All-trans-retinoic acid generation is an antidotal clearance pathway for all-trans-retinal in the retina  

全反式维甲酸的生成是视网膜中全反式视黄醛的一种解毒代谢途径（英文）

作　　者：Qing-qing XIA Ling-min ZHANG Ying-ying ZHOU Ya-lin WU Jie LI 

机构地区：[1]Central Laboratory,Department of Laboratory Medicine,Huangyan Hospital of Wenzhou Medical University,Taizhou First People rs Hospital,Taizhou 318020,China [2]Eye Institute of Xiamen University,Fujian Provincial Key Laboratory of Ophthalmology and Visual Science,School of Medicine,Xiamen University,Xiamen 361102,China

出　　处：《Journal of Zhejiang University-Science B(Biomedicine & Biotechnology)》2019年第12期960-971,共12页浙江大学学报（英文版）B辑（生物医学与生物技术）

基　　金：Project supported by the Zhejiang Provincial Natural Science Foundation of China(No.LQ17H120001);the Medical Science and Technology Program of Zhejiang Province(Nos.2016KYA195 and 2017KY714);the National Natural Science Foundation of China(No.81801424);the 211 Talents Training Program of Taizhou,China

摘　　要：The present study was designed to analyze the metabolites of all-trans-retinal(atRal) and compare the cytotoxicity of atRal versus its derivative all-trans-retinoic acid(atRA) in human retinal pigment epithelial(RPE) cells. We confirmed that atRA was produced in normal pig neural retina and RPE. The amount of all-trans-retinol(atROL) converted from atRal was about 2.7 times that of atRal-derived atRA after incubating RPE cells with 10 μmol/L atRal for 24 h, whereas atRA in medium supernatant is more plentiful(91 vs. 29 pmol/mL), suggesting that atRA conversion ? facilitates elimination of excess atRal in the retina. Moreover, we found that mRNA expression of retinoic acid-specific hydroxylase CYP26 b1 was dose-dependently up-regulated by atRal exposure in RPE cells, indicating that atRA inactivation may be also initiated in atRal-accumulated RPE cells. Our data show that atRA-caused viability inhibition was evidently reduced compared with the equal concentration of its precursor atRal. Excess accumulation of atRal provoked intracellular reactive oxygen species(ROS) overproduction, heme oxygenase-1(HO-1) expression, and increased cleaved poly(ADP-ribose) polymerase 1(PARP1) expression in RPE cells. In contrast, comparable dosage of atRA-induced oxidative stress was much weaker, and it could not activate apoptosis in RPE cells. These results suggest that atRA generation is an antidotal metabolism pathway for atRal in the retina. Moreover, we found that in the eyes of ABCA4-/-RDH8-/-mice, a mouse model with atRal accumulation in the retina, the atRA content was almost the same as that in the wild type. It is possible that atRal accumulation simultaneously and equally promotes atRA synthesis and clearance in eyes of ABCA4-/-RDH8-/-mice, thus inhibiting the further increase of atRA in the retina. Our present study provides further insights into atRal clearance in the retina.目的:探讨视网膜中全反式视黄醛(atRal)能否代谢生成全反式维甲酸(atRA),并比较两者对视网膜色素上皮细胞(RPE)的细胞毒性作用,以阐明atRA生成的意义。创新点:建立atRA的超高效液相串联质谱(UPLCMS/MS)检测方法,并证明atRA的生成是视网膜中atRal的重要解毒代谢通路。方法:利用UPLC-MS/MS分别检测猪眼神经视网膜及RPE层中atRA的含量;利用ARPE-19细胞系模拟atRal在RPE中累积,用UPLC-MS/MS检测细胞内及培养基中atRA的含量,并用定量聚合酶链反应(qP CR)检测CYP26b1的表达;利用CCK8、DCFH-DA染色、qP CR、westernblot等方法对比等浓度atRA和atRal在RPE细胞中所诱导的细胞毒性、氧化应激、凋亡相关蛋白表达水平;用UPLC-MS/MS检测视网膜atRal清除障碍的ABCA4-/-RDH8-/-小鼠眼球中atRA及全反式视黄醇。结论:明确atRA在正常视网膜中能够代谢产生;证明其形成有利于RPE细胞中累积的atRal迅速代谢消除;其自身诱导细胞氧化应激的能力显著低于atRal,因而能显著减弱后者的细胞毒性。

关 键 词：All-trans-retinal All-trans-retinoic acid Antidotal pathway Human retinal pigment epithelial cell Oxidative stress 

分 类 号：R777.1[医药卫生—眼科]