Liu Li, Jiang Youde, Steinle Jena J
Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University School of Medicine, Detroit, MI 48201, USA.
J Clin Med. 2022 Mar 30;11(7):1915. doi: 10.3390/jcm11071915.
Diabetic retinopathy is associated with increased inflammatory mediator levels. In these studies, we focused on prohibitin 1. We performed western blotting for retinal lysates from diabetic mice and Epac1 floxed and cdh5Cre-Epac1 mice. We also grew primary retinal endothelial cells (REC) in normal (5 mM) and high (25 mM) glucose, and treated some cells with an Epac 1 agonist or prohibitin 1 siRNA. Western blotting was done to confirm knockdown of prohibitin 1 and Epac 1 agonism. We measured the tumor necrosis factor alpha (TNFα), interleukin-1-beta (IL-1β), phosphorylated prohibitin 1, phosphorylated nuclear factor kappa beta (NFkB), high mobility group box 1 (HMGB1) and reactive oxygen species (ROS) levels in REC after transfection with prohibitin 1 siRNA. Results showed that high glucose increased the inflammatory mediators, as well as HMGB1 and ROS. The levels of ROS, HMGB1, and inflammatory pathways were all reduced after cells were transfected with prohibitin 1 siRNA. Epac1 reduced prohibitin 1 phosphorylation. In conclusion, decreased prohibitin 1 significantly reduced the inflammatory mediator and ROS levels in REC. Epac1 regulates the prohibitin 1 levels in REC.
糖尿病性视网膜病变与炎症介质水平升高有关。在这些研究中，我们聚焦于抑制素1。我们对糖尿病小鼠以及Epac1基因敲除和cdh5Cre-Epac1小鼠的视网膜裂解物进行了蛋白质免疫印迹分析。我们还将原代视网膜内皮细胞（REC）分别培养在正常（5 mM）和高糖（25 mM）环境中，并对一些细胞用Epac 1激动剂或抑制素1小干扰RNA（siRNA）进行处理。通过蛋白质免疫印迹分析来确认抑制素1的敲低和Epac 1的激动作用。在用抑制素1 siRNA转染REC后，我们检测了肿瘤坏死因子α（TNFα）、白细胞介素-1β（IL-1β）、磷酸化抑制素1、磷酸化核因子κB（NFkB）、高迁移率族蛋白B1（HMGB1）和活性氧（ROS）的水平。结果显示，高糖增加了炎症介质以及HMGB1和ROS的水平。在用抑制素1 siRNA转染细胞后，ROS、HMGB1和炎症信号通路的水平均降低。Epac1降低了抑制素1的磷酸化水平。总之，抑制素1水平降低显著降低了REC中的炎症介质和ROS水平。Epac1调节REC中的抑制素1水平。
