Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2015 Feb 15;29(4):409–425. doi: 10.1101/gad.255331.114

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© 2015 Miyazaki et al.; Published by Cold Spring Harbor Laboratory Press

This article, published in Genes & Development, is available under a Creative Commons License (Attribution 4.0 International), as described at http://creativecommons.org/licenses/by/4.0.

PMC Copyright notice

Figure 3. — An expanding population of CXCR5⁺ cells in thymi derived from Id2^fl/flId3^fl/flIL7R^Cre mice. (A) Flow cytometric analysis of CD4 versus CD8 expression in total thymocytes and CD69 versus CD5 expression and CD69 versus TCRβ expression gated on DP cells derived from 2-wk-old Id2^fl/flId3^fl/flIL7R^Cre or littermate control mice. The graph shows the frequency of CD5⁺CD69⁻, CD69⁺TCRβ⁻, and CD69⁺TCRβ⁺ cells in the DP compartment. Numbers adjacent to the outlined areas or in quadrants indicate the percentages of cells in the population. Data are representative of one experiment with 2-wk-old mice (mean ± SD; n = 5 [Ctrl] and 3 [Id2^fl/flId3^fl/flIL7R^Cre] biological replicates). (B) The frequency of CD4SP cells (top) and the number of CXCR5⁺PD-1⁺ CD4SP cells (bottom) are plotted. Data were derived from five independent experiments (mean ± SD; n = 5 or 3 [2-wk], 3 [4-wk], and 4 [5- or 8-wk] biological replicates). (C) Intracellular staining of Ki67 in CD4SP cells derived from 8-wk-old Id2^fl/flId3^fl/flIL7R^Cre or littermate control mice. Numbers above lines indicate percentages of Ki67-expressing cells. The graph shows the frequency of Ki67-expressing cells derived from 6- to 8-wk-old Id2^fl/flId3^fl/flIL7R^Cre or littermate control mice. Data were derived from two independent experiments (mean ± SD; n = 4 biological replicates). (D) Flow cytometric analysis of BrdU incorporation and CXCR5 expression gated on CD4SP (TCRβ⁺CD25⁻) cells derived from 8-wk-old Id2^fl/flId3^fl/flIL7R^Cre or littermate control mice. Mice were injected intraperitoneally with 1 mg of PBS (BrdU⁻) or BrdU twice (24 h and 2 h prior to harvesting). Numbers in quadrants indicate BrdU-incorporated cells. The graph shows the frequency of BrdU⁺ cells in DP and CD4SP cells. Data were derived from two independent experiments (mean ± SD; n = 4 biological replicates). (E) Flow cytometric analysis of cell size (FSC) in DP and CD4SP cells derived from 5- or 8-wk-old Id2^fl/flId3^fl/flIL7R^Cre or littermate control mice. Data were derived from two (5-wk) or four (8-wk) independent experiments. (F) Representative hematoxylin and eosin (H&E) staining of thymi derived from 6-mo-old Id2^fl/flId3^fl/flIL7R^Cre or littermate control mice. Original magnification, 200×. Data are representative of three independent experiments. (G,H) RNA-seq data for the TCRα V locus (H) and TCRβ V locus (G). mRNA was isolated from sorted CXCR5⁺CD44^hi CD4 T cells from SRBC-immunized wild-type spleens (T_FH) and CD4SP thymocytes derived from 8-wk-old Id2^fl/flId3^fl/flIL7R^Cre, Id3^−/−, and Id3^+/− thymi and analyzed using RNA-seq. Numbers of reads are indicated for each of the tracks. (G,H) Two independent experiments with three mice each. (*) P < 0.05; (**) P < 0.01 (Student’s t-test).