IBD is characterized by inflammation involving a complex interplay between innate and adaptive immunity, non-immune cells, and intestinal microbes. To understand the contributions of innate immunity to inflammation during colitis we have developed a chronic model of innate immune-mediated colitis wherein RAG1-/- mice are crossed with mice expressing TNFAIP3 in intestinal epithelial cells (villin-TNFAIP3 mice). These villin-TNFAIP3 x RAG1-/- (TRAG) mice develop early onset, 100% penetrant, chronic colitis driven by gut microbes. To understand the inflammatory signals that lead to innate colitis in this model, we crossed TRAG mice with TNFα-/- mice (FRAG mice) and compared the severity of colitis in TRAG vs. FRAG mice. In addition, we characterized TNFα-producing myeloid populations from the lamina propria of TRAG mice. Finally, we assessed the expression and localization of proteins involved in cell death pathways, to unravel the mechanisms driving innate-immune mediated colitis. Our results show that, compared to RAG1-/- mice, TRAG mice have significantly increased numbers of lamina propria leukocytes, including increased numbers of neutrophils, inflammatory monocytes, macrophages, dendritic cells, and innate lymphoid cells. TRAG mice lacking TNFα (FRAG mice) had significantly lower numbers of all these leukocyte subsets. Consistent with this, FRAG mice had significantly lower histological colitis scores, compared to TRAG mice. Inflammatory monocytes, monocytes, macrophages, and neutrophils were found to be the main sources of TNFα production, while TNFα production was null or minimal in dendritic cells, natural killer cells, ILC1, ILC3 and IC2. Macrophages and inflammatory monocytes in TRAG mice were characterized by the expression of iNOS, whereas macrophages and monocytes of FRAG and RAG1-/- mice were notably marked by Arginase1 expression. Localization of active caspase3, phosphor-MLKL and TUNEL positive cells indicated extensive epithelial crypt cell death in TRAG mice that was markedly suppressed in FRAG mice. Together our results show that TNFα drives innate immune mediated colitis, potentially through induction of IEC necroptosis. TNFAIP3 is well known as a negative regulator of receptor-mediated NFκB and MAPK activation and therefore attempts to increase TNFAIP3 expression are considered promising avenues for prevention of inflammation. The present study, indicating that intestinal epithelial cell expression of TNFAIP3 drives TNFα-induced IEC death and innate colitis, highlights potential tissue-specific roles for TNFAIP3 that should be considered in the paradigm of TNFAIP3 as a target for prevention of inflammation.
While the cause of Inflammatory bowel disease (IBD) is unknown, certain symptoms may be altered by a multitude of factors including genetics, lifestyle choices, and diet choices. High fat diets (HFDs) are diets with fat accounting for ≥20% of total caloric intake. These diets are known risk factors for IBD onset and worsening symptoms. Mice which overexpress the NF-kB inhibitor TNFAIP3 (A20) were crossed with RAG1-/- mice. These A20 x RAG1-/- (TRAG) mice develop 100% penetrant colitis by eight weeks of age which is not observed in A20 or RAG1-/- littermates. This study examined the effects of a 45% Kcal from fat diet (Research Diets D12451), 10% Kcal fat or “low fat” diet (LFD; Research Diets D12450H), and 10% Kcal fat with cornstarch and maltodextrin for carbohydrates instead of sucrose or “no sucrose” diet (NSD; Research Diets D12450K) on TRAG mice, focusing on changes in inflammation histologically, the gut microbiome, blood glucose, and fat content. All diets were matched for carbohydrates, proteins, and micronutrients. Diets were irradiated to prevent outside bacterial exposure. HFDs have been implicated in worsening colitis symptoms in clinical studies, but no such studies have been completed using the TRAG model of colitis. Four-week-old mice consumed these non-standard chows for four weeks and were humanely euthanized at eight weeks. Severity of colitis was assessed histologically, and changes in the microbiome were assessed via rRNA sequencing. Male and female mice experienced greater weight gain over time on the HFD compared to mice on the LFD (male p=0.0007, female p=0.002) and NSD (male p=0.005, female p=0.003). No significant difference in food consumption was observed in male or female mice on any of the three diets. Both male and female mice had higher blood glucose levels on the HFD than on the LFD and NSD. Male and female mice also had shorter colon lengths on the HFD compared to the LFD and NSD. Male mice on the HFD showed a significant increase in IG and PG weights p=0.0095 and p=0.0015, respectively), as well as liver weights (p=0.005). Female mice also saw a significant increase in PG weight when consuming the HFD relative to the LFD (p=0.045), but no significant difference was observed between the HFD and NSD. This investigation suggests that diets high in fat may worsen colitis symptoms such as inflammation and shortened colon length. HFDs may also increase WAT throughout the body, including in the liver, particularly in males. For future studies TRAG mice should be placed on the same diets for 8-12 weeks to see if results persist as mice age. In addition, TRAG mice could be placed on these diets starting at eight weeks of age to observe how HFDs alter colitis post-onset. Male and female sex differences should be noted, as it appears male mice have more significant changes in their overall WAT than females while consuming a HFD.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.