The inhibitory effect of small interference RNA protein kinase C-alpha on the experimental proliferative vitreoretinopathy induced by dispase in mice

Abstract

Aim:To evaluate the effects of small interference RNA protein kinase C-alpha (siRNA-PKCα) on experimental proliferative vitreoretinopathy (PVR) induced by dispase in mice. Methods: C57BL/6 mice PVR models (4-6 weeks old) were induced by intravitreal injection of dispase and then equally divided into six groups. After 1 week, the five treatment groups received 2 µL intravitreal injections of siRNA-PKCα at a concentration of 250 nM, 500 nM, 750 nM, 1000 nM, and 1500 nM, respectively, while the negative control group received 2 µL of 500 nM no-silencing siRNA. SiRNA-PKCα was transfected by a square wave electroporator. Postoperative ophthalmic observations of lens clarity and the fundus of the eyes were performed periodically. The eyeballs of the mice were enucleated and imbedded in optimal cutting temperature to perform histological and immunofluorescence analysis at the end of a 4-week observation period. Results: Four weeks after the siRNA-PKCα injections, there are 100% lens dissolution and 100% PVR in the 250 nM group and 70%, 70%, 70%, and 50% PVR in the 500 nM, 750 nM, 1000 nM, and 1500 nM groups, respectively, which is significantly different from the negative group. Abnormalities in fundus appearance were related to the concentrations of siRNA-PKCα; a higher concentration of siRNA-PKCα resulted in a more normal fundus. Histological sections by hematoxylin-eosin staining of the eyes support the clinical observation. Immunofluorescence analysis showed that RPE65, glutamine synthase, glial acidic fibrillary protein, and α-smooth muscle actin were increasing in the retina with the decreasing concentration of siRNA-PKCα, indicating that intraocular siRNA-PKCα can partly inhibit changes of markers for glia cells, fibroblast cells, retinal pigment epithelium cells, and Müller cells in the process of PVR. Conclusion: Gene therapy with siRNA-PKCα could effectively inhibit PVR in mice and provide us with a novel therapeutic target on PVR.

A Mouse Model of Proliferative Vitreoretinopathy Induced by Intravitreal Injection of Gas and RPE Cells

“…27 Currently, the most popular mouse PVR model uses injection of the proteolytic enzyme dispase. [28][29][30][31] Although dispase injection may trigger events that lead to PVR formation, such as cells invading the vitreous cavity, retinal folds, and the appearance of intravitreal membranes, 28 the integrity of the retina is severely affected. The dispase enzyme breaks down the retina structure, producing marked hemorrhage, which does not mimic human PVR pathogenesis or progression.…”

Small Molecular-Sized Artesunate Attenuates Ocular Neovascularization via VEGFR2, PKCα and PDGFR Targets

“…Downregulation of VEGFR2 might be a possible target of the anti-NV action of ART. PKCα is involved in the pathophysiological processes of DR 26 27 and was also found to promote retinal pigment epithelial (RPE) cell proliferation in proliferative vitreoretinopathy (PVR) in our previous study 28 29 . The PKCα inhibitor midostaurin (PKC412A) has been used in a phase IIA study of metastatic melanoma 30 .…”

In VitroVitamin K3Effect on Conjunctival Fibroblast Migration and Proliferation

“…Corticosteroids [ 1 – 6 ] antiproliferative agents (5-fluoro-uracil and other fluoropyrimidines, taxol, doxorubicin, mycophenolate mofetil…, alone or in combination or with different delivery systems) [ 3 – 12 ], systemic, periocular, intraocular steroidal, and nonsteroidal anti-inflammatory agents [ 5 , 13 – 16 ], colchicine [ 8 ], daunomycin [ 8 ], tissue plasminogen activator [ 17 ], heparin [ 12 , 18 – 20 ], interferon-gamma [ 21 , 22 ], calcium channel blockers [ 23 ], prolyl and lysyl hydroxylase inhibitors [ 19 , 24 – 26 ], retinoic acid [ 27 , 28 ], alpha-tocopherol [ 29 – 31 ], disintegrins [ 32 ], siRNA-PKC α [ 33 ]… are some of the useful drugs that have been used in the treatment of conditions such as proliferative vitreoretinopathy, bleb scarring after trabeculectomy, and other disorders with cell proliferation (progressive conjunctival or extraocular cicatrization).…”