2. The Role of TCR gamma-delta T cells in Celiac Disease Columbia University, College of Physicians and Surgeons By: Nertila Ujkaj Littin Kandoth Sandra Mentor: Dr. Manavalan

3. Celiac disease (CD) a digestive disease that damages the small intestine and interferes with absorption of nutrients from food.a digestive disease that damages the small intestine and interferes with absorption of nutrients from food. when people with CD consume gluten, their immune system responds by damaging the small intestinewhen people with CD consume gluten, their immune system responds by damaging the small intestine a genetic disordera genetic disorder symptoms: diarrhea, weight loss, malnutrition, etc…symptoms: diarrhea, weight loss, malnutrition, etc… diagnosis is done by intestinal biopsy histological grading and presence of serum auto- antibodies.diagnosis is done by intestinal biopsy histological grading and presence of serum auto- antibodies. CD is considered an autoimmune disorder because the body's own immune system causes the damageCD is considered an autoimmune disorder because the body's own immune system causes the damage

4. The Gut Lumen

5. AntigenPresentingCell B cell LP CD4 + CD4 + T cell Anti-gliadin and anti-TTG2 antibodies HLA-DQ2 or DQ8 IEL  T cell Gluten MICA/B NKG2DCD94/NKG2C IL-15 IEL  T cell IFN-  & Cytotoxic granules ? IMMUNOPATHOGENESIS OF CELIAC DISEASE Gliadin Peptide INTRAEPITHELIAL SPACE LAMINA PROPRIA

6. Objective To determine if the TCR gamma-delta IELs in patients with CD are Regulatory/Suppressor T cellsTo determine if the TCR gamma-delta IELs in patients with CD are Regulatory/Suppressor T cells –Regulatory T cells (also known as suppressor T cells) are a specialized subpopulation of T cells that act to suppress activation of the immune system and thereby maintain immune system homeostasis and tolerance to self. –They are characterized by having some of the following markers Intracellular FOXP3Intracellular FOXP3 TGF-betaTGF-beta IL-10IL-10 CTLA 4CTLA 4 GITRGITR

7. Methods Isolation of Lymphocytes from the Duodenal biopsies of 30 patients with Celiac Disease Immunostaining for the following extracellular and intracellular markers: –T-Cell markers CD3 CD8 TCR alpha-beta TCR-gamma-delta –Markers for Regulatory/Suppressor cells: Intra-cellular FOXP3 Intracellular TGF-beta GITR CTLA-4 Intra-cellular IL-10 Flow cytometry- technique for counting, examining and sorting microscopic particles (such as cells) suspended in a stream of fluid

8. http://www.flow-cytometry.de/img/fcm.gif

9. A. Immunohistochemistry EPITHELIAL CELLS B. Flow Cytometry INTRAEPITHELIAL LYMPHOCYTES (IEL)

10. Results

13. AntigenPresentingCell B cell LP CD4 + CD4 + T cell Anti-gliadin and anti-TTG2 antibodies HLA-DQ2/8 IEL  T cell TTG Gliadin MICA/B NKG2DCD94/NKG2C IL-15 IFN-  & Cytotoxic granules TGF  IEL  T cell FOXP3

14. Conclusion The TCR gamma-delta T cells have FOXP3 and secrete TGF-beta and are probably Regulator/ Suppressor T cells Future studies To prove that TCR gamma delta T cells can suppress the immune response To determine if CD patients who are on therapy have more suppressor TCR gamma-delta T cells than Active CD patients

15. References Green, P.H. and B. Jabri, Celiac disease. Annu Rev Med, 2006. 57: p. 207-21. Shan, L., et al., Structural basis for gluten intolerance in celiac sprue. Science, 2002. 297(5590): p. 2275-9. Meresse, B., et al., Coordinated induction by IL15 of a TCR-independent NKG2D signaling pathway converts CTL into lymphokine-activated killer cells in celiac disease. Immunity, 2004. 21(3): p. 357-66. Chen, Y., et al., Protection of the intestinal mucosa by intraepithelial gamma delta T cells. Proc Natl Acad Sci U S A, 2002. 99(22): p. 14338-43. Peng, S.L., et al., Propagation and regulation of systemic autoimmunity by gammadelta T cells. J Immunol, 1996. 157(12): p. 5689-98. Roncador, G., et al., Analysis of FOXP3 protein expression in human CD4+CD25+ regulatory T cells at the single-cell level. Eur J Immunol, 2005. 35(6): p. 1681-91. Manavalan, J.S., et al., Alloantigen specific CD8+CD28- FOXP3+ T suppressor cells induce ILT3+ ILT4+ tolerogenic endothelial cells, inhibiting alloreactivity. Int Immunol, 2004. 16(8): p. 1055-68.

16. Acknowledgements Dr. Manavalan Dr. Sat Bhattacharya Harlem Children Society Allison Shannon Harold Celiac Disease Center of Columbia University

17. Thank You!!!