Regulation of Cyr61/CCN1 expression by hypoxia through cooperation of c-Jun/AP-1 and HIF-1α in retinal vascular endothelial cells.

Hypoxia is the most important factor in the pathogenesis of diabetic retinopathy. Cysteine-rich 61 (Cyr61) is one of the angiogenic factors involved in the development of proliferative diabetic retinopathy (PDR). The aim of this study was to investigate the mechanism of hypoxia-induced Cyr61 expression in retinal vascular endothelial cells. The hypoxia-induced expression of mRNA and protein of Cyr61 was studied in monkey choroidal retinal vascular endothelial (RF/6A) cells. Luciferase reporter assays and electrophoretic mobility shift assays were used to identify the hypoxia responsible region and transcription factors in the Cyr61 promoter. Chromatin immunoprecipitation and immunoprecipitation were performed to study the role of hypoxia-inducible factor (HIF)-1α and c-Jun/activator protein-1 (AP-1) in Cyr61 transcriptional regulation. The results showed that hypoxia significantly induced Cyr61 mRNA and protein expression in RF/6A cells. The effect was mediated through phosphorylation of c-Jun. Luciferase assays, electrophoretic mobility shift assays, chromatin immunoprecipitation and immunoprecipitation showed that HIF-1α interacted with c-Jun/AP-1 and their binding on the AP-1 binding motif within the Cyr61 promoter induced the expression of Cyr61. In conclusion, hypoxia controlled the transcriptional regulation of the Cyr61 gene in RF/6A cells by cooperation of HIF-1α and c-Jun/AP-1. Cyr61 might play an important role in ischemic retinal diseases, such as PDR.

You JJ ，Yang CM ，Chen MS ，Yang CH ... -  《-》

Cysteine-rich 61, a member of the CCN family, as a factor involved in the pathogenesis of proliferative diabetic retinopathy.

Cysteine-rich 61 (Cyr61/CCN1) is reported to mediate angiogenesis. In this study, its role in ocular angiogenesis and proliferative diabetic retinopathy (PDR) was investigated. The effects of Cyr61 were evaluated by determining proliferation and chemotaxis and in an assay of capillary tube formation in synthetic matrix by chorioretinal endothelial cells (RF/6A). In the same cells, Cyr61 expression under hypoxic conditions was then investigated. Interactions between Cyr61 and vascular endothelial growth factor (VEGF) were examined using endothelial cell chemotaxis, tube-formation assay, and cross-stimulation assay. A mouse model of oxygen-induced retinopathy (OIR) and a rat model of streptozocin-induced diabetes were used to evaluate Cyr61 expression in the retina. Cyr61 levels were also measured and chemotactic effects were evaluated in vitreous samples from patients with PDR. Cyr61 significantly induced proliferation, migration, and synthetic matrix tube formation of RF/6A cells. Hypoxia significantly induced Cyr61 mRNA and protein expression. Cyr61 induced expression of VEGF and vice versa. Anti-Cyr61 or anti-VEGF could inhibit the effects of both Cyr61 and VEGF. Intravitreal injection of anti-Cyr61 antibody significantly inhibited retinal neovascularization in the mouse OIR model. Cyr61 mRNA and protein were significantly expressed in the retina of streptozocin-induced diabetic rats. Vitreous levels of Cyr61 were elevated in patients with PDR when compared with nondiabetic patients. Cyr61 acts as an angiogenic mediator of ocular neovascularization in vitro and in vivo. It may interact with VEGF in a synergetic manner. Vitreous levels of Cyr61 are elevated in PDR, and it may play an important role in the disease's pathogenesis.

You JJ ，Yang CH ，Chen MS ，Yang CM ... -  《-》

Cysteine-rich 61 (CYR61) is up-regulated in proliferative diabetic retinopathy.

To investigate the role of CYR61 as a retinal angiogenic factor in proliferative diabetic retinopathy (PDR). Effects of CYR61 on RF/6A cell proliferation, migration and angiogenesis were observed by MTT assay, Transwell assay, and tube formation assay. The expression and distribution of CYR61 on retina layers of diabetic mouse were demonstrated by immunohistochemistry. The expression of Cyr61 mRNA in diabetic mouse retina was evaluated by reverse transcription-polymerase chain reaction (RT-PCR). Vitreous CYR61 levels of PDR and non-diabetic patients were measured by enzyme-linked immunosorbent assay (ELISA). Expression and distribution of CYR61 on epiretinal membrane of PDR, proliferative vitreoretinopathy (PVR) and idiopathic epiretinal membrane were evaluated by immunohistochemistry. RF/6A cell proliferation, migration and tube formation capacity increased with increased concentration of CYR61 (p = 0.000). Anti-CYR61 antibody could inhibit cell migration and tube formation promoted by CYR61. In diabetic mouse, CYR61 was expressed in retina layers just as normal mouse, but the staining was stronger than normal in ganglion cell layer and inner plexiform layer. The Cyr61 mRNA expression in retina of diabetic mouse was more than that in normal mouse (p = 0.009). Vitreous CYR61 level was higher in patients with PDR than non-diabetic patients (p = 0.000). PDR patients with plenty of neovasculature on retina and epiretinal membranes had higher level of vitreous CYR61 than patients with little neovasculature (p = 0.001). CYR61 expressed in the cytoplasm of epiretinal membranes in PDR, especially in the wall cells of the tube-like structure. CYR61 are likely to be involved in the pathogenesis of diabetic retinopathy, and may play a role in the course of neovasculation.

Zhang X ，Yu W ，Dong F 《-》

Coordinate activation of HIF-1 and NF-kappaB DNA binding and COX-2 and VEGF expression in retinal cells by hypoxia.

Lukiw WJ ，Ottlecz A ，Lambrou G ，Grueninger M ，Finley J ，Thompson HW ，Bazan NG ... -  《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》

The regulation of hypoxic genes by calcium involves c-Jun/AP-1, which cooperates with hypoxia-inducible factor 1 in response to hypoxia.

Salnikow K ，Kluz T ，Costa M ，Piquemal D ，Demidenko ZN ，Xie K ，Blagosklonny MV ... -  《-》