Jiantao Wang

Inhibition of tumor growth and vasculogenic mimicry by curcumin through down-regulation of the EphA2/PI3K/MMP pathway in a murine choroidal melanoma model.

This study aims to investigate the underlying mechanism by which curcumin inhibits tumor growth and reduces vasculogenic mimicry (VM) in a murine choroidal melanoma model. Sixty mice were given subretinal injection with B16F10 cells and divided into a treatment and a control group. Curcumin was administered to the treatment group once a day at a dose of 100 mg/kg for 18 days starting at d3 (the day of inoculation is designated as d0); an equivalent volume of poloxamer-F68 was administered to the control group. Immunohistochemical and histochemical double staining were ued to detect the different blood supply patterns. The amounts of epithelial cell kinase (EphA2), phosphatidylinositol-3-kinase (PI3K), and matrixmetalloproteinase-2 and -9 (MMP-2, MMP-9) proteins expressed in the tumor tissue were analyzed using immunohistochemical staining; mRNA levels were measured using real-time PCR analysis. Results indicate that the tumor volume is reduced (P=0.000) and that the numbers of VM (P=0.000), mosaic vessels (P=0.031), and endothelium-dependent vessels (P=0.000) are significantly decreased by curcumin (P=0.001). The expression levels of EphA2, PI3K, MMP-2, and -9 are also lower in the treatment group than in the control group (P=0.001); similarly, mRNA levels in the treatment group are lower than those in the control group (P=0.000). In conclusion, curcumin has the ability to inhibit the growth of engrafted melanoma VM channels through the regulation of vasculogenic factors that could be related to the down-regulation of the EphA2/PI3K/MMPs signaling pathway. Thus, curcumin has the potential of being a clinical inhibitor of VM of choroidal melanoma. See commentary: Curcumin: A component of the golden spice, targets multiple angiogenic pathways

Mesenchymal Stem Cells Ameliorate Experimental Autoimmune Uveoretinitis by Comprehensive Modulation of Systemic Autoimmunity

PURPOSE The authors studied the therapeutic effect of rat mesenchymal stem cells (MSCs) on experimental autoimmune uveoretinitis (EAU) induced in rats by peptide 1169-1191 of the interphotoreceptor retinoid-binding protein (IRBP). METHODS The authors intravenously injected syngeneic (isolated from Lewis rats) or allogeneic (isolated from Wistar rats) MSCs into IRBP-induced EAU Lewis rats, either before disease onset (simultaneous with immunization, preventive protocol) or at different time points after disease onset (therapeutic protocol). T-cell response to IRBP 1169-1191 from MSC-treated rats was evaluated, Th1/Th2/Th17 cytokines produced by lymphocytes were measured, and CD4(+)CD25(+) regulatory T cells (Treg) were detected. RESULTS MSC administration before disease onset not only strikingly reduced the severity of EAU, it also delayed the onset of the disease. MSC administration was also effective after disease onset and at the peak of disease, but not after disease stabilization. Clinical efficacy for all treatments was consistent with reduced cellular infiltrates and milder uveal and retinal impairment. T-cell response to IRBP 1169-1191 from MSC-treated rats was inhibited. MSCs significantly decreased the production of IFN-γ and IL-17 and increased the production of IL-10 of T lymphocytes from EAU rats either in vivo or in vitro. Allogeneic and syngeneic MSCs showed a similar immunosuppression potential with regard to clinical effect, T cell proliferation, and cytokine secretion, and MSC therapy upregulated Treg cells. CONCLUSIONS These data suggest that the immunoregulatory properties of MSCs effectively interfere with the autoimmune attack in the course of EAU through the comprehensive modulation of systemic autoimmunity.

MicroRNA-483-3p Inhibits Extracellular Matrix Production by Targeting Smad4 in Human Trabecular Meshwork Cells.

PURPOSE This study investigated the effects of microRNA-483-3p (miR-483-3p) on extracellular matrix (ECM) production, and clarified the regulatory mechanism of microRNA-483-3p in human trabecular meshwork cells (HTMCs) under oxidative stress. METHODS The expression levels of ECM (fibronectin, laminin, collagen I) in HTMCs under oxidative stress were measured by Western blot. Changes of miR-483-3p expression in HTMCs were evaluated by quantitative polymerase chain reaction (qPCR). After using lentivirus stably expressing pri-miR-483, the effects of miR-483-3p on the ECM were assessed by qPCR and Western blot. Smad4, the potential target of miR-483-3p according to mRNA target-predicting algorithms, was confirmed by luciferase assay and Western blot. Furthermore, the effects of Smad4 knockdown on ECM expression were investigated by qPCR and Western blot. RESULTS The mRNA and protein levels of ECM (fibronectin, laminin, collagen I) were upregulated in HTMCs induced by oxidative stress. The expression level of miR-483-3p decreased in HTMCs under oxidative stress, and the ectopic expression of miR-483-3p decreased the levels of ECM. In addition, miR-483-3p targeted Smad4 through two binding sites, resulting in a decrease of Smad4 expression. Furthermore, knockdown of Smad4 reduced the levels of ECM in HTMCs. CONCLUSIONS MicroRNA-483-3p has an inhibitory effect on ECM production in HTMCs through downregulating Smad4, which indicates that miR-483-3p may serve as a potential therapeutic target in glaucoma.

Relationship between the Pathogenesis of Glaucoma and miRNA

Small RNA (microRNA or miRNA) is a kind of small noncoding single-stranded RNA that regulates complementary mRNA at the posttranscriptional level in eukaryotic organisms. As important regulatory factors, miRNAs play an important role in the occurrence and development of glaucoma and widely participate in regulating biological processes of glaucoma-related genes. This article reviews the connection between the aqueous humor, trabecular meshwork, the apoptosis of retinal ganglion cells, and miRNA.

HES1 promotes extracellular matrix protein expression and inhibits proliferation and migration in human trabecular meshwork cells under oxidative stress

Glaucoma is the leading cause of irreversible blindness. The most prevalent form of glaucoma is primary open-angle glaucoma (POAG). Oxidative stress is one of the major pathogenic factors of the POAG, and can elicit molecular and functional changes in trabecular meshwork cells, causing increased aqueous humor outflow resistance and elevated intraocular pressure. However, the regulatory mechanisms underlying oxidative stress-induced cell phenotypic changes remain elusive. Herein, we exposed primary human trabecular meshwork cells to the oxidative stress induced by 300 μM H2O2 for 2 h, and found significantly up-regulated expression of extracellular matrix proteins and a transcription factor, hairy and enhancer of split-1 (HES1). The cell functions, including migration and proliferation, were impaired by the oxidative stress. Furthermore, HES1 shRNA abrogated the extracellular matrix protein up-regulation and rescued the functional defects caused by the oxidative stress; conversely, HES1 overexpression resulted in the molecular and functional changes similar to those induced by H2O2. These results suggest that HES1 promotes extracellular matrix protein expression and inhibits proliferative and migratory functions in the trabecular meshwork cells under oxidative stress, thereby providing a novel pathogenic mechanism underlying and a potential therapeutic target to the POAG.

Video microscope recording of the dynamic course of thrombosis and thrombolysis of the retinal vein in rabbits.

Purpose:The purpose of this study was to investigate the dynamic course of experimental thrombosis and thrombolysis of the retinal veins. Methods:Dynamic changes in the blood flow in the retinal veins were documented on a digital recorder through a microscope-mounted video camera and were analyzed on a monitor by video playback. Photochemical thrombus formation was induced by intravenous injection of 30% Rose Bengal followed by endoillumination of individual branch retinal veins of the eyes of 20 anesthetized pigmented rabbits. Subsequently, 10 rabbits were treated with an infusion in an ear vein of 3 mg/kg recombinant tissue plasminogen activator, whereas 10 control rabbits were administered with similar volumes of normal saline solution. Occlusion and recanalization were confirmed histologically and assessed by video microscopy. Results:At the site exposed to light, photochemical injury resulted in the formation of a white thrombus, stagnation of blood flow, and subsequent complete venous occlusion in 20 rabbits. In this study, of the 10 animals in the recombinant tissue plasminogen activator treatment group, 9 (90%) showed evident thrombolysis, whereas none of the control group animals showed evident thrombolysis. The video showed that the massive thrombus disrupted into nonuniform fragments, which were quickly washed away by the blood flow, and the circulation was reestablished with no recurrence of secondary obstruction. Conclusion:In vivo data suggest that video microscopy may provide a visual approach for observing the dynamic changes occurring during experimental thrombus formation and lysis by the early administration of recombinant tissue plasminogen activator; this approach may assist in future investigation of thrombus research of ocular fundus.

Α-Melanocyte-Stimulating Hormone Protects Early Diabetic Retina from Blood-Retinal Barrier Breakdown and Vascular Leakage via MC4R

Background/Aims: Blood-retinal barrier (BRB) breakdown and vascular leakage is the leading cause of blindness of diabetic retinopathy (DR). Hyperglycemia-induced oxidative stress and inflammation are primary pathogenic factors of this severe DR complication. An effective interventional modality against the pathogenic factors during early DR is needed to curb BRB breakdown and vascular leakage. This study sought to examine the protective effects of α-Melanocyte-stimulating hormone (α-MSH) on early diabetic retina against vascular hyperpermeability, electrophysiological dysfunction, and morphological deterioration in a rat model of diabetes and probe the mechanisms underlying the α-MSH’s anti-hyperpermeability in both rodent retinas and simian retinal vascular endothelial cells (RF6A). Methods: Sprague Dawley rats were injected through tail vein with streptozotocin to induce diabetes. The rats were intravitreally injected with α-MSH or saline at Week 1 and 3 after hyperglycemia. In another 2 weeks, Evans blue assay, transmission electron microscopy, electroretinogram (ERG), and hematoxylin and eosin (H&E) staining were performed to examine the protective effects of α-MSH in diabetic retinas. The expression of pro-inflammatory factors and tight junction at mRNA and protein levels in retinas was analyzed. Finally, the α-MSH’s anti-hyperpermeability was confirmed in a high glucose (HG)-treated RF6A cell monolayer transwell culture by transendothelial electrical resistance (TEER) measurement and a fluorescein isothiocyanate-Dextran assay. Universal or specific melanocortin receptor (MCR) blockers were also employed to elucidate the MCR subtype mediating α-MSH’s protection. Results: Evans blue assay showed that BRB breakdown and vascular leakage was detected, and rescued by α-MSH both qualitatively and quantitatively in early diabetic retinas; electron microscopy revealed substantially improved retinal and choroidal vessel ultrastructures in α-MSH-treated diabetic retinas; scotopic ERG suggested partial rescue of functional defects by α-MSH in diabetic retinas; and H&E staining revealed significantly increased thickness of all layers in α-MSH-treated diabetic retinas. Mechanistically, α-MSH corrected aberrant transcript and protein expression of pro-inflammatory factor and tight junction genes in the diseased retinas; moreover, it prevented abnormal changes in TEER and permeability in HG-stimulated RF6A cells, and this anti-hyperpermeability was abolished by a universal MCR blocker or an antagonist specific to MC4R. Conclusions: This study showed previously undescribed protective effects of α-MSH on inhibiting BRB breakdown and vascular leakage, improving electrophysiological functions and morphology in early diabetic retinas, which may be due to its down-regulating pro-inflammatory factors and augmenting tight junctions. α-MSH acts predominantly on MC4R to antagonize hyperpermeability in retinal microvessel endothelial cells.