Figure 5. Activation of the HGF/c-Met pathway in RPEΔMT mice is critical for RPE cell survival.

(A) Immunoblot detects increased HGF and phosphorylated c-Met (Y1234/1235) in RPE cells from pigmented RPEΔMT mice at 14 weeks, compared with that in control. (B–E) Original magnification, ×400. Immunostaining of retinal sections shows increased reactivity of HGF (C) and p-c-Met (Y1234/1235) (E) in 22-week-old albino RPEΔMT mice. (F) Immuno­blot shows diminished p-c-Met (Y1234/1235) reactivity in 26-week-old albino RPEΔMT mice at 2 days PI of PHA-665752, a selective c-Met inhibitor. (G) Quantification of total remaining RPE cells at 2 days PI (normalized by the number of cells in vehicle-injected Tfam^loxp/loxp control mice) reveals a significant loss of RPE cells in RPEΔMT mice treated with PHA-665752 (PHA) (n = 3) versus RPEΔMT mice treated with vehicle (Veh) (n = 3). ^§P < 0.01. (H and I) Immunostaining detects activated caspase-9–reactive RPE cells (green) in PHA-665752–injected RPEΔMT mice (I, arrows; inset is higher magnification image from another area). (J and K) Staining for phalloidin and cre on RPE flat mounts. (K) Z-scanning images (20-μm stack) show large areas of atrophic RPE with attenuated phalloidin staining and few, condensed cre-expressing RPE cells (arrows) in PHA-665752–treated RPEΔMT mice. (J) No effect is observed in vehicle-treated RPEΔMT mice. (H–K) Original magnification, ×200. (L and M) Light microscopy shows a denuded Bruch’s membrane and tiny pyknotic RPE cells (arrows) in PHA-665752–treated RPEΔMT mice (M), whereas RPE remains thickened and intact in vehicle-treated RPEΔMT mice (L). Dotted red lines in L and M denote the approximate edge of the outer segment layer. Original magnification, ×630.