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Modulation of Aire regulates the expression of tissue-restricted antigens.

Kont V, Laan M, Kisand K, Merits A, Scott HS, Peterson P - Mol. Immunol. (2007)

Bottom Line: Aire had an allele dose-dependent effect on TRA expression in the thymuses of mice from two strains, C57BL/6J and Balb/c, but had no effect on TRA expression in the lymph nodes.By manipulating in vitro organ-cultures we showed that thymic microenvironment plays a dominant role in Aire expression whereas TRAs follow the same pattern.The data underline a direct role for Aire in TRA expression and suggest that modulation of Aire has a potential to control central tolerance and autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Molecular Pathology, Biomedicum, Tartu University, Ravila 19, 50411 Tartu, Estonia.

ABSTRACT
Intrathymic expression of tissue-restricted antigens (TRAs) has been viewed as the key element in the induction of central tolerance and recently, a central role for the autoimmune regulator (Aire) has been suggested in this process. The aim of this study was to establish whether down or up-regulation of Aire leads to alterations in TRA expression and whether this is limited to thymic epithelial cells. This study also characterized whether TRAs follow Aire expression during normal development, and whether thymic microenvironment plays a role in the expression of Aire and TRAs. We did several in vivo and in vitro experiments to manipulate Aire expression and measured expression of four TRAs (Trefoil factor-3, Insulin-2, Major urinary protein-1 and Salivary protein-1) by real-time RT-PCR. Aire had an allele dose-dependent effect on TRA expression in the thymuses of mice from two strains, C57BL/6J and Balb/c, but had no effect on TRA expression in the lymph nodes. In the thymus, Aire and TRAs were both localized in the medulla and were co-expressed during normal development and involution. In the primary stromal cells as well as thymic epithelial cell line, the adenoviral over-expression of Aire resulted in an increase in TRA expression. By manipulating in vitro organ-cultures we showed that thymic microenvironment plays a dominant role in Aire expression whereas TRAs follow the same pattern. The data underline a direct role for Aire in TRA expression and suggest that modulation of Aire has a potential to control central tolerance and autoimmunity.

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TRA expression in mTEC vs. cTEC populations of WT and Aire KO mice. Thymuses were stained with anti-EpCAM and anti-Aire antibodies and analyzed by immunofluorescent microscopy (A) or were enzyme digested and FACS-sorted according to the expression of EpCAM and analyzed for the expression level of TRAs by real-time PCR (B). Medullary compartment of thymus was distinctly characterized by high-EpCAM expression and by the presence of Aire-positive cells. TRAs were highly expressed in the thymic medulla but not in cortex of the WT mice. Aire KO mice showed virtually no expression of TRAs either in medulla or cortex. Data in (B) are mean with S.E.M. of triplicate measurements of one out of two representative experiments.
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fig3: TRA expression in mTEC vs. cTEC populations of WT and Aire KO mice. Thymuses were stained with anti-EpCAM and anti-Aire antibodies and analyzed by immunofluorescent microscopy (A) or were enzyme digested and FACS-sorted according to the expression of EpCAM and analyzed for the expression level of TRAs by real-time PCR (B). Medullary compartment of thymus was distinctly characterized by high-EpCAM expression and by the presence of Aire-positive cells. TRAs were highly expressed in the thymic medulla but not in cortex of the WT mice. Aire KO mice showed virtually no expression of TRAs either in medulla or cortex. Data in (B) are mean with S.E.M. of triplicate measurements of one out of two representative experiments.

Mentions: In order to establish whether Aire co-localizes in the thymus with TRAs, we purified the thymic mTEC based on the cell-surface marker EpCAM (Fig. 3A) and analyzed the expression of the TRA genes. As seen in Fig. 3B, the expression of the Tff3, Ins2, Mup1 and Spt1 antigens was limited to the mTEC population, i.e. the cell population of Aire expression. The cTEC population showed a very low expression for all four TRAs both in the WT as well as Aire KO mouse. Collectively these data show that Aire dose-dependently regulates TRA expression in thymus but not in the lymph nodes, and confirms by real-time PCR the previously published microarray data, suggesting that both Aire and TRAs are predominantly expressed in thymus medullary epithelium.


Modulation of Aire regulates the expression of tissue-restricted antigens.

Kont V, Laan M, Kisand K, Merits A, Scott HS, Peterson P - Mol. Immunol. (2007)

TRA expression in mTEC vs. cTEC populations of WT and Aire KO mice. Thymuses were stained with anti-EpCAM and anti-Aire antibodies and analyzed by immunofluorescent microscopy (A) or were enzyme digested and FACS-sorted according to the expression of EpCAM and analyzed for the expression level of TRAs by real-time PCR (B). Medullary compartment of thymus was distinctly characterized by high-EpCAM expression and by the presence of Aire-positive cells. TRAs were highly expressed in the thymic medulla but not in cortex of the WT mice. Aire KO mice showed virtually no expression of TRAs either in medulla or cortex. Data in (B) are mean with S.E.M. of triplicate measurements of one out of two representative experiments.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC1994210&req=5

fig3: TRA expression in mTEC vs. cTEC populations of WT and Aire KO mice. Thymuses were stained with anti-EpCAM and anti-Aire antibodies and analyzed by immunofluorescent microscopy (A) or were enzyme digested and FACS-sorted according to the expression of EpCAM and analyzed for the expression level of TRAs by real-time PCR (B). Medullary compartment of thymus was distinctly characterized by high-EpCAM expression and by the presence of Aire-positive cells. TRAs were highly expressed in the thymic medulla but not in cortex of the WT mice. Aire KO mice showed virtually no expression of TRAs either in medulla or cortex. Data in (B) are mean with S.E.M. of triplicate measurements of one out of two representative experiments.
Mentions: In order to establish whether Aire co-localizes in the thymus with TRAs, we purified the thymic mTEC based on the cell-surface marker EpCAM (Fig. 3A) and analyzed the expression of the TRA genes. As seen in Fig. 3B, the expression of the Tff3, Ins2, Mup1 and Spt1 antigens was limited to the mTEC population, i.e. the cell population of Aire expression. The cTEC population showed a very low expression for all four TRAs both in the WT as well as Aire KO mouse. Collectively these data show that Aire dose-dependently regulates TRA expression in thymus but not in the lymph nodes, and confirms by real-time PCR the previously published microarray data, suggesting that both Aire and TRAs are predominantly expressed in thymus medullary epithelium.

Bottom Line: Aire had an allele dose-dependent effect on TRA expression in the thymuses of mice from two strains, C57BL/6J and Balb/c, but had no effect on TRA expression in the lymph nodes.By manipulating in vitro organ-cultures we showed that thymic microenvironment plays a dominant role in Aire expression whereas TRAs follow the same pattern.The data underline a direct role for Aire in TRA expression and suggest that modulation of Aire has a potential to control central tolerance and autoimmunity.

View Article: PubMed Central - PubMed

Affiliation: Molecular Pathology, Biomedicum, Tartu University, Ravila 19, 50411 Tartu, Estonia.

ABSTRACT
Intrathymic expression of tissue-restricted antigens (TRAs) has been viewed as the key element in the induction of central tolerance and recently, a central role for the autoimmune regulator (Aire) has been suggested in this process. The aim of this study was to establish whether down or up-regulation of Aire leads to alterations in TRA expression and whether this is limited to thymic epithelial cells. This study also characterized whether TRAs follow Aire expression during normal development, and whether thymic microenvironment plays a role in the expression of Aire and TRAs. We did several in vivo and in vitro experiments to manipulate Aire expression and measured expression of four TRAs (Trefoil factor-3, Insulin-2, Major urinary protein-1 and Salivary protein-1) by real-time RT-PCR. Aire had an allele dose-dependent effect on TRA expression in the thymuses of mice from two strains, C57BL/6J and Balb/c, but had no effect on TRA expression in the lymph nodes. In the thymus, Aire and TRAs were both localized in the medulla and were co-expressed during normal development and involution. In the primary stromal cells as well as thymic epithelial cell line, the adenoviral over-expression of Aire resulted in an increase in TRA expression. By manipulating in vitro organ-cultures we showed that thymic microenvironment plays a dominant role in Aire expression whereas TRAs follow the same pattern. The data underline a direct role for Aire in TRA expression and suggest that modulation of Aire has a potential to control central tolerance and autoimmunity.

Show MeSH
Related in: MedlinePlus