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Targeted expression of major histocompatibility complex (MHC) class II molecules demonstrates that dendritic cells can induce negative but not positive selection of thymocytes in vivo.

Brocker T, Riedinger M, Karjalainen K - J. Exp. Med. (1997)

Bottom Line: Using the CD 11c promoter we expressed MHC class II I-E molecules specifically on DC of all tissues, but not on other cell types.In contrast, it only DC expressed I-E in a class II-deficient background, positive selection of CD4+ T cells could not be observed.Thus negative, but not positive, selection events can be induced by DC in vivo.

View Article: PubMed Central - PubMed

Affiliation: Basel Institute for Immunology, Switzerland.

ABSTRACT
It is well established that lymphoid dendritic cells (DC) play an important role in the immune system. Beside their role as potent inducers of primary T cell responses, DC seem to play a crucial part as major histocompatibility complex (MHC) class II+ "interdigitating cells" in the thymus during thymocyte development. Thymic DC have been implicated in tolerance induction and also by some authors in inducing major histocompatibility complex restriction of thymocytes. Most of our knowledge about thymic DC was obtained using highly invasive and manipulatory experimental protocols such as thymus reaggregation cultures, suspension cultures, thymus grafting, and bone marrow reconstitution experiments. The DC used in those studies had to go through extensive isolation procedures or were cultured with recombinant growth factors. Since the functions of DC after these in vitro manipulations have been reported to be not identical to those of DC in vivo, we intended to establish a system that would allow us to investigate DC function avoiding artificial interferences due to handling. Here we present a transgenic mouse model in which we targeted gene expression specifically to DC. Using the CD 11c promoter we expressed MHC class II I-E molecules specifically on DC of all tissues, but not on other cell types. We report that I-E expression on thymic DC is sufficient to negatively select I-E reactive CD4+ T cells, and to a less complete extent, CD8+ T cells. In contrast, it only DC expressed I-E in a class II-deficient background, positive selection of CD4+ T cells could not be observed. Thus negative, but not positive, selection events can be induced by DC in vivo.

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The majority of peritoneal lavage cells do not express the  CD11c-Eαd transgene. Peritoneal washes of the mice indicated were performed 5 d after an initial intraperitoneal injection of 2 ml 3% thioglycollate. The isolated cells were incubated for 48 h in IFN-γ–containing medium (1,000 U/ml) to induce MHC class II expression, and then stained  with mAbs specific for I-E (PE) and I-A (FITC). Shown are all cells from  the cultures with the gates set on live cells only. The percentages of I-E  positive cells in B6CD11c-Eαd mice varied from 2–7% in the four experiments done.
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Figure 6: The majority of peritoneal lavage cells do not express the CD11c-Eαd transgene. Peritoneal washes of the mice indicated were performed 5 d after an initial intraperitoneal injection of 2 ml 3% thioglycollate. The isolated cells were incubated for 48 h in IFN-γ–containing medium (1,000 U/ml) to induce MHC class II expression, and then stained with mAbs specific for I-E (PE) and I-A (FITC). Shown are all cells from the cultures with the gates set on live cells only. The percentages of I-E positive cells in B6CD11c-Eαd mice varied from 2–7% in the four experiments done.

Mentions: Further, when we analyzed peritoneal macrophages after IFN-γ treatment for class II expression, only 2–7% of the cells in these preparations showed I-E transgene expression, while the majority of macrophages were I-E–negative like the B6 control (Fig. 6, B6, B6CD11c-Eαd). Analysis of these I-E–positive cells showed that they were Mac-1+, Fcγ III/II receptor+, B220−, CD19−, and CD5− (data not shown). This indicates that, most likely, some peritoneal macrophages, but not peritoneal B cells, partially express the I-E transgene. Since macrophages are equally distributed in thymic cortex where we could not detect I-E staining in the B6CD11c-Eαd mice (see Fig. 2 e) and medulla (36), the cells expressing the I-E transgene in the thymus of the B6CD11c-Eαd transgenic animals are most likely thymic DC.


Targeted expression of major histocompatibility complex (MHC) class II molecules demonstrates that dendritic cells can induce negative but not positive selection of thymocytes in vivo.

Brocker T, Riedinger M, Karjalainen K - J. Exp. Med. (1997)

The majority of peritoneal lavage cells do not express the  CD11c-Eαd transgene. Peritoneal washes of the mice indicated were performed 5 d after an initial intraperitoneal injection of 2 ml 3% thioglycollate. The isolated cells were incubated for 48 h in IFN-γ–containing medium (1,000 U/ml) to induce MHC class II expression, and then stained  with mAbs specific for I-E (PE) and I-A (FITC). Shown are all cells from  the cultures with the gates set on live cells only. The percentages of I-E  positive cells in B6CD11c-Eαd mice varied from 2–7% in the four experiments done.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2196043&req=5

Figure 6: The majority of peritoneal lavage cells do not express the CD11c-Eαd transgene. Peritoneal washes of the mice indicated were performed 5 d after an initial intraperitoneal injection of 2 ml 3% thioglycollate. The isolated cells were incubated for 48 h in IFN-γ–containing medium (1,000 U/ml) to induce MHC class II expression, and then stained with mAbs specific for I-E (PE) and I-A (FITC). Shown are all cells from the cultures with the gates set on live cells only. The percentages of I-E positive cells in B6CD11c-Eαd mice varied from 2–7% in the four experiments done.
Mentions: Further, when we analyzed peritoneal macrophages after IFN-γ treatment for class II expression, only 2–7% of the cells in these preparations showed I-E transgene expression, while the majority of macrophages were I-E–negative like the B6 control (Fig. 6, B6, B6CD11c-Eαd). Analysis of these I-E–positive cells showed that they were Mac-1+, Fcγ III/II receptor+, B220−, CD19−, and CD5− (data not shown). This indicates that, most likely, some peritoneal macrophages, but not peritoneal B cells, partially express the I-E transgene. Since macrophages are equally distributed in thymic cortex where we could not detect I-E staining in the B6CD11c-Eαd mice (see Fig. 2 e) and medulla (36), the cells expressing the I-E transgene in the thymus of the B6CD11c-Eαd transgenic animals are most likely thymic DC.

Bottom Line: Using the CD 11c promoter we expressed MHC class II I-E molecules specifically on DC of all tissues, but not on other cell types.In contrast, it only DC expressed I-E in a class II-deficient background, positive selection of CD4+ T cells could not be observed.Thus negative, but not positive, selection events can be induced by DC in vivo.

View Article: PubMed Central - PubMed

Affiliation: Basel Institute for Immunology, Switzerland.

ABSTRACT
It is well established that lymphoid dendritic cells (DC) play an important role in the immune system. Beside their role as potent inducers of primary T cell responses, DC seem to play a crucial part as major histocompatibility complex (MHC) class II+ "interdigitating cells" in the thymus during thymocyte development. Thymic DC have been implicated in tolerance induction and also by some authors in inducing major histocompatibility complex restriction of thymocytes. Most of our knowledge about thymic DC was obtained using highly invasive and manipulatory experimental protocols such as thymus reaggregation cultures, suspension cultures, thymus grafting, and bone marrow reconstitution experiments. The DC used in those studies had to go through extensive isolation procedures or were cultured with recombinant growth factors. Since the functions of DC after these in vitro manipulations have been reported to be not identical to those of DC in vivo, we intended to establish a system that would allow us to investigate DC function avoiding artificial interferences due to handling. Here we present a transgenic mouse model in which we targeted gene expression specifically to DC. Using the CD 11c promoter we expressed MHC class II I-E molecules specifically on DC of all tissues, but not on other cell types. We report that I-E expression on thymic DC is sufficient to negatively select I-E reactive CD4+ T cells, and to a less complete extent, CD8+ T cells. In contrast, it only DC expressed I-E in a class II-deficient background, positive selection of CD4+ T cells could not be observed. Thus negative, but not positive, selection events can be induced by DC in vivo.

Show MeSH