Intestinal lamina propria supports acquired eTreg suppressor function

Abstract

The intestinal immune system must maintain tolerance to commensal microbiota and self antigens whilst defending against invading pathogens. Recognising how homeostasis is established and maintained in a complex immune environment such as the gut is critical to understanding how to re-establish tolerance once broken in inflammatory disorders. Peripherally induced regulatory T cells (Tregs) play a key role in homeostasis. In intestinal tissue, Tregs work in concert with a diverse network of cells but which cellular interactions occur to instruct Treg adaptation and acquisition of distinct Treg suppressor function is not clear. We used two-photon in vivo live imaging and NICHE-seq to deep phenotype Helicobacter hepaticus (Hh)- specific Tregs with shared specificity but distinct spatially compartmentalised functions in the tissue. We show transcriptionally distinct central Treg (cTreg) and effector Treg (eTreg) populations in lymphoid versus gut tissue. The lamina propria (LP), and not embedded lymphoid aggregates (LA), is the key location of acquired immune suppressor eTreg function. Tregs recruited to the LP compartment are the dominant interacting cell type and acquired a more effector Treg profile with upregulation of Areg, Gzmb, Icos, Tigit, Tnfrsf4 (OX40), and Tnfrsf18 (GITR). We identify IL-1B+ macrophages, CD206+ macrophages, and ILC2 in the LP niche as the key players governing Treg survival and function. In contrast, LA, dominated by interactions with ILC3s and populations of IL-6+ DCs, are equipped to tip the balance towards a pro- inflammatory response. By functionally isolating the gut tissue from secondary lymphoid organs, we show that eTregs maintain their phenotype in the context of inflammatory insult. Blocking their key effector molecule, IL-10, results in locally differentiated Th17 cell proliferation without overt inflammation due to local IL-10 independent mechanisms that constrain inflammation. Our results reveal a previously unrecognised spatial mechanism of tolerance, and demonstrate how knowledge of local interactions can guide cell function and potentially be manipulated for the next generation of tolerance-inducing therapies.

Blimp-1 and c-Maf regulate immune gene networks to protect against distinct pathways of pathobiont-induced colitis

“…Given that mice with T cell-specific deletion of Prdm1, Maf or both transcription factors did not exhibit any signs of intestinal inflammation, this provided us with an excellent baseline to determine their role in regulating the intestinal immune response to a defined pathobiont. No intestinal pathology was observed in the control H. hepaticus-infected mice unless they were co-administered anti-IL-10R mAb, in keeping with previous reports 8, [34][35][36] . As no increase in pathology was observed in the Prdm1 fl/fl Maf fl/fl Cd4 Cre H. hepaticus-infected mice when co-administered anti-IL-10R mAb, this suggests that the high level a, Modules of co-expressed genes were derived from human adult IBD colonic biopsies (GSE193677) using the R package WGCNA and b, tested in an independent human pediatric IBD dataset (GSE126124).…”

“…Given that CD4 + T cells from LPLs of H. hepaticus-infected control mice showed a similar level of expression of Il17a, although not Il22, to that in Maf fl/fl Cd4 Cre mice, this suggests that microbiota may be maintaining Il17a expression at the steady state as has been previously described 17,32,33 . The source of the increased Il17a and Il22 in the total LPLs from H. hepaticus Maf fl/fl Cd4 Cre mice above that of infected control mice may be attributed to the increased abundance of T H 17 cells or to other IL-17A producers such as ILC3 or γδ-T cells 34 and to the heterogeneity of T H 17 cells.…”

Blimp-1 and c-Maf regulate common and unique gene networks to protect against distinct pathways of pathobiont-induced colitis

“…We now show that infection with the pathobiont H. hepaticus led to graded increases in colitis, with the Prdm1 fl/fl Cd4 Cre showing mild colitis, the Maf fl/fl Cd4 Cre showing moderate colitis and the double deficient Prdm1 fl/fl Maf fl/fl Cd4 Cre showing severe colitis. No intestinal pathology was observed in the control H. hepaticus infected mice unless they were co-administered anti-IL-10R mAb in keeping with previous reports 8, 45,46,47,48 . Since no increase in pathology was observed in the Prdm1 fl/fl Maf fl/fl Cd4 Cre mice H. hepaticus infected mice when co-administered anti-IL-10R mAb, this suggested that the high level of intestinal pathology resulted from effects of both Prdm1 and Maf on other immune factors, in addition to their co-dominant role in Il10 gene regulation.…”

“…However, since RNA-Seq from sorted CD4 + T cells and scRNA-Seq analysis of the ‘T cells Ctla4 high ’ cluster from LPL from H. hepaticus infected control mice showed a similar level of expression to that of Il17a in uninfected mice, this suggests that microbiota may be maintaining TH17 cells at the steady state as has been described 16, 43, 44 . The source of the increased Il17a in the LPL from H. hepaticus Maf fl/fl Cd4 Cre mice above that of infected control mice may be attributed to the increased abundance of TH17 cells, or to other IL-17A producers such as ILC3 or ψ8 T cells as recently suggested 45 ..…”

“…However, although Foxp3 + RORgt + Treg were almost completely abolished in the LPL from infected Maf fl/fl Cd4 Cre mice in keeping with a previous report 31 , the LPL from the infected double deficient Prdm1 fl /fl Maf fl/ fl Cd4 Cre mice expressed the lowest levels of Il10 and exhibited the maximum pathology indicating that Foxp3 + RORgt + T cells are not the only IL-10 producing Foxp3 + Treg population regulating intestinal pathology. Intestinal Tregs have been recently referred to as Foxp3 + Treg effector cells which exert their effects in lymphoid aggregates in the LP, since they show increased expression of Areg, Gzmb, Icos, Tigit, Tnfrsf4 (OX40), and Tnfrsf18 (GITR), in addition to IL-10 45 . Whilst our findings support the increased expression of these effector genes in the Foxp3 + Tregs, we show that these genes were also increased in Foxp3 - CD4 + T cells, in the LPL from H. hepaticus infected Prdm1 fl/fl Cd4 Cre , Maf fl/fl Cd4 Cre and double deficient Prdm1 fl /fl Maf fl/fl Cd4 Cre mice as compared to controls, although to a much greater extent in the Foxp3- T cells (data not shown).…”