The effect of polypyrimidine tract binding protein-associated splicing factor on hydrogen peroxide induced apoptosis of retinal pigment epithelial_Chinese Journal of Ocular Fundus Diseases

Authors：

Tian Fang , Li Wenbo , Huang Liangyu , Gao Meizi , Zhao Jinzhi , Hu Bojie , Zhang Xiaomin , Li Xiaorong ,  Dong Lijie

Tianjin Medical University Eye Hospital, Tianjin Medical University Eye Institute , College of Optometry and Ophthalmology, Tianjin 300384, China;

Corresponding author：

Dong Lijie, Email: aitaomubang@126.com

Keywords：

Retinal pigment epithelium; Polypyrimidine tract-binding protein; Apoptosis; Hydrogen peroxide

DOI：

10.3760/cma.j.issn.1005-1015.2018.02.012

Video：

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Objective To observe the effect of polypyramidine tract binding protein-associated splicing factor (PSF) on hydrogen peroxide (H₂O₂) induced apoptosis of retinal pigment epithelial (RPE) cells in vitro. Methods RPE cells were cultured and divided into a normal group, normal+H₂O₂ group, Vec+H₂O₂group, PSF+H₂O₂ group according to the experimental design. Overexpression of PSF in RPE cells were achieved by pEGFP-PSF plasmid transient transfection into RPE cells, then RPE cells were exposed to H₂O₂. The morphological changes were observed by hematoxylin-eosin (HE) staining and Live/Dead staining while the survival rate of cells was detected by MTT assay. The effect of PSF on H₂O₂-induced RPE apoptosis was analyzed by Cell Death Detection ELISA kit. Meanwhile, intracellular reactive oxygen species (ROS) level was detected by using DCFH-DA method. Results Overexpression of PSF could effectively alleviate the morphological changes induced by H₂O₂ stimulation shown by HE staining, and effectively reduce dead cells number shown by Live/Dead staining. After H₂O₂ stimulation, the survival rate, apoptosis rate and ROS production level in PSF overexpression group were 0.68±0.12, 0.44±0.08 and 18 616±3 382.54 respectively, showing significant difference in comparison with the vector plasmid group and normal group (P＜0.05). Conclusion PSF overexpression plays a protective role in H₂O₂-induced apoptosis by inhibiting the production of ROS in RPE cells.

Citation： Tian Fang, Li Wenbo, Huang Liangyu, Gao Meizi, Zhao Jinzhi, Hu Bojie, Zhang Xiaomin, Li Xiaorong, Dong Lijie. The effect of polypyrimidine tract binding protein-associated splicing factor on hydrogen peroxide induced apoptosis of retinal pigment epithelial. Chinese Journal of Ocular Fundus Diseases, 2018, 34(2): 159-163. doi: 10.3760/cma.j.issn.1005-1015.2018.02.012 Copy

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5.	漆晨, 东莉洁, 乐毅, 等.多聚嘧啶序列结合蛋白相关剪接因子对体外培养的视网膜色素上皮细胞磷脂酰肌醇3激酶/丝氨酸-苏氨酸蛋白激酶信号通路的调控作用[J].中华眼底病杂志, 2015, 31(4): 363-367. DOI: 10.3760/cma.j.issn.1005-1015.2015.04.013.Qi C, Dong LJ, Le Y , et al.The regulation of PTB-associated splicing factor on phosphatidylinositol 3 kinase/Akt signaling pathway in retinal pigment epithelial cells[J].Chin J Ocul Fundus Dis, 2015, 31(4): 363-367. DOI: 10.3760/cma.j.issn.1005-1015.2015.04.013.
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15.	Dong L, Hong N, Shao Y, et al. PTB-associatedsp licing 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.
16.	田芳, 东莉洁, 周玉, 等.重组腺相关病毒-多聚嘧啶序列结合蛋白相关剪接因子对氧诱导视网膜新生血管形成的抑制作用[J].中华眼底病杂志, 2014, 30(5): 504-508. DOI: 10.3760/cma.j.issn.1005-1015.2014.05.019.Tian F, Dong LJ, Zhou Y, et al.Inhibition of oxygen induced retinal neovascularization by recombinant adeno-associated virus-polypyrimidine tract-binding protein-associated splicing factor intraocular injection in mice[J].Chin J Ocul Fundus Dis, 2014, 30(5): 504-508. DOI: 10.3760/cma.j.issn.1005-1015.2014.05.019.
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20.	Kim YW, West XZ, Byzova TV.Inflammation and oxidative stress in angiogenesis and vascular disease[J]. J Mol Med (Berl), 2013, 91(3): 323-328. DOI: 10.1007/s00109-013-1007-3.
21.	Storz G, Imlay JA.Oxidative stress[J].Curr Opin Microbiol, 1999, 2(2): 188-194.
22.	Santschi C, Moos NV, Koman VB, et al. Non-invasive continuous monitoring of pro-oxidant effects of engineered nanoparticles on aquatic microorganisms[J]. J Nanobiotechnology, 2017, 15(1): 19. DOI: 10.1186/s12951-017-0253-x.
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24.	Kim DI, Park MJ, Choi JH, et al.PRMT1 and PRMT4 regulate oxidative stress-induced retinal pigment epithelial cell damage in sirt1-dependent and sirt1-independent manners[J/OL].Oxid Med Cell Longev, 2015, 2015: 617919[2015-10-25]. http://dx.doi.org/10.1155/2015/617919. DOI: 10.1155/2015/617919.
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26.	Jarrett SG, Lin H, Godley BF, et al.Mitochondrial DNA damage and its potential role in retinal degeneration[J].Prog Retin Eye Res, 2008, 27(6): 596-607. DOI: 10.1016/j.preteyeres.2008.09.001.
27.	Indo HP, Yen HC, Nakanishi I, et al.A mitochondrial superoxide theory for oxidative stress diseases and aging[J].J Clin Biochem Nutr, 201556(1): 1-7. DOI: 10.3164/jcbn.14-42.

1. Minasyan L, Sreekumar PG, Hinton DR, et al. Protective mechanisms of the mitochondrial- derived peptide humanin in oxidative and endoplasmic reticulum stress in RPE cells[J/OL].Oxid Med Cell Longev, 2017, 2017: 1675230[2017-07-26]. https://doi.org/10.1155/2017/1675230. DOI: 10.1155/2017/1675230.
2. Shadrach KG, Rayborn ME, Hollyfield JG, et al.DJ-1-dependent regulation of oxidative stress in the retinal pigment epithelium (RPE)[J/OL].PLoS One, 2013, 8(7): 67983[2013-07-02]. https://doi.org/10.1371/journal.pone.0067983. DOI: 10.1371/journal.pone.0067983.
3. Tysoncapper AJ, Shiells EA, Robson SC. Interplay between polypyrimidine tract binding protein-associated splicing factor and human myometrial progesterone receptors[J].J Mol Endocrinol, 2009, 43(1): 29-41. DOI: 10.1677/JME-09-0001.
4. Greco-Stewart VS, Thibault CS, Pelchat M. Binding of the polypyrimidine tract-binding protein-associated splicing factor (PSF) to the hepatitis delta virus RNA[J]. Virology, 2006, 356(1-2): 35-44. DOI: 10.1016/j.virol.2006.06.040.
5. 漆晨, 东莉洁, 乐毅, 等.多聚嘧啶序列结合蛋白相关剪接因子对体外培养的视网膜色素上皮细胞磷脂酰肌醇3激酶/丝氨酸-苏氨酸蛋白激酶信号通路的调控作用[J].中华眼底病杂志, 2015, 31(4): 363-367. DOI: 10.3760/cma.j.issn.1005-1015.2015.04.013.Qi C, Dong LJ, Le Y , et al.The regulation of PTB-associated splicing factor on phosphatidylinositol 3 kinase/Akt signaling pathway in retinal pigment epithelial cells[J].Chin J Ocul Fundus Dis, 2015, 31(4): 363-367. DOI: 10.3760/cma.j.issn.1005-1015.2015.04.013.
6. Mitter SK, Chunjuan S, Xiaoping Q, et al. Dysregulated autophagy in the RPE is associated with increased susceptibility to oxidative stress and AMD[J]. Autophagy, 2014, 10(11): 1989-2005. DOI: 10.4161/auto.36184.
7. Mitter SK, Chunjuan S, Xiaoping Q, et al. Dysregulated autophagy in the RPE is associated with increased susceptibility to oxidative stress and AMD[J]. Autophagy, 2014, 10(11): 1989-2005. DOI: 10.1167/iovs.14-14696. DOI: 10.1167/iovs.14-14696.
8. 何津岩, 东莉洁, 葛林, 等.重组pEGFP-C2-p100质粒构建及表达[J].天津医科大学学报, 2008, 14(2): 135-137. DOI: 10.3969/j.issn.1006-8147.2008.02.001.He JY, Dong LJ, Ge L, et al. Construction and expression of a pEGFP-C2-p100 recombinant plasmid[J].Journal of Tianjin Medical University, 2008, 14(2): 135-137. DOI: 10.3969/j.issn.1006-8147.2008.02.001.
9. Grigoryan E, Markitantova Y. Cellular and molecular preconditions for retinal pigment epithelium (RPE) natural reprogramming during retinal regeneration in urodela[J/OL]. Biomedicines, 2016, 4(4): 28[2016-12-01]. http://www.mdpi.com/2227-9059/4/4/28. DOI: 10.3390/biomedicines4040028.
10. Beatty S, Koh H, Phil M, et al. The role of oxidative stress in the pathogenesis of age-related macular degeneration[J]. Surv Ophthalmol, 2000, 45(2): 115-134.
11. Cai J, Nelson KC, Wu M, et al. Oxidative damage and protection of the RPE[J]. Prog Retin Eye Res, 2000, 19(2): 205-221.
12. Datta S, Cano M, Ebrahimi K, et al. The impact of oxidative stress and inflammation on RPE degeneration in non-neovascular AMD[J].Prog Retin Eye Res, 2017, 60: 201-218. DOI: 10.1016/j.preteyeres.2017.03.002.
13. Thurman JM, Renner B, Kunchithapautham K, et al.Oxidative stress renders retinal pigment epithelial cells susceptible to complement-mediated injury[J].J Biol Chem, 2009, 284(25): 16939-16947. DOI: 10.1074/jbc.M808166200.
14. Bandello F, Sacconi R, Querques L, et al. Recent advances in the management of dry age-related macular degeneration: a review[J]. F1000Res, 2017, 6: 245. DOI: 10.12688/f1000research.10664.1.
15. Dong L, Hong N, Shao Y, et al. PTB-associatedsp licing 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.
16. 田芳, 东莉洁, 周玉, 等.重组腺相关病毒-多聚嘧啶序列结合蛋白相关剪接因子对氧诱导视网膜新生血管形成的抑制作用[J].中华眼底病杂志, 2014, 30(5): 504-508. DOI: 10.3760/cma.j.issn.1005-1015.2014.05.019.Tian F, Dong LJ, Zhou Y, et al.Inhibition of oxygen induced retinal neovascularization by recombinant adeno-associated virus-polypyrimidine tract-binding protein-associated splicing factor intraocular injection in mice[J].Chin J Ocul Fundus Dis, 2014, 30(5): 504-508. DOI: 10.3760/cma.j.issn.1005-1015.2014.05.019.
17. Kai K, Tokarz P, Koskela A, et al. Autophagy regulates death of retinal pigment epithelium cells in age-related macular degeneration[J].Cell Biol Toxicol, 2017, 33(2): 113-128. DOI: 10.1007/s10565-016-9371-8.
18. Xia C, Meng Q, Liu LZ, et al.Reactive oxygen species regulate angiogenesis and tumor growththrough vascular endothelial growth factor[J].Cancer Res, 2007, 67: 10823-10830. DOI: 10.1158/0008-5472.CAN-07-0783.
19. Ushio-Fukai M, Nakamura Y. Reactive oxygen species and angiogenesis: NADPH oxidase as target for cancer therapy[J].Cancer Lett., 2008, 266(1): 37-52. DOI: 10.1016/j.canlet.2008.02.044.
20. Kim YW, West XZ, Byzova TV.Inflammation and oxidative stress in angiogenesis and vascular disease[J]. J Mol Med (Berl), 2013, 91(3): 323-328. DOI: 10.1007/s00109-013-1007-3.
21. Storz G, Imlay JA.Oxidative stress[J].Curr Opin Microbiol, 1999, 2(2): 188-194.
22. Santschi C, Moos NV, Koman VB, et al. Non-invasive continuous monitoring of pro-oxidant effects of engineered nanoparticles on aquatic microorganisms[J]. J Nanobiotechnology, 2017, 15(1): 19. DOI: 10.1186/s12951-017-0253-x.
23. Kim KS, Lee D, Song CG, et al.Reactive oxygen species-activated nanomaterials as theranostic agents[J].Nanomedicine (Lond), 2015, 10(17): 2709-2723. DOI: 10.2217/nnm.15.108.
24. Kim DI, Park MJ, Choi JH, et al.PRMT1 and PRMT4 regulate oxidative stress-induced retinal pigment epithelial cell damage in sirt1-dependent and sirt1-independent manners[J/OL].Oxid Med Cell Longev, 2015, 2015: 617919[2015-10-25]. http://dx.doi.org/10.1155/2015/617919. DOI: 10.1155/2015/617919.
25. Jarrett SG, Lin H, Godley BF, et al. Mitochondrial DNA damage and its potential role in retinal degeneration[J].Prog Retin Eye Res, 2008, 27(6): 596-607. DOI: 10.1016/j.preteyeres.2008.09.001.
26. Jarrett SG, Lin H, Godley BF, et al.Mitochondrial DNA damage and its potential role in retinal degeneration[J].Prog Retin Eye Res, 2008, 27(6): 596-607. DOI: 10.1016/j.preteyeres.2008.09.001.
27. Indo HP, Yen HC, Nakanishi I, et al.A mitochondrial superoxide theory for oxidative stress diseases and aging[J].J Clin Biochem Nutr, 201556(1): 1-7. DOI: 10.3164/jcbn.14-42.

Chinese Journal of Ocular Fundus Diseases

The effect of polypyrimidine tract binding protein-associated splicing factor on hydrogen peroxide induced apoptosis of retinal pigment epithelial

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