摘要
目的研究黄斑变性方对蓝光损伤的视网膜色素上皮细胞(human retinal pigmental epithelial cell,hRPE)凋亡以及线粒体膜电位、线粒体活性氧表达的影响,阐释黄斑变性方对蓝光损伤的hRPE细胞的保护作用及可能的机制,为临床应用黄斑变性方治疗年龄相关性黄斑变性提供理论依据。方法黄斑变性方由熟地、山茱萸、山药、茯苓、白术、当归、黄精、枸杞子等组成,具有补益肝肾、养阴明目的功效,但其具体作用机制尚不明确。本研究采用血清药理学方法,分为A组(空白对照组)、B组(蓝光照射未加中药组)、C组(中药低剂量组,10%含药血清处理)、D组(中药中剂量组,20%含药血清处理)、E组(中药高剂量组,30%含药血清处理)5个组,不同浓度的含药血清分别干预蓝光照射的hRPE细胞,流式细胞仪检测细胞的凋亡及线粒体膜电位、线粒体活性氧表达的影响。结果经黄斑变性方干预后的细胞,A组空白对照组、B组蓝光照射未加中药组、C组中药低剂量组、D组中药中剂量组、E组中药高剂量组,hRPE细胞凋亡数分别为(1.09±0.71)%、(18.94±4.32)%、(4.76±1.76)%、(2.29±1.34)%和(1.17±0.57)%,各组比较差异有统计学意义(F=23.723,P<0.01),两两比较差异有显著统计学意义(P<0.01),且hRPR细胞凋亡率随含药血清浓度的增加而逐渐减少,具有浓度依赖性。经黄斑变性方干预的细胞线粒体活性氧阳性细胞率在C组中药低剂量组、D组中药中剂量组、E组中药高剂量组分别为(3.22±0.13)%,(2.96±0.20)%,(1.91±0.17)%,而未经含药血清干预的B组单纯蓝光照射组细胞线粒体活性氧阳性细胞率为(5.43±0.27)%,各组比较差异有统计学意义(F=25.245,P<0.01),两两比较差异有统计学意义(P<0.01);在经黄斑变性方含药血清低剂量、中剂量和高剂量干预后,各组细胞线粒体膜电位阳性细胞百分率分别为(76.56±1.13)%,(80.31±2.20)%和(95.32±3.17)%,而未经含药血清干预的细胞线粒体膜电位阳性的细胞率为(69.89±2.27)%,各组比较差异有统计学意义(F=47.834,P<0.01),两两比较差异有统计学意义(P<0.01)。结论黄斑变性方可通过线粒体凋亡途径保护蓝光照射引起的视网膜色素上皮细胞的凋亡,我们的研究结果为黄斑变性方的临床应用提供了理论依据。
Objective To study the effect of Macular Degeneration Formula(黄斑变性方)on apoptosis,mitochondrial membrane potential and mitochondrial reactive oxygen species expression of human retinal pigment epithelial(hRPE)cells injured by blue light,and elucidate the protective effect and its possible mechanism of Macular Degeneration Formula on hRPE cells injured by blue light.To provide a theoretical basis for the clinical application of Macular Degeneration Formula in the treatment of age-related macular degeneration.Methods Macular Degeneration Formula is composed of Shudihuang(Rehmanniae Radix Praeparata),Shanyurou(Corni Fructus),Shanyao(Dioscoreae Rhizoma),Fuling(Poria),Baizhu(Atractylodis Macrocephalae Rhizoma),Danggui(Angelicae Sinensis Radix),Huanging(Polygonati Rhizoma)and Gouqizi(Lycii Fructus),and has the effects of tonifying liver and kidney,nourishing Yin and improving eyesight,but its specific mechanism is not clear.In this study,the serum pharmacology method was used to divide the rats into five groups:group A(blank control group),group B(blue light irradiation group),group C(low dose of traditional Chinese medicine group,10% medicated serum),group D(medium dose of traditional Chinese medicine group,20% medicated serum)and group E(high dose of traditional Chinese medicine group,30% medicated serum).Flow cytometry was used to detect cell apoptosis,mitochondrial membrane potential and mitochondrial reactive oxygen species expression.Results The number of apoptosis of hRPE cells in the group A(blank control group),group B(blue light irradiation group),group C(low dose of traditional Chinese medicine group),group D(medium dose of traditional Chinese medicine group)and group E(high dose of traditional Chinese medicine group)were(1.09±0.71)%,(18.94±4.32)%,(4.76±1.76)%,(2.29±1.34)% and(1.17±0.57)%,respectively(F=23.723,P<0.01).There was significant difference between every two groups(P<0.01),and the number of apoptosis of hRPE cells gradually decreased with the concentration of traditional Chinese medicine in a concentration dependent manner.The number of mitochondrial ROS positive cells in low-dose group,medium dose group and high-dose group was(3.22±0.13)%,(2.96±0.20)% and(1.91±0.17)%,respectively,while the number of mitochondrial ROS positive cells in group B(blue light irradiation group was(5.43±0.27)%(F=25.245,P<0.01).There was significant difference between the two groups(P<0.01).The number of mitochondrial membrane potential positive cells intervened by Macular Degeneration Formula was(76.56±1.13)%,(80.31±2.20)% and(95.32±3.17)%,while the number of mitochondrial membrane potential positive cells not intervened by Macular Degeneration Formula was(69.89±2.27)%,the difference was statistically significant(F=47.834,P<0.01).The difference was statistically significant between every two groups(P<0.01).Conclusion Macular Degeneration Formula can protect hRPE cells from apoptosis induced by blue light through mitochondrial apoptosis pathway,which provides theoretical basis for clinical application of Macular Degeneration Formula.
作者
杜秀娟
高延娥
郭大东
DU Xiujuan;GAO Yan’e;GUO Dadong(Affiliated Eye Hospital of Shandong University of Traditional Chinese Medicine,Jinan 250002,Shandong,China;Eye Institute of Shandong University of Traditional Chinese Medicine,Jinan 250002,Shandong,China)
出处
《辽宁中医杂志》
CAS
2023年第7期230-233,I0006,共5页
Liaoning Journal of Traditional Chinese Medicine
基金
国家自然科学基金面上项目(81974584)
陕西省重点研发计划项目(2019SF-295)
山东省中医药科技发展计划项目(2019-0251)
山东省医药卫生科技发展计划项目(2019WS566)。
关键词
黄斑变性方
凋亡
线粒体膜电位
线粒体活性氧
Macular Degeneration Formula(黄斑变性方)
apoptosis
mitochondrial membrane potential
mitochondrial reactive oxygen species
