Figure 3.

Brain-specific SIRT1 inactivation amplifies the proliferating OPC and NSC populations. (a) Mice lacking active SIRT1 specifically in NSCs and neural progenitors during embryonic development were generated by breeding mice in which exon 4 of Sirt1 is flanked by loxP sites (Sirt1^lox/lox) with mice carrying a transgene in which Cre recombinase is expressed under the Nestin promoter (NestinCre). NestinCre;Sirt1^lox/lox and control Sirt1^{lox / lox} adult mice were injected with bromodeoxyuridine (BrdU) for 7 days to assess proliferation. SVZ, subventricular zone; LV, lateral ventricle; Sep, septum; Str, striatum; CC, corpus callosum. Schematic not drawn to scale. (b) Proliferating NG2-positive cells were observed in the striatum and septum of both NestinCre;Sirt1^lox/lox and Sirt1^lox/lox mice injected with BrdU for 7 days. Arrowheads indicate a double-labelled cell that is a dividing OPC. Scale bar, 50 μm. (c) The density of proliferating OPCs in the striatum and septum is greater in NestinCre;Sirt1^lox/lox mice when compared with control mice. Mean±s.e.m. of n = 5 Sirt1^lox/lox and n 4 = NestinCre;Sirt1^lox/lox mice. Two-tailed, unpaired Student’s t-test; striatum, *P = 0.0139; septum, P = 0.041. (d) Absence of active SIRT1 increases the percentage of NG2-positive cells in proliferating NSC and neural progenitor cultures. Primary NSC and neural progenitor cultures from Sirt1^lox/lox and NestinCre;Sirt1^lox/lox 8-week-old mice were grown for a week in proliferation medium, and then immunostained for the OPC marker NG2. Arrowheads indicate NG2-positive cells. Mean±s.e.m. of 4 independent experiments. Two-tailed, unpaired Student’s t-test, *P = 0.0209. Scale bar, 50 μm. (e) Adult mice lacking SIRT1 in the nervous system have increased numbers of proliferating cells in the SVZ. Mice were injected with ethynyldeoxyuridine (EdU) for 2 h. Mean±s.e.m. of 4 mice per genotype. Two-tailed, unpaired Student’s t-test, *P = 0.0257. (f) The density of DCX-positive proliferating neuroblasts, but not NeuN-positive newly formed neurons, in the olfactory bulb (OB) granule cell layer is greater in NestinCre;Sirt1^lox/lox mice when compared with control Sirt1^lox/lox mice. Mice were injected with BrdU daily for 7 days before analysis. Mean±s.e.m. of 3 mice per genotype. Two-tailed, unpaired Student’s t-test, ***P = 0.0009.