摘要
目的探讨驻景丸加减方对干性年龄相关性黄斑变性(AMD)模型小鼠视网膜的保护作用以及对核因子E2相关因子2(Nrf2)通路和自噬的影响。方法取6月龄健康雄性C57BL/6小鼠90只,将小鼠随机分为年龄对照组(正常饮食)、溶媒对照组(高脂饮食+氢醌)、对照药物组(高脂饮食+氢醌+莱视盯)及中药组(高脂饮食+氢醌+驻景丸加减方,分低、中、高三个剂量组)6组,每组15只。莱视盯主要成分叶黄素、β-胡萝卜素、葡萄糖酸锌(每100 g含叶黄素2.20 g、β-胡萝卜素0.86 g、锌1.50 g)。各组小鼠灌胃给药,每天1次,持续3个月。动物观察期满处死并摘取眼球,透射电镜观察视网膜组织,测量视网膜色素上皮(RPE)下沉积物面积及Bruch膜厚度,检测视网膜匀浆活性氧自由基(ROS)、丙二醛(MDA)含量,Western blot检测Nrf2、血红素加氧酶-1(HO-1)、醌氧化还原酶-1(NQO-1)、p62和LC3蛋白表达水平。结果与年龄对照组相比,溶媒对照组RPE下沉积物面积显著增大、Bruch膜明显增厚,差异均有统计学意义(均为P<0.01);与溶媒对照组相比,中药中、高剂量组RPE下沉积物面积减小、Bruch膜厚度变薄,差异均有统计学意义(均为P<0.01);与溶媒对照组相比,中药低剂量组和对照药物组RPE下沉积物面积也均减小,差异均有统计学意义(均为P<0.05),但Bruch膜厚度变化不明显。与溶媒对照组相比,中药中、高剂量组ROS、MDA含量显著降低(均为P<0.01),中药低剂量组和对照药物组ROS、MDA含量也均降低(均为P<0.05);与溶媒对照组相比,中药中、高剂量组细胞质Nrf2蛋白表达显著下调,细胞核Nrf2蛋白表达上调(均为P<0.01),中药低剂量组和对照药物组细胞质Nrf2蛋白表达下调,细胞核Nrf2蛋白表达上调(均为P<0.05);与溶媒对照组相比,中药中、高剂量组HO-1、NQO-1、p62和LC3 II蛋白表达均显著上调(均为P<0.01),中药低剂量组HO-1、p62和LC3 II蛋白表达均上调(均为P<0.05),对照药物组HO-1和LC3 II蛋白表达均上调(均为P<0.05)。结论驻景丸加减方可激活Nrf2通路,上调下游靶基因酶HO-1、NQO-1表达,快速清除ROS、MDA;增强自噬,加快清除氧化损伤的细胞器,对干性AMD模型小鼠视网膜有保护作用。
Objective To investigate the protective effect of modified prescription of Zhujingwan on the retina of mice with dry age-related macular degeneration(AMD)and the involvements of the nuclear factor E2-related factor 2(Nrf2)pathway and autophagy.Methods Ninety 6-month-old male C57BL/6 mice were randomly divided into 6 groups aging control group(normal diet),vehicle control group(high-fat diet+hydroquinone),drug control group(high-fat diet+hydroquinone+Laishiding),and Chinese herbs groups(high-fat diet+hydroquinone+low,medium and high dose of modified prescription of Zhujingwan),with 15 mice in each group.Main ingredients of Laishiding included 2.20 g lutein,0.86 gβ-carotene,and 1.50 g zinc gluconate per 100 g.Drugs were given once daily through intragastric administration for 3 months.At the end of the experimental period,animals were sacrificed for collecting eyeballs.The sediment area under RPE and Bruch membrane(BrM)thickness were evaluated by electron microscopy.ROS and MDA levels of mouse retina were measured.In addition,protein expressions of Nrf2,heme oxygenase-1(HO-1),quinone oxidoreductase-1(NQO-1),p62 and LC3 of mouse retina were detected by Western blot.Results Compared with the aging control group,significantly increased sediment area under RPE and BrM were detected in the vehicle control group(both P<0.01).Compared with the vehicle control group,the sediment area under RPE reduced and the BrM thickness were significantly lower in Chinese herbs groups with the medium and high dose of modified prescription of Zhujingwan(all P<0.01).In addition,the sediment area under RPE also significantly decreased in Chinese herbs group with the low dose of modified prescription of Zhujingwan(P<0.05),while the change of BrM thickness was not obvious.Compared with the vehicle control group,ROS and MDA levels significantly decreased in Chinese herbs group with three doses of modified prescription of Zhujingwan(low dose group P<0.05,medium and high dose groups P<0.01).Compared with the vehicle control group,the protein level of Nrf2 was significantly down-regulated in cytoplasm and up-regulated in nucleus in Chinese herbs group with the medium and high doses of modified prescription of Zhujingwan(all P<0.01),and the same trends were observed in the low dose group and positive drug group as well(P<0.05).Protein expressions of HO-1,NQO-1,p62 and LC3 II were up-regulated in Chinese herbs group with the medium and high doses of modified prescription of Zhujingwan(all P<0.01),and those of HO-1,p62 and LC3 II were up-regulated in the low dose group(all P<0.05).Meanwhile,protein levels of HO-1 and LC3 II were up-regulated in positive drug group(P<0.05).Conclusion The modified prescription of Zhujingwan can activate the Nrf2 pathway and up-regulate target genes HO-1 and NQO-1,thus quickly eliminating ROS and MDA.In addition,it also enhances autophagy and accelerates the removal of oxidative damage organelles,and thus plays a protective role in retina of mice with dry age-related macular degeneration.
作者
柯玲玲
周欣
张元钟
刘雨露
仲路
孙化萍
杭丽
金青子
徐新荣
KE Lingling;ZHOU Xin;ZHANG Yuanzhong;LIU Yulu;ZHONG Lu;SUN Huaping;HANG Li;JIN Qingzi;XU Xinrong(Department of Ophthalmology,Jiujiang Hospital of Chinese Medicine,Jiujiang 332000,Jiangxi Province,China;Department of Ophthalmology,Jiangsu Provincial Hospital of Chinese Medicine(the Affiliated Hospital of Nanjing University of Chinese Medicine),Nanjing 210029,Jiangsu Province,China;Department of Ophthalmology,Nanjing Hospital of Chinese Medicine,Nanjing 210022,Jiangsu Province,China)
出处
《眼科新进展》
CAS
北大核心
2021年第9期812-816,共5页
Recent Advances in Ophthalmology
基金
国家自然科学基金资助(编号82074177)
江苏省重点研发计划项目(编号BE2018757)。
关键词
年龄相关性黄斑变性
驻景丸加减方
氧化损伤
Nrf2通路
自噬
age-related macular degeneration
modified prescription of Zhujingwan
oxidative damage
Nrf2 pathway
autophagy
