Fig. 4. The negative selection of AQP4-specific thymocytes is dependent on AQP4-sufficient B cells in the thymus.

a, Representative cytograms of P41-10–I-A^b+CD4⁺ single-positive thymocytes in naive WT, Aqp4^−/−, Aqp4^ΔTEC, Aqp4^ΔB, Mb1-cre^KI/KI and Cd40^−/− mice (top). Bottom, corresponding fractions of FOXP3 in AQP4-specific CD4⁺ single-positive thymocytes. Data are mean ± s.d. b, Quantification of the absolute numbers of P41-10–I-A^b+ CD4⁺ single-positive thymocytes. Data are mean ± s.d. (symbols indicate biological replicates). Statistical analysis was performed using one-way ANOVA with Dunnett’s post test. c, Mixed bone marrow chimeras were generated by grafting congenically marked Rag1^−/− bone marrow engineered to retrogenically express an AQP4-specific TCR (clone 6) along with bone marrow from either WT or Aqp4^ΔB mice (4:1) into lethally irradiated Aqp4^−/− recipients and tested 6 weeks after engraftment. The diagram was created using Servier Medical Art under a Creative Commons license CC BY 3.0. d,e, Representative cytograms of the polyclonal and the retrogenic (retro; AQP4-specific) thymic compartment facing a thymic environment equipped with either WT (n = 6 biological replicates) or AQP4-deficient B cells (n = 7 biological replicates) (d) and quantification of the thymic CD4⁺ single-positive thymocyte fraction (e). Data are mean ± s.d. Statistical analysis was performed using two-way ANOVA with Sidak’s post test. f, Correlation of thymic CD45.2 B cell counts with the retrogenic AQP4-specific TCR clone 6 CD4⁺ single-positive fraction in the thymus. Statistical analysis was performed using Pearson’s R² and simple linear regression. The individual symbols represent biological replicates.

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