title: Ex vivo immunosuppressive effects of mesenchymal stem cells on Crohn's disease mucosal T cells are largely dependent on indoleamine 2,3-dioxygenase activity and cell-cell contact.
abstract: BACKGROUND
  Crohn's disease (CD) is a disabling chronic enteropathy sustained by a harmful T-cell response toward antigens of the gut microbiota in genetically susceptible subjects. Growing evidence highlights the safety and possible efficacy of mesenchymal stem cells (MSCs) as a new therapeutic tool for this condition. Therefore, we aimed to investigate the effects of bone marrow-derived MSCs on pathogenic T cells with a view to clinical application.
  
  
  METHODS
  T-cell lines from both inflamed and non-inflamed colonic mucosal specimens of CD patients and from healthy mucosa of control subjects were grown with the antigen muramyl-dipeptide in the absence or presence of donors' MSCs. The MSC effects were evaluated in terms of T-cell viability, apoptotic rate, proliferative response, immunophenotype, and cytokine profile. The role of the indoleamine 2,3-dioxygenase (IDO) was established by adding a specific inhibitor, the 1-methyl-DL-tryptophan, and by using MSCs transfected with the small interfering RNA (siRNA) targeting IDO. The relevance of cell-cell contact was evaluated by applying transwell membranes.
  
  
  RESULTS
  A significant reduction in both cell viability and proliferative response to muramyl-dipeptide, with simultaneous increase in the apoptotic rate, was found in T cells from both inflamed and non-inflamed CD mucosa when co-cultured with MSCs and was reverted by inhibiting IDO activity and expression. A reduction of the activated CD4(+)CD25(+) subset and increase of the CD3(+)CD69(+) population were also observed when T-cell lines from CD mucosa were co-cultured with MSCs. In parallel, an inhibitory effect was evident on the expression of the pro-inflammatory cytokines tumor necrosis factor-α, interferon-γ, interleukin-17A and -21, whereas that of the transforming growth factor-β and interleukin-6 were increased, and production of the tolerogenic molecule soluble HLA-G was high. These latter effects were almost completely eliminated by blocking the IDO, whose activity was upregulated in MSCs co-cultured with CD T cells. The use of a semipermeable membrane partially inhibited the MSC immunosuppressive effects. Finally, hardly any effects of MSCs were observed when T cells obtained from control subjects were used.
  
  
  CONCLUSIONS
  MSCs exert potent immunomodulant effects on antigen-specific T cells in CD through a complex paracrine and cell-cell contact-mediated action, which may be exploited for widespread therapeutic use.
authors.affiliation: Clinica Medica I, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. rachele.ciccocioppo@unipv.it.|Centre for the Study and Cure of Inflammatory Bowel Disease, Clinica Medica I, IRCCS San Matteo Hospital Foundation, University of Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. rachele.ciccocioppo@unipv.it.
authors.forename: Rachele
authors.initials: R
authors.lastname: Ciccocioppo
authors.affiliation: Clinica Medica I, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. cangemi.giusy@gmail.com.
authors.forename: Giuseppina C
authors.initials: GC
authors.lastname: Cangemi
authors.affiliation: International Clinical Research Center, St. Anne's University Hospital and Masaryk University, Pekarska 53, Brno, 656 91, Czech Republic. peter.kruzliak@savba.sk.
authors.forename: Peter
authors.initials: P
authors.lastname: Kruzliak
authors.affiliation: Clinica Medica I, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. ale_gallia@hotmail.com.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Gallia
authors.affiliation: Clinica Medica I, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. elena.betti19@gmail.com.
authors.forename: Elena
authors.initials: E
authors.lastname: Betti
authors.affiliation: Servizio di Immunogenetica, Immunoematologia e Medicina Trasfusionale, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. c.badulli@smatteo.pv.it.
authors.forename: Carla
authors.initials: C
authors.lastname: Badulli
authors.affiliation: Servizio di Immunogenetica, Immunoematologia e Medicina Trasfusionale, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. m.martinetti@smatteo.pv.it.
authors.forename: Miryam
authors.initials: M
authors.lastname: Martinetti
authors.affiliation: Servizio di Immunogenetica, Immunoematologia e Medicina Trasfusionale, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. marila.cervio@unipv.it.
authors.forename: Marila
authors.initials: M
authors.lastname: Cervio
authors.affiliation: Clinica Medica III, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. alessandro.pecci@unipv.it.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Pecci
authors.affiliation: Clinica Medica III, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. bozzivaleria@libero.it.
authors.forename: Valeria
authors.initials: V
authors.lastname: Bozzi
authors.affiliation: Chirurgia Generale I, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. paolo.dionigi@unipv.it.
authors.forename: Paolo
authors.initials: P
authors.lastname: Dionigi
authors.affiliation: Dipartimento di Medicina Occupazionale, Ergonomia e Disabilità, Laboratorio di Nanotecnologia, Fondazione IRCCS Salvatore Maugeri, Università di Pavia, Via Maugeri 8-10, Pavia, 27100, Italy. livia.visai@unipv.it.|Dipartimento di Medicina Molecolare, Centro di Ingegneria Tissutale, INSTM UdR Pavia, Università di Pavia, Pavia, 27100, Italy. livia.visai@unipv.it.
authors.forename: Livia
authors.initials: L
authors.lastname: Visai
authors.affiliation: Laboratori di Oncoematologia Pediatrica, Fondazione IRCCS Policlinico San Matteo, Piazzale Golgi 19, Pavia, 27100, Italy. ti.ri@libero.it.
authors.forename: Antonella
authors.initials: A
authors.lastname: Gurrado
authors.affiliation: Clinica Medica I, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. c.alvisi@smatteo.pv.it.
authors.forename: Costanza
authors.initials: C
authors.lastname: Alvisi
authors.affiliation: Laboratorio di Ematologia, Fondazione IRCCS Policlinico San Matteo, Piazzale Golgi 19, Pavia, 27100, Italy. c.picone@smatteo.pv.it.
authors.forename: Cristina
authors.initials: C
authors.lastname: Picone
authors.affiliation: Centro di Ricerca di Medicina Rigenerativa, Fondazione IRCCS Policlinico San Matteo, Piazzale Golgi 19, Pavia, 27100, Italy. m.monti@smatteo.pv.it.
authors.forename: Manuela
authors.initials: M
authors.lastname: Monti
authors.affiliation: Dipartimento di Onco-Ematologia Pediatrica e Medicina Trasfusionale, Ospedale Pediatrico Bambino Gesù, Via Sant'Onofrio 4, Rome, 00165, Italy. mebernardo@gmail.com.
authors.forename: Maria E
authors.initials: ME
authors.lastname: Bernardo
authors.affiliation: Clinica Medica I, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. gobbipg@smatteo.pv.it.
authors.forename: Paolo
authors.initials: P
authors.lastname: Gobbi
authors.affiliation: Clinica Medica I, Dipartimento di Medicina Interna, Fondazione IRCCS Policlinico San Matteo, Università di Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. gr.corazza@smatteo.pv.it.|Centre for the Study and Cure of Inflammatory Bowel Disease, Clinica Medica I, IRCCS San Matteo Hospital Foundation, University of Pavia, Piazzale Golgi 19, Pavia, 27100, Italy. gr.corazza@smatteo.pv.it.
authors.forename: Gino R
authors.initials: GR
authors.lastname: Corazza
mesh_terms: D000119:Acetylmuramyl-Alanyl-Isoglutamine / Q000494:pharmacology; D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000954:Antigens, Surface / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D001854:Bone Marrow Cells / Q000166:cytology; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival; D002478:Cells, Cultured; D018920:Coculture Techniques; D003424:Crohn Disease / Q000473:pathology*; D016207:Cytokines / Q000378:metabolism; D005260:Female; D059951:HLA-G Antigens / Q000378:metabolism; D006801:Humans; D016130:Immunophenotyping; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000166:cytology; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism; D008875:Middle Aged; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D013601:T-Lymphocytes / Q000166:cytology* / Q000187:drug effects / Q000276:immunology; D059008:Time-Lapse Imaging; D014364:Tryptophan / Q000031:analogs & derivatives / Q000494:pharmacology; D055815:Young Adult
chemical_list: D000954:Antigens, Surface; D016207:Cytokines; D059951:HLA-G Antigens; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D034741:RNA, Small Interfering; D000119:Acetylmuramyl-Alanyl-Isoglutamine; D014364:Tryptophan; C525396:1-methyltryptophan