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GILT Accelerates Autoimmunity to the Melanoma Antigen Tyrosinase-Related Protein 1

The Journal of Immunology, 2010. GILT Accelerates Autoimmunity to the Melanoma Antigen Tyrosinase-Related Protein 1. Xu Zhenjie. 2011,08,31. 1. Background. GILT ( Gamma-IFN inducible lysosomal thiol reductase) is expressed in APCs, where it localizes to MHC class II-loading compartments.

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GILT Accelerates Autoimmunity to the Melanoma Antigen Tyrosinase-Related Protein 1

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  1. The Journal of Immunology, 2010 GILT Accelerates Autoimmunity to the Melanoma Antigen Tyrosinase-Related Protein 1 Xu Zhenjie 2011,08,31

  2. 1. Background • GILT ( Gamma-IFN inducible lysosomal thiol reductase) is expressed in APCs, where it localizes to MHC class II-loading compartments. • GILT can facilitates the generation of MHC class II-restricted epitopes from disulfide bond containing Ags. • Melanocyte differentiation Ags are melanosomal integral membrane proteins involved in melanin pigment synthesis. These Ags contain a dileucine-based sorting signal that targets them to the endosomal system where they can be processed for MHC class II-restricted presentation. • However, it is not known how GILT would influence the development of immune responses to these Ags in vivo.

  3. 2. Materials 95-10: an I-Ab-restricted T cell hybridoma recognizing murine TRP1 Cell lines B16.F10: Mel (Melanoma line) PVD: SCC (squamous cell carcinoma) line C57BL/6 : wild-type mice GILT-/- mice mice TRP1tg mice TRP1Bw RAG-/- TRP1tg mice

  4. 3. Methods and results • GILT is required for efficient MHC class II-restricted processing of a TRP1 epitope in vitro. • GILT accelerates the onset of vitiligo in TRP1-specific TCR transgenic mice. • Increased percentage of TRP1-specific transgenic T cells in GILT-deficient mice. • Effector memory cells are increased in the presence of GILT and following development of vitiligo. GILT facilitated class II-restricted processing of TRP1, thereby enhancing T cell activation and accelerating vitiligo.

  5. 3.1. GILT is required for efficient MHC class II-restricted processing of TRP1 B16 cells Wild-type mice GILT-/- mice freeze-thaw splenic B cells Stained by CD45R and I-Ab lysates Immunoblotted with anti-TRP1 mAb Flow cytometry Coculture with B16 cells lysates and 95-10 cells control Measuring IL-2 production by ELISA

  6. Positive control, TRP1 peptide Negative control, SCC lysates There was a statistically significant difference between the ability of wild-type and GILT-/- B cells to present the TRP1 epitope.

  7. Wild-type mice GILT-/- mice DCs Stained by CD11c and I-Ab Flow cytometry Coculture with B16 cells lysates and 95-10 cells Measuring IL-2 production by ELISA control There was a statistically significant difference between the ability of wild-type and GILT-/- DCs to present the TRP1 epitope.

  8. 3. 2. GILT accelerates the onset of vitiligo mediated by TRP1-specific CD4+ T cells 3.2.1 To determine whether GILT would impact the development of a TRP1-specific autoimmune response in vivo, 3.2.2 To demonstrate that GILT in APCs is necessary for the accelerated onset of vitiligo,

  9. Wild type (n=127) TRP1tg mice GILT+/- (n = 56) GILT-/- (n =76) A

  10. A Transfer into CD4+ TRP1-specific T cells RAG-/- mice GILT-/- RAG-/- mice GILT in APCs increases the severity and accelerates the onset of vitiligo.

  11. 3. 3. Increased percentage of TRP1-specific transgenic T cells in GILT-deficient mice RAG-/- mice CD4+ TRP1-specific T cells Transfer into GILT-/- RAG-/- mice Total cell numbers isolated from thymi, spleens, and lymph nodes of the GILT-positive and -negative strains were identical. CD4:CD8 T cell ratio were observed in the thymus, spleen, or lymph node.

  12. Transgenic T cells were identified by Vβ14 and CD4 staining. The percentage of transgenic T cells in GILT-/-TRP1tgmice increased. To exclude the absence of GILT in T cells as the cause of increased T cells, we evaluated the percentage of nontransgenic T cells.

  13. ? TCR was expressed in both GILT-/-TRP1tg mice and TRP1tg mice. The influence of T regulatory (Treg) cell was exclude. The expansion of T cells was limited to the TRP1-specific CD4+Vb14+ T cells.

  14. 3.4 Effector memory cells are increased in the presence of GILT and following development of vitiligo. GILT-/-TRP1tg mice and TRP1tg mice CD4+Vβ14+ T cells CD62L-CD44+ (effector memory) CD62L+CD44+ (central memory) CD62L+CD44- (naive) GILT facilitated class II-restricted processing of TRP1, thereby enhancing T cell activation and accelerating vitiligo.

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