Mixtures of the two major conjugated linoleic acid (CLA) isomers trans-10,cis-12-CLA and cis-9,trans-11-CLA are used as over the counter supplements for weight loss. Because of the reported adverse effects of CLA on insulin sensitivity in some mouse studies, we sought to compare the impact of dietary t10c12-CLA and c9t11-CLA on liver, adipose tissue, and systemic metabolism of adult lean mice. We fed 8 week-old C57Bl/6J male mice with low fat diets (10.5% Kcal from fat) containing 0.8% t10c12-CLA or c9t11-CLA for 9 or 38 days. Diets containing c9t11-CLA had minimal impact on the endpoints studied. However, 7 days after starting the t10c12-CLA diet, we observed a dramatic reduction in fat mass measured by NMR spectroscopy, which interestingly rebounded by 38 days. This rebound was apparently due to a massive accumulation of lipids in the liver, because adipose tissue depots were visually undetectable. Hepatic steatosis and the disappearance of adipose tissue after t10c12-CLA feeding was associated with elevated plasma insulin levels and insulin resistance, compared to mice fed a control diet or c9t11-CLA diet. Unexpectedly, despite being insulin resistant, mice fed t10c12-CLA had normal levels of blood glucose, without signs of impaired glucose clearance. Hepatic gene expression and fatty acid composition suggested enhanced hepatic de novo lipogenesis without an increase in expression of gluconeogenic genes. These data indicate that dietary t10c12-CLA may alter hepatic
Commensal bacteria are critical regulators of both tissue homeostasis and the development and exacerbation of autoimmunity. However, it remains unclear how the intestinal microbiota contributes to inflammation in tissues such as the central nervous system (CNS) where these microbes are typically absent and whether T cell receptor (TCR) specificity for commensal-derived antigens is important to the development of tissue inflammation-related outcomes. Here, we found that ileum- and cecum-colonizing segmented filamentous bacteria (SFB)-specific T cells (clone TCR7B8) can infiltrate the CNS wherein they can be reactivated and produce high levels of inflammatory cytokines including IFNg, IL-17A, TNFa, and GM-CSF in the absence of regulatory T cells. In contrast, other SFB-specific T cells (clone TCR1A2) recognizing an epitope in which 8/9 amino acids overlap with those recognized by TCR7B8 failed to induce such neuroinflammation. Despite their similar SFB-derived peptide antigen targets, TCR7B8 was found to recognize peptides derived from host proteins including receptor tyrosine-protein kinase ErbB2, trophinin 1, and anaphase-promoting complex subunit 2 in vitro, whereas TCR1A2 did not, indicating that TCR7B8 induces CNS inflammation via molecular mimicry. Immune checkpoint blockade accelerated TCR7B8-mediated CNS inflammation, suggesting a potential cause of immune-related adverse events induced in cancer patients undergoing such treatment. Together, our findings reveal a potential mechanism whereby gut commensal-specific T cells are dysregulated and contribute to extraintestinal inflammation.
Down Regulated in Adenoma (DRA), the major chloride transporter in the mammalian intestine has recently emerged as a critical gene contributing to the pathogenesis of Ulcerative Colitis (UC). We have previously shown that DRA knockout (KO) mice display several features of UC including compromised barrier function and dysbiosis. Further, DRA KO mice show increased susceptibility to intestinal inflammation. To elucidate the mechanisms by which loss of DRA could impact gut inflammation, gene expression between DRA KO and wild type littermate controls (WT) by RNAseq was compared. The data showed that out of 1100 genes differentially expressed in DRA KO colon, majority were related to immune responses. Upon conducting more specific immune based analysis using nanoString nCounter® Immunology Panel, we observed a significant upregulation of genes promoting effector T cell expansion, specifically Th2. Interleukin‐33 (IL‐33), an alarmin cytokine is known to be significantly upregulated in colons of UC patients and is a key driver of type 2 immune cell expansion. Since alterations in epithelial‐immune cell crosstalk can trigger inflammatory responses, we hypothesized that DRA KO mice may have aberrant epithelial‐immune cell crosstalk which could trigger changes in Lamina Propria Lymphocytes (LPLs). Following isolation of colonic LPLs and staining for various innate and adaptive immune cells, we detected a significant increase (> 2‐fold) in type 2 innate lymphoid cells (ILC2) and in Th2 cells(~8‐fold) in DRA KO mice which were both GATA3+. These observations were specific to the colon and not seen in the ileal LP or mesenteric lymph nodes. We also observed a significant induction of type 2 effector protein amphiregulin, demonstrating an increased type‐2 immune tone in DRA KO colons. Since IL‐33 is important for immune cell activation following tissue injury at mucosal surfaces we analyzed IL‐33 protein levels in the colon. IL‐33 was markedly elevated in DRA KO colons and appeared to be independent of microbiome changes as this increase in IL‐33 still persisted even after cohousing with WT mice for over 4 weeks (allowing exchange of the microbiome). Confocal imaging of colonic tissues and colonoids further confirmed the increase in epithelial IL‐33 and its dissemination from the nucleus to the cytoplasm in DRA KO colonocytes. Since IL‐33 is known to specifically expand a subset of T regs which are also GATA3+; we gated LPLs double positive for FOXP3 and GATA3 using FACS. Interestingly, total T regs and GATA3+ T regs were both markedly elevated in DRA KO colonic LP compared to WT, confirming the functional relevance of increased IL‐33. These results were further validated in human colonoid derived monolayers where DRA was significantly reduced parallel to upregulation of IL‐33 protein in UC, mirroring the results from DRA KO mice. In conclusion, loss of DRA from colonocytes triggers the release of the IL‐33 to drive a type 2 immune response. These observations emphasize the critical importance of loss of DRA and its implicati...
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