A Novel Strategy to Study the Invasive Capability of Adherent-Invasive Escherichia coli by Using Human Primary Organoid-Derived Epithelial Monolayers.

Over the last decades, Adherent-Invasive Escherichia coli (AIEC) has been linked to the pathogenesis of Crohn's Disease. AIEC's characteristics, as well as its interaction with the gut immune system and its role in intestinal epithelial barrier dysfunction, have been extensively studied. Nevertheless, the currently available techniques to investigate the cross-talk between this pathogen and intestinal epithelial cells (IECs) are based on the infection of immortalized cell lines. Despite their many advantages, cell lines cannot reproduce the conditions in tissues, nor do they reflect interindividual variability or gut location-specific traits. In that sense, the use of human primary cultures, either healthy or diseased, offers a system that can overcome all of these limitations. Here, we developed a new infection model by using freshly isolated human IECs. For the first time, we generated and infected monolayer cultures derived from human colonic organoids to study the mechanisms and effects of AIEC adherence and invasion on primary human epithelial cells. To establish the optimal conditions for AIEC invasion studies in human primary organoid-derived epithelial monolayers, we designed an infection-kinetics study to assess the infection dynamics at different time points, as well as with two multiplicities of infection (MOI). Overall, this method provides a model for the study of host response to AIEC infections, as well as for the understanding of the molecular mechanisms involved in adhesion, invasion and intracellular replication. Therefore, it represents a promising tool for elucidating the cross-talk between AIEC and the intestinal epithelium in healthy and diseased tissues.

Torque Teno Virus Is Associated With the State of Immune Suppression Early After Liver Transplantation.

The development of noninvasive biomarkers that reflect the state of immunosuppression (IS) remains an unmet need in liver transplantation (LT). Torque Teno virus (TTV) is a highly prevalent, nonpathogenic DNA virus whose plasma levels may be associated with the immune status of the host. The aim of this study was to assess the role of TTV as a biomarker of IS in LT recipients. TTV DNA in plasma was quantified by real-time polymerase chain reaction at different time points during the first year after transplant in a prospectively followed cohort of 63 de novo LT recipients, and any correlation between TTV DNA and biopsy-proven acute cellular rejection (ACR) and opportunistic infections was then evaluated. In addition, TTV DNA was studied in 10 longterm LT recipients in monotherapy with tacrolimus, 10 tolerant recipients, and 10 healthy controls. TTV was detected in the plasma of all patients. Among the 63 LT recipients, 20 episodes of ACR were diagnosed, and there were 28 opportunistic infections, 26 of them being cytomegalovirus (CMV) infections. TTV viremia was significantly lower during ACR (4.41 versus 5.95 log10 copies/mL; P = 0.002) and significantly higher during CMV infections (5.79 versus 6.59 log10 copies/mL; P = 0.009). The area under the receiver operating characteristic curve of TTV viral load for the diagnosis of moderate ACR was 0.869, with a sensitivity and negative predictive value of 100%, respectively, for a cutoff point of 4.75 log10 copies/mL. There were no statistically significant differences in TTV DNA in either longterm or tolerant patients and healthy controls. In conclusion, plasma TTV DNA levels are associated with immune-related events after LT and could constitute a potential biomarker of the state of IS during the first months after transplant.

Iron Deficiency Impairs Intra-Hepatic Lymphocyte Mediated Immune Response.

Hepatic expression of iron homeostasis genes and serum iron parameters predict the success of immunosuppression withdrawal following clinical liver transplantation, a phenomenon known as spontaneous operational tolerance. In experimental animal models, spontaneous liver allograft tolerance is established through a process that requires intra-hepatic lymphocyte activation and deletion. Our aim was to determine if changes in systemic iron status regulate intra-hepatic lymphocyte responses. We used a murine model of lymphocyte-mediated acute liver inflammation induced by Concanavalin A (ConA) injection employing mice fed with an iron-deficient (IrDef) or an iron-balanced diet (IrRepl). While the mild iron deficiency induced by the IrDef diet did not significantly modify the steady state immune cell repertoire and systemic cytokine levels, it significantly dampened inflammatory liver damage after ConA challenge. These findings were associated with a marked decrease in T cell and NKT cell activation following ConA injection in IrDef mice. The decreased liver injury observed in IrDef mice was independent from changes in the gut microflora, and was replicated employing an iron specific chelator that did not modify intra-hepatic hepcidin secretion. Furthermore, low-dose iron chelation markedly impaired the activation of isolated T cells in vitro. All together, these results suggest that small changes in iron homeostasis can have a major effect in the regulation of intra-hepatic lymphocyte mediated responses.