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Thymic medullary epithelial cell differentiation, thymocyte emigration, and the control of autoimmunity require lympho-epithelial cross talk via LTbetaR.

Boehm T, Scheu S, Pfeffer K, Bleul CC - J. Exp. Med. (2003)

Bottom Line: The nature of these signals has been elusive so far.We show that thymocytes and medullary epithelial cells (MECs) communicate via the lymphotoxin beta receptor (LTbetaR) signaling axis.Impaired lympho-epithelial cross talk in the absence of the LTbetaR causes aberrant differentiation and reduced numbers of thymic MECs, leads to the retention of mature T lymphocytes, and is associated with autoimmune phenomena, suggesting an unexpected role for LTbetaR signaling in central tolerance induction.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute for Immunobiology, Stuebeweg 51, 79108 Freiburg, Germany.

ABSTRACT
Thymocytes depend on the interaction with thymic epithelial cells for the generation of a diverse, nonautoreactive T cell repertoire. In turn, thymic epithelial cells acquire their three-dimensional cellular organization via instructive signals from developing thymocytes. The nature of these signals has been elusive so far. We show that thymocytes and medullary epithelial cells (MECs) communicate via the lymphotoxin beta receptor (LTbetaR) signaling axis. Normal differentiation of thymic MECs requires LTbetaR ligand on thymocytes and LTbetaR together with nuclear factor-kappaB-inducing kinase (Nik) in thymic epithelial cells. Impaired lympho-epithelial cross talk in the absence of the LTbetaR causes aberrant differentiation and reduced numbers of thymic MECs, leads to the retention of mature T lymphocytes, and is associated with autoimmune phenomena, suggesting an unexpected role for LTbetaR signaling in central tolerance induction.

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The absence of LTβR signaling leads to the accumulation of thymocytes with the phenotype of recent thymic emigrants. (A) FACS® analysis of thymocytes in LTβR−/− mice and heterozygous controls. Thymocytes of 12-wk-old mice were stained with the indicated antibodies and gated on lymphocytes for the top panels and on CD4 single-positive cells (CD4SP) for the middle and bottom panels. Total thymocyte numbers did not differ significantly between mutant mice and appropriate controls (1.19 ± 0.27 × 108 per LTβR−/− thymus and 1.68 ± 0.37 × 108 per LTβR+/− control thymus; 1.43 ± 0.58 × 108 per aly/aly thymus and 1.77 ± 0.46 × 108 per aly/+ control thymus). Percent of cells in each quadrant are indicated. For the bottom panel, the percent of integrin β7+ CD69− cells among CD4SP in the indicated box is given. (B) Quantification of thymocyte subsets in LTβR−/−, aly/aly, and heterozygous control mice. Thymocytes from 12-wk-old mice were stained as shown in A. Results are shown as percent of total thymocytes. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar. (C) 12-wk-old LTβR−/− mice and appropriate wild-type controls were injected with BrdU once and kept on BrdU containing drinking water for 3 d, after which the indicated thymocyte subpopulations were sorted by FACS® and stained with a FITC-labeled anti-BrdU monoclonal antibody. The percentage of cells that incorporated BrdU was determined. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar.
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fig6: The absence of LTβR signaling leads to the accumulation of thymocytes with the phenotype of recent thymic emigrants. (A) FACS® analysis of thymocytes in LTβR−/− mice and heterozygous controls. Thymocytes of 12-wk-old mice were stained with the indicated antibodies and gated on lymphocytes for the top panels and on CD4 single-positive cells (CD4SP) for the middle and bottom panels. Total thymocyte numbers did not differ significantly between mutant mice and appropriate controls (1.19 ± 0.27 × 108 per LTβR−/− thymus and 1.68 ± 0.37 × 108 per LTβR+/− control thymus; 1.43 ± 0.58 × 108 per aly/aly thymus and 1.77 ± 0.46 × 108 per aly/+ control thymus). Percent of cells in each quadrant are indicated. For the bottom panel, the percent of integrin β7+ CD69− cells among CD4SP in the indicated box is given. (B) Quantification of thymocyte subsets in LTβR−/−, aly/aly, and heterozygous control mice. Thymocytes from 12-wk-old mice were stained as shown in A. Results are shown as percent of total thymocytes. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar. (C) 12-wk-old LTβR−/− mice and appropriate wild-type controls were injected with BrdU once and kept on BrdU containing drinking water for 3 d, after which the indicated thymocyte subpopulations were sorted by FACS® and stained with a FITC-labeled anti-BrdU monoclonal antibody. The percentage of cells that incorporated BrdU was determined. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar.

Mentions: The defect causing abnormal differentiation of MECs in LTβR−/− and aly/aly mice lies in the stroma compartment. (A) Thymus sections of the indicated bone marrow chimeras were analyzed 6–8 wk after bone marrow transfer. The middle and right panels in the top row represent consecutive sections to demonstrate the presence of wild-type thymocytes. The staining reagent is indicated in parentheses. The left panels in the top and the bottom row are low magnification images of the subsequent panels. (B) UEA-1–positive MECs in LTβR−/− and wild-type mice were counted on 10 high power fields for each genotype. (C) MECs in preparations of thymic epithelial cells of single thymi from LTβR−/− and wild-type mice were identified and counted by FACS® analysis. The overall size of LTβR−/− thymi as measured by thymocyte numbers did not significantly differ from control thymi (see Fig. 6). Histogram plots are electronically gated on cells negative for CD45 and CDR1, and the marker indicates the fluorescence intensity scored as high level UEA-1 expression. MEC numbers of four individual thymi per genotype were determined. (D) The absolute number of CD45−G8.8+CDR1−B7.1+ MECs from preparations of epithelial cells of pools from five mice were determined. The gate that identifies these MECs among CD45−G8.8+ thymic epithelial cells is shown in Fig. 7. The results of three independent experiments for each genotype are shown. Error bars represent standard deviations from the mean and statistical analysis was done using Student's t test.


Thymic medullary epithelial cell differentiation, thymocyte emigration, and the control of autoimmunity require lympho-epithelial cross talk via LTbetaR.

Boehm T, Scheu S, Pfeffer K, Bleul CC - J. Exp. Med. (2003)

The absence of LTβR signaling leads to the accumulation of thymocytes with the phenotype of recent thymic emigrants. (A) FACS® analysis of thymocytes in LTβR−/− mice and heterozygous controls. Thymocytes of 12-wk-old mice were stained with the indicated antibodies and gated on lymphocytes for the top panels and on CD4 single-positive cells (CD4SP) for the middle and bottom panels. Total thymocyte numbers did not differ significantly between mutant mice and appropriate controls (1.19 ± 0.27 × 108 per LTβR−/− thymus and 1.68 ± 0.37 × 108 per LTβR+/− control thymus; 1.43 ± 0.58 × 108 per aly/aly thymus and 1.77 ± 0.46 × 108 per aly/+ control thymus). Percent of cells in each quadrant are indicated. For the bottom panel, the percent of integrin β7+ CD69− cells among CD4SP in the indicated box is given. (B) Quantification of thymocyte subsets in LTβR−/−, aly/aly, and heterozygous control mice. Thymocytes from 12-wk-old mice were stained as shown in A. Results are shown as percent of total thymocytes. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar. (C) 12-wk-old LTβR−/− mice and appropriate wild-type controls were injected with BrdU once and kept on BrdU containing drinking water for 3 d, after which the indicated thymocyte subpopulations were sorted by FACS® and stained with a FITC-labeled anti-BrdU monoclonal antibody. The percentage of cells that incorporated BrdU was determined. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar.
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Related In: Results  -  Collection

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fig6: The absence of LTβR signaling leads to the accumulation of thymocytes with the phenotype of recent thymic emigrants. (A) FACS® analysis of thymocytes in LTβR−/− mice and heterozygous controls. Thymocytes of 12-wk-old mice were stained with the indicated antibodies and gated on lymphocytes for the top panels and on CD4 single-positive cells (CD4SP) for the middle and bottom panels. Total thymocyte numbers did not differ significantly between mutant mice and appropriate controls (1.19 ± 0.27 × 108 per LTβR−/− thymus and 1.68 ± 0.37 × 108 per LTβR+/− control thymus; 1.43 ± 0.58 × 108 per aly/aly thymus and 1.77 ± 0.46 × 108 per aly/+ control thymus). Percent of cells in each quadrant are indicated. For the bottom panel, the percent of integrin β7+ CD69− cells among CD4SP in the indicated box is given. (B) Quantification of thymocyte subsets in LTβR−/−, aly/aly, and heterozygous control mice. Thymocytes from 12-wk-old mice were stained as shown in A. Results are shown as percent of total thymocytes. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar. (C) 12-wk-old LTβR−/− mice and appropriate wild-type controls were injected with BrdU once and kept on BrdU containing drinking water for 3 d, after which the indicated thymocyte subpopulations were sorted by FACS® and stained with a FITC-labeled anti-BrdU monoclonal antibody. The percentage of cells that incorporated BrdU was determined. Each dot indicates one mouse and the mean percentage is indicated by a horizontal bar.
Mentions: The defect causing abnormal differentiation of MECs in LTβR−/− and aly/aly mice lies in the stroma compartment. (A) Thymus sections of the indicated bone marrow chimeras were analyzed 6–8 wk after bone marrow transfer. The middle and right panels in the top row represent consecutive sections to demonstrate the presence of wild-type thymocytes. The staining reagent is indicated in parentheses. The left panels in the top and the bottom row are low magnification images of the subsequent panels. (B) UEA-1–positive MECs in LTβR−/− and wild-type mice were counted on 10 high power fields for each genotype. (C) MECs in preparations of thymic epithelial cells of single thymi from LTβR−/− and wild-type mice were identified and counted by FACS® analysis. The overall size of LTβR−/− thymi as measured by thymocyte numbers did not significantly differ from control thymi (see Fig. 6). Histogram plots are electronically gated on cells negative for CD45 and CDR1, and the marker indicates the fluorescence intensity scored as high level UEA-1 expression. MEC numbers of four individual thymi per genotype were determined. (D) The absolute number of CD45−G8.8+CDR1−B7.1+ MECs from preparations of epithelial cells of pools from five mice were determined. The gate that identifies these MECs among CD45−G8.8+ thymic epithelial cells is shown in Fig. 7. The results of three independent experiments for each genotype are shown. Error bars represent standard deviations from the mean and statistical analysis was done using Student's t test.

Bottom Line: The nature of these signals has been elusive so far.We show that thymocytes and medullary epithelial cells (MECs) communicate via the lymphotoxin beta receptor (LTbetaR) signaling axis.Impaired lympho-epithelial cross talk in the absence of the LTbetaR causes aberrant differentiation and reduced numbers of thymic MECs, leads to the retention of mature T lymphocytes, and is associated with autoimmune phenomena, suggesting an unexpected role for LTbetaR signaling in central tolerance induction.

View Article: PubMed Central - PubMed

Affiliation: Max Planck Institute for Immunobiology, Stuebeweg 51, 79108 Freiburg, Germany.

ABSTRACT
Thymocytes depend on the interaction with thymic epithelial cells for the generation of a diverse, nonautoreactive T cell repertoire. In turn, thymic epithelial cells acquire their three-dimensional cellular organization via instructive signals from developing thymocytes. The nature of these signals has been elusive so far. We show that thymocytes and medullary epithelial cells (MECs) communicate via the lymphotoxin beta receptor (LTbetaR) signaling axis. Normal differentiation of thymic MECs requires LTbetaR ligand on thymocytes and LTbetaR together with nuclear factor-kappaB-inducing kinase (Nik) in thymic epithelial cells. Impaired lympho-epithelial cross talk in the absence of the LTbetaR causes aberrant differentiation and reduced numbers of thymic MECs, leads to the retention of mature T lymphocytes, and is associated with autoimmune phenomena, suggesting an unexpected role for LTbetaR signaling in central tolerance induction.

Show MeSH
Related in: MedlinePlus