School of Pharmacy, Faculty of Health Sciences, University of Eastern Finland, 70210, Kuopio, Finland.
Department of Ophthalmology, Institute of Clinical Medicine, University of Eastern Finland, 70210, Kuopio, Finland.
Immunol Res. 2022 Oct;70(5):678-687. doi: 10.1007/s12026-022-09300-0. Epub 2022 Jun 4.
In addition to hypoxia, inflammation is capable of inducing vascular endothelial growth factor (VEGF) expression in human retinal pigment epithelial (RPE) cells. Excessive levels of VEGF promote choroidal neovascularization and thereby contribute to the pathogenesis of wet age-related macular degeneration (AMD). Intravitreal anti-VEGF injections ameliorate pathological vessel neoformation in wet AMD but excessive dampening of VEGF can result in a degeneration of the RPE. In the present study, we induced VEGF production by exposing human ARPE-19 cells to the pro-inflammatory IL-1α and subsequently to hydroquinone, a component of tobacco smoke that is a major environmental risk factor for AMD. Effects were monitored by measuring the levels of VEGF and anti-angiogenic pigment epithelium-derived factor (PEDF) using an enzyme-linked immunosorbent assay (ELISA) technique. In addition, we measured the production of reactive oxygen species (ROS) using the 2',7'-dichlorofluorescin diacetate (H2DCFDA) probe and studied the effects of two anti-oxidants, ammonium pyrrolidinedithiocarbamate (APDC) and N-acetyl-cysteine (NAC), on VEGF production. Cellular and secreted VEGF as well as secreted PEDF levels were reduced at all tested hydroquinone concentrations (10, 50, or 200 µM); these effects were evident prior to any reduction of cell viability evoked by hydroquinone. Cell viability was carefully explored in our previous study and verified by microscoping in the present study. APDC further reduced the VEGF levels, whereas NAC increased them. The 50 μM concentration of hydroquinone increased ROS production in ARPE-19 cells primed with IL-1α. Hydroquinone disturbs the regulatory balance of VEGF and PEDF in inflammatory conditions. These data support the idea that hydroquinone mediates RPE degeneration by reducing VEGF levels and may predispose to dry AMD since VEGF is as well important for retinal integrity.
除了缺氧，炎症也能够诱导人视网膜色素上皮（RPE）细胞中血管内皮生长因子（VEGF）的表达。过量的 VEGF 促进脉络膜新生血管形成，从而导致湿性年龄相关性黄斑变性（AMD）的发病机制。玻璃体内抗 VEGF 注射可改善湿性 AMD 中的病理性血管新生，但 VEGF 的过度抑制可导致 RPE 退化。在本研究中，我们通过用促炎细胞因子白细胞介素 1α（IL-1α）处理人 ARPE-19 细胞，然后再用烟草烟雾的成分对其进行处理，从而诱导 VEGF 的产生，该成分是 AMD 的主要环境风险因素。通过酶联免疫吸附试验（ELISA）技术测量 VEGF 和抗血管生成的色素上皮衍生因子（PEDF）的水平来监测效果。此外，我们使用 2'，7'-二氯荧光素二乙酸酯（H2DCFDA）探针测量活性氧（ROS）的产生，并研究了两种抗氧化剂，吡咯烷二硫代氨基甲酸盐（APDC）和 N-乙酰半胱氨酸（NAC）对 VEGF 产生的影响。在所有测试的氢醌浓度（10、50 或 200 μM）下，细胞和分泌的 VEGF 以及分泌的 PEDF 水平均降低；这些作用在氢醌引起的细胞活力下降之前就已经显现。在我们之前的研究中，我们仔细研究了细胞活力，并在本研究中通过显微镜检查进行了验证。APDC 进一步降低了 VEGF 水平，而 NAC 则增加了它们的水平。IL-1α 预处理的 ARPE-19 细胞中，50 μM 浓度的氢醌增加了 ROS 的产生。在炎症条件下，氢醌扰乱了 VEGF 和 PEDF 的调节平衡。这些数据支持这样一种观点，即氢醌通过降低 VEGF 水平来介导 RPE 退化，并可能导致干性 AMD，因为 VEGF 对视网膜完整性也很重要。
