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YangKyung Cho, Hironori Uehara, Ling Luo, Bala Ambati; Anti- Angiogenesis Effect Of Morpholino Antisense Oligomer In Murine Corneal Suture Model. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4875.
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© ARVO (1962-2015); The Authors (2016-present)
VEGF receptor 1(VEGFR1) is known to be one of the main receptors for VEGF-A. VEGFR1 gene produces two kinds of isoform, soluble and membrane bound VEGFR1. By stimulation of VEGF-A, membrane bound VEGFR1 leads angiogenesis. On the other hand, soluble VEGFR1 sequestered VEGF-A activity by binding to VEGF-A, so results in anti-angiogenetic effect. These isoforms are generated by alternative splicing. Previously, we succeeded to convert membrane VEGFR1 to soluble VEGFR1 by blocking splicing event with morpholino antisense oligomer(sVEGFR1_MO, Flt morpholino) using culture cells. In this study, we examined Flt morpholino can inhibit angiogenesis in vivo murine corneal suture model
We used a murine corneal suture model to investigate the effect of morpholino promoting sFlt in suppressing neovascularization and lymphangiogenesis. In an inhibition experiment, we placed 3 corneal sutures on day 0, and then injected Flt morpholino, nonspecific(STD) morpholino, and PBS into corneal stroma of each group on day 1 and 4. On day 7, we harvested the cornea . In a regression experiment, we placed 3 corneal suture on day 0 and we injected Flt morpholino, STD morpholino, and PBS into corneal stroma of each group on day 7 and 10 and then we harvested corneas on day 14. After staining of vessels(CD31) and lymphatic vessels(LYVE1) of cornea, we checked with fluorescent microscope and digitally quantified neovascularization.
In analysis of angiogenesis(in regression experiment), we can find significant difference between groups(mean area of angiogenesis/triangular corneal area around suture is 0.262±0.182 in Flt morpholino group(N=22), 0.535±0.256 in STD morpholino group(N=29), and 0.364±0.184 in PBS group(N=28)). Flt morpholino group showed less neovascularization than STD morpholino group(p=0.000). In analysis of lymphangiogenesis(in both inhibition and regression experiment), we can find significant difference between groups(in inhibition, mean area of lymphangiogenesis/triangular corneal area around suture is 0.272±0.181 in Flt group(N=29), 0.470±0.206 in STD group(N=20), and 0.347±0.194 in PBS group(N=30)). Flt morpholino group showed less lymphangiogensis than STD morpholino group(p=0.002 in inhibition, p=0.000 in regression)
Morpholinos promoting sFlt reduced angiogenesis and lymphangiogenesis compared with nonspecific morpholino and had a tendency of reducing angiogenesis and lymphangiogenesis compared with control PBS
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