Departments of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, Tennessee, United States.
Ophthalmology and Visual Sciences, Vanderbilt University School of Medicine, Nashville, Tennessee, United States.
Invest Ophthalmol Vis Sci. 2014 Jun 10;55(7):4253-60. doi: 10.1167/iovs.14-14216.
The purpose of this study was to investigate the role(s) of cytochrome P450 epoxygenases (CYPs) and their products, the epoxyeicosatrienoic acids (EETs), in hypoxia-induced VEGF production and pathologic retinal angiogenesis.
Human retinal astrocytes, Müller cells, and retinal microvascular endothelial cells (HRMEC) were exposed to hypoxia, and relative CYP2C expression was measured by RT-PCR. Astrocyte and Müller cell VEGF production was measured by ELISA after exposure to hypoxia and treatment with the general CYP inhibitor, SKF-525a. Human retinal microvascular endothelial cells were treated with the CYP product, 11,12-epoxyeicosatrienoic acid [EET], or SKF-525a in the presence or absence of VEGF. Proliferation of HRMEC and tube formation were assayed. Oxygen-induced retinopathy (OIR) was induced in newborn rats. Retinal CYP2C11 and CYP2C23 expression were measured by RT-PCR. The OIR rats received SKF-525a by intravitreal injection and preretinal neovascularization (NV) was quantified. Retinal VEGF protein levels were measured by ELISA.
Human retinal astrocytes were the only cells to exhibit significant induction of CYP2C8 and CYP2C9 mRNA expression by hypoxia. Astrocytes, but not Müller cells, exhibited reduced hypoxia-induced VEGF production when treated with SKF-525a. 11,12-EET induced HRMEC proliferation and tube formation, and SKF-525a inhibited VEGF-induced proliferation. Oxygen-induced retinopathy induced expression of CYP2C23, but had no effect on CYP2C11. SKF-525a inhibited retinal NV and reduced retinal VEGF levels in OIR rats.
The CYP-derived 11,12-EET may exhibit a proangiogenic biological function in the retina following stimulation by hypoxia in astrocytes. Inhibition of CYP may provide a rational therapy against retinal NV, because it can reduce VEGF production and VEGF-induced angiogenic responses in endothelial cells.
本研究旨在探讨细胞色素 P450 加单氧酶（CYPs）及其产物环氧二十碳三烯酸（EETs）在缺氧诱导的 VEGF 产生和病理性视网膜血管生成中的作用。
将人视网膜星形胶质细胞、Müller 细胞和视网膜微血管内皮细胞（HRMEC）暴露于缺氧环境中，并通过 RT-PCR 测量相对 CYP2C 表达。在缺氧和用通用 CYP 抑制剂 SKF-525a 处理后，通过 ELISA 测量星形胶质细胞和 Müller 细胞的 VEGF 产生。用 CYP 产物 11,12-环氧二十碳三烯酸[EET]或 SKF-525a 处理 HRMEC，并在存在或不存在 VEGF 的情况下检测细胞增殖和管形成。在新生大鼠中诱导氧诱导的视网膜病变（OIR）。通过 RT-PCR 测量视网膜 CYP2C11 和 CYP2C23 的表达。OIR 大鼠通过玻璃体内注射 SKF-525a，并定量视网膜前新生血管（NV）。通过 ELISA 测量视网膜 VEGF 蛋白水平。
只有人视网膜星形胶质细胞在缺氧刺激下表现出 CYP2C8 和 CYP2C9 mRNA 表达的显著诱导。用 SKF-525a 处理时，星形胶质细胞而非 Müller 细胞表现出缺氧诱导的 VEGF 产生减少。11,12-EET 诱导 HRMEC 增殖和管形成，而 SKF-525a 抑制 VEGF 诱导的增殖。OIR 诱导 CYP2C23 的表达，但对 CYP2C11 没有影响。SKF-525a 抑制 OIR 大鼠的视网膜 NV 并降低视网膜 VEGF 水平。
CYP 衍生的 11,12-EET 在星形胶质细胞受到缺氧刺激后可能在视网膜中表现出促血管生成的生物学功能。抑制 CYP 可能为治疗视网膜 NV 提供合理的治疗方法，因为它可以减少 VEGF 产生和内皮细胞中 VEGF 诱导的血管生成反应。
