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Quantitative impact of thymic clonal deletion on the T cell repertoire.

van Meerwijk JP, Marguerat S, Lees RK, Germain RN, Fowlkes BJ, MacDonald HR - J. Exp. Med. (1997)

Bottom Line: Interactions between major histocompatibility complex (MHC) molecules expressed on stromal cells and antigen-specific receptors on T cells shape the repertoire of mature T lymphocytes emerging from the thymus.The quantitative impact of negative selection on the potentially available repertoire is currently unknown.To address this issue, we have constructed radiation bone marrow chimeras in which MHC molecules are present on radioresistant thymic epithelial cells (to allow positive selection) but absent from radiosensitive hematopoietic elements responsible for negative selection.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, University of Lausanne, Epalinges, Switzerland.

ABSTRACT
Interactions between major histocompatibility complex (MHC) molecules expressed on stromal cells and antigen-specific receptors on T cells shape the repertoire of mature T lymphocytes emerging from the thymus. Some thymocytes with appropriate receptors are stimulated to undergo differentiation to the fully mature state (positive selection), whereas others with strongly autoreactive receptors are triggered to undergo programmed cell death before completing this differentiation process (negative selection). The quantitative impact of negative selection on the potentially available repertoire is currently unknown. To address this issue, we have constructed radiation bone marrow chimeras in which MHC molecules are present on radioresistant thymic epithelial cells (to allow positive selection) but absent from radiosensitive hematopoietic elements responsible for negative selection. In such chimeras, the number of mature thymocytes was increased by twofold as compared with appropriate control chimeras This increase in steady-state numbers of mature thymocytes was not related to proliferation, increased retention, or recirculation and was accompanied by a similar two- to threefold increase in the de novo rate of generation of mature cells. Taken together, our data indicate that half to two-thirds of the thymocytes able to undergo positive selection die before full maturation due to negative selection.

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Positive selection depends on expression of MHC molecules  on radioresistant thymic epithelial cells. (A) Thymi from wild-type→   wild-type, MHC I°II°→ I°II°, and wild-type→ I°II° chimeras were analyzed by flow cytometry 6 wk after grafting. Thymocytes were stained  with anti-CD8, anti-TCR, and anti-CD4 antibodies and analyzed on a  FACScan® using LYSYS II software. Contour plots are 75% logarithmic  and TCR histograms are from total thymus and from electronically gated  cells using the gates indicated in the figure. (B) Percentages of CD4SP  (CD4+CD8− TCRhigh) and CD8SP cells (CD4−CD8+TCRhigh) in thymi  from the indicated chimeras were determined and depicted as percentage  of an age- and sex-matched nonchimeric control mouse ± SD.
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Figure 1: Positive selection depends on expression of MHC molecules on radioresistant thymic epithelial cells. (A) Thymi from wild-type→ wild-type, MHC I°II°→ I°II°, and wild-type→ I°II° chimeras were analyzed by flow cytometry 6 wk after grafting. Thymocytes were stained with anti-CD8, anti-TCR, and anti-CD4 antibodies and analyzed on a FACScan® using LYSYS II software. Contour plots are 75% logarithmic and TCR histograms are from total thymus and from electronically gated cells using the gates indicated in the figure. (B) Percentages of CD4SP (CD4+CD8− TCRhigh) and CD8SP cells (CD4−CD8+TCRhigh) in thymi from the indicated chimeras were determined and depicted as percentage of an age- and sex-matched nonchimeric control mouse ± SD.

Mentions: Wild-type mice were C57BL/6 (H-2b haplotype) (see Fig. 1; Jackson Laboratories, Bar Harbor, ME; other experiments, Harlan Netherlands, Zeist, The Netherlands). Mice deficient in MHC class I expression (MHC I°) because of a targeted disruption of the β2-microglobulin gene (26) were obtained from Dr. B. Koller and Genpharm (Mountain View, CA). Mice of H-2b haplotype (I-Eα−) with an introduced mutation in the I-Aβb gene (27), and therefore deficient in MHC class II expression (MHC II°), were obtained from Dr. L. Glimcher and Genpharm and used in the experiment shown in Fig. 1. In all other experiments, mice deficient in MHC class II expression due to an induced disruption of the I-Aαb gene (28) in C57BL/6 stem cells (I-Eα−) were used (provided by Dr. H. Bluethmann, Basel, Switzerland). MHC I° and MHC II° animals were interbred in our facilities to obtain MHC I°II° mice. The β2-microglobulin–deficient mice, as well as the I-Aβb mutants, were crossed at least six generations to C57BL/6 mice, after which intercrossing yielded mice homozygous for the disrupted alleles.


Quantitative impact of thymic clonal deletion on the T cell repertoire.

van Meerwijk JP, Marguerat S, Lees RK, Germain RN, Fowlkes BJ, MacDonald HR - J. Exp. Med. (1997)

Positive selection depends on expression of MHC molecules  on radioresistant thymic epithelial cells. (A) Thymi from wild-type→   wild-type, MHC I°II°→ I°II°, and wild-type→ I°II° chimeras were analyzed by flow cytometry 6 wk after grafting. Thymocytes were stained  with anti-CD8, anti-TCR, and anti-CD4 antibodies and analyzed on a  FACScan® using LYSYS II software. Contour plots are 75% logarithmic  and TCR histograms are from total thymus and from electronically gated  cells using the gates indicated in the figure. (B) Percentages of CD4SP  (CD4+CD8− TCRhigh) and CD8SP cells (CD4−CD8+TCRhigh) in thymi  from the indicated chimeras were determined and depicted as percentage  of an age- and sex-matched nonchimeric control mouse ± SD.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196036&req=5

Figure 1: Positive selection depends on expression of MHC molecules on radioresistant thymic epithelial cells. (A) Thymi from wild-type→ wild-type, MHC I°II°→ I°II°, and wild-type→ I°II° chimeras were analyzed by flow cytometry 6 wk after grafting. Thymocytes were stained with anti-CD8, anti-TCR, and anti-CD4 antibodies and analyzed on a FACScan® using LYSYS II software. Contour plots are 75% logarithmic and TCR histograms are from total thymus and from electronically gated cells using the gates indicated in the figure. (B) Percentages of CD4SP (CD4+CD8− TCRhigh) and CD8SP cells (CD4−CD8+TCRhigh) in thymi from the indicated chimeras were determined and depicted as percentage of an age- and sex-matched nonchimeric control mouse ± SD.
Mentions: Wild-type mice were C57BL/6 (H-2b haplotype) (see Fig. 1; Jackson Laboratories, Bar Harbor, ME; other experiments, Harlan Netherlands, Zeist, The Netherlands). Mice deficient in MHC class I expression (MHC I°) because of a targeted disruption of the β2-microglobulin gene (26) were obtained from Dr. B. Koller and Genpharm (Mountain View, CA). Mice of H-2b haplotype (I-Eα−) with an introduced mutation in the I-Aβb gene (27), and therefore deficient in MHC class II expression (MHC II°), were obtained from Dr. L. Glimcher and Genpharm and used in the experiment shown in Fig. 1. In all other experiments, mice deficient in MHC class II expression due to an induced disruption of the I-Aαb gene (28) in C57BL/6 stem cells (I-Eα−) were used (provided by Dr. H. Bluethmann, Basel, Switzerland). MHC I° and MHC II° animals were interbred in our facilities to obtain MHC I°II° mice. The β2-microglobulin–deficient mice, as well as the I-Aβb mutants, were crossed at least six generations to C57BL/6 mice, after which intercrossing yielded mice homozygous for the disrupted alleles.

Bottom Line: Interactions between major histocompatibility complex (MHC) molecules expressed on stromal cells and antigen-specific receptors on T cells shape the repertoire of mature T lymphocytes emerging from the thymus.The quantitative impact of negative selection on the potentially available repertoire is currently unknown.To address this issue, we have constructed radiation bone marrow chimeras in which MHC molecules are present on radioresistant thymic epithelial cells (to allow positive selection) but absent from radiosensitive hematopoietic elements responsible for negative selection.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, University of Lausanne, Epalinges, Switzerland.

ABSTRACT
Interactions between major histocompatibility complex (MHC) molecules expressed on stromal cells and antigen-specific receptors on T cells shape the repertoire of mature T lymphocytes emerging from the thymus. Some thymocytes with appropriate receptors are stimulated to undergo differentiation to the fully mature state (positive selection), whereas others with strongly autoreactive receptors are triggered to undergo programmed cell death before completing this differentiation process (negative selection). The quantitative impact of negative selection on the potentially available repertoire is currently unknown. To address this issue, we have constructed radiation bone marrow chimeras in which MHC molecules are present on radioresistant thymic epithelial cells (to allow positive selection) but absent from radiosensitive hematopoietic elements responsible for negative selection. In such chimeras, the number of mature thymocytes was increased by twofold as compared with appropriate control chimeras This increase in steady-state numbers of mature thymocytes was not related to proliferation, increased retention, or recirculation and was accompanied by a similar two- to threefold increase in the de novo rate of generation of mature cells. Taken together, our data indicate that half to two-thirds of the thymocytes able to undergo positive selection die before full maturation due to negative selection.

Show MeSH
Related in: MedlinePlus