Figure 2. Network analysis and extracellular signal regulating kinase (Erk) phosphorylation upon Dock5 inhibition. A: A network analysis of the affected gene expressions in the lens epithelial cells (LECs) of rupture of lens cataract (RLC) mice was performed using Ingenuity Pathway Analysis (IPA) software. In the highest-scored network in the LECs from the RLC mice, “Antimicrobial Response, Inflammatory Response, Dermatological Diseases and Conditions,” the protein–protein associations are shown. Nodes are displayed using various shapes that indicate the functional class of the genes: squares, growth factors and cytokines; double circles, group or complex; inverted triangles, kinases; triangles, phosphatases; vertical diamonds, enzymes; horizontal diamonds, peptidases; trapezoids, transporters; vertical ellipses, transmembrane receptors; horizontal ellipses, transcription regulators; and circles, other molecules. The color density of each node indicates the degree of upregulation (red) or downregulation (green) of the respective gene expression. Edges indicate the relationship between the nodes. Arrowheads denote the directionality of the interaction. Perpendicular bars denote an inhibitory interaction. Edges with solid lines indicate a direct interaction, and those with dotted lines indicate an indirect interaction. B: The eyes from 21-day-old wild-type (WT; C57BL/6) mice were used for the immunostaining of phosphorylated Erk (pErk). C,D: Higher magnification of lenses at the equator from a 21-day-old RLC (C) and Dock5-knockout (KO) (D) mouse and corresponding WT mice (C, BALB/c; D, C57BL/6). The photographs are representative images from three different mice. E,F: pErk was quantified as described in the Methods, Results, and Discussion sections. Three mice, that is, six lenses (indicated as black circles, squares, and triangles), were analyzed. In (E), the red and green bars denote the average and standard error of the mean (SEM). In (F), the pErk positivity is compared in the paired mice. G: Madin–Darby canine kidney (MDCK) cells were cultured on a plastic dish and treated with or without N-(3,5-dichlorophenyl) benzenesulfonamide (C21) and dimethyl sulfoxide (DMSO) for 48 h. Cells were washed and subjected to SDS–PAGE and western blotting, with the antibodies indicated at the bottom. H: The intensity of the corresponding bands was quantified, and the value of pErk/totak Erk was normalized by the value of the cells without drug treatment (denoted as “none”). The graph shows the averages from six independent experiments with the standard error of the mean (SEM), and the p value (0.0011) was calculated using the t test. The average ± SEM values were 5.83 ± 1.05 for C21 and 1.08 ± 0.13 for DMSO.