Figure 7.

IL-5 improves the cognitive function in aged mice. (A) The Object Placement Test was performed with aged mice after injection of IL-5, IL-13, or PBS daily for 2 d. Percentage of time spent with the displaced objects was quantified. Data are from 9 or 10 mice per group and are representative of two independent experiments. (B) The Morris Water Maze test was performed with aged mice after PBS, IL-5, or IL-13 treatment for 2 d. Percentage of time spent in the target quadrant was quantified. Data are from 9 or 10 mice per group and are representative of two independent experiments. (C) IL-5 was deleted in activated ILC2/b6 cells using the CRISPR-mediated gene knockout technique. The Object Placement Test was performed with aged mice 1 wk after i.c.v injection with control or IL-5–deficient ILC2/b6 cells. Percentage of time spent with displaced objects was quantified. Data are from 9 or 10 mice per group, pooled from two independent experiments. (D) Aged mice received i.c.v injection of 10 µg anti–IL-5 antibody or isotype control, followed by i.p. injection of IL-33 or PBS daily for 2 d. The Object Placement Test was performed after 1 wk of i.c.v injection. Percentage of time spent with the displaced objects was quantified. Data are from 8–11 mice per group, pooled from two independent experiments. (E) IL-5- or PBS-treated aged mice were injected with EdU in vivo. EdU+ cells in the HC DG regions was examined and quantified by immunofluorescence at 2 d after the last dose of IL-5 or PBS treatment. Data are from four mice per group. (F) Heatmap depicts the gene expression profiles of CD3+ T cells from the brains of PBS- or IL-5–treated aged mice by RNA-Seq. (G) Numbers of CD4+ and CD8+ T cells in the brains of young (Y) or aged (A) mice treated with PBS or IL-5. Data are from four mice per group and are representative of three independent experiments. (H) Representative flow cytometry profiles of TNFα expression by CD4+ or CD8+ T cells in the brains of aged mice treated with PBS or IL-5. (I) MFI of TNFα in brain CD4+ or CD8+ T cells of aged mice treated with PBS or IL-5. Data are from three to five mice per group and are representative of three independent experiments. (J) Representative flow cytometry profiles of CD8+ cells in the brains of aged mice that were injected i.v. with anti-CD45.2 PE antibody and euthanized 3 min later. PB, pacific blue. (K) The number of noncirculating brain CD8+ T cells that were not labeled with i.v. injected anti-CD45.2 antibody. Data are from three mice per group and are representative of two independent experiments. (L) The number of noncirculating CD8+ T cells in the CP of young or aged mice treated with PBS or IL-5. Data are from four mice per group and are representative of two independent experiments. (M) The number of microglia in IL-5– or PBS-treated aged mice. Data are from three mice per group and are representative of two independent experiments. (N) Representative flow cytometry profiles of CD68 expression by microglia in IL-5– or PBS-treated aged mice. (O) MFI of CD68 on microglia of IL-5– or PBS-treated aged mice. Data are from three mice per group and are representative of two independent experiments. (P) Heatmap depicts the gene expression profiles of microglia from PBS- or IL-5–treated aged mice by RNA-Seq. (Q) CD4+ or CD8+ T cells from the brains of aged mice were sort-purified and cultured in the presence or absence of IL-5 for 24 h. Tnf expression was examined by qPCR. Data are from six mice per group and are representative of two independent experiments. (R) Expression of the indicated genes in CD8+ T cells sorted from the aged brain and cultured in the presence or absence of IL-5 for 24 h. Data are from six mice per group and are representative of two independent experiments. (S) Annexin V staining was performed at 3 d of culture. Data are from four mice per group and are representative of two independent experiments. Error bars are mean ± SEM. *, P < 0.05; **, P < 0.01. n.s., not significant.