TUNEL staining in the inferior retinas after PONT.

<p>(A) The number of positive-staining cells increased significantly in the inferior retinas 1 week after PONT, compared with the normal retinas (P<0.01). There were no significant differences between normal and any other group. (B) The positive cells in the GCL of the inferior retina 1 week after PONT (red). The positive staining was located in the nuclei (C, D), which is shown with DAPI (blue). (n = 4, 3, 3, 5 and 4 in normal, 12 h, 1d, 4d and 1w groups respectively.).</p

Schematic diagrams showing the procedures for the estimation of RGC survival.

<p>Rats were fed with PBS or LBP 1 week before CONT or PONT until sacrifice. (A) In CONT experiments, CONT was performed and then a piece of gelatin soaked with FG was placed close to the ON stump to label RGCs on day 0. Rats were sacrificed 1 week or 2 weeks after surgery. (B) In PONT experiments, SC labeling was performed 1 week before PONT and rats were sacrificed 1 week or 4 weeks after surgery.</p

Western-blot analysis of the inferior retinas after PONT.

<p>Each band from one animal. (A) The expression levels of TNF-α in the inferior retinas did not change after PONT at different time-points (P>0.05). (B) MnSOD level increased significantly 1 day after PONT and had returned to normal 4 days after PONT (*P<0.05) (C) P-JNK1 level did not change after PONT (P>0.05). P-JNK2/3 levels increased significantly from 1 day until 1 week after PONT (*P<0.05). (D) The levels of p-c-jun increased from 1 day until 1 week after PONT (*P<0.05). (n = 3 in each group.).</p

Western-blot analysis of the inferior retinas after treatment with PBS or LBP.

<p>Each band is from one animal. (A) The expression of MnSOD 1 day after PONT both in PBS and LBP groups. (B, C, D, E) The expressions of p-JNK2/3, p-c-jun, BDNF, IGF-1 both 1 day and 1 week after PONT both in PBS and LBP groups. (n = 3, 3, 5 and 4 in PBS 1 day, LBP 1 day, PBS 1 week and LBP 1 week groups respectively.).</p

Effects of LBP on survival of RGCs 1 week and 2 weeks after CONT.

<p>RGCs were labeled by FG. The arrows indicate microglia which were easily distinguished from RGCs and not counted. The blue arrowheads indicate RGCs. (A, C, D) Orally feeding of 0.1 mg/kg, 1 mg/kg and 10 mg/kg LBP showed no significant effects on the survival of RGCs 1 week after CONT (compared with PBS group) and no significant difference among the three different dosages of LBP groups was detected. (A, E, F) 1 mg/kg LBP showed no significant effects on the survival of RGCs 2 weeks after CONT (compared with PBS group). (n = 10, 8, 16, 12 in PBS, 0.1 mg/kg LBP, 1 mg/kg LBP, 10 mg/kg LBP groups sacrificed 1 week after CONT and n = 7 and 6 in PBS and 1 mg/kg LBP groups sacrificed 2 weeks after CONT.).</p

Western-blot analysis of the inferior retinas after treatment with PBS or LBP.

<p>(A) LBP increased the expression of MnSOD 1 day after PONT (*P<0.05). (B, C) LBP decreased the expression of p-JNK2/3 and p-c-jun (*P<0.05). (D) LBP did not change the expression of BDNF either 1 day or 1 week after PONT (P>0.05). (E) LBP increased the expression of IGF-1 1 day after PONT (*P<0.05), but did not change the expression of IGF-1 1 week after PONT (P>0.05). (n = 3, 3, 5 and 4 in PBS 1 day, LBP 1 day, PBS 1 week and LBP 1 week groups respectively.).</p

Level of drebrin was increased in the smoking group. Brain sections were stained with anti-drebrin antibody.

<p>The CA3 region of the control (a) and smoking (b) group were shown, magnification, 100X. Western blot analysis showed that there was a trend of increase in the band intensity of drebrin but it is not statistically significant (c and d).</p

Smoking induced activation of stress kinases.

<p>The total lysate of the hippocampus of the rats were subjected to Western blotting analysis. The levels of p-ERK1/2, total ERK1/2 (a), p-JNK and total JNK (b), p-GSK3β (Ser9), p-GSK3β (pY216), p-PP2A (pY307) (e) and cleaved caspase-3 (i) were detected. α-Tubulin was used as loading control. Quantitative analysis of the band intensity of the detected kinases was shown in (b, d, f–g, and j). *P<0.05 compared to control.</p

Level of acetylated-tubulin was decreased in the hippocampus of the smoking group.

<p>Brain sections were stained with anti-acetylated-tubulin antibody. The CA1 and CA3 regions of control group (a and b), smoking group (c and d) were shown, magnification, 200X. Western blot analysis confirmed the decrease of acetylated-tubulin in the hippocampus of the smoking group (e); quantitative analysis of the band intensity was shown in (f). *P<0.05 compared to control.</p

Level of 8-OHG was elevated in rat hippocampus exposed to cigarette smoking.

<p>Brain sections were stained with anti-8-OHG antibody to detect the level of oxidative stress. Representative photos were selected from the control group (a and c) and the smoking group (b and d), magnification, 100X. An enlarged photo of the CA3 region of the smoking group was shown in (e), magnification, 400X.</p