Regulation of Peripheral Inflammation by a Non-Viable, Non-Colonizing Strain of Commensal Bacteria.

The gastrointestinal tract represents one of the largest body surfaces that is exposed to the outside world. It is the only mucosal surface that is required to simultaneously recognize and defend against pathogens, while allowing nutrients containing foreign antigens to be tolerated and absorbed. It differentiates between these foreign substances through a complex system of pattern recognition receptors expressed on the surface of the intestinal epithelial cells as well as the underlying immune cells. These immune cells actively sample and evaluate microbes and other particles that pass through the lumen of the gut. This local sensing system is part of a broader distributed signaling system that is connected to the rest of the body through the enteric nervous system, the immune system, and the metabolic system. While local tissue homeostasis is maintained by commensal bacteria that colonize the gut, colonization itself may not be required for the activation of distributed signaling networks that can result in modulation of peripheral inflammation. Herein, we describe the ability of a gut-restricted strain of commensal bacteria to drive systemic anti-inflammatory effects in a manner that does not rely upon its ability to colonize the gastrointestinal tract or alter the mucosal microbiome. Orally administered EDP1867, a gamma-irradiated strain of Veillonella parvula, rapidly transits through the murine gut without colonization or alteration of the background microbiome flora. In murine models of inflammatory disease including delayed-type hypersensitivity (DTH), atopic dermatitis, psoriasis, and experimental autoimmune encephalomyelitis (EAE), treatment with EDP1867 resulted in significant reduction in inflammation and immunopathology. Ex vivo cytokine analyses revealed that EDP1867 treatment diminished production of pro-inflammatory cytokines involved in inflammatory cascades. Furthermore, blockade of lymphocyte migration to the gut-associated lymphoid tissues impaired the ability of EDP1867 to resolve peripheral inflammation, supporting the hypothesis that circulating immune cells are responsible for promulgating the signals from the gut to peripheral tissues. Finally, we show that adoptively transferred T cells from EDP1867-treated mice inhibit inflammation induced in recipient mice. These results demonstrate that an orally-delivered, non-viable strain of commensal bacteria can mediate potent anti-inflammatory effects in peripheral tissues through transient occupancy of the gastrointestinal tract, and support the development of non-living bacterial strains for therapeutic applications.

OX40L-JAG1-Induced Expansion of Lineage-Stable Regulatory T Cells Involves Noncanonical NF-κB Signaling.

Foxp3+T regulatory cells (Tregs) control autoimmune response by suppressing proliferation and effector functions of self-reactive Foxp3-CD4+/CD8+ T cells and thereby maintain the critical balance between self-tolerance and autoimmunity. Earlier, we had shown that OX40L-JAG1 cosignaling mediated through their cognate receptors OX40 and Notch3 preferentially expressed on murine Tregs can selectively induce their proliferation in the absence of TCR stimulation. However, the differential molecular mechanisms regulating TCR-independent versus TCR-dependent Treg proliferation and lineage stability of the expanded Tregs remained unknown. In this study, we show that OX40L-JAG1 treatment induced TCR-independent proliferation of Tregs in the thymus and periphery. The use of Src kinase inhibitor permitted us to demonstrate selective inhibition of TCR-dependent T cell proliferation with little to no effect on OX40L-JAG1-induced TCR-independent Treg expansion in vitro, which was critically dependent on noncanonical NF-κB signaling. OX40L-JAG1-expanded Tregs showed sustained lineage stability as indicated by stable demethylation marks in Treg signature genes such as Foxp3, Il2ra, Ctla4, Ikzf2, and Ikzf4. Furthermore, OX40L-JAG1 treatment significantly increased CTLA4+ and TIGIT+ Tregs and alleviated experimental autoimmune thyroiditis in mice. Relevance of our findings to humans became apparent when human OX40L and JAG1 induced TCR-independent selective expansion of human Tregs in thymocyte cultures and increased human Tregs in the liver tissue of humanized NSG mice. Our findings suggest that OX40L-JAG1-induced TCR-independent Treg proliferation is a conserved mechanism that can be used to expand lineage-stable Tregs to treat autoimmune diseases.

Critical role of OX40 signaling in the TCR-independent phase of human and murine thymic Treg generation.

Regulatory T cells (Tregs) play a pivotal role in immune-tolerance, and loss of Treg function can lead to the development of autoimmunity. Natural Tregs generated in the thymus substantially contribute to the Treg pool in the periphery, where they suppress self-reactive effector T cells (Teff) responses. Recently, we showed that OX40L (TNFSF4) is able to drive selective proliferation of peripheral Tregs independent of canonical antigen presentation (CAP-independent) in the presence of low-dose IL-2. Therefore, we hypothesized that OX40 signaling might be integral to the TCR-independent phase of murine and human thymic Treg (tTreg) development. Development of tTregs is a two-step process: Strong T-cell receptor (TCR) signals in combination with co-signals from the TNFRSF members facilitate tTreg precursor selection, followed by a TCR-independent phase of tTreg development in which their maturation is driven by IL-2. Therefore, we investigated whether OX40 signaling could also play a critical role in the TCR-independent phase of tTreg development. OX40-/- mice had significantly reduced numbers of CD25-Foxp3low tTreg precursors and CD25+Foxp3+ mature tTregs, while OX40L treatment of WT mice induced significant proliferation of these cell subsets. Relative to tTeff cells, OX40 was expressed at higher levels in both murine and human tTreg precursors and mature tTregs. In ex vivo cultures, OX40L increased tTreg maturation and induced CAP-independent proliferation of both murine and human tTregs, which was mediated through the activation of AKT-mTOR signaling. These novel findings show an evolutionarily conserved role for OX40 signaling in tTreg development and proliferation, and might enable the development of novel strategies to increase Tregs and suppress autoimmunity.

Dichotomous effect of caffeine, curcumin, and naringenin on genomic DNA of normal and diabetic subjects.

Nutraceutical compounds show antioxidant and prooxidant properties under stress conditions like cancer, diabetes, and other diseases. The objective of this study is to find the dichotomic behavior of caffeine, curcumin, and naringenin on DNA of diabetic and normal subjects in the presence and absence of copper, hydrogen peroxide, and complex of copper-hydrogen peroxide. Hydrogen peroxide releases hydroxyl free radicals ((•)OH) on oxidation of Cu (I) to Cu (II) through Fenton-type reaction to cause DNA damage. In the results, agarose gel electrophoretic pattern speculates the prooxidant effect of caffeine and antioxidant effect of curcumin on DNA in the presence of copper and hydrogen peroxide. UV-Vis spectral analysis shows hyperchromism on addition of DNA to caffeine, hypochromism with curcumin, and subtle changes with naringenin. The chosen nutraceuticals act as inducers and quenchers of oxidative free radicals arising from diabetes.