The Effect of Scyllo-inositol on Adult Hippocampal Neurogenesis in the TgCRND8 Model of Alzheimer's Disease

I hypothesize that scyllo-inositol treatment, through the reduction of amyloid-beta peptide (Abeta) pathology, increases neurogenesis in the hippocampus of TgCRND8 mice. Adult hippocampal neurogenesis in the dentate gyrus is altered in both young (100/121-day old) TgCRND8 mice and old (200/221-day old) TgCRND8 mice. Treatment of young TgCRND8 mice with the anti-Abeta aggregation compound, scyllo-inositol, rescues alterations in cell proliferation, increases the survival and neuronal differentiation of newborn cells while reducing hippocampal Abeta pathology. scyllo-Inositol treatment of old TgCRND8 mice also reduces hippocampal Abeta pathology, but has no effect on the deficits in cell proliferation and neuronal differentiation. Reduction of hippocampal Abeta pathology in young TgCRND8 mice is sufficient to rescue alterations in neurogenesis. In contrast, the reduction of hippocampal Abeta pathology in the old TgCRND8 mice is not sufficient to rescue neurogenesis, suggesting that the neurogenic niche may be dysfunctional or damaged.MAS

Effects of Neurotrophic Support and Amyloid-Targeted Combined Therapy on Adult Hippocampal Neurogenesis in a Transgenic Model of Alzheimer's Disease

<div><p>Although it is recognized that multi-drug therapies may be necessary to combat AD, there is a paucity of preclinical proof of concept studies. We present a combination treatment paradigm, which temporally affects different aspects of Alzheimer’s disease (AD)-like pathology, specifically Aβ-toxicity and neurogenesis. At early stages of AD-like pathology, in TgCRND8 mice, we found that combating Aβ pathology with <i>scyllo</i>-inositol ameliorated deficits in neurogenesis. Older TgCRND8 mice with established amyloid load had decreased progenitor cell proliferation and survival compared to non-transgenic mice, regardless of <i>scyllo</i>-inositol treatment. The prolonged exposure to Aβ-pathology leads to deficits in the neurogenic niche, thus targeting Aβ alone is insufficient to rescue neurogenesis. To support the neurogenic niche, we combined <i>scyllo</i>-inositol treatment with leteprinim potassium (neotrofin), the latter of which stimulates neurotrophin expression. We show that the combination treatment of <i>scyllo</i>-inositol and neotrofin enhances neuronal survival and differentiation. We propose this proof of concept combination therapy of targeting Aβ-pathology and neurotrophin deficits as a potential treatment for AD.</p></div

Hippocampal cell proliferation, differentiation and survival in Tg mice with early AD-like pathology and age-matched Tg littermates.

<p>(A) A representative image of the dentate gyrus stained for BrdU, demonstrates the distribution of proliferating BrdU+ cells. (B) A representative image of BrdU (green), DCX (red) and GFAP (white) positive cells in the dentate gyrus at 100 days of age. The arrow highlights a representative BrdU+/DCX+ cell. (C) A representative image of BrdU (green), NeuN (red) and GFAP (white) positive cells in the dentate gyrus. The arrow highlights a representative BrdU+/NeuN+ cell. Cell proliferation was examined at 100 days of age (D,E) while differentiation and survival was examined at 121 days of age (F,G) as a function of <i>scyllo</i>-inositol treatment. (D) The number of proliferating BrdU+ cells in the dentate gyrus of NTg (n = 7), NTg-SI (n = 6), Tg (n = 7) and Tg-SI (n = 5) mice were compared. This demonstrates more BrdU+ cells in Tg compared to NTg and to Tg-SI. (E) The percentage of BrdU+/DCX+ cells in the dentate gyrus showed less neuronal BrdU+ cells in Tg compared to the other three cohorts. (F) The number of BrdU+ cells surviving in NTg (n = 6), NTg-SI (n = 5), Tg (n = 6) and Tg-SI (n = 6) mice demonstrates more cell survival in Tg-SI mice. (G) The percentage of BrdU+/NeuN+ cells shows no difference between cohorts. Scale bars indicate 100 μm (A) or 25 μm (B,C). Data are mean ± SEM. One-way ANOVA with Fisher’s Post-hoc test, * represents p< 0.05 and ** represents p< 0.01.</p

Phenotypic analysis of CR+ cells within the granular cell layer and subgranular zone of the dentate gyrus in 200 day old Tg mice treated with <i>scyllo</i>-inositol, neotrofin and <i>scyllo</i>-inositol/neotrofin combination.

<p>(A) Representative image of the dentate gyrus stained for CR (red), DCX (green) and NeuN (blue). Arrow indicates a CR+/DCX+/NeuN- cell. Arrowhead indicates a CR+/DCX+/NeuN+ cell. (B) Orthogonal projection of CR+/DCX+/NeuN- immature neurons. (C) Orthogonal projection CR+/DCX+/NeuN+ immature neurons. (D) The number of CR+ cells in Tg-SI (n = 6), Tg-NEO (n = 6) and Tg-SI/NEO (n = 7) mice demonstrated no differences between cohorts. (E) Graphical representation of the changes in the percentage of CR+/DCX-/NeuN-, CR+/DCX+/NeuN-, CR+/DCX+/NeuN+ and CR+/DCX-/NeuN+ out of total CR+ cells in Tg-SI, Tg-NEO and Tg-SI/NEO mice. Scale bar indicates 100 μm (A) or 20 μm (B/C). Data are mean ± SEM (D) and mean (E). One-way ANOVA with Fisher’s Post-hoc test.</p

Hippocampal cell proliferation in <i>scyllo</i>-inositol, neotrofin, and <i>scyllo</i>-inositol/neotrofin treated 200 day old Tg mice.

<p>(A) The number of proliferating BrdU+ cells in Tg-SI (n = 7), Tg-NEO (n = 6) and Tg-SI/NEO (n = 7) mice demonstrated no difference between cohorts. (B) The percentage of BrdU+/DCX+ cells (Tg-SI (n = 5), Tg-NEO (n = 6) and Tg-SI/NEO (n = 7)]) and (C) BrdU+/GFAP+ (Tg-SI (n = 7), Tg-NEO (n = 6) and Tg-SI/NEO (n = 7)) in the dentate gyrus of all groups were assessed and showed no changes between treatment groups. (D) Representative image of the dentate gyrus stained with BrdU (red) and GFAP (green) demonstrate the distribution of BrdU+ proliferating cells and astrocytes. (E) Representative BrdU+/GFAP+ cells are indicated by arrows. Scale bar indicates 100 μm (D) or 25 μm (E). Data are mean ± SEM. One-way ANOVA with Fisher’s Post-hoc test.</p

Phenotypic characterization of total CR cells in the subgranular zone and granular cell layer of the dentate gyrus in Tg mice treated with <i>scyllo</i>-inositol, neotrofin or <i>scyllo</i>-inositol/neotrofin.

<p>Phenotypic characterization of total CR cells in the subgranular zone and granular cell layer of the dentate gyrus in Tg mice treated with <i>scyllo</i>-inositol, neotrofin or <i>scyllo</i>-inositol/neotrofin.</p

Timeline of therapeutic interventions.

<p>(A) TgCRND8 (Tg) mice with early AD-like pathology and age-matched non-transgenic (NTg) mice were treated or untreated from 72 days of age until sacrifice at either 100 or 121 days, for proliferation and differentiation/survival, respectively. BrdU was intraperitoneally injected in these cohorts of mice for 5 days from day 96–100. (B) Tg mice with late AD-like pathology and age-matched NTg mice were treated or untreated from day 172 days of age until sacrifice at either 200 or 221 days. Tg mice were treated with <i>scyllo</i>-inositol (Tg-SI), neotrofin (Tg-NEO) or a combination of both (Tg-SI/NEO) for 28 days. BrdU was injected in these cohorts of mice for 5 days from day 196–200.</p

Hippocampal cell differentiation and survival in <i>scyllo</i>-inositol, neotrofin, and <i>scyllo</i>-inositol/neotrofin treated 200 day old Tg mice.

<p>(A) The number of DCX+ cells in Tg-SI (n = 6), Tg-NEO (n = 6) and Tg-SI/NEO (n = 7) mice were not significantly different between cohorts. (B) The percentage of DCX+ cells that were DCX+/CR+ immature neurons were assessed and showed no difference between treatment groups. (C) The percentage of DCX+/CR+ cells that were NeuN+ represents immature neurons approaching maturity. Tg-SI/NEO mice had a significantly greater percentage of DCX+/CR+/NeuN+ cells than Tg-SI and Tg-NEO mice. Tg-SI mice had a greater percentage than Tg-NEO mice. (D/E) Representative images of DCX (green) and CR (red) positive cells in the hippocampus, showing DCX+/CR- and DCX+/CR+ cells in Tg-SI mice and in Tg-SI/NEO mice. (E) Arrows indicate DCX+/CR+ cells. Scale bar indicates 20 μm. Data are mean ± SEM. One-way ANOVA with Fisher’s Post-hoc test, *** represents p<0.001.</p

Hippocampal cell proliferation in Tg mice with late AD-like pathology and age-matched NTg littermates.

<p>Cell proliferation was assessed at 200 days of age (A,B). (A) The number of proliferating BrdU+ cells in NTg (n = 6), NTg-SI (n = 6), Tg (n = 6) and Tg-SI (n = 5) mice was compared, demonstrating less proliferation in Tg and Tg-SI vs. NTg-SI. (B) The percentage of BrdU+/DCX+ cells in the dentate gyrus was not different between cohorts. (C) Dentate gyrus stained with BrdU (red) and DCX (green) demonstrating the distribution of proliferating cells, as well as neuroblasts and immature neurons, respectively. (D) Representative orthogonal projection of a BrdU+/DCX+ cell. Scale bar indicates 100 μm (C) or 25 μm (D). Data are mean ± SEM. One-way ANOVA with Fisher’s Post-hoc test, ** represents p< 0.01.</p

Hippocampal Aβ plaques in 121 and 221 day old <i>scyllo</i>-inositol treated and untreated Tg mice.

<p>(A) Percent hippocampal area covered in plaques in 121 day old Tg (n = 5) and Tg-SI (n = 6) mice showed decreased plaques after treatment. (B) Representative images of Aβ plaques in the hippocampus of 121 day old mice demonstrate the reduction in plaques after <i>scyllo</i>-inositol treatment. (C) Percent hippocampal area covered in plaques in 221 day old Tg (n = 5) and Tg-SI (n = 6) mice also showed less plaques after treatment. (D) Representative images of Aβ plaques in the hippocampus of 221 day old mice demonstrate the reduction in plaques after <i>scyllo</i>-inositol treatment. Scale bar indicates 200 μm. Data are mean ± SEM. Unpaired t-test, * represents p< 0.05.</p