Neurogenic and Neurotrophic Effects of BDNF Peptides in Mouse Hippocampal Primary Neuronal Cell Cultures
Figure 5
(A) BDNF peptides B-5, B-4 and B-3 induce the expression of BDNF.
Western blot analyses of cells treated with peptides B-5, B-4 and B-3, or with BDNF, or vehicle for 5 days, showed an increase in BDNF expression in cells treated with the peptides. A sample of adult mouse brain was included as a control for the migration of the bands corresponding to pro-BDNF and BDNF. (B) Densitometric quantitation of the Western blots developed with anti-BDNF. Data was normalized to GAPDH as loading control and then to 5 days control vehicle treated cells, C 5d. (C) Peptides B-5 and B-3 activated TrkB receptor in primary E18 hippocampal cells. Western blots showing phosphorylation of TrkB at Tyr 706 on treatment with Peptide B-5 (1 µM), Peptide B-3 (1 µM) or BDNF (20 ng/ml, 0.79 nM) for 1 h as compared to control treated cells, C. Lower blots show the levels of TrkB receptor and the levels of GAPDH as a loading control. (D) Densitometric analysis of the Western-blots for pTrkB normalized to TrkB, and TrkB normalized to GAPDH. Control was taken as 100 percent in each case. (E) BDNF peptides (B-5 and B-3) increased the expression of TrkB. Increase in expression of TrkB in TrkB stably-expressing NIH-3T3 fibroblast cells, as a function of time. Cells were treated for 5, 15 or 60 min with B-5 (1 µM), B-3 (1 µM), BDNF (20 ng/ml), or vehicle only (Control, C). Western blots of total TrkB, and GAPDH included as a loading control. (F) Densitometric quantitation of the Western blots for TrkB normalized to GAPDH. Data are shown as mean ± standard deviation, n = 3. *p<0.05, **p<0.01, ***p<0.001. One-way ANOVA/post-hoc test/Student’s t test. Dashed line in (D) denotes that B-5 induction of TrkB expression almost approaches significance (p = 0.057, one-way ANOVA).