Influence of Plasminogen Activator Inhibitor Type 1 on Choroidal Neovascularization

peer reviewedHigh levels of the plasminogen activators, but also their inhibitor, plasminogen activator inhibitor 1 (PAI-1), have been documented in neovascularization of severe ocular pathologies such as diabetic retinopathy or age-related macular degeneration (AMD). AMD is the primary cause of irreversible photoreceptors loss, and current therapies are limited. PAI-1 has recently been shown to be essential for tumoral angiogenesis. We report here that deficient PAI-1 expression in mice prevented the development of subretinal choroidal angiogenesis induced by laser photocoagulation. When systemic and local PAI-1 expression was achieved by intravenous injection of a replication-defective adenoviral vector expressing human PAI-1 cDNA, the wild-type pattern of choroidal angiogenesis was restored. These observations demonstrate the proangiogenic activity of PAI-1 not only in tumoral models, but also in choroidal experimental neovascularization sharing similarities with human AMD. They identify therefore PAI-1 as a potential target for therapeutic ocular anti-angiogenic strategies

Estrogens reduce the expression of YKL-40 in the retina: Implications for eye and joint diseases

PURPOSE. To identify modifications in the gene expression profile of the ocular posterior segment in ovariectomized (OVX) mice with and without substitutive estradiol therapy and to select differentially expressed genes that could be relevant to the natural history of human age-related macular degeneration (AMD). METHODS. Chorioretinal tissues from two groups of 25 treated and untreated OVX mice were analyzed by using cDNA array technology. The expression level of selected genes was confirmed in triplicate by RT-PCR and related to the estrogenic status of the animals. Expression of the YKL-40 gene was further investigated in intact or diseased human retinas and in a murine model of experimental choroidal neovascularization (CNV), using laser pressure catapulting. RESULTS. Of the approximately, 10,000 genes screened, only YKL-40 expression was significantly downregulated by 17-beta-estradiol. YKL-40 was expressed in intact human neural retina and in the RPE. The expression of YKL-40 was upregulated in experimental CNV and in neovascular membranes extracted from patients affected by the exudative form of AMD. CONCLUSIONS. These observations indicate that YKL-40 expression in the retina is modulated by serum levels of estradiol. This protein could be relevant to the development of AMD and is also a new mediator to take into account when evaluating the broad consequences of hormonal replacement therapy

Heterogeneous RNA editing and influence of ADAR2 on mesothelioma chemoresistance and the tumor microenvironment

We previously observed increased levels of adenosine-deaminase-acting-on-dsRNA (Adar)-dependent RNA editing during mesothelioma development in mice exposed to asbestos. The aim of this study was to characterize and assess the role of ADAR-dependent RNA editing in mesothelioma. We found that tumors and mesothelioma primary cultures have higher ADAR-mediated RNA editing compared to mesothelial cells. Unsupervised clustering of editing in different genomic regions revealed heterogeneity between tumor samples as well as mesothelioma primary cultures. ADAR2 expression levels are higher in BRCA1-associated protein 1 wild-type tumors, with corresponding changes in RNA editing in transcripts and 3'UTR. ADAR2 knockdown and rescue models indicated a role in cell proliferation, altered cell cycle, increased sensitivity to antifolate treatment, and type-1 interferon signaling upregulation, leading to changes in the microenvironment in vivo. Our data indicate that RNA editing contributes to mesothelioma heterogeneity and highlights an important role of ADAR2 not only in growth regulation in mesothelioma but also in chemotherapy response, in addition to regulating inflammatory response downstream of sensing nucleic acid structures

New insights into the classification and nomenclature of cortical GABAergic interneurons.

A systematic classification and accepted nomenclature of neuron types is much needed but is currently lacking. This article describes a possible taxonomical solution for classifying GABAergic interneurons of the cerebral cortex based on a novel, web-based interactive system that allows experts to classify neurons with pre-determined criteria. Using Bayesian analysis and clustering algorithms on the resulting data, we investigated the suitability of several anatomical terms and neuron names for cortical GABAergic interneurons. Moreover, we show that supervised classification models could automatically categorize interneurons in agreement with experts' assignments. These results demonstrate a practical and objective approach to the naming, characterization and classification of neurons based on community consensus

Combined analysis of single cell RNA-Seq and ATAC-Seq data reveals putative regulatory toggles operating in native and iPS-derived retina.

We report the generation and analysis of single-cell RNA-Seq data (> 38,000 cells) from native and iPSC-derived murine retina at four matched developmental stages spanning the emergence of the major retinal cell types. We combine information from temporal sampling, visualization of 3D UMAP manifolds, pseudo-time and RNA velocity analyses, to show that iPSC-derived 3D retinal aggregates broadly recapitulate the native developmental trajectories. However, we show relaxation of spatial and temporal transcriptome control, premature emergence and dominance of photoreceptor precursor cells, and susceptibility of dynamically regulated pathways and transcription factors to culture conditions in iPSC-derived retina. We generate bulk ATAC-Seq data for native and iPSC-derived murine retina identifying ~125,000 peaks. We combine single-cell RNA-Seq with ATAC-Seq information and obtain evidence that approximately half the transcription factors that are dynamically regulated during retinal development may act as repressors rather than activators. We propose that sets of activators and repressors with cell-type specific expression constitute regulatory toggles that lock cells in distinct transcriptome states underlying differentiation. We provide evidence supporting our hypothesis from the analysis of publicly available single-cell ATAC-Seq data for adult mouse retina. We identify subtle but noteworthy differences in the operation of such toggles between native and iPSC-derived retina particularly for the Etv1, Etv5, Hes1 and Zbtb7a group of transcription factors

Serotonin 3A Receptor Subtype as an Early and Protracted Marker of Cortical Interneuron Subpopulations

To identify neocortical neurons expressing the type 3 serotonergic receptor, here we used transgenic mice expressing the enhanced green fluorescent protein (GFP) under the control of the 5-HT3A promoter (5-HT3A:GFP mice). By means of whole-cell patch-clamp recordings, biocytin labeling, and single-cell reversed-transcriptase polymerase chain reaction on acute brain slices of 5-HT3A:GFP mice, we identified 2 populations of 5-HT3A-expressing interneurons within the somatosensory cortex. The first population was characterized by the frequent expression of the vasoactive intestinal peptide and a typical bipolar/bitufted morphology, whereas the second population expressed predominantly the neuropeptide Y and exhibited more complex dendritic arborizations. Most interneurons of this second group appeared very similar to neurogliaform cells according to their electrophysiological, molecular, and morphological properties. The combination of 5-bromo-2-deoxyuridine injections with 5-HT3A mRNA detection showed that cortical 5-HT3A interneurons are generated around embryonic day 14.5. Although at this stage the 5-HT3A receptor subunit is expressed in both the caudal ganglionic eminence and the entopeduncular area, homochronic in utero grafts experiments revealed that cortical 5-HT3A interneurons are mainly generated in the caudal ganglionic eminence. This protracted expression of the 5-HT3A subunit allowed us to study specific cortical interneuron populations from their birth to their final functional phenotype

MicroRNA-21 Exhibits Antiangiogenic Function by Targeting RhoB Expression in Endothelial Cells

BACKGROUND: MicroRNAs (miRNAs) are endogenously expressed small non-coding RNAs that regulate gene expression at post-transcriptional level. The recent discovery of the involvement of these RNAs in the control of angiogenesis renders them very attractive in the development of new approaches for restoring the angiogenic balance. Whereas miRNA-21 has been demonstrated to be highly expressed in endothelial cells, the potential function of this miRNA in angiogenesis has never been investigated. METHODOLOGY/PRINCIPAL FINDINGS: We first observed in endothelial cells a negative regulation of miR-21 expression by serum and bFGF, two pro-angiogenic factors. Then using in vitro angiogenic assays, we observed that miR-21 acts as a negative modulator of angiogenesis. miR-21 overexpression reduced endothelial cell proliferation, migration and the ability of these cells to form tubes whereas miR-21 inhibition using a LNA-anti-miR led to opposite effects. Expression of miR-21 in endothelial cells also led to a reduction in the organization of actin into stress fibers, which may explain the decrease in cell migration. Further mechanistic studies showed that miR-21 targets RhoB, as revealed by a decrease in RhoB expression and activity in miR-21 overexpressing cells. RhoB silencing impairs endothelial cell migration and tubulogenesis, thus providing a possible mechanism for miR-21 to inhibit angiogenesis. Finally, the therapeutic potential of miR-21 as an angiogenesis inhibitor was demonstrated in vivo in a mouse model of choroidal neovascularization. CONCLUSIONS/SIGNIFICANCE: Our results identify miR-21 as a new angiogenesis inhibitor and suggest that inhibition of cell migration and tubulogenesis is mediated through repression of RhoB

Placental Growth Factor Contributes to Micro-Vascular Abnormalization and Blood-Retinal Barrier Breakdown in Diabetic Retinopathy

OBJECTIVE: There are controversies regarding the pro-angiogenic activity of placental growth factor (PGF) in diabetic retinopathy (DR). For a better understanding of its role on the retina, we have evaluated the effect of a sustained PGF over-expression in rat ocular media, using ciliary muscle electrotransfer (ET) of a plasmid encoding rat PGF-1 (pVAX2-rPGF-1). MATERIALS AND METHODS: pVAX2-rPGF-1 ET in the ciliary muscle (200 V/cm) was achieved in non diabetic and diabetic rat eyes. Control eyes received saline or naked plasmid ET. Clinical follow up was carried out over three months using slit lamp examination and fluorescein angiography. After the control of rPGF-1 expression, PGF-induced effects on retinal vasculature and on the blood-external barrier were evaluated respectively by lectin and occludin staining on flat-mounts. Ocular structures were visualized through histological analysis. RESULTS: After fifteen days of rPGF-1 over-expression in normal eyes, tortuous and dilated capillaries were observed. At one month, microaneurysms and moderate vascular sprouts were detected in mid retinal periphery in vivo and on retinal flat-mounts. At later stages, retinal pigmented epithelial cells demonstrated morphological abnormalities and junction ruptures. In diabetic retinas, PGF expression rose between 2 and 5 months, and, one month after ET, rPGF-1 over-expression induced glial activation and proliferation. CONCLUSION: This is the first demonstration that sustained intraocular PGF production induces vascular and retinal changes similar to those observed in the early stages of diabetic retinopathy. PGF and its receptor Flt-1 may therefore be looked upon as a potential regulatory target at this stage of the disease