Genetic variation in TNFA predicts protection from severe bacterial infections in patients with end-stage liver disease awaiting liver transplantation

Background & Aims: Augmented susceptibility to infections increases mortality in patients with end-stage liver disease (ESLD). We sought to determine the contribution of selected genetic variants involved in inflammatory signalling downstream of the Toll-like receptor 4 (TLR4) to severe bacterial infections (SBIs) in patients with ESLD. Methods: We retrospectively assessed incidence of SBIs in 336 adult ESLD patients enlisted for orthotopic liver transplantation (OLT) and genotyped them for TLR4 c.+1196C/T, CD14 c.-159C/T, TNFA c. 238G/A, TNFA c. 863C/A, IL1B c. 31C/T and IL1RN variable number of tandem repeats allelic variants. Principal findings were validated in an independent cohort of 332 ESLD patients. Results: Thirty-four percent of patients from the identification cohort and 40% of patients from the validation cohort presented with SBI while enlisted for OLT. The presence of the variant allele TNFA c. -238A (rs361525) was associated with lower serum levels of TNF-alpha, and with significantly decreased risk of SBI in both cohorts. Multivariate analysis showed that the relative protection from SBI associated with this allele almost completely negated the increased susceptibility to SBI owed to advanced ESLD. Although not predictive of overall mortality, the presence of the TNFA c. -238A allele was associated with a complete prevention of SBI-related pre-transplant deaths. Conclusions: Our results suggest that genetic variability in inflammatory signalling is associated with the development of SBI in patients with ESLD. Specifically, we identified the importance of the TNFA c. -238A allele as a strong predictor of protection from SBI, and as a genetic marker associated with significantly improved pre-transplant survival in patients with SBI. (C) 2013 European Association for the Study of the Liver. Published by Elsevier B. V. All rights reserved

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

Introduction: 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-alpha, interferon-gamma, interleukin-17A and -21, whereas that of the transforming growth factor-beta 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. Conclusion: 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