Preliminary discussion on the potential mechanism of follistatin-like protein 1 in the process of proliferative diabetic retinopathy_Chinese Journal of Ocular Fundus Diseases

Authors：

Niu Rui , Dong Lijie , Du Xueli , He Yanhua , Nie Zetong , Cui Weina , Chen Qiong ,  Hu Bojie

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

Corresponding author：

Hu Bojie, Email: bhu07@tmu.edu.cn

Keywords：

Follistatin-related proteins; Human umbilical vein endothelial cells; Diabetic retinopathy

DOI：

10.3760/cma.j.cn511434-20190115-00024

Video：

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Objective To observe the changes of follistatin-like protein 1 (FSTL1) in serum of patients with proliferative diabetic retinopathy (PDR).Methods Twenty PDR patients confirmed by clinical examination and 20 normal people were included in the study. Human retinal vascular endothelial cells (HRCEC) were divided into HRCEC blank control group, 3 h hypoxia group, 6 h hypoxia group. Human umbilical vein endothelial cell (HUVEC) were divided into HUVEC blank control group, 3h hypoxia group, 6h hypoxia group. Real-time quantitative PCR (RT-PCR) and ELISA were used to determine the expression of FSTL1, TGF-β, VEGF, connective tissue growth factor (CTGF) mRNA and protein in peripheral blood and cells of all groups from all subjects.Results The expressions of FSTL1, TGF-β1, CTGF, VEGF mRNA in blood samples of patients with PDR were 1.79±0.58, 0.97±0.21, 1.85±0.69 and 1.38±0.44. The expressions of FSTL1, TGF-β1 protein were 1.19±0.50, 0.71±0.24 ng/ml and 734.03±116.45, 649.36±44.23 ng/L. Compared with normal people, the differences were statistically significant (t_mRNA=0.90, 0.21, 2.85, 1.77; P=0.00, 0.00, 0.04, 0.02. t_protein=1.88, 7.68; P=0.00, 0.02). The cell viability of HRCEC cells in the 3 h hypoxia group and the 6 h hypoxia group were 0.66±0.05 and 0.64±0.04, respectively. Compared with the blank control group, the difference was statistically significant (F=13.02, P=0.00). The cell viability of HUVEC cells in the 3 h hypoxia group and the 6 h hypoxia group were 0.63±0.06 and 0.68±0.06, respectively. Compared with the blank control group, the difference was statistically significant (F=26.52, P=0.00). Comparison of FSTL1, TGF-β1, CTGF, and VEGF mRNA expression in HRCEC blank control group and 3 h hypoxia group, the differences were statistically significant (F=14.75, 44.93, 85.54, 6.23; P=0.01, 0.00, 0.00, 0.03). Compared with the HRCEC blank control and 3 h hypoxia group, the expressions of FSTL1 and TGF-β1 protein were statistically significant (P＜0.05). There was a statistically significant difference in TGF-β1 protein expression in the hypoxic 6 h group (P=0.03) and no significant difference in FSTL1 protein expression (P=0.68). Comparison of FSTL1, TGF-β1, CTGF, and VEGF mRNA expression in HUVEC blank control group and 3h hypoxia group, the differences were statistically significant (F=19.08, 25.12, 22.89, 13.07; P=0.00, 0.00, 0.00, 0.01). Immunofluorescence staining results showed that FSTL1, TGF-β1, CTGF, and VEGF proteins were positively expressed in cells in the 3h hypoxia and 6h hypoxia groups.Conclusion The expression of FSTL1 gene and protein in serum of PDR patients was significantly higher than that of normal people.

Citation： Niu Rui, Dong Lijie, Du Xueli, He Yanhua, Nie Zetong, Cui Weina, Chen Qiong, Hu Bojie. Preliminary discussion on the potential mechanism of follistatin-like protein 1 in the process of proliferative diabetic retinopathy. Chinese Journal of Ocular Fundus Diseases, 2020, 36(3): 220-226. doi: 10.3760/cma.j.cn511434-20190115-00024 Copy

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1. Zhu X, Bai Y, Yu W, et al. The effects of pleiotrophin in proliferative diabetic retinopathy[J/OL]. PLoS One, 2015, 10(1): 0115523[2015-01-24]. 10.1371/journal.pone.0115523">https://doi.org/10.1371/journal.pone.0115523. DOI: 10.1371/journal.pone.0115523.
2. Crawford TN, Alfaro DV 3rd, Kerrison JB, et al. Diabetic retinopathy and angiogenesis[J]. Curr Diabetes Rev, 2009, 5(1): 8-13. DOI: 10.2174/157339909787314149.
3. Babapoor-Farrokhran S, Jee K, Puchner B, et al. Angiopoietin-like 4 is a potent angiogenic factor and a novel therapeutic target for patients with proliferative diabetic retinopathy[J]. Proc Natl Acad Sci USA, 2015, 112(23): 3030-3039. DOI: 10.1073/pnas.1423765112.
4. Lange CAK, Stavrakas P, Luhmann UFO, et al. Intraocular oxygen distribution in advanced proliferative diabetic retinopathy[J]. Am J Ophthalmol, 2011, 152(3): 406-412. DOI: 10.1016/j.ajo.2011.02.014.
5. Deng X, Yang Y, Sun H, et al. Analysis of whole genome-wide methylation and gene expression profiles in visceral omental adipose tissue of pregnancies with gestational diabetes mellitus[J]. J Chin Med Assoc, 2018, 81(7): 623-630. DOI: 10.1016/j.jcma.2017.06.027.
6. Shibanuma M, Mashimo J, Mita A, et al. Cloning from a mouse osteoblastic cell line of a set of transforming-growth-factor-beta 1-regulated genes, one of which seems to encode a follistatin-related polypeptide[J]. Eur J Biochem, 2010, 217(1): 13-19. DOI: 10.1111/j.1432-1033.1993.tb18212.x.
7. Zhang W, Wang W, Liu J, et al. Follistatin-like 1 protects against hypoxia-induced pulmonary hypertension in mice[J/OL]. Sci Rep, 2017, 7: 45820[2017-03-31]. https://www.nature.com/articles/srep45820. DOI: 10.1038/srep45820.
8. Yimin X, Yanxia Z, Yueqiu C, et al. Inhibition of microRNA-9-5p protects against cardiac remodeling following myocardial infarction in mice[J]. Hum Gene Ther, 2019, 30(3): 286-301. DOI: 10.1089/hum.2018.059.
9. 中华医学会眼科学会眼底病学组. 我国糖尿病视网膜病变临床诊疗指南(2014年)[J]. 中华眼科杂志, 2014, 50(11): 851-865. DOI: 10.3760/cma.j.issn.0412-4081.2014.11.014.Chinese Ocular Fundus Diseases Society, Ophthalmology Branch of Chinese Medical Association. Guidelines for clinical diagnosis and treatment of diabetic retinopathy in China[J]. Chin J Ophthalmol, 2014, 50(11): 851-865. DOI: 10.3760/cma.j.issn.0412-4081.2014.11.014.
10. 田芳, 胡博杰, 李文博, 等. 高表达多聚嘧啶序列结合蛋白相关剪接因子对糖基化终产物诱导下视网膜Müller细胞凋亡的影响[J]. 中华眼底病杂志, 2019, 35(1): 70-75. DOI: 10.3760/cma.j.issn.1005-1015.2019.01.015.Tian F, Hu BJ, Li WB, et al. Effects of polypyramidine tract binding protein-associated splicing factor overexpression on apoptosis of human Müller cells under advanced glycation end products treatment[J]. Chin J Ocul Fundus Dis, 2019, 35(1): 70-75. DOI: 10.3760/cma.j.issn.1005-1015.2019.01.015.
11. Ma F, Hu L, Yu M, et al. Emodin decreases hepatic hypoxia-inducible factor-1[formula: see text] by inhibiting its biosynthesis[J]. Am J Chin Med, 2016, 44(5): 997-1008. DOI: 10.1142/S0192415X16500555.
12. 牛瑞, 东莉洁, 马腾, 等. 结缔组织生长因子重组干扰载体慢病毒颗粒的构建及其对视网膜血管内皮细胞内源性结缔组织生长因子表达的抑制作用[J]. 中华眼底病杂志, 2018, 34(6): 580-585. DOI: 10.3760/cma.j.issn.1005-1015.2018.06.011.Niu R, Dong LJ, Ma T, et al. Construction of connective tissue growth factor recombinant interference vector lentiviral particle and its inhibitory effect on endogenous connective tissue growth factor expression in retinal vascular endothelial cells[J]. Chin J Ocul Fundus Dis, 2018, 34(6): 580-585. DOI: 10.3760/cma.j.issn.1005-1015.2018.06.011.
13. Ouchi N, Asaumi Y, Ohashi K, et al. DIP2A functions as a FSTL1 receptor[J]. J Biol Chem, 2010, 285(10): 7127-7134. DOI: 10.1074/jbc.M109.069468.
14. Hambrock HO, Kaufmann B, Müller S, et al. Structural characterization of TSC-36/Flik: analysis of two charge isoforms[J]. J Biol Chem, 2004, 279(12): 11727-11735. DOI: 10.1074/jbc.M309318200.
15. Balemans W, van Hul W. Extracellular regulation of BMP signaling in vertebrates: a cocktail of modulators[J]. Dev Biol, 2002, 250(2): 231-250. DOI: 10.1006/dbio.2002.0779.
16. Oshima Y, Ouchi N, Sato K, et al. Follistatin-like 1 is an Akt-regulated cardioprotective factor that is secreted by the heart[J]. Circulation, 2008, 117(24): 3099-3108. DOI: 10.1161/CIRCULATIONAHA.108.767673.
17. Jin N, Hatton N, Swartz DR, et al. Hypoxia activates jun-N-terminal kinase, extracellular signal-regulated protein kinase, and p38 kinase in pulmonary arteries[J]. Am J Respir Cell Mol Biol, 2000, 23(5): 593-601. DOI: 10.1165/ajrcmb.23.5.3921.
18. Liu Y, Li F, Gao F, et al. Periostin promotes tumor angiogenesis in pancreatic cancer via Erk/VEGF signaling[J]. Oncotarget, 2016, 7(26): 40148-40159. DOI: 10.18632/oncotarget.9512.
19. Wang C, Liu W, Zhang X, et al. MEK/ERK signaling is involved in the role of VEGF and IGF1 in cardiomyocyte differentiation of mouse adipose tissue-derived stromal cells[J]. Int J Cardiol, 2017, 228: 427-434. DOI: 10.1016/j.ijcard.2016.11.199.
20. Chen H, Cong Q, Du Z, et al. Sulfated fucoidan FP08S2 inhibits lung cancer cell growth in vivo by disrupting angiogenesis via targeting VEGFR2/VEGF and blocking VEGFR2/Erk/VEGF signaling[J]. Cancer Lett, 2016, 382(1): 44-52. DOI: 10.1016/j.canlet.2016.08.020.
21. Iwamoto MA, Park JE. Vascular endothelial growth factor in ocular fluid of patients with diabetic retinopathy and other retinal disorders[J]. N Engl J Med, 1994, 331(22): 1480-1487. DOI: 10.1056/NEJM199412013312203.
22. Du L, Roberts JD Jr. Transforming growth factor-β downregulates sGC subunit expression in pulmonary artery smooth muscle cells via MEK and ERK signaling[J]. Am J Physiol Lung Cell Mol Physiol, 2019, 316(1): 20-34. DOI: 10.1152/ajplung.00319.2018.
23. Yang K, Gao K, Hu G, et al. CTGF enhances resistance to 5-FU-mediating cell apoptosis through FAK/MEK/ERK signal pathway in colorectal cancer[J]. Onco Targets Ther, 2016, 9: 7285-7295. DOI: 10.2147/OTT.S108929.
24. Chen HY, Lin CH, Chen BC. ADAM17/EGFR-dependent ERK activation mediates thrombin-induced CTGF expression in human lung fibroblasts[J]. Exp Cell Res, 2018, 370(1): 39-45. DOI: 10.1016/j.yexcr.2018.06.008.
25. Chang CH, Ou TT, Yang MY, et al. Nelumbo nucifera Gaertn leaves extract inhibits the angiogenesis and metastasis of breast cancer cells by downregulation connective tissue growth factor (CTGF) mediated PI3K/AKT/ERK signaling[J]. J Ethnopharmacol, 2016, 188: 111-122. DOI: 10.1016/j.jep.2016.05.012.

Chinese Journal of Ocular Fundus Diseases

Preliminary discussion on the potential mechanism of follistatin-like protein 1 in the process of proliferative diabetic retinopathy

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