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PTPN2 deficiency exacerbates epithelial barrier dysfunction in vivo and in vitro via redistribution of occludin and ZO-1. Ronald R. Marchelletta 1 , Lucas Bernts 1 , Stephan Myers 1 , Taylaur Smith 1 , Young Park 1 , Kim E Barrett 1 , Declan F McCole 2 1 University of California, San Diego

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  1. PTPN2 deficiency exacerbates epithelial barrier dysfunction in vivo and in vitro via redistribution of occludin and ZO-1 Ronald R. Marchelletta1, Lucas Bernts1, Stephan Myers1, Taylaur Smith1, Young Park1, Kim E Barrett1, Declan F McCole2 1University of California, San Diego 2University of California, Riverside

  2. Disclosures • Nothing to disclose

  3. Gastrointestinal Barrier • Aids in the absorption of nutrients while excluding harmful molecules and bacteria. • Defective epithelial barrier is a hallmark of UC and CD. • Barrier dysfunction is in part driven by IFNgsignaling. • Genome-Wide Association Studies show that SNP in Protein Tyrosine Phosphatase N2 is Associated with increase incidence of CD and UC. Abreu, Reviews Immunology 10, 131-144 (February 2010)

  4. PTPN2 Restricts IFNg-Mediated Barrier Dysfunction • PTPN2 is a negative regular of IFNgsignaling. • PTPN2 protects epithelial barrier function. • Loss of PTPN2 causes: • Increased epithelial permeability • Decreased epithelial resistance Scharl, M et al., Gastroenerology, 2009 McCole, DF, NY Academy of Science, 2012

  5. Tight-Junction Turner JR. Nat Rev Immunol 2009;9:799-809. Ivanov AI. Am J Pathol. 2010 Aug;177(2):512-24

  6. PTPN2 KO Mice Show Increased Permeability in Proximal Colon • Mount tissue in Ussing Chamber • Applied FD4 to the mucosal side • Sampled at the serosal side (2 hours). humanbiology.wzw.tum.de

  7. Mislocalization of Occludin & ZO-1 from tight junctions in PTPN2 KO mouse colon Arrows indicate location of tight junctions. Knock-out exhibits internalization and mislocalization of occludin.

  8. C216S Mutant Cell line • HCA7 intestinal epithelial cell line • 216 Cysteine to Serine (C216S) • Lentiviral Vector • Doxycycline activation

  9. Detection of HA Tag in C216S-Transduced Cells • HA-TC45 expression confirmed in DOX induced condition • Nearly 3-fold increase in PTPN2 with increased transactivator protein

  10. C216S expressing cells show increased phospho-STAT1

  11. ZO-1 and Occludin are reduced in C216S mutant cell line

  12. PTPN2 mutant cells show reduced occludin/ZO-1 co-localization *

  13. Conclusion • PTPN2 KO mice show increased colonic FITC-dextran permeability. • Occludin appears mislocalized from the tight junctions in PTPN2 KO. • C216S mutant cells have altered tight junction composition. • Therefore, loss of PTPN2 expression or activity in vivo and in vitro compromises barrier function possibly through occludin mislocalization.

  14. Acknowledgements • Michel Tremblay – McGill University • Tony Tiganis – Monash University

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