Single-cell transcriptomics identifies aberrant glomerular angiogenic signalling in the early stages of WT1 kidney disease.

WT1 encodes a podocyte transcription factor whose variants can cause an untreatable glomerular disease in early childhood. Although WT1 regulates many podocyte genes, it is poorly understood which of them are initiators in disease and how they subsequently influence other cell-types in the glomerulus. We hypothesised that this could be resolved using single-cell RNA sequencing (scRNA-seq) and ligand-receptor analysis to profile glomerular cell-cell communication during the early stages of disease in mice harbouring an orthologous human mutation in WT1 (Wt1).

Novel Approach to Measure Transepithelial Electrical Resistance in Intestinal Cells.

The technique electric cell-substrate impedance sensing (ECIS) can be used to detect and monitor the behavior of intestinal cells. The methodology presented was designed to achieve results within a short time frame, and it was tailored to use a colonic cancer cell line. Differentiation of intestinal cancer cells has previously been reported to be regulated by retinoic acid (RA).

Performance evaluation of a non-invasive one-step multiplex RT-qPCR assay for detection of SARS-CoV-2 direct from saliva.

Polymerase chain reaction (PCR) has proven to be the gold-standard for SARS-CoV-2 detection in clinical settings. The most common approaches rely on nasopharyngeal specimens obtained from swabs, followed by RNA extraction, reverse transcription and quantitative PCR. Although swab-based PCR is sensitive, swabbing is invasive and unpleasant to administer, reducing patient compliance for regular testing and resulting in an increased risk of improper sampling.

Direct RT-qPCR Assay for the Detection of SARS-CoV-2 in Saliva Samples.

Since mid-2020 there have been complexities and difficulties in the standardisation and administration of nasopharyngeal swabs. Coupled with the variable and/or poor accuracy of lateral flow devices, this has led to increased societal 'testing fatigue' and reduced confidence in test results. Consequently, asymptomatic individuals have developed reluctance towards repeat testing, which remains the best way to monitor COVID-19 cases in the wider population.

Endothelial Cell RNA-Seq Data: Differential Expression and Functional Enrichment Analyses to Study Phenotypic Switching.

RNA-seq is a common approach used to explore gene expression data between experimental conditions or cell types and ultimately leads to information that can shed light on the biological processes involved and inform further hypotheses. While the protocols required to generate samples for sequencing can be performed in most research facilities, the resulting computational analysis is often an area in which researchers have little experience. Here we present a user-friendly bioinformatics workflow which describes the methods required to take raw data produced by RNA sequencing to interpretable results.

Simple Gene Knockdown in Endothelial Cells Using Short Interfering RNA Oligonucleotides.

Cell transfection using short interfering RNAs (siRNAs) is a widely used technique to perform loss of function studies by "knocking down" genes of interest. Oftentimes, primary cells can be difficult to transfect, but here we provide a simple and robust method using cultured endothelial cells and routine transfection reagents. Knockdown studies can be used to complement overexpression studies and validate biochemical pathway analysis, as well as functional assays.

Quantification of Angiogenesis in Laser Choroidal Neovascularization.

Murine laser-induced laser choroidal neovascularization is a widely used and robust model of wet (exudative) age-related macular degeneration (wAMD). wAMD is one of the leading causes of blindness in the Western world. In brief, a focused laser beam is used to penetrate Bruch's membrane, which separates the choriocapillaris (well-vascularized choroid layer) from the pigmented layers of the retina.

Transcription Factor Control of Lymphatic Quiescence and Maturation of Lymphatic Neovessels in Development and Physiology.

The lymphatic system is a vascular system comprising modified lymphatic endothelial cells, lymph nodes and other lymphoid organs. The system has diverse, but critical functions in both physiology and pathology, and forms an interface between the blood vascular and immune system. It is increasingly evident that remodelling of the lymphatic system occurs alongside remodelling of the blood microvascular system, which is now considered a hallmark of most pathological conditions as well as being critical for normal development.

Hydrogen Sulfide Is a Novel Protector of the Retinal Glycocalyx and Endothelial Permeability Barrier.

Significantly reduced levels of the anti-inflammatory gaseous transmitter hydrogen sulfide (HS) are observed in diabetic patients and correlate with microvascular dysfunction. HS may protect the microvasculature by preventing loss of the endothelial glycocalyx. We tested the hypothesis that HS could prevent or treat retinal microvascular endothelial dysfunction in diabetes.

METTL3 Regulates Angiogenesis by Modulating let-7e-5p and miRNA-18a-5p Expression in Endothelial Cells.

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Bioinspired artificial exosomes based on lipid nanoparticles carrying let-7b-5p promote angiogenesis in vitro and in vivo.

MicroRNAs (miRNAs) regulate gene expression by post-transcriptional inhibition of target genes. Proangiogenic small extracellular vesicles (sEVs; popularly identified with the name "exosomes") with a composite cargo of miRNAs are secreted by cultured stem cells and present in human biological fluids. Lipid nanoparticles (LNPs) represent an advanced platform for clinically approved delivery of RNA therapeutics.

Comparative effects of viral-transport-medium heat inactivation upon downstream SARS-CoV-2 detection in patient samples.

The COVID-19 pandemic, which began in 2020 is testing economic resilience and surge capacity of healthcare providers worldwide. At the time of writing, positive detection of the SARS-CoV-2 virus remains the only method for diagnosing COVID-19 infection. Rapid upscaling of national SARS-CoV-2 genome testing presented challenges: (1) Unpredictable supply chains of reagents and kits for virus inactivation, RNA extraction and PCR-detection of viral genomes.

Phenotypic and functional translation of IL33 genetics in asthma.

Background: Asthma is a complex disease with multiple phenotypes that may differ in disease pathobiology and treatment response. IL33 single nucleotide polymorphisms (SNPs) have been reproducibly associated with asthma. IL33 levels are elevated in sputum and bronchial biopsies of patients with asthma.

Blocking endothelial apoptosis revascularizes the retina in a model of ischemic retinopathy.

Aberrant, neovascular retinal blood vessel growth is a vision-threatening complication in ischemic retinal diseases. It is driven by retinal hypoxia frequently caused by capillary nonperfusion and endothelial cell (EC) loss. We investigated the role of EC apoptosis in this process using a mouse model of ischemic retinopathy, in which vessel closure and EC apoptosis cause capillary regression and retinal ischemia followed by neovascularization.

Enhanced notch signaling modulates unproductive revascularization in response to nitric oxide-angiopoietin signaling in a mouse model of peripheral ischemia.

Introduction: Arteriolargenesis can be induced by concomitant stimulation of nitric Oxide (NO)-Angiopoietin receptor (Tie)-Vascular Endothelial Growth Factor (VEGF) signaling in the rat mesentery angiogenesis assay. We hypothesized that the same combination of exogenously added growth factors would also have a positive impact on arteriolargenesis and, consequently, the recovery of blood flow in a model of unilateral hindlimb ischemia.

Results And Methods: NO-Tie mice had faster blood flow recovery compared to control mice, as assessed by laser speckle imaging.

Activation of Notch signaling by soluble Dll4 decreases vascular permeability via a cAMP/PKA-dependent pathway.

The Notch ligand delta-like ligand 4 (Dll4), upregulated by VEGF, is a key regulator of vessel morphogenesis and function, controlling tip and stalk cell selection during sprouting angiogenesis. Inhibition of Dll4 results in hypersprouting, nonfunctional, poorly perfused vessels, suggesting a role for Dll4 in the formation of mature, reactive, functional vessels, with low permeability and able to restrict fluid and solute exchange. We tested the hypothesis that Dll4 controls transvascular fluid exchange.

BDNF (Brain-Derived Neurotrophic Factor) Promotes Embryonic Stem Cells Differentiation to Endothelial Cells Via a Molecular Pathway, Including MicroRNA-214, EZH2 (Enhancer of Zeste Homolog 2), and eNOS (Endothelial Nitric Oxide Synthase).

Objective- The NTs (neurotrophins), BDNF (brain-derived neurotrophic factor) and NT-3 promote vascular development and angiogenesis. This study investigated the contribution of endogenous NTs in embryonic stem cell (ESC) vascular differentiation and the potential of exogenous BDNF to improve the process of ESC differentiation to endothelial cells (ECs). Approach and Results- Mouse ESCs were differentiated into vascular cells using a 2-dimensional embryoid body (EB) model.

Physiological Role of Vascular Endothelial Growth Factors as Homeostatic Regulators.

The vascular endothelial growth factor (VEGF) family of proteins are key regulators of physiological systems. Originally linked with endothelial function, they have since become understood to be principal regulators of multiple tissues, both through their actions on vascular cells, but also through direct actions on other tissue types, including epithelial cells, neurons, and the immune system. The complexity of the five members of the gene family in terms of their different splice isoforms, differential translation, and specific localizations have enabled tissues to use these potent signaling molecules to control how they function to maintain their environment.

VEGFR2 promotes central endothelial activation and the spread of pain in inflammatory arthritis.

Chronic pain can develop in response to conditions such as inflammatory arthritis. The central mechanisms underlying the development and maintenance of chronic pain in humans are not well elucidated although there is evidence for a role of microglia and astrocytes. However in pre-clinical models of pain, including models of inflammatory arthritis, there is a wealth of evidence indicating roles for pathological glial reactivity within the CNS.

Can the co-dependence of the immune system and angiogenesis facilitate pharmacological targeting of tumours?

Tumours elicit a number of mechanisms to induce a reprogramming of innate and adaptive immune cells to their advantage, inducing a pro-angiogenic phenotype. Investigation of these events is now leading to the identification of specific myeloid and lymphoid cell-targeted therapies, as well as of unexplored off-target activities of clinically relevant chemotherapeutic and metabolic drugs. It is also leading to an enhanced understanding of the interplay between angiogenesis and the immune system, and the value of novel co-targeting approaches using both immunotherapy and anti-angiogenic therapy.

Measurement of Angiogenesis, Arteriolargenesis, and Lymphangiogenesis Phenotypes by Use of Two-Dimensional Mesenteric Angiogenesis Assay.

Successful therapeutic angiogenesis requires an understanding of how the myriad interactions of growth factors released during angiogenesis combine to form a mature vascular bed. This requires a model in which multiple physiological and cell biological parameters can be identified. The adenoviral-mediated mesenteric angiogenesis assay as described here is ideal for that purpose.

The Orphan Receptor Tie1 Controls Angiogenesis and Vascular Remodeling by Differentially Regulating Tie2 in Tip and Stalk Cells.

Tie1 is a mechanistically poorly characterized endothelial cell (EC)-specific orphan receptor. Yet, Tie1 deletion is embryonic lethal and Tie1 has been implicated in critical vascular pathologies, including atherosclerosis and tumor angiogenesis. Here, we show that Tie1 does not function independently but exerts context-dependent effects on the related receptor Tie2.

Licensed human natural killer cells aid dendritic cell maturation via TNFSF14/LIGHT.

Interactions between natural killer (NK) cells and dendritic cells (DCs) aid DC maturation and promote T-cell responses. Here, we have analyzed the response of human NK cells to tumor cells, and we identify a pathway by which NK-DC interactions occur. Gene expression profiling of tumor-responsive NK cells identified the very rapid induction of TNF superfamily member 14 [TNFSF14; also known as homologous to lymphotoxins, exhibits inducible expression, and competes with HSV glycoprotein D for HVEM, a receptor expressed by T lymphocytes (LIGHT)], a cytokine implicated in the enhancement of antitumor responses.