Journal: Scientific Reports

Article Title: Deletion of p75 NTR prevents vaso-obliteration and retinal neovascularization via activation of Trk- A receptor in ischemic retinopathy model

doi: 10.1038/s41598-018-30029-0

Figure Lengend Snippet: Deletion of p75 NTR preserved hypoxia-induced VEGF expression and VEGFR2 activation. ( A ) Representative Western blotting and bar graph of retinal VEGF protein expression from WT and p75 NTR−/− pups exposed to OIR at p14. Two-way ANOVA showed significant effect of hypoxia increasing VEGF both in WT and p75 NTR−/− pups (*significant compared to WT normoxic group, p < 0.05, n = 6–12). ( B ) Representative Western blotting and bar graph of pVEGFR2 compared to its total protein VEGFR2 level. Two-way ANOVA showed marked gene deletion effect where p75 NTR−/− showed significant increase in pVEGFR2 under normoxia or hypoxia compared to WT-normoxic controls (*significant compared to WT normoxic group using, p < 0.05, n = 4–5).

Article Snippet: Membranes were blocked with 5% milk or BSA in PBS-tween and incubated overnight in 4 °C with the following primary antibodies: p75 NTR (kind gift from Dr. Bruce Carter, Department of Biochemistry, Vanderbilt University), Akt (#9272, Cell Signaling), p-Akt (#9275 S, Cell Signaling), cleaved-PARP (#5625, Cell Signaling), total PARP (#9532, Cell Signaling), cleaved caspase-3 (#9664, Cell Signaling), TrkA (#76291, Abcam), phospho-TrkA (#1445, Abcam), NGF (# AN-240, Alomone), proNGF (#ANT-005, Alomone), sortilin (#16640, Abcam), BDNF and proBDNF (SC-546, Santa Cruz), VEGF (#ABS82, Millipore), VEGFR2 (#2472, Cell Signaling), phopho-VEGFR2 (#2474, Cell Signaling), then re-probed with the primary antibodies for the house-keeping proteins; actin (#A5441, Sigma) or tubulin (#ab4074, Abcam) to confirm equal loading.

Techniques: Expressing, Activation Assay, Western Blot