175163.fig.002a
175163.fig.002b
175163.fig.002c
Figure 2: Impact of fasudil on PDR vitreous-induced collagen gel contraction and MLC phosphorylation. After pretreatment with or without anti-TGF-β mAb or fasudil, hyalocytes were stimulated with vitreous with PDR. (a) In hyalocyte-containing collagen gels, fasudil almost completely suppressed the contraction of collagen gels treated with PDR vitreous. The diameter of the gels was measured and statistically analyzed (*P<.05;  **P<.01; NS, not significant, n=3 each). (b) Western blot analysis was performed to detect phosphorylated MLC (pMLC). Fasudil abolished MLC phosphorylation, induced by PDR vitreous. Lane-loading differences were normalized by MLC. Signal intensities were quantified and expressed as percentages of the pMLC/MLC ratio compared with control (*P<.05;  **P<.01,  n=3 each). Experimental PVR in rabbit eyes. (c) Therapeutic potential of fasudil in reducing the progression of experimental PVR. PVR was classified into six stages (0–5). Rhombus, vehicle (n=5); purple square, fasudil 10 μM (n=  5); trigone, fasudil 30 μM from stage 2 (n=6); blue square, fasudil 30 μM (n=5) (*P<.05,  **P<.01, not significant versus vehicle). (d) Tractional retinal detachment because of formation and cicatricial contraction of preretinal proliferative membrane was observed by stereomicroscopy in vehicle-treated eyes (stage 5 PVR). (g) In contrast, intravitreal membranes adhered to the retina without causing retinal detachment (arrowhead) in 30 μM fasudil-treated eyes with stage 2 PVR. Micrographs depict α-SMA expression (brown) in preretinal proliferative membrane with stage 5 PVR (e) and stage 2 PVR (h) by immunohistochemical analysis. (Scale bar, 200 μm). (f and i) Magnified images of (e) and (h),respectively, (Scale bar, 10 μm).