P005 The solute carrier LC22A4/Organic Cation Transporter (OCTN)-1 as a novel Inflammatory Bowel Disease determinant at the microbe-host interface.
Masi, L.(1);Petito, V.(1);Fidaleo, M.(2);Palucci, I.(3);Del Chierico, F.(4);Ivagnes, V.(3);Mercuri, A.(5);Graziani, C.(1);Lopetuso, L.R.(1);Armuzzi, A.(1);Gasbarrini, A.(1);Pani, G.(5);Scaldaferri, F.(1);
(1)Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Gastroenterology and Internal Medicine, Rome, Italy;(2)Università La Sapienza, Department of Biology and Biotechnologies Charles Darwin, Rome, Italy;(3)Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Microbiology, Rome, Italy;(4)Children Hospital Bambino Gesù IRCCS, Unit of Human Microbiome, Rome, Italy;(5)Catholic University of the Sacred Heart, Department of Translational Medicine, Rome, Italy;
The putative Ergothioneine Transporter, SLC22A4/OCTN1, is expressed in the intestine and monocytes/macrophages, and its widely represented missense variant L503F (0.4 Minor Allele Frequency in Caucasians) has been found to be associated with Crohn’s disease, sporadic colorectal cancer in early age and ulcerative colitis (UC) patients. The leucine-to-phenylalanine substitution at amino acid 503 affects OCTN1 capacity to transport ergothioneine, acetylcholine and likely other substrates of endogenous or microbial origin; however, whether this defect contributes to inflammatory bowel disease (IBD) pathogenesis, and the mechanistic underpinnings, remain elusive.
In order to gain mechanistic insight on the role of SLC22A4 in IBD, we first analyzed primary monocytes from healthy donors and UC patients of varying OCTN1 genotypes. Then we stably knocked down or overexpressed OCTN1 and its disease-associated variant in the monocytic cell line THP-1. Finally, we compared wild-type and OCTN1 knockout C57BL/6 mice in a chemical colitis paradigm in vivo.
Primary adherent monocytes from healthy donors homozygous for the IBD-associated allele (TT) displayed a significant enhancement of interleukin 1 beta (IL-1β) response to peptidoglycan (PGN) compared to the other genotypes. Likewise, cells from UC patients bearing the 503F variant released more IL-1β in response to live bacteria and displayed reduced expression of autophagy markers (e.g. p62, LC3), compared to patients negative for the variant.
In agreement with these findings, OCTN1-deficient THP-1 macrophages secreted reduced amounts of IL-1β when challenged in vitro with PGN or live bacteria. Most relevant, THP1 cells engineered to overexpress the 503F variant of OCTN1 displayed a much stronger IL-1β response to muramyl dipeptide and PGN challenge, compared to cells transduced with the “wild type” transporter. Also consistent with a role for OCTN1 variants in inflammatory cascades relevant to IBD, in vivo studies revealed milder DSS-induced colitis in OCTN1 knockout mice compared to wild-type animals, both at the peak of disease severity and at the end of the recovery period.
Collectively our initial evidence supports the view that OCTN1 may have a causative role in IBD via deranged bacterial sensing and cytokine production, with 503F likely behaving as a gain-of-function variant. Thus, targeting OCTN1 variant may provide unprecedented opportunities for the personalized treatment of IBD patients and a better understanding of disease pathogenesis.