OTX008 inhibits retinal neovascularization in oxygen-induced retinopathy mice_Chinese Journal of Ocular Fundus Diseases

Authors：

Yang Ning , Zhang Wenxi , He Tao ,  Xing Yiqiao

Department of Ophthalmology, Remin Hospital of Wuhan University, Wuhan 430070, China;

Corresponding author：

Xing Yiqiao, Email: xing_yiqiao@aliyun.com

Keywords：

Retinal neovascularization/prevention & control; Galectin 1/antagonists & inhibitors; Animal experimentation

DOI：

10.3760/cma.j.issn.1005-1015.2017.02.016

Video：

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Objective To investigate the inhibitory effects and possible related mechanism of OTX008 [a selective inhibitor of galectin-1 (Galectin-1)] on retinal neovascularization (RNV) in mouse model of oxygen-induced retinopathy (OIR). Methods 7-day-old (P7) C57BL/6J mice were randomly (according to random number table) divided into 4 groups including normal group, OIR group, OIR-OTX008 group and OIR-phosphate buffered saline (PBS) group. To establish the OIR mouse model, mice from all groups except normal group were expose to (75±2)% oxygen for 5 days and then to room air. OIR-OTX008 group received an intravitreal injection of 1 μl (0.25 μg/μl) OTX008 at P12, OIR-PBS group received the equal volume (1 μl) of PBS injection. Mice from 4 groups were euthanized at P17, and retinas were collected for molecular biological analysis and morphological study. RNV was evaluated by counting the number of pre-retinal neovascular nuclei and the whole-mount immunofluorescent staining of mouse retina. Cyrosections of retinas were imaged via confocal microscopy to observe the enrichment of staining of Galectin-1. Protein levels of Galectin-1, Neuropilin-1 and phosphorylation of vascular endothelial growth factor receptor 2 (pVEGFR2) were determined with Western blot. Results At P17, Galectin-1 expressed higher in retinal ganglion cell layer, inner plexiform layer and inner nuclear layer from OIR group and OIR-PBS group than normal group. Galectin-1 expressed less in cryosection retinas from OIR-OTX008 group than OIR group and OIR-PBS group. The numbers of pre-retinal neovascular cell nuclei from OIR group and OIR-PBS group were obviously more than that from normal group (t=9.314,P＜0.05). The number of pre-retinal neovascular cell nuclei from OIR-OTX008 group were obviously lower than those from OIR group and OIR-PBS group (t=8.038, 7.774;P＜0.05). The RNV tufts area (t=13.250, 12.570), non-perfusion area (t=15.590, 12.430) and hypoxic area (t=9.542, 9.928) from OIR-OTX008 group were significantly smaller than those in OIR group and OIR-PBS group (P＜0.05). Protein levels of Galectin-1 (t=24.800, 23.060), Neuropilin-1 (t=4.120, 3.530) and pVEGFR2 (t=25.880, 15.480)　in the OIR-OTX008 group were significantly down-regulated than those from OIR group and OIR-PBS group (P＜0.05). Conclusion Intravitreal injection of OTX008 inhibits RNV and ameliorates retinal hypoxia in mice model of OIR possibly through down-regulating Galectin-1, Neurolinpin-1 and pVEGFR2.

Citation： Yang Ning, Zhang Wenxi, He Tao, Xing Yiqiao. OTX008 inhibits retinal neovascularization in oxygen-induced retinopathy mice. Chinese Journal of Ocular Fundus Diseases, 2017, 33(2): 181-185. doi: 10.3760/cma.j.issn.1005-1015.2017.02.016 Copy

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11.	Connor KM, Krah NM, Dennison RJ, et al. Quantification of oxygen-induced retinopathy in the mouse: a model of vessel loss, vessel regrowth and pathological angiogenesis [J]. Nat Protoc, 2009, 4(11): 1565-1573. DOI: 10.1038/nprot.2009.187.
12.	Smith LE, Wesolowski E, McLellan A, et al. Oxygen-induced retinopathy in the mouse [J]. Invest Ophthalmol Vis Sci, 1994, 35(1): 101-111.
13.	Hsieh SH, Ying NW, Wu MH, et al. Galectin-1, a novel ligand of neuropilin-1, activates VEGFR-2 signaling and modulates the migration of vascular endothelial cells [J]. Oncogene, 2008, 27(26): 3746-3753. DOI: 10.1038/sj.onc.1211029.

1. Yamaguchi M, Nakao S, Arita R, et al. Vascular normalization by rock inhibitor: therapeutic potential of ripasudil (K-115) eye drop in retinal angiogenesis and hypoxia [J]. Invest Ophthalmol Vis Sci, 2016, 57(4): 2264-2276. DOI: 10.1167/iovs.15-17411.
2. Chen J, Smith LE. Retinopathy of prematurity [J]. Angiogenesis, 2007, 10(2): 133-140. DOI: 10.1007/s10456-007-9066-0.
3. Bucher F, Stahl A, Agostini HT, et al. Hyperoxia causes reduced density of retinal astrocytes in the central avascular zone in the mouse model of oxygen-induced retinopathy [J]. Mol Cell Neurosci, 2013, 56: 225-233. DOI: 10.1016/j.mcn.2013.06.001.
4. Astorgues-Xerri L, Riveiro ME, Tijeras-Raballand A, et al. Unraveling galectin-1 as a novel therapeutic target for cancer [J]. Cancer Treat Rev, 2014, 40(2): 307-319. DOI: 10.1016/j.ctrv. 2013.07.007.
5. Ito K, Stannard K, Gabutero E, et al. Galectin-1 as a potent target for cancer therapy: role in the tumor microenvironment [J]. Cancer Metastasis Rev, 2012, 31(3-4): 763-778. DOI: 10.1007/s10555-012-9388-2.
6. Camby I, Le Mercier M, Lefranc F, et al. Galectin-1: a small protein with major functions [J]. Glycobiology, 2006, 16(11): 137R-157R. DOI: 10.1093/glycob/cwl025.
7. Tang D, Gao J, Wang S, et al. Cancer-associated fibroblasts promote angiogenesis in gastric cancer through galectin-1 expression [J]. Tumour Biol, 2016, 37(2): 1889-1899. DOI: 10.1007/s13277-015-3942-9.
8. Zhang P, Zhang P, Shi B, et al. Galectin-1 overexpression promotes progression and chemoresistance to cisplatin in epithelial ovarian cancer [J]. Cell Death Dis, 2014, 5: 991. DOI: 10.1038/cddis. 2013.526.
9. Croci DO, Cerliani JP, Dalotto-Moreno T, et al. Glycosylation-dependent lectin-receptor interactions preserve angiogenesis in anti-VEGF refractory tumors [J]. Cell, 2014, 156(4): 744-758. DOI: 10.1016/j.cell.2014.01.043.
10. Astorgues-Xerri L, Riveiro ME, Tijeras-Raballand A, et al. OTX008, a selective small-molecule inhibitor of galectin-1, downregulates cancer cell proliferation, invasion and tumour angiogenesis [J]. Eur J Cancer, 2014, 50(14): 2463-2477. DOI: 10.1016/j.ejca.2014.06.015.
11. Connor KM, Krah NM, Dennison RJ, et al. Quantification of oxygen-induced retinopathy in the mouse: a model of vessel loss, vessel regrowth and pathological angiogenesis [J]. Nat Protoc, 2009, 4(11): 1565-1573. DOI: 10.1038/nprot.2009.187.
12. Smith LE, Wesolowski E, McLellan A, et al. Oxygen-induced retinopathy in the mouse [J]. Invest Ophthalmol Vis Sci, 1994, 35(1): 101-111.
13. Hsieh SH, Ying NW, Wu MH, et al. Galectin-1, a novel ligand of neuropilin-1, activates VEGFR-2 signaling and modulates the migration of vascular endothelial cells [J]. Oncogene, 2008, 27(26): 3746-3753. DOI: 10.1038/sj.onc.1211029.

Chinese Journal of Ocular Fundus Diseases

OTX008 inhibits retinal neovascularization in oxygen-induced retinopathy mice

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