Ocular Fibrosis Group

P.I.;Keijiro Ishikawa

1. Development of molecular-targeting therapies for intraocular proliferative diseases including Diabetic retinopathy, Proliferative vitreoretinopathy and Age-related macular degeneration

Intraocular proliferative diseases such as diabetic retinopathy (DR), age-related macular degeneration (AMD) and proliferative vitreoretinopathy (PVR) are a leading cause of decreased vision and blindness in Japan. In those diseases, retinal fibro(vascular) membrane formation above and beneath the retina plays a pivotal role in the primary pathology (Figure 1).
In order to identify genes responsible for intraocular proliferation, we first determined the gene expression profiling of human retina, ERMs associated with proliferative diabetic retinopathy (PDR-ERMs), and PVR (PVR-ERMs) (Figure 2). We next determined "highly expressed genes in PDR- and in PVR-ERMs" by comparing the gene expression profiles between PDR-, PVR-ERMs and the retina. Subsequent analyses identified matricellular proteins, including periostin and tenascin C as important molecules whose expressions are enhanced specifically in proliferating ERMs compared to the retina (Ishikawa K et al. IOVS 2015).
We found increased periostin and tenascin C expression in the vitreous of patients with both PDR and PVR. Immunohistochemical analysis showed colocalization of periostin and α-SMA in PDR- and PVR-ERMs (Kobayashi Y et al. Mol Vis 2016; Ishikawa K et al. FASEB J 2014; Yoshida S et al. IOVS 2012). In vitro, both periostin and tenascin C increased proliferation, adhesion, migration and collagen production in RPE cells. Periostin blockade suppressed migration and adhesion induced by transforming growth factor-β2 (TGF-β2) and PVR vitreous. In vivo, periostin and tenascin C inhibition had the inhibitory effect on experimental retinal and choroidal fibrovascular formation, and progression of experimental PVR without affecting the viability of retinal cells (Kobayashi Y et al. Lab Invest 2016; Ishikawa K et al. FASEB J 2014). These results identified periostin and tenascin C as a pivotal molecule for fibro(vascular) formation. Thus, developing the novel antibody and/or innovative could be a potential therapeutic strategy for inhibiting the progression of intraocular proliferative diseases including DR and AMD.

2. Exploration of the underlying mechanisms in EMT of RPE；Establishment of EMT targeting therapy to prevent fibrosis associated with AMD and PVR.

In the pathogenesis of intraocular fibrosis associated with AMD and PVR, epithelial to mesenchymal transition (EMT) of RPE is one of the important steps; that is, RPE that undergo EMT acquire a fibroblast phenotype with increased capacity to proliferate and migrate and with the ability to produce ECM, which facilitate the formation of fibrotic membrane (Ishikawa K et al. Am J Pathol. 2016). Our comprehensive gene expression analyses of surgically resected human fibrous membranes associated with PDR and PVR revealed the significant EMT-related molecules (Ishikawa K et al. IOVS 2015).
We study the underlying mechanisms in EMT of RPE by investigating the expression and functions of those EMT-related molecules in human samples and/or the animal models of both pre- and subretinal fibrosis (rabbit PVR model and mouse laser-induced CNV model) by real-time PCR, ELISA, Western blot and IHC, etc (Figure 3) (Ishikawa K et al. Sci Rep. 2015; Ishikawa K et al. Exp Eye Res. 2016). Moreover, we explore the biological function and the underlying molecular pathways related to EMT in vitro and seek to establish EMT targeting therapy to prevent fibrosis associated with AMD and PVR (Figure 4).

1. Human samples or clinical information can be collected after approval by the Ethics Committee of the Kyushu University Hospital, and obtainment of informed consent for the surgery and the use of specimens from patients. The expression of the molecules can be examined using those surgically-resected specimens and vitreous samples.
2. The pathways mediated by those molecules can be further examined by in vitro assay using primary RPE cells.
3. Check the inhibitory effect of those molecules in both animal models, and determine whether the molecules can be novel therapeutic targets.

Selected publications

1. TNFRSF10A downregulation induces retinal pigment epithelium degeneration during the pathogenesis of age-related macular degeneration and central serous chorioretinopathy
   Mori K, Ishikawa K, Fukuda Y, Rui ji, Wada I, Kubo Y, Akiyama M, Notomi Shoji, Murakami Y, Nakao S, Arakawa S, Shiose Satomi, Hisatomi T, Yoshida S, Ram Kannan, Sonoda KH;
   Hum Mol Genet. 2022/02/01
2. Drainage retinotomy confers risk of epiretinal membrane formation following vitrectomy for rhegmatogenous retinal detachment repair
   Ishikawa K, Akiyama M, Mori K, Nakama T, Notomi S, Nakao S, Kohno RI,Takeda A, Sonoda KH;
   Am J Ophthalmol. 234:20-27, 2021/07/30
3. Changes in metamorphopsia after the treatand-extend regimen of anti-VEGF therapy for macular edema associated with branch retinal vein occlusion.
   Mori K, Ishikawa K, Wada I, Kubo Y, Kobayashi Y, Nakama T, Masatoshi Haruta, Akiyama M, Nakao S, Yoshida S, Sonoda KH.
   PLoS One. 15:10, 2020/10/28
4. Periostin and tenascin-C interaction promotes angiogenesis in ischemic proliferative retinopathy.
   Kubo Y, Ishikawa K, Mori K, Kobayashi Y, Nakama T, Arima M, Nakao S, Hisatomi T, Haruta M, Yoshida S, Sonoda KH.
   Sci Rep. 10(1):9299, 2020/06/09
5. Vitrectomy with peripapillary internal limiting membrane peeling for macular retinoschisis associated with normal-tension glaucoma.
   Ishikawa K, Fukui T, Nakao S, Shiose S, Sonoda KH.
   Am J Ophthalmol Case Rep. 2020 Mar 17;18:100663.
6. Increased expression of periostin and tenascin-C in eyes with neovascular glaucoma secondary to PDR.
   Ishikawa K, Kohno RI, Mori K, Murakami Y, Nakao S, Akiyama M, Yoshida S, Sonoda KH.
   Graefes Arch Clin Exp Ophthalmol. 2020 Mar ;258(3):621-628.
7. Decrease in the number of microaneurysms in diabetic macular edema after anti-vascular endothelial growth factor therapy: implications for indocyanine green angiography-guided detection of refractory microaneurysms.
   Mori K, Yoshida S, Kobayashi Y, Ishikawa K, Nakao S, Hisatomi T, Haruta M, Isihibashi T, Sonoda KH.
   Graefes Arch Clin Exp Ophthalmol. 2020 Apr;258(4):735-741.
8. Preoperative estimation of distance between retinal break and limbus with wide-field fundus imaging: Potential clinical utility for conventional scleral buckling.
   Ishikawa K, Kohno RI, Hasegawa E, Nakao S, Yoshida S, Sonoda KH.
   PLoS One. 2019 Feb 12;14(2):e0212284.
9. αB-crystallin regulates subretinal fibrosis by modulation of epithelial-mesenchymal transition.
   Ishikawa K, Sreekumar PG, Spee C, Nazari H, Zhu D, Kannan R, Hinton DR.
   Am J Pathol. 2016;186:859-73.
10. Molecular mechanisms of subretinal fibrosis in age-related macular degeneration.
    Ishikawa K, Kannan R, Hinton DR.
    Exp Eye Res. 2016;142:19-25.
11. Tenascin-C secreted by transdifferentiated retinal pigment epithelial cells promotes choroidal neovascularization via integrin alphaV.
    Kobayashi Y, Yoshida S, Zhou Y, Nakama T, Ishikawa K, Kubo Y, Arima M, Nakao S, Hisatomi T, Matsuda A, Sonoda KH, Ishibashi T.
    Lab Invest. 2016;96:1178-1188.
12. Different roles played by periostin splice variants in retinal neovascularization.
    Nakama T, Yoshida S, Ishikawa K, Kobayashi Y, Abe T, Kiyonari H, Shioi G, Katsuragi N, Ishibashi T, Morishita R, Taniyama Y.
    Exp Eye Res. 2016;153:133-140.
13. Resveratrol inhibits epithelial-mesenchymal transition of retinal pigment epithelium and development of proliferative vitreoretinopathy.
    Ishikawa K, He S, Terasaki H, Nazari H, Zhang H, Spee C, Kannan R, Hinton DR.
    Sci Rep. 2015;10;5:16386.
14. Microarray analysis of gene expression in fibrovascular membranes excised from patients with proliferative diabetic retinopathy.
    Ishikawa K, Yoshida S, Kobayashi Y, Zhou Y, Nakama T, Nakao S, Sassa Y, Oshima Y, Niiro H, Akashi K, Kono T, Ishibashi T.
    Invest Ophthalmol Vis Sci. 2015;56:932-946.
15. Periostin promotes the generation of fibrous membranes in proliferative vitreoretinopathy.
    Ishikawa K, Yoshida S, Nakao S, Nakama T, Kita T, Asato R, Sassa Y, Arita R, Miyazaki M, Enaida H, Oshima Y, Murakami N, Niiro H, Ono J, Matsuda A, Goto Y, Akashi K, Izuhara K, Kudo A, Kono T, Hafezi-Moghadam A, Ishibashi T.
    FASEB J. 2014;28:131-142.