Figure 1.
(+)- And (−)-phenserine induce neurotrophic actions, as assessed by increasing cellular proliferation of SH-SY5Y cells.
SH-SY5Y cells were exposed for 24 hr to increased concentrations (3–300 µM) of (+)-phenserine (A), and 30 µM concentration of (−)- and (+)-phenserine, (+)-N1-norphenserine, (+)-N8-norphenserine and (+)-N1,N8-bisnorphenserine (B). Inhibitors of PKC (GF109203X, 2.5 µM), MEK1/2 (U0126, 5 µM) or MEK1 (PD98059, 10 µM) were added to the cells 30 min before adding (+)-phenserine (C). After 24 hr, cell proliferation was determined by MTS assay. *P<0.05, **p<0.01, and ***p<0.001 compared to control samples without drug, #p<0.05 compared to samples with (+)-phenserine only (one-way ANOVA followed by Bonferronís multiple comparison post hoc test). Results are expressed as mean ± SEM. Chemical structures of (−)- and (+)-phenserine, (+)-N1-norphenserine, (+)-N8-norphenserine and (+)-N1,N8-bisnorphenserine are shown in (D).
Figure 2.
Neurotrophic actions retained in APPswe transfected and wt SH-SY5Y cells challenged with Aβ42 and H2O2.
Cells were exposed to (+)-phenserine (30 µM) for 24 hr before measuring cell proliferation using MTS assay in (A). 10 or 30 µM (+)-phenserine was added 24 hr before adding 0.1, 1 or 10 µM Aβ42 or 10 µM H2O2 before measuring cell proliferation in untransfected SH-SY5Y cells (B). **p<0.01, and ***p<0.001 compared to control samples without drug, ##p<0.01 and ###p<0.001 compared to samples with (+)-phenserine only (one-way ANOVA followed by Bonferronís multiple comparison post hoc test). Results are expressed as mean ± SEM.
Figure 3.
Phenserine and primary metabolites exert neuroprotective actions against H2O2 and glutamate-induced toxicity in SH-SY5Y cells.
Cells were exposed to 3, 10 or 30 µM (+)-phenserine 24 hr before addition of 30–100 µM H2O2 or to 100 mM glutamate in (A), or to (−)-phenserine, (+)-N1-norphenserine, (+)-N8-norphenserine or (+)-N1,N8-bisnorphenserine 24 hr before addition of 100 µM H2O2 in (B). In (C), inhibitors of PKC (GF109203X, 2.5 µM), MEK1/2 (U0126, 5 µM) or MEK1 (PD98059, 10 µM) were added to the cells 30 min before adding 30 µM (+)-phenserine. 100 µM H2O2 was added after 24 hr and after additional 24 hr incubation, cell proliferation was determined by MTS assay. *P<0.05 **p<0.01, and ***p<0.001 compared to control samples without drug, #p<0.05, ##p<0.01 and ###p<0.001 compared to samples with (+)-phenserine only, °°° compared to samples exposed to (+)-phenserine and H2O2 (one-way ANOVA followed by Bonferronís multiple comparison post hoc test). Results are expressed as mean ± SEM.
Figure 4.
Enhanced subventricular progenitor cell survival and increased size of Tg2576 mouse neurospheres after (+)-phenserine exposure.
Representative images show SVZ progenitor cells exposed to vehicle or 0.01 µM (+)-phenserine (A). Effects of the MEK1/2 inhibitor U0126, MEK1 inhibitor PD98059, and PKC inhibitor GF109203X on neurosphere size and neurosphere density in the presence of (+)-phenserine are shown in (B). ***P<0.001 compared to control samples without (+)-phenserine or inhibitor, ##P<0.01 and ###P<0.001 compared to samples with (+)-phenserine only (one-way ANOVA followed by Bonferronís multiple comparison post hoc test). The number of neurospheres composed of SVZ progenitor cells, isolated from the lateral (LGE) and medial (MGE) ganglionic eminence at embryonic day 13.5, were measured 7 days after exposure to 0.05, 0.1 and 1 µM (+)-phenserine (C). *p<0.05, **p<0.01, and ***p<0.001 compared to samples without (+)-phenserine. Data were analysed with one-way ANOVA followed by Dunnetts post hoc test. Results are expressed as mean ± SEM.
Figure 5.
Enhanced doublecortin reactivity in the SVZ of 4–6 months old APPswe mice after (+)-phenserine treatment.
Doublecortin (DCX) immunoreactivity in the subventricular zone (SVZ) in 4–6 months old mice are shown in A and B. Cortical BDNF levels in wt 4–6 months old mice are shown in C. **p<0.01 compared to wt mice treated with saline. Results are expressed as mean ± SEM.