Silencing of S100A4, a metastasis-associated protein, inhibits retinal neovascularization via the downregulation of BDNF in oxygen-induced ischaemic retinopathy.

BackgroundTo investigate the underlying mechanism of S100A4 function and whether it has a role in retinal neovascularization (RNV) in a mouse model of oxygen-induced retinopathy (OIR).MethodsRetinas from a mouse model of OIR were treated with and without an intravitreous injection of adenoviral-S100A4-RNAi or adenoviral green fluorescence protein (GFP) at postnatal day 12 (P12). At P17, the efficacy of adenoviral gene transfer was assessed using fluorescence microscopy and western blot analysis. RNV was evaluated by whole-mount immunofluorescence staining of the mouse retina and by counting the number of pre-retinal neovascular cells. Protein and mRNA expression levels of S100A4, brain-derived growth factor (BDNF), and vascular endothelial growth factor (VEGF) were measured using western blot analysis and real-time PCR.ResultsRetinal S100A4 levels were positively correlated with the progression of RNV. In the OIR-S100A4-RNAi group, both protein and mRNA expression levels of S100A4 in the retina significantly decreased at P17 compared with those in the OIR group. Ad-S100A4-RNAi transfer was clearly demonstrated by GFP fluorescence in many layers of the retina 5 days after the Ad-S100A4-RNAi transfer. Whole-mount immunofluorescence staining of the retina and quantification of the pre-retinal neovascular cells demonstrated that RNV was significantly inhibited. Meanwhile, the levels of the transcription and translation of BDNF, VEGF, and hypoxia-inducible factor-1α (HIF-1α) significantly decreased in the OIR-S100A4-RNAi group.ConclusionsAd-S100A4-RNAi transfer ameliorates RNV. The related mechanism may involve silencing S100A4 to decrease the activation of BDNF, which downregulates VEGF expression via HIF-1α. This finding could provide a new therapeutic target for the treatment of ocular neovascularization diseases.

Cheng G ，He T ，Xing Y 《-》

S100A4 gene silencing in oxygen-induced ischemic retinopathy inhibits retinal neovascularization via down-regulation of CREB expression.

Cheng G ，Tian K ，Zhang L ，Yang N ，Xing Y ，He T ... -  《-》

Overexpression of 15-lipoxygenase-1 in oxygen-induced ischemic retinopathy inhibits retinal neovascularization via downregulation of vascular endothelial growth factor-A expression.

15-Lipoxygenase-1 (15-LOX-1) plays an important role in regulating angiogenesis, but the mechanism to date is controversial, even contradictory. The goal of our study was to investigate whether 15-LOX-1 plays a role in inhibiting retinal neovascularization (RNV) in a mouse model of oxygen-induced retinopathy (OIR) and the underlying mechanism. Experiments were performed using retinas from a mouse model of OIR that was treated with and without intravitreous injection of adenoviral-15-lipoxygenase-1 (Ad-15-LOX-1) or adenoviral-green fluorescence protein (Ad-GFP) at postnatal day 12 (P12). At P17, the efficacy of the gene transfer was assessed with immunofluorescence staining. RNV was evaluated with fluorescein angiography on flatmounted retinas and quantified by counting the preretinal neovascular cells. Expression of 15-LOX-1 and vascular endothelial growth factor-A (VEGF-A) were determined with real-time PCR and western blot. RNV during OIR was associated with decreased 15-LOX-1 expression, and retinal 15-LOX-1 levels were negatively correlated with the progression of RNV. In the intravitreous injected Ad-15-LOX-1 mice with OIR, retinal 15-LOX-1 expression was significantly increased at the protein and mRNA levels at P17. 15-LOX-1 expression was clearly demonstrated, primarily in the outer plexiform layer, inner nuclear layer, and ganglion cell layer retinas, five days after gene delivery. Fluorescein retinal angiography and quantification of the preretinal neovascular cells demonstrated that RNV was significantly inhibited. Meanwhile, the expression levels of VEGF-A were significantly decreased at the transcriptional and translational levels. Our results suggest that overexpression of 15-LOX-1 inhibits RNV in a mouse model of OIR via downregulation of VEGF-A expression, and 15-LOX-1 may be a novel therapeutic target for ocular neovascularization diseases.

Li Z ，He T ，Du K ，Xing YQ ，Yan Y ，Chen Z ，Zhang H ，Shen Y ... -  《MOLECULAR VISION》

Silencing of galectin-1 inhibits retinal neovascularization and ameliorates retinal hypoxia in a murine model of oxygen-induced ischemic retinopathy.

Aberrant neovascularization is a consequence of inappropriate angiogenic signaling and contributes to several diseases. Although many regulators of pathogenic angiogenesis have been identified, the understanding of this process remains incomplete. Galectin-1 (Gal-1), as a homodimeric protein with a single carbohydrate-recognition domain, is implicated in several pathologic processes, including angiogenesis; however, its involvement in retinal neovascularization (RNV) remains unknown. Here, we investigated the anti-angiogenic effect of silencing Gal-1 through intravitreal injection in a mouse model of oxygen-induced retinopathy (OIR). Our results revealed that Gal-1 was overexpressed and closely related to retinal neo-vessels in OIR retinas. After silencing Gal-1 via intravitreal injection of adenoviral-Gal-1-RNA interference (Ad-Gal-1-RNAi), RNV and retinal hypoxia were significantly attenuated, indicating the anti-angiogenic effect of Gal-1 inhibition. Western blot analysis and real-time polymerase chain reaction indicated that the expression of both neuropilin-1 (Nrp-1) and B cell lymphoma-2 (Bcl-2) decreased after intravitreal injection of Ad-Gal-1-RNAi, implying the possible involvement of Nrp-1 and Bcl-2 in Gal-1-related angiogenic processes. Additionally, whole-mount fluorescence and hematoxylin and eosin staining showed that intravitreal injection of Ad-Gal-1-RNAi did not significantly disrupt the retinal vasculature and neuronal structure of room air mice. Moreover, Ad-Gal-1-RNAi transfer promoted retinal vascular sprouting and increased retinal vascular perfusion, likely through decreased phosphorylation of myosin phosphatase target protein-1. Collectively, our results demonstrated that Gal-1 functions as an important regulator in RNV and offers a promising strategy for the treatment of RNV diseases, such as proliferative diabetic retinopathy and retinopathy of prematurity.

Yang N ，Zhang W ，He T ，Xing Y ... -  《-》

Celecoxib attenuates retinal angiogenesis in a mouse model of oxygen-induced retinopathy.

This study aimed to investigate the anti-angiogenic effects of Celecoxib on the expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible transcription factor 1α (HIF-1α) in a mouse model for oxygen-induced retinopathy (OIR). The OIR mice were exposed to 75% oxygen from postnatal day 7 (P7) to P12, after which the mice were randomly assigned to two groups (Celecoxib and vehicle) and were brought to room air for additional five days. Celecoxib or vehicle was administered from P12 to P17. Age-matched mice maintained in room air from birth to P17 were administered vehicle from P12 to P17 (RA group). Blood vessel profiles in the retina were used to count by histologic methods. Retina protein and mRNA of VEGF and HIF-1α were assessed by immunohistochemistry, western-blot and RT-PCR. Compared with the RA group, the OIR mice exhibited over-expression in VEGF and HIF-1α mRNA and protein. In addition, they had a positive and spatial correlation. Celecoxib- treated OIR mice reduced the retinal neovascular tufts and the levels of VEGF and HIF-1α. These data suggest that Celecoxib inhibits retinal pathogenic angiogenesis through down-regulating HIF-1α expression which suppressing VEGF transcription. Celecoxib could potentially serve as a portent pharmaceutical agent to inhibit retinal angiogenesis.

Liu N ，Chen L ，Cai N 《International Journal of Clinical and Experimental Pathology》