[1]易彩霞,孙馨,印双红,等.褪黑素联合锌对H2O2诱导的人视网膜色素上皮细胞氧化损伤的保护作用[J].眼科新进展,2022,42(3):189-193.[doi:10.13389/j.cnki.rao.2022.0038]
　YI Caixia,SUN Xin,YIN Shuanghong,et al.Protective effects of melatonin and zinc on retinal pigment epithelial cells against H2O2-induced oxidative damage[J].Recent Advances in Ophthalmology,2022,42(3):189-193.[doi:10.13389/j.cnki.rao.2022.0038]
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褪黑素联合锌对H2O2诱导的人视网膜色素上皮细胞氧化损伤的保护作用/HTML
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《眼科新进展》[ISSN:1003-5141/CN:41-1105/R]

卷:
42卷
期数:
2022年3期
页码:
189-193
栏目:
实验研究
出版日期:
2022-03-05

文章信息/Info

Title:
Protective effects of melatonin and zinc on retinal pigment epithelial cells against H2O2-induced oxidative damage
作者:
易彩霞; 孙馨; 印双红; 黄啸
554300　贵州省铜仁市，铜仁学院大健康学院
Author(s):
YI Caixia; SUN Xin; YIN Shuanghong; HUANG Xiao
School of Sports and Health Science,Tongren University,Tongren 554300,Guizhou Province,China
关键词:
视网膜色素上皮细胞; 褪黑素; 锌; 氧化损伤; 炎症反应
Keywords:
retinal pigment epithelial cells;  melatonin;  zinc;  oxidative damage;  inflammatory response
分类号:
R774.1
DOI:
10.13389/j.cnki.rao.2022.0038
文献标志码:
A
摘要:
目的　探讨褪黑素（MT）联合锌（Zn）对H2O2诱导的人视网膜色素上皮（ARPE）细胞氧化损伤的保护作用。方法　常规培养ARPE细胞株（ARPE-19），利用不同浓度H2O2处理ARPE-19细胞，将细胞存活率接近50%的H2O2浓度作为氧化损伤浓度。将ARPE-19细胞分为：不同浓度MT（10-5 mol·L-1、10-6mol·L-1、10-7mol·L-1）+ZnCl2（15 μmol·L-1）组，ZnCl2组（采用15 μmol·L-1 ZnCl2培养细胞），H2O2组（不添加MT和ZnCl2培养细胞），空白对照组（细胞不做任何处理）。采用流式细胞仪检测细胞凋亡情况。采用试剂盒测量各组细胞内活性氧（ROS）和丙二醛（MDA）的含量。ELISA检测各组细胞内白细胞介素-6（IL-6）和肿瘤坏死因子-α（TNF-α）的表达量。结果　当H2O2浓度为300 μmol·L-1时，细胞存活率为(54.31±1.91)%，此时细胞存活率接近50%，后续实验中均选择该浓度作为氧化损伤浓度。ZnCl2组、10-5 mol·L-1 MT+ZnCl2组、10-6 mol·L-1 MT+ZnCl2组、10-7 mol·L-1 MT+ZnCl2组细胞凋亡率分别为3.05%、1.17%、1.50%、1.71%，与空白对照组（0）相比差异均无统计学意义（均为P>0.05）。H2O2组、ZnCl2组、10-7mol·L-1 MT+ZnCl2组细胞内ROS含量分别为空白对照组的1.76倍、1.59倍、1.41倍，差异均有统计学意义（均为P<0.05）；10-5 mol·L-1 MT+ZnCl2组、10-6 mol·L-1 MT+ZnCl2组细胞内ROS含量分别为空白对照组的1.13倍和1.25倍，差异均无统计学意义（均为P>0.05）。H2O2组细胞内MDA的含量较空白对照组显著提高 （P<0.05）；ZnCl2组、10-5 mol·L-1 MT+ZnCl2组、10-6 mol·L-1 MT+ZnCl2组、10-7 mol·L-1 MT+ZnCl2组细胞内MDA的含量均较空白对照组稍增加，差异均无统计学意义（均为P>0.05）。H2O2组、ZnCl2组、10-7 mol·L-1 MT+ZnCl2组细胞内IL-6和TNF-α的表达量均显著高于空白对照组，差异均有统计学意义（均为P<0.05）；10-5 mol·L-1MT+ZnCl2组和10-6 mol·L-1 MT+ZnCl2组细胞内IL-6和TNF-α的表达量均略高于空白对照组，差异均无统计学意义（均为P>0.05）。结论　MT联合Zn能有效降低H2O2损伤所致ARPE-19细胞内ROS的含量，随之减少细胞内MDA的含量，同时也能抑制IL-6和TNF-α的表达量，从而起到保护RPE细胞的作用。
Abstract:
Objective　To investigate the protective effects of melatonin (MT) and zinc (Zn) on retinal pigment epithelial (RPE) cells against H2O2-induced oxidative damage.Methods　ARPE-19 cells were treated with H2O2 of different concentrations, and finally the H2O2 concentration that ensured the cell viability close to 50% was chosen to be used in other experiments. ARPE-19 cells were divided into the MT (10-5 mol·L-1, 10-6 mol·L-1, and 10-7 mol·L-1)+ZnCl2 (15 μmol·L-1) group, ZnCl2 group (cells were cultured with 15 μmol·L-1 ZnCl2), H2O2 group (cells were cultured without MT and ZnCl2), and blank control group (cells received no treatment). Cell apoptosis was detected by flow cytometry. The content of reactive oxygen species (ROS) and malondialdehyde (MDA) in the cells was determined by assay kits. The expression levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were measured by ELISA kits.Results　When the concentration of H2O2 was 300 μmol·L-1, the cell survival rate was (54.31±1.91) %, close to 50%. Thus, this concentration was selected in subsequent experiments for oxidative damage. The apoptosis rates in the ZnCl2 group, 10-5 mol·L-1 MT+ZnCl2 group, 10-6 mol·L-1 MT+ZnCl2 group, and 10-7 mol·L-1 MT+ZnCl2 group were 3.05%, 1.17%, 1.50%, and 1.71%, respectively, without statistically significant differences compared with the blank control group (all P>0.05). The ROS content in the H2O2 group, ZnCl2 group, and 10-7 mol·L-1 MT+ZnCl2 group was 1.76, 1.59, and 1.41 times higher than that in the blank control group, respectively, and the differences were statistically significant (all P<0.05). The ROS content in the 10-5 mol·L-1 MT+ZnCl2 group and 10-6 mol·L-1 MT+ZnCl2 group were 1.13 and 1.25 times higher than that in the blank control group, respectively, but the differences were not statistically significant (all P>0.05). Compared with the blank control group, the MDA content in the H2O2 group was significantly increased (P<0.05), while in the ZnCl2 group, 10-5 mol·L-1 MT+ZnCl2 group, 10-6 mol·L-1 MT+ZnCl2 group, and 10-7 mol·L-1 MT+ZnCl2 group, the MDA content was slightly increased, without statistically significant differences (all P>0.05). The expression levels of IL-6 and TNF-α in the H2O2 group, ZnCl2 group, and 10-7 mol·L-1 MT+ZnCl2 group were significantly higher than those in the blank control group, and the differences were statistically significant (all P<0.05). The expression levels of IL-6 and TNF-α in the 10-5 mol·L-1 MT+ZnCl2 group and 10-6 mol·L-1 MT+ZnCl2 group were slightly higher than those in the blank control group, and the differences were not statistically significant (all P>0.05).Conclusion　MT and Zn can effectively reduce the ROS and MDA content in ARPE-19 cells induced by H2O2 damage, and meanwhile inhibit the expression of IL-6 and TNF-α, thus protecting RPE cells.

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备注/Memo

备注/Memo:
国家自然科学基金资助（编号：31800839）；贵州省教育厅普通本科高校青年科技人才成长项目（编号：黔教合KY字［2022］069号）
更新日期/Last Update: 2022-03-05