Figure 2: Tet2 regulates the balance of aNSC proliferation and differentiation both in vitro and in vivo.

(a–d) Representative BrdU staining in both WT (a) and Tet2^−/− (c) aNSCs to access their proliferation states. Tet2 depletion promoted aNSC proliferation. (b,d) Overlay between BrdU and DAPI staining. Scale bar, 50 μm. n=3. (e) Quantification of BrdU⁺ aNSCs from WT and Tet2^−/− mice in a–d indicated that depletion of Tet2 significantly promoted aNSC proliferation (n=3; unpaired t-test, P<0.05). (f–m) Representative IF staining in WT and Tet2^−/− differentiated neural cells from isolated in vivo aNSCs with neuronal cell marker Tuj1 (f,j) and glial marker GFAP (g,k). The loss of Tet2 leads to a decrease of Tuj1⁺ neurons and GFAP⁺ glial cells. DAPI staining (h,l) and merged overlay (i,m) are indicated. Scale bar, 50 μm. n=4. (n,o) Quantification of Tuj1⁺ cells (n) and GFAP⁺ glial cells (o) in WT and Tet2^−/− neural cells differentiated from aNSCs (n=4; unpaired t-test, P<0.01 or 0.05). Fewer Tuj1⁺ neuronal cells and GFAP⁺ glial cells were found Tet2^−/−. (p) A diagram shows the experimental scheme to investigate adult neurogenesis in WT and Tet2^−/− mice in vivo. WT and Tet2^−/− mice were administered BrdU and killed at 1 or 4 weeks afterwards to assess the aNSC proliferation and differentiation states in the presence or absence of Tet2. (q) Quantification of BrdU⁺ cells in the hippocampus of WT and Tet2^−/− mice killed 7 days after final BrdU administration. Depletion of Tet2 resulted in a significant increase of aNSC proliferation (WT, n=4; Tet2^−/−, n=3; unpaired t-test, P<0.05). (r) Quantification of BrdU⁺ cells in the hippocampus of WT and Tet2^−/− mice 4 weeks after the final BrdU administration. Depletion of Tet2 resulted in a significant increase of aNSC proliferation (n=5; unpaired t-test, P<0.05). (s) Quantification of BrdU⁺/NeuN⁺ cells in the hippocampus of WT and Tet2^−/− mice 4 weeks after the final BrdU administration (n=4; unpaired t-test, *P<0.05). (t) Quantification of the percentage of BrdU⁺/NeuN⁺ cells among total BrdU⁺ cells in the hippocampus of WT and Tet2^−/− mice 4 weeks after the final BrdU administration (n=4,; unpaired t-test, *P<0.05).