Protective effect of polypyrimidine bundle-binding protein-related splicing factor on retinal pigment epithelial cell injury induced by advanced glycation end products_Chinese Journal of Ocular Fundus Diseases

Authors：

Qi Chen , Zhang Hui , Lin Tingting , Ke Yifeng , Ren Xinjun , Bu Shaochong , Huang Liangyu , Wang Yong , Jiao Mingfei , Hu Liying , Wang Qiong , Hong Yaru , Li Xiaorong ,  Dong Lijie

Tianjin Medical University Eye Hospital , Tianjin Medical University Eye Institute , The College of Optometry & Ophthalmology , Tianjin 300384, China;
Qi Chen and Zhang Hui are contributed equally to the article;

Corresponding author：

Dong Lijie, Email: aitaomubang@126.com

Keywords：

Retinal pigment epithelium; Polypyrimidine tract-binding protein; Glycosylation end products, advanced; Heme oxygenase (decyclizing)

DOI：

10.3760/cma.j.issn.1005-1015.2020.01.011

Video：

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Objective To observe the protective effect of polypyrimidine bundle-binding protein-related splicing factor (PSF) over-expression on RPE cell injury induced by advanced glycation end products (AGEs).Methods The human RPE cells cultured in vitro were divided into three groups: normal control group (N group), blank control group (N + AGEs group), empty vector control group (Vec + AGEs group), and PSF high expression group (PSF + AGEs). group). RPE cells in N group were routinely cultured; RPE cells in N + AGEs group were only transfected but did not introduce any exogenous genes combined with AGEs induction; Vec +AGEs group and PSF + AGEs group were transfected with pcDNA The empty vector or pcDNA-PSF eukaryotic expression plasmid was introduced into RPE cells and induced by AGEs. Except the N group, the other 3 groups of cells were transfected accordingly, and were stimulated with 150 μg/ml AGEs for 72 h after 24 h. HE staining and Hoechst 33258 staining were used to observe the effect of high PSF expression on the morphological changes of RPE cells; ROS level detection was used to analyze the effect of PSF high expression on the ROS expression of RPE cells induced by AGEs; MTT colorimetric method was used to detect the high PSF expression Effects on the viability of RPE cells; Western blot was used to detect the effects of different time and dose of PSF on the expression of heme oxygenase 1 (HO-1).Results HE staining and Hoechst 33258 staining observation showed that the cells in group N were full in shape, the nucleus was round, the cytoplasm was rich, and the staining was uniform; the cells in N + AGEs group and Vec + AGEs group were reduced in size, the eosinophilic staining was enhanced, and the nucleus was densely densely stained. Pyrolysis and even fragmentation; the morphology of cells in the PSF + AGEs group was still full, the cytoplasm staining was more uniform, and the nucleus staining was uniform. The results of MTT colorimetry showed that high expression of PSF can effectively improve the viability of RPE cells, but this effect can be effectively antagonized by ZnPP, and the difference is statistically significant (F=33.26, P＜0.05). DCFH-DA test results showed that compared with the N + AGEs group and Vec + AGEs group, the ROS production in PSF + AGEs group decreased, the difference was statistically significant (F=11.94, P＜0.05). Western blot analysis showed that PSF protein up-regulated HO-1 expression in a time- and dose-dependent manner. The relative expression level of HO-1 at 24, 48, and 72 h after PSF protein was significantly higher than that at 0 h, and the difference was statistically significant (F=164.91, P＜0.05). The relative expression level of HO-1 under the action of 0.1, 0.5, 1.0, 1.5, and 2.0 μg PSF protein was significantly higher than 0.0 μg, and the difference was statistically significant (F=104.82, P＜0.05).Conclusion PSF may inhibit the production of ROS by up-regulating the expression of HO-1, thus protecting the RPE cells induced by AGEs.

Citation： Qi Chen, Zhang Hui, Lin Tingting, Ke Yifeng, Ren Xinjun, Bu Shaochong, Huang Liangyu, Wang Yong, Jiao Mingfei, Hu Liying, Wang Qiong, Hong Yaru, Li Xiaorong, Dong Lijie. Protective effect of polypyrimidine bundle-binding protein-related splicing factor on retinal pigment epithelial cell injury induced by advanced glycation end products. Chinese Journal of Ocular Fundus Diseases, 2020, 36(1): 46-52. doi: 10.3760/cma.j.issn.1005-1015.2020.01.011 Copy

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2. Dong L, Nian H, Shao Y, et al. PTB-associated splicing factor inhibits IGF-1-induced VEGF upregulation in a mouse model of oxygen-induced retinopathy[J]. Cell Tissue Res, 2015, 360(2): 233-243. DOI: 10.1007/s00441-014-2104-5.
3. Dong L, Zhang X, Fu X, et al. PTB-associated splicing factor (PSF) functions as a repressor of STAT6-mediated Ig epsilon gene transcription by recruitment of HDAC1[J]. J Biol Chem, 2011, 286(5): 3451-3459. DOI: 10.1074/jbc.M110.168377.
4. 田芳, 李文博, 黄亮瑜, 等. 聚嘧啶束结合蛋白相关剪接因子对过氧化氢诱导下视网膜色素上皮细胞凋亡的影响[J]. 中华眼底病杂志, 2018, 34(2): 159-163. DOI: 10.3760/cma.j.issn.1005-1015.2018.02.012.Tian F, Li WB, Huang LY, et al. The effect of polypyrimidine tract binding protein-associated splicing factor on hydrogen peroxide induced apoptosis of retinal pigment epithelial[J]. Chin J Ocul Fundus Dis, 2018, 34(2): 159-163. DOI: 10.3760/cma.j.issn.1005-1015.2018.02.012.
5. Zhang L, Dong L, Liu X, et al. Alpha-melanocyte-stimulating hormone protects retinal vascular endothelial cells from oxidative stress and apoptosis in a rat model of diabetes[J/OL]. PLoS One, 2014, 9(4): 93433[2014-04-02]. http://dx.plos.org/10.1371/journal.pone.0093433. DOI: 10.1371/journal.pone.0093433.
6. Shi Y, Su C, Wang JT, et al. Temporal and spatial changes in VEGF, αA- and αB-crystallin expression in a mouse model of oxygen-induced retinopathy[J]. Int J Clin Exp Med, 2015, 8(3): 3349-3359.
7. Wei R, Dong L, Xiao Q, et al. Engagement of Toll-like receptor 2 enhances interleukin (IL)-17(+) autoreactive T cell responses via p38 mitogen-activated protein kinase signalling in dendritic cells[J]. Clin Exp Immunol, 2014, 178(2): 353-363. DOI: 10.1111/cei.12405.
8. 邢小丽, 黄亮瑜, 苏睿虹, 等. 丁基苯酞对过氧化氢诱导下视网膜Müller细胞凋亡的影响[J]. 中华眼底病杂志, 2018, 34(5): 481-486. DOI: 10.3760/cma.j.issn.1005-1015.2018.05.014.Xing XL, Huang LY, Su RH, et al. Effect of dl-3-n-Butylphthalide on apoptosis of retinal müller cells induced by hydrogen peroxide[J]. Chin J Ocul Fundus Dis, 2018, 34(5): 481-486. DOI: 10.3760/cma.j.issn.1005-1015.2018.05.014.
9. 邢小丽, 黄亮瑜, 张哲, 等. 丁基苯酞对H2O2诱导下视网膜色素上皮细胞凋亡的保护作用[J]. 中华眼底病杂志, 2019, 35(5): 480-487. DOI: 10.3760/cma.j.issn.1005-1015.2019.05.011.Xing XL, Huang LY, Zhang Z, et al. Effects of butylphthalide on hydrogen peroxide induced retinal pigment epithelial cells injury[J]. Chin J Ocul Fundus Dis, 2019, 35(5): 480-487. DOI: 10.3760/cma.j.issn.1005-1015.2019.05.011.
10. 张哲, 刘巨平, 东莉洁, 等. 高糖状态下视网膜血管内皮细胞基因表达谱的RNA-Seq分析[J]. 中华眼底病杂志, 2018, 34(4): 377-381. DOI: 10.3760/cma.j.issn.1005-1015.2018.04.014.Zhang Z, Liu JP, Dong LJ, et al. RNA-Seq analysis of gene expression profiling in retinal vascular endothelial cells under high glucose condition[J]. Chin J Ocul Fundus Dis, 2018, 34(4): 377-381. DOI: 10.3760/cma.j.issn.1005-1015.2018.04.014.
11. 牛瑞, 东莉洁, 马腾, 等. 抗血管内皮生长因子药物治疗后视网膜血管内皮细胞基因表达谱的RNA-Seq分析[J]. 中华眼底病杂志, 2018, 34(3): 275-280. DOI: 10.3760/cma.j.issn.1005-1015.2018.03.016.Niu R, Dong LJ, Ma T, et al. RNA-Seq analysis of gene expression profiling in human retinal vascular endothelial cells after anti-vascular endothelial growth factor treatment[J]. Chin J Ocul Fundus Dis, 2018, 34(3): 275-280. DOI: 10.3760/cma.j.issn.1005-1015.2018.03.016.
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Chinese Journal of Ocular Fundus Diseases

Protective effect of polypyrimidine bundle-binding protein-related splicing factor on retinal pigment epithelial cell injury induced by advanced glycation end products

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