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Galectin-9-mediated protection from allo-specific T cells as a mechanism of immune privilege of corneal allografts.

Shimmura-Tomita M, Wang M, Taniguchi H, Akiba H, Yagita H, Hori J - PLoS ONE (2013)

Bottom Line: The eye is an immune-privileged organ, and corneal transplantation is therefore one of the most successful organ transplantation.T-cell immunoglobulin and mucin domain (Tim)-3 is a regulatory molecule for T-cell function, and galectin (Gal)-9 is a Tim-3 ligand.Blockade of Tim-3 or Gal-9 did not abolish anterior chamber-associated immune deviation.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Nippon Medical School, Tokyo, Japan. machiko@jichi.ac.jp

ABSTRACT
The eye is an immune-privileged organ, and corneal transplantation is therefore one of the most successful organ transplantation. The immunosuppressive intraocular microenvironment is known as one of the mechanisms underlying immune privilege in the eye. T-cell immunoglobulin and mucin domain (Tim)-3 is a regulatory molecule for T-cell function, and galectin (Gal)-9 is a Tim-3 ligand. We investigated the role of this pathway in establishing the immune-privileged status of corneal allografts in mice. Gal-9 is constitutively expressed on the corneal epithelium, endothelium and iris-ciliary body in normal mouse eyes and eyes bearing surviving allografts, and Tim-3 was expressed on CD8 T cells infiltrating the allografts. Allograft survival in recipients treated with anti-Tim-3 monoclonal antibody (mAb) or anti-Gal-9 mAb was significantly shorter than that in control recipients. In vitro, destruction of corneal endothelial cells by allo-reactive T cells was enhanced when the cornea was pretreated with anti-Gal-9 mAb. Blockade of Tim-3 or Gal-9 did not abolish anterior chamber-associated immune deviation. We propose that constitutive expression of Gal-9 plays an immunosuppressive role in corneal allografts. Gal-9 expressed on corneal endothelial cells protects them from destruction by allo-reactive T cells within the cornea.

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Expression of Tim-3 and Gal-9 in corneal allografts and secondary lymphoid organs.Normal corneas of C57BL/6 mice were transplanted into normal eyes of BALB/c mice. Graft-bearing eyes were isolated at 2, 4, and 8 weeks. (A, C, and E) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with anti-Gal-9 mAb followed by FITC-conjugated anti-rat IgG mAb or streptavidin-FITC (green). Nuclei were stained with PI (red). (B, D, F, and G) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with biotinylated anti-Tim-3 Ab followed by streptavidin-FITC (green), and PE-conjugated anti-CD8 mAb (red). Nuclei were stained with DAPI (blue). Arrows in D show the Tim-3+ CD8+ cells, and G represents the magnification of one of the double positive cells. Cep, corneal epithelium; Cst, corneal stroma; Ced, corneal endothelium. Original magnification, ×40. (H, I, J, and K) Whole spleen (SP) or LN cells from recipients bearing surviving (Allo Sur) or rejected (Allo Rej) allografts and syngeneic (Syn) grafts were analyzed by flow cytometry. Data are displayed as the proportion of Tim-3+ CD4+ cells in the whole SP and LN (H, I), Tim-3+ CD8+ cells in the whole SP and LN (J, K). Data are presented as the mean ± standard deviation of five recipients in each group. N.S., not significant.
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pone-0063620-g003: Expression of Tim-3 and Gal-9 in corneal allografts and secondary lymphoid organs.Normal corneas of C57BL/6 mice were transplanted into normal eyes of BALB/c mice. Graft-bearing eyes were isolated at 2, 4, and 8 weeks. (A, C, and E) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with anti-Gal-9 mAb followed by FITC-conjugated anti-rat IgG mAb or streptavidin-FITC (green). Nuclei were stained with PI (red). (B, D, F, and G) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with biotinylated anti-Tim-3 Ab followed by streptavidin-FITC (green), and PE-conjugated anti-CD8 mAb (red). Nuclei were stained with DAPI (blue). Arrows in D show the Tim-3+ CD8+ cells, and G represents the magnification of one of the double positive cells. Cep, corneal epithelium; Cst, corneal stroma; Ced, corneal endothelium. Original magnification, ×40. (H, I, J, and K) Whole spleen (SP) or LN cells from recipients bearing surviving (Allo Sur) or rejected (Allo Rej) allografts and syngeneic (Syn) grafts were analyzed by flow cytometry. Data are displayed as the proportion of Tim-3+ CD4+ cells in the whole SP and LN (H, I), Tim-3+ CD8+ cells in the whole SP and LN (J, K). Data are presented as the mean ± standard deviation of five recipients in each group. N.S., not significant.

Mentions: When normal corneas of C57BL/6 were transplanted orthotopically into normal eyes of BALB/c mice, Gal-9 was strongly expressed on corneal epithelium, endothelial cells, and superficial stromal cells in the surviving corneal allografts (Fig. 3A, C, E). Tim-3-expressing cells were present in corneal epithelium and stroma at the center of the surviving allograft at 4 weeks after transplantation (Fig. 3D). To identify the character of these Tim-3 positive cells, we performed double-staining with CD4, CD8, CD11b, and CD11c. These cells were only CD8 positive (Fig. 3G). Tim-3 positive cells were not present in short or long term surviving corneal allografts (Fig. 3B, F). Proportions of Tim-3 expressing CD4+ or CD8+ T cells in splenocytes and lymph node (LN) cells were statistically indistinguishable among recipients bearing surviving allografts, those bearing rejected allografts, and those bearing syngeneic allografts (Fig. 3H, I, J, K), indicating that expression of Tim-3 in the spleen and lymph nodes does not correlate with the fate of corneal grafts.


Galectin-9-mediated protection from allo-specific T cells as a mechanism of immune privilege of corneal allografts.

Shimmura-Tomita M, Wang M, Taniguchi H, Akiba H, Yagita H, Hori J - PLoS ONE (2013)

Expression of Tim-3 and Gal-9 in corneal allografts and secondary lymphoid organs.Normal corneas of C57BL/6 mice were transplanted into normal eyes of BALB/c mice. Graft-bearing eyes were isolated at 2, 4, and 8 weeks. (A, C, and E) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with anti-Gal-9 mAb followed by FITC-conjugated anti-rat IgG mAb or streptavidin-FITC (green). Nuclei were stained with PI (red). (B, D, F, and G) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with biotinylated anti-Tim-3 Ab followed by streptavidin-FITC (green), and PE-conjugated anti-CD8 mAb (red). Nuclei were stained with DAPI (blue). Arrows in D show the Tim-3+ CD8+ cells, and G represents the magnification of one of the double positive cells. Cep, corneal epithelium; Cst, corneal stroma; Ced, corneal endothelium. Original magnification, ×40. (H, I, J, and K) Whole spleen (SP) or LN cells from recipients bearing surviving (Allo Sur) or rejected (Allo Rej) allografts and syngeneic (Syn) grafts were analyzed by flow cytometry. Data are displayed as the proportion of Tim-3+ CD4+ cells in the whole SP and LN (H, I), Tim-3+ CD8+ cells in the whole SP and LN (J, K). Data are presented as the mean ± standard deviation of five recipients in each group. N.S., not significant.
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Related In: Results  -  Collection

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pone-0063620-g003: Expression of Tim-3 and Gal-9 in corneal allografts and secondary lymphoid organs.Normal corneas of C57BL/6 mice were transplanted into normal eyes of BALB/c mice. Graft-bearing eyes were isolated at 2, 4, and 8 weeks. (A, C, and E) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with anti-Gal-9 mAb followed by FITC-conjugated anti-rat IgG mAb or streptavidin-FITC (green). Nuclei were stained with PI (red). (B, D, F, and G) Cryostat sections of surviving graft centers were examined by immunofluorescent staining with biotinylated anti-Tim-3 Ab followed by streptavidin-FITC (green), and PE-conjugated anti-CD8 mAb (red). Nuclei were stained with DAPI (blue). Arrows in D show the Tim-3+ CD8+ cells, and G represents the magnification of one of the double positive cells. Cep, corneal epithelium; Cst, corneal stroma; Ced, corneal endothelium. Original magnification, ×40. (H, I, J, and K) Whole spleen (SP) or LN cells from recipients bearing surviving (Allo Sur) or rejected (Allo Rej) allografts and syngeneic (Syn) grafts were analyzed by flow cytometry. Data are displayed as the proportion of Tim-3+ CD4+ cells in the whole SP and LN (H, I), Tim-3+ CD8+ cells in the whole SP and LN (J, K). Data are presented as the mean ± standard deviation of five recipients in each group. N.S., not significant.
Mentions: When normal corneas of C57BL/6 were transplanted orthotopically into normal eyes of BALB/c mice, Gal-9 was strongly expressed on corneal epithelium, endothelial cells, and superficial stromal cells in the surviving corneal allografts (Fig. 3A, C, E). Tim-3-expressing cells were present in corneal epithelium and stroma at the center of the surviving allograft at 4 weeks after transplantation (Fig. 3D). To identify the character of these Tim-3 positive cells, we performed double-staining with CD4, CD8, CD11b, and CD11c. These cells were only CD8 positive (Fig. 3G). Tim-3 positive cells were not present in short or long term surviving corneal allografts (Fig. 3B, F). Proportions of Tim-3 expressing CD4+ or CD8+ T cells in splenocytes and lymph node (LN) cells were statistically indistinguishable among recipients bearing surviving allografts, those bearing rejected allografts, and those bearing syngeneic allografts (Fig. 3H, I, J, K), indicating that expression of Tim-3 in the spleen and lymph nodes does not correlate with the fate of corneal grafts.

Bottom Line: The eye is an immune-privileged organ, and corneal transplantation is therefore one of the most successful organ transplantation.T-cell immunoglobulin and mucin domain (Tim)-3 is a regulatory molecule for T-cell function, and galectin (Gal)-9 is a Tim-3 ligand.Blockade of Tim-3 or Gal-9 did not abolish anterior chamber-associated immune deviation.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology, Nippon Medical School, Tokyo, Japan. machiko@jichi.ac.jp

ABSTRACT
The eye is an immune-privileged organ, and corneal transplantation is therefore one of the most successful organ transplantation. The immunosuppressive intraocular microenvironment is known as one of the mechanisms underlying immune privilege in the eye. T-cell immunoglobulin and mucin domain (Tim)-3 is a regulatory molecule for T-cell function, and galectin (Gal)-9 is a Tim-3 ligand. We investigated the role of this pathway in establishing the immune-privileged status of corneal allografts in mice. Gal-9 is constitutively expressed on the corneal epithelium, endothelium and iris-ciliary body in normal mouse eyes and eyes bearing surviving allografts, and Tim-3 was expressed on CD8 T cells infiltrating the allografts. Allograft survival in recipients treated with anti-Tim-3 monoclonal antibody (mAb) or anti-Gal-9 mAb was significantly shorter than that in control recipients. In vitro, destruction of corneal endothelial cells by allo-reactive T cells was enhanced when the cornea was pretreated with anti-Gal-9 mAb. Blockade of Tim-3 or Gal-9 did not abolish anterior chamber-associated immune deviation. We propose that constitutive expression of Gal-9 plays an immunosuppressive role in corneal allografts. Gal-9 expressed on corneal endothelial cells protects them from destruction by allo-reactive T cells within the cornea.

Show MeSH
Related in: MedlinePlus