CCR2⁺CD103⁻ intestinal dendritic cells develop from DC-committed precursors and induce interleukin-17 production by T cells

The identification of intestinal macrophages (m phi s) and dendritic cells (DCs) is a matter of intense debate. Although CD103(+) mononuclear phagocytes (MPs) appear to be genuine DCs, the nature and origins of CD103(-) MPs remain controversial. We show here that intestinal CD103(-)CD11b(+) MPs can be separated clearly into DCs and m phi s based on phenotype, gene profile, and kinetics. CD64(-)CD103(-)CD11b(+) MPs are classical DCs, being derived from Flt3 ligand-dependent, DC-committed precursors, not Ly6C hi monocytes. Surprisingly, a significant proportion of these CD103(-)CD11b(+) DCs express CCR2 and there is a selective decrease in CD103(-)CD11b(+) DCs in mice lacking this chemokine receptor. CCR2(+)CD103(-) DCs are present in both the murine and human intestine, drive interleukin (IL)-17a production by Tcells in vitro, and show constitutive expression of IL-12/IL-23p40. These data highlight the heterogeneity of intestinal DCs and reveal a bona fide population of CCR2(+) DCs that is involved in priming mucosal T helper type 17 (Th17) responses

The ancient drug salicylate indirectly targets fructose-1,6-bisphosphatase to suppress liver glucose production in diet-induced obese mice

Aims: The benefit of salicylate in the treatment of diabetes has been recognized for over a century; however, challenging side effects have prevented widespread use. A better understanding of the relevant enzyme targets mediating its anti-hyperglycaemic effect may lead to the development of novel therapies for diabetes. Here, we investigated the contribution of 5′-adenosine monophosphate (AMP)-dependent inhibition of fructose-1,6-bisphosphatase 1 (FBP1) to the anti-hyperglycaemic action of salicylate. Methods: We studied AMP-insensitive FBP1 G27P knockin (KI) mice through a variety of cellular approaches, including proteomics, Seahorse metabolic analysis, glucose production, and other assays, in addition to a detailed assessment of metabolic responses in vivo. Results: Compared with wild-type littermates, AMP-insensitive FBP1 KI mice were resistant to the effects of the drug on body weight, glucose tolerance, pyruvate disposal, liver lipid content and hepatic glucose production. Compared with wild-type, KI hepatocytes exhibited baseline differences in glycolytic, TCA cycle and fatty acid oxidation enzyme levels, potentially linking gluconeogenic dysregulation and its reversal to non-carbohydrate fuel management. Conclusion: Collectively, our data highlight a novel mechanism of action for the effects of salicylate on glycaemia and weight gain, which depends on AMP-mediated allosteric inhibition of FBP1.</p

Identification and characterization of murine glycoprotein 2‐expressing intestinal dendritic cells

The intestinal lamina propria (LP) contains distinct subsets of classical dendritic cells (cDC), each playing key non-redundant roles in intestinal immune homeostasis. Here, we show that glycoprotein 2 (GP2), a GPI-anchored protein and receptor for bacterial type-I fimbriae, is selectively expressed by CD103+CD11b+ cDC in the murine small intestine (SI). GP2 expression was induced on CD103+CD11b+ cDC within the SI-LP and was regulated by IRF4, TGFβR1- and retinoic acid signalling. Mice selectively lacking Gp2 on CD103+CD11b+ cDC (huLang-Cre.gp2fl/fl mice) had normal numbers and proportions of innate and adaptive immune cells in the SI-LP suggesting that GP2 expression by CD103+CD11b+ cDC is not required for intestinal immune homoeostasis

Intestinal CD103CD11b cDC2 Conventional Dendritic Cells Are Required for Primary CD4 T and B Cell Responses to Soluble Flagellin

Systemic immunization with soluble flagellin (sFliC) from Salmonella Typhimurium induces mucosal responses, offering potential as an adjuvant platform for vaccines. Moreover, this engagement of mucosal immunity is necessary for optimal systemic immunity, demonstrating an interaction between these two semi-autonomous immune systems. Although TLR5 and CD103+CD11b+ cDC2 contribute to this process, the relationship between these is unclear in the early activation of CD4+ T cells and the development of antigen-specific B cell responses. In this work, we use TLR5-deficient mice and CD11c-cre.Irf4fl/fl mice (which have reduced numbers of cDC2, particularly intestinal CD103+CD11b+ cDCs), to address these points by studying the responses concurrently in the spleen and the mesenteric lymph nodes (MLN). We show that CD103+CD11b+ cDC2 respond rapidly and accumulate in the MLN after immunization with sFliC in a TLR5-dependent manner. Furthermore, we identify that whilst CD103+CD11b+ cDC2 are essential for the induction of primary T and B cell responses in the mucosa, they do not play such a central role for the induction of these responses in the spleen. Additionally, we show the involvement of CD103+CD11b+ cDC2 in the induction of Th2-associated responses. CD11c-cre.Irf4fl/fl mice showed a reduced primary FliC-specific Th2-associated IgG1 responses, but enhanced Th1-associated IgG2c responses. These data expand our current understanding of the mucosal immune responses promoted by sFliC and highlights the potential of this adjuvant for vaccine usage by taking advantage of the functionality of mucosal CD103+CD11b+ cDC2.</p

Immune functions of intestinal dendritic cells

The intestinal surface is daily challenged with tremendous amount of foreign material derived our diet and from the commensal bacteria that densely populate the mucosal surface. Occasionally the gut mucosa is exposed to pathogens trying to enter our body. The important task of the intestinal immune system is to remain tolerant toward innocuous luminal constituents and to elicit defense responses towards invading pathogens. Conventional dendritic cells (cDC) play a central role in the initiation of such tolerogenic and defense responses. They scan and sample the local environment, migrate to the draining lymph nodes, where they activate adaptive immune cells specifically recognizing the presented luminal antigens. Several subsets of cDC populate the intestinal mucosa, but their role in the adaptive immune response is incompletely defined. The present Ph.D. thesis aimed to identify some of the in vivo functions of intestinal cDC subsets in the generation of adaptive immune responses. In paper 1, we demonstrated that cDC1 play a critical role in the homoeostasis of various subsets of intestinal T cells. In particular, we could show that cDC1 are crucial for the differentiation of Th1 cells and for the generation of conventional CD8αβ+ and cytotoxic-like CD4+CD8αα+ IEL, the latter requiring β8-integrin expression on cDC1. Lack of cDC1 in the MLN, was associated with impaired priming and reduced induction of small intestinal homing receptors on responding antigen-specific CD8αβ+ T cells. Collectively these results highlight multiple non-redundant roles for cDC1 in the maintenance of intestinal T cell homeostasis.In paper II, we demonstrate that a fraction of small intestinal CD103+CD11b+ cDC specifically expresses glycoprotein 2 (GP2). GP2 is a GPI-anchored protein previously found to be expressed by M-cells in the intestinal epithelium and to act as a receptor for type-I fimbriae on bacteria. The presence of GP2+ cDC in the intestine was dependent on expression of the transcription factor IRF4 and was regulated by the microbiota and aryl hydrocarbon receptor signaling. cDC-derived GP2 was not required for normal intestinal T cell homeostasis. The role of this molecule on cDC remained unidentified, but it may potentially be involved in uptake of type-I fimbriated bacteria and in regulation of adaptive immune responses.In paper III, we addressed the role of intestinal cDC subsets in the initiation of B cell responses toward systemically administered flagellin (FliC). Mice lacking cDC2, but not cDC1, failed to induce the differentiation FliC-specific IgA and IgG plasma cell in the MLN, correlating with abrogated formation of FliC-specific germinal centers. Thus, intestinal cDC2 play a crucial role in the generation of antibody responses toward FliC.Collectively, the work performed in present Ph.D. thesis has broadened our understanding on the in vivo functionality of intestinal cDC subsets in the generation of adaptive immune responses

The Ancient Drug Salicylate Indirectly Targets Fructose-1,6-Bisphosphatase to Suppress Liver Glucose Production in Diet-Induced Obese Mice.

AIMS: The benefit of salicylate in the treatment of diabetes has been recognized for over a century; however, challenging side effects have prevented widespread use. A better understanding of the relevant enzyme targets mediating its anti-hyperglycaemic effect may lead to the development of novel therapies for diabetes. Here, we investigated the contribution of 5'-adenosine monophosphate (AMP)-dependent inhibition of fructose-1,6-bisphosphatase 1 (FBP1) to the anti-hyperglycaemic action of salicylate.METHODS: We studied AMP-insensitive FBP1 G27P knockin (KI) mice through a variety of cellular approaches, including proteomics, Seahorse metabolic analysis, glucose production, and other assays, in addition to a detailed assessment of metabolic responses invivo.RESULTS: Compared with wild-type littermates, AMP-insensitive FBP1 KI mice were resistant to the effects of the drug on body weight, glucose tolerance, pyruvate disposal, liver lipid content and hepatic glucose production. Compared with wild-type, KI hepatocytes exhibited baseline differences in glycolytic, TCA cycle and fatty acid oxidation enzyme levels, potentially linking gluconeogenic dysregulation and its reversal to non-carbohydrate fuel management.CONCLUSION: Collectively, our data highlight a novel mechanism of action for the effects of salicylate on glycaemia and weight gain, which depends on AMP-mediated allosteric inhibition of FBP1

Migration of intestinal dendritic cell subsets upon intrinsic and extrinsic TLR3 stimulation

AbstractInitiation of adaptive immunity to particulate antigens in lymph nodes largely depends on their presentation by migratory dendritic cells (DCs). DC subsets differ in their capacity to induce specific types of immunity, allowing subset-specific DC-targeting to influence vaccination and therapy outcomes. Faithful drug design however requires exact understanding of subset-specific versus global activation mechanisms. cDC1, the subset of DCs that excel in supporting immunity towards viruses, intracellular bacteria and tumors, express uniquely high levels of the pattern recognition receptor TLR3. Using various genetic models, we show here that both the cDC1 and cDC2 subsets of cDCs are activated and migrate equally well in response to TLR3 stimulation in a cell extrinsic and TNFα dependent manner, but that cDC1 show a unique requirement for type I interferon signaling. Our findings reveal common and differing pathways regulating DC subset migration, offering important insights for the design of DC-based vaccination and therapy approaches.</jats:p