac functions controlled by ischemia-induced lncRNA H19.

Myocardial ischemia induces a multifaceted remodeling process in the heart. Novel therapeutic entry points to counteract maladaptive signalling include the modulation of non-coding RNA molecules such as long non-coding RNA (lncRNA). We here questioned if the lncRNA candidate H19 exhibits regulatory potential in the setting of myocardial infarction. Initial profiling of H19 expression revealed a dynamic expression profile of H19 with upregulation in the acute phase after murine cardiac ischemia. In vitro, we found that oxygen deficiency leads to H19 upregulation in several cardiac cell types. Repression of endogenous H19 caused multiple phenotypes in cultivated murine cardiomyocytes including enhanced cardiomyocyte apoptosis, at least partly through attenuated vitamin D signalling. Unbiased proteome analysis revealed further involvement of H19 in mRNA splicing and translation as well as inflammatory signalling pathways. To study H19 function more precisely, we investigated the phenotype of systemic H19 loss in a genetic mouse model of H19 deletion (H19 KO). Infarcted heart tissue of H19 KO mice showed a massive increase of pro-inflammatory cytokines after ischemia-reperfusion injury (I/R) without significant effects on scar formation or cardiac function but exaggerated cardiac hypertrophy indicating pathological cardiac remodeling. H19-dependent changes in cardiomyocyte-derived extracellular vesicle release and alterations in NF-κB signalling were evident. Cardiac cell fractionation experiments revealed that enhanced H19 expression in the proliferative phase after MI derived mainly from cardiac fibroblasts. Here further research is needed to elucidate its role in fibroblast activation and function. In conclusion, the lncRNA H19 is dynamically regulated after MI and involved in multiple pathways of different cardiac cell types including cardiomyocyte apoptosis and cardiac inflammation.

Non-coding RNAs: key players in cardiac disease.

Molecular mechanisms underlying heart failure (HF) are only partly understood. Non-coding RNAs (ncRNAs) have been reported to control function and signalling routes in the myocardium. As ncRNAs such as microRNAs (miRNAs), long non-coding RNAs (lncRNAs) or circular RNAs (circRNAs) can be selectively targeted via pharmacological approaches, this opens new avenues for diagnostic and therapeutic approaches. Here, we review the main ncRNA classes and how they influence cardiac biology. In addition we provide insight into the role of ncRNAs in chemotherapy-induced cardiac dysfunction. To provide a better understanding of ncRNAs in cardiovascular biology we present an outlook on specialized functions such as chromatin remodelling, biomarker potential and the recently discovered ncRNA-derived micropeptides.

Cardiac endurance training alters plasma profiles of circular RNA MBOAT2.

Marathon running is an extreme physical activity, which determines cardiopulmonary adaption of athletes. Circular RNAs (circRNAs) as potential biomarkers in the blood stream have so far not been tested after such strenuous activities. In silico approaches were performed to identify the potential candidate circRNA MBOAT2. Next, we demonstrated high stability and conservation of circRNA MBOAT2 as well as its abundancy in human plasma. In addition to Sanger sequencing of the circRNA specific head-to-tail junction, or back-splice site, we established a synthetic plasmid standard which allowed exact copy number calculations of circRNA MBOAT2. We then analyzed plasmatic circRNA MBOAT2 and observed a significantly lower level 24 h after the marathon. Such alterations were correlated to physical exercise parameters confirming the role of circRNA MBOAT2 as a promising noncoding RNA biomarker detecting cardiopulmonary adaption.NEW & NOTEWORTHY In brief, we herein report a timeline of circulating circular RNA (circRNA) MBOAT2 in a cohort of marathon runners. Time-course analysis of plasmatic circRNA MBOAT2 demonstrated a significantly lowered level 24 h after the marathon. Abundancy of circRNA was correlated to physical exercise parameters highlighting the role of circRNA MBOAT2 as a valuable noncoding RNA biomarker detecting and following up cardiopulmonary adaption.

Quaking Inhibits Doxorubicin-Mediated Cardiotoxicity Through Regulation of Cardiac Circular RNA Expression.

RATIONALE: RBPs (RNA-binding proteins) have been described to be expressed and regulated in various organs including the heart. Little is known about the role of RBPs in heart failure induced by the chemotherapy drug doxorubicin and their interaction with circular RNAs. OBJECTIVE: We aimed to identify key RBPs involved in doxorubicin-mediated heart failure and to elucidate their function. METHODS AND RESULTS: Global transcriptome profiling from murine myocardium exposed to doxorubicin identified 5 differentially expressed RBPs. Expression of the RBP QKI (Quaking) in response to doxorubicin was strongly downregulated in rodent cardiomyocytes and human induced pluripotent stem cell-derived cardiomyocytes in vitro and in vivo in mice. Knockdown of Qki in primary cardiomyocytes increased apoptosis and atrophy after treatment with doxorubicin, whereas lentiviral mediated overexpression of Qki5 inhibited the doxorubicin-induced apoptosis in cardiomyocytes. In vivo, AAV9 (adeno-associated virus serotype 9)-mediated cardiac overexpression of Qki5 prevented cardiac apoptosis and cardiac atrophy induced by doxorubicin and improved cardiac function. Mechanistically, by lentiviral-based overexpression and CRISPR/Cas9-mediated silencing of Qki5, we identified regulated expression of specific circular RNAs derived from Ttn (Titin), Fhod3 (Formin homology 2 domain containing 3), and Strn3 (Striatin, calmodulin-binding protein 3). Moreover, inhibition of Ttn-derived circular RNA increased the susceptibility of cardiomyocytes to doxorubicin. CONCLUSIONS: We here show that overexpression of Qki5 strongly attenuates the toxic effect of doxorubicin via regulating a set of circular RNAs. Qki5 is, thus, an interesting target molecule to combat doxorubicin-induced cardiotoxicity.

Rhenium Complexes of Pyridyl-Mesoionic Carbenes: Photochemical Properties and Electrocatalytic CO2 Reduction.

Mesoionic carbenes have found wide use as components of homogeneous catalysts. Recent discoveries have, however, shown that metal complexes of such ligands also have huge potential in photochemical research and in the activation of small molecules. We present here three ReI complexes with mesoionic pyridyl-carbene ligands. The complexes display reduction steps which were investigated via UV-vis-NIR-IR spectro-electrochemistry, and these results point toward an EC mechanism. The ReI compounds emit in the visible range in solution at room temperature with excited state lifetimes that are dependent on the substituents of the mesoionic carbenes. These complexes are also potent electrocatalysts for the selective reduction of CO2 to CO. Whereas the substituents on the carbenes have no influence on the reduction potentials, the electrocatalytic efficiency is strongly dependent on the substituents. This fact is likely a result of catalyst instability. The results presented here thus introduce mesoionic carbenes as new potent ligands for the generation of emissive ReI complexes and for electrocatalytic CO2 reduction.

Circulating non-coding RNAs in biomarker-guided cardiovascular therapy: a novel tool for personalized medicine?

Current clinical guidelines emphasize the unmet need for technological innovations to guide physician decision-making and to transit from conventional care to personalized cardiovascular medicine. Biomarker-guided cardiovascular therapy represents an interesting approach to inform tailored treatment selection and monitor ongoing efficacy. However, results from previous publications cast some doubts about the clinical applicability of biomarkers to direct individualized treatment. In recent years, the non-coding human transcriptome has emerged as a new opportunity for the development of novel therapeutic strategies and biomarker discovery. Non-coding RNA (ncRNA) signatures may provide an accurate molecular fingerprint of patient phenotypes and capture levels of information that could complement traditional markers and established clinical variables. Importantly, ncRNAs have been identified in body fluids and their concentrations change with physiology and pathology, thus representing promising non-invasive biomarkers. Previous publications highlight the translational applicability of circulating ncRNAs for diagnosis and prognostic stratification within cardiology. Numerous independent studies have also evaluated the potential of the circulating non-coding transcriptome to predict and monitor response to cardiovascular treatment. However, this field has not been reviewed in detail. Here, we discuss the state-of-the-art research into circulating ncRNAs, specifically microRNAs and long non-coding RNAs, to support clinical decision-making in cardiovascular therapy. Furthermore, we summarize current methodological and conceptual limitations and propose future steps for their incorporation into personalized cardiology. Despite the lack of robust population-based studies and technical barriers, circulating ncRNAs emerge as a promising tool for biomarker-guided therapy.

miR-21-KO Alleviates Alveolar Structural Remodeling and Inflammatory Signaling in Acute Lung Injury.

Acute lung injury (ALI) is characterized by enhanced permeability of the air-blood barrier, pulmonary edema, and hypoxemia. MicroRNA-21 (miR-21) was shown to be involved in pulmonary remodeling and the pathology of ALI, and we hypothesized that miR-21 knock-out (KO) reduces injury and remodeling in ALI. ALI was induced in miR-21 KO and C57BL/6N (wildtype, WT) mice by an intranasal administration of 75 µg lipopolysaccharide (LPS) in saline (n = 10 per group). The control mice received saline alone (n = 7 per group). After 24 h, lung function was measured. The lungs were then excised for proteomics, cytokine, and stereological analysis to address inflammatory signaling and structural damage. LPS exposure induced ALI in both strains, however, only WT mice showed increased tissue resistance and septal thickening upon LPS treatment. Septal alterations due to LPS exposure in WT mice consisted of an increase in extracellular matrix (ECM), including collagen fibrils, elastic fibers, and amorphous ECM. Proteomics analysis revealed that the inflammatory response was dampened in miR-21 KO mice with reduced platelet and neutrophil activation compared with WT mice. The WT mice showed more functional and structural changes and inflammatory signaling in ALI than miR-21 KO mice, confirming the hypothesis that miR-21 KO reduces the development of pathological changes in ALI.

Comprehensive Bioinformatics Identifies Key microRNA Players in ATG7-Deficient Lung Fibroblasts.

BACKGROUND: Deficient autophagy has been recently implicated as a driver of pulmonary fibrosis, yet bioinformatics approaches to study this cellular process are lacking. Autophagy-related 5 and 7 (ATG5/ATG7) are critical elements of macro-autophagy. However, an alternative ATG5/ATG7-independent macro-autophagy pathway was recently discovered, its regulation being unknown. Using a bioinformatics proteome profiling analysis of ATG7-deficient human fibroblasts, we aimed to identify key microRNA (miR) regulators in autophagy. METHOD: We have generated ATG7-knockout MRC-5 fibroblasts and performed mass spectrometry to generate a large-scale proteomics dataset. We further quantified the interactions between various proteins combining bioinformatics molecular network reconstruction and functional enrichment analysis. The predicted key regulatory miRs were validated via quantitative polymerase chain reaction. RESULTS: The functional enrichment analysis of the 26 deregulated proteins showed decreased cellular trafficking, increased mitophagy and senescence as the major overarching processes in ATG7-deficient lung fibroblasts. The 26 proteins reconstitute a protein interactome of 46 nodes and miR-regulated interactome of 834 nodes. The miR network shows three functional cluster modules around miR-16-5p, miR-17-5p and let-7a related to multiple deregulated proteins. Confirming these results in a biological setting, serially passaged wild-type and autophagy-deficient fibroblasts displayed senescence-dependent expression profiles of miR-16-5p and miR-17-5p. CONCLUSIONS: We have developed a bioinformatics proteome profiling approach that successfully identifies biologically relevant miR regulators from a proteomics dataset of the ATG-7-deficient milieu in lung fibroblasts, and thus may be used to elucidate key molecular players in complex fibrotic pathological processes. The approach is not limited to a specific cell-type and disease, thus highlighting its high relevance in proteome and non-coding RNA research.

Integrative Bioinformatic Analyses of Global Transcriptome Data Decipher Novel Molecular Insights into Cardiac Anti-Fibrotic Therapies.

Integrative bioinformatics is an emerging field in the big data era, offering a steadily increasing number of algorithms and analysis tools. However, for researchers in experimental life sciences it is often difficult to follow and properly apply the bioinformatical methods in order to unravel the complexity and systemic effects of omics data. Here, we present an integrative bioinformatics pipeline to decipher crucial biological insights from global transcriptome profiling data to validate innovative therapeutics. It is available as a web application for an interactive and simplified analysis without the need for programming skills or deep bioinformatics background. The approach was applied to an ex vivo cardiac model treated with natural anti-fibrotic compounds and we obtained new mechanistic insights into their anti-fibrotic action and molecular interplay with miRNAs in cardiac fibrosis. Several gene pathways associated with proliferation, extracellular matrix processes and wound healing were altered, and we could identify micro (mi) RNA-21-5p and miRNA-223-3p as key molecular components related to the anti-fibrotic treatment. Importantly, our pipeline is not restricted to a specific cell type or disease and can be broadly applied to better understand the unprecedented level of complexity in big data research.

(Spectro)electrochemical and Electrocatalytic Investigation of 1,1'-Dithiolatoferrocene-Hexacarbonyldiiron.

Hexacarbonyldiiron bridged by a 1,1'-dithiolatoferrocene, [Fe(C5H4S)2{Fe(CO)3}2] (1), was synthesized, and the electrochemistry showed reversible oxidation at the Fe(C5H4S)2 site and quasi-reversible reduction at the hexacarbonyldiiron site. Spectroelectrochemical techniques showed reduction-induced ligand isomerization, where the thiolate ligand went from bridging to terminal and one carbon monoxide ligand moved to a quasi-bridging position; this mechanism was further supported by cyclic voltammetry simulation and density functional theory calculations. Complex 1 showed electrocatalytic activity toward hydrogen-evolving reaction.

At the Borderline between Metal-Metal Mixed Valency and a Radical Bridge Situation: Four Charge States of a Diruthenium Complex with a Redox-Active Bis( mer-tridentate) Ligand.

The complex ions [L3Ru(µ,η3:η3-BL)RuL3] n+ (1 n+, L3 = 4,4',4″-tri- tert-butyl-2,6,2',6″-terpyridine and H2BL2- = 1,2-bis(salicyloyl)hydrazide(2-)) were isolated with PF6- or ClO4- counterions ( n = 1) and as bis(hexafluorophosphate) ( n = 2). Structural, electrochemical, and spectroscopic characterization reveals the monocation as intermediate ( Kc = 108.2) in the three-step reversible redox system 10/+/2+/3+. The 1+ ion has the molecule-bridged (Ru- - -Ru 4.727 Å) ruthenium centers involved in five- and six-membered chelate rings, and it exhibits long-wavelength absorptions at λmax 2240, 1660, and 1530 nm (εmax = 1000, 3000, and 8000 M-1 cm-1, respectively), which would be compatible with a RuIIIRuII mixed-valent situation or with a coordinated radical ion bridge. In fact, EPR and DFT analysis of 1+ reveals that the spin is equally distributed over the ligand bridge and over both metals. The oxidized paramagnetic ions 12+ and 13+ have been studied by 1H NMR and EPR and by TD-DFT supported UV-vis-NIR and MIR (mid-IR) spectroelectrochemistry. The capacity of various kinds of bis( mer-tridentate) bridging ligands (π donors or π acceptors, cyclometalated or noncyclometalated) for mediating metal-metal interactions is discussed.

MicroRNA-Based Therapy of GATA2-Deficient Vascular Disease.

BACKGROUND: The transcription factor GATA2 orchestrates the expression of many endothelial-specific genes, illustrating its crucial importance for endothelial cell function. The capacity of this transcription factor in orchestrating endothelial-important microRNAs (miRNAs/miR) is unknown. METHODS: Endothelial GATA2 was functionally analyzed in human endothelial cells in vitro. Endogenous short interfering RNA-mediated knockdown and lentiviral-based overexpression were applied to decipher the capacity of GATA2 in regulating cell viability and capillary formation. Next, the GATA2-dependent miR transcriptome was identified by using a profiling approach on the basis of quantitative real-time polymerase chain reaction. Transcriptional control of miR promoters was assessed via chromatin immunoprecipitation, luciferase promoter assays, and bisulfite sequencing analysis of sites in proximity. Selected miRs were modulated in combination with GATA2 to identify signaling pathways at the angiogenic cytokine level via proteome profiler and enzyme-linked immunosorbent assays. Downstream miR targets were identified via bioinformatic target prediction and luciferase reporter gene assays. In vitro findings were translated to a mouse model of carotid injury in an endothelial GATA2 knockout background. Nanoparticle-mediated delivery of proangiogenic miR-126 was tested in the reendothelialization model. RESULTS: GATA2 gain- and loss-of-function experiments in human umbilical vein endothelial cells identified a key role of GATA2 as master regulator of multiple endothelial functions via miRNA-dependent mechanisms. Global miRNAnome-screening identified several GATA2-regulated miRNAs including miR-126 and miR-221. Specifically, proangiogenic miR-126 was regulated by GATA2 transcriptionally and targeted antiangiogenic SPRED1 and FOXO3a contributing to GATA2-mediated formation of normal vascular structures, whereas GATA2 deficiency led to vascular abnormalities. In contrast to GATA2 deficiency, supplementation with miR-126 normalized vascular function and expression profiles of cytokines contributing to proangiogenic paracrine effects. GATA2 silencing resulted in endothelial DNA hypomethylation leading to induced expression of antiangiogenic miR-221 by GATA2-dependent demethylation of a putative CpG island in the miR-221 promoter. Mechanistically, a reverted GATA2 phenotype by endogenous suppression of miR-221 was mediated through direct proangiogenic miR-221 target genes ICAM1 and ETS1. In a mouse model of carotid injury, GATA2 was reduced, and systemic supplementation of miR-126-coupled nanoparticles enhanced miR-126 availability in the carotid artery and improved reendothelialization of injured carotid arteries in vivo. CONCLUSIONS: GATA2-mediated regulation of miR-126 and miR-221 has an important impact on endothelial biology. Hence, modulation of GATA2 and its targets miR-126 and miR-221 is a promising therapeutic strategy for treatment of many vascular diseases.

Podocytes regulate the glomerular basement membrane protein nephronectin by means of miR-378a-3p in glomerular diseases.

The pathophysiology of many proteinuric kidney diseases is poorly understood, and microRNAs (miRs) regulation of these diseases has been largely unexplored. Here, we tested whether miR-378a-3p is a novel regulator of glomerular diseases. MiR-378a-3p has two predicted targets relevant to glomerular function, the glomerular basement membrane matrix component, nephronectin (NPNT), and vascular endothelial growth factor VEGF-A. In zebrafish (Danio rerio), miR-378a-3p mimic injection or npnt knockdown by a morpholino oligomer caused an identical phenotype consisting of edema, proteinuria, podocyte effacement, and widening of the glomerular basement membrane in the lamina rara interna. Zebrafish vegf-A protein could not rescue this phenotype. However, mouse Npnt constructs containing a mutated 3'UTR region prevented the phenotype caused by miR-378a-3p mimic injection. Overexpression of miR-378a-3p in mice confirmed glomerular dysfunction in a mammalian model. Biopsies from patients with focal segmental glomerulosclerosis and membranous nephropathy had increased miR-378a-3p expression and reduced glomerular levels of NPNT. Thus, miR-378a-3p-mediated suppression of the glomerular matrix protein NPNT is a novel mechanism for proteinuria development in active glomerular diseases.