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Crohn's disease (CD) is a complex chronic inflammatory disorder with both gastrointestinal and extra-intestinal manifestations associated immune dysregulation. Analyzing 202,359 cells from 170 specimens across 83 patients, we identify a distinct epithelial cell type in both terminal ileum and ascending colon (hereon as 'LND') with high expression of LCN2, NOS2, and DUOX2 and genes related to antimicrobial response and immunoregulation. LND cells, confirmed by in-situ RNA and protein imaging, are rare in non-IBD controls but expand in active CD, and actively interact with immune cells and specifically express IBD/CD susceptibility genes, suggesting a possible function in CD immunopathogenesis. Furthermore, we discover early and late LND subpopulations with different origins and developmental potential. A higher ratio of late-to-early LND cells correlates with better response to anti-TNF treatment. Our findings thus suggest a potential pathogenic role for LND cells in both Crohn's ileitis and colitis.
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Colon , Enfermedad de Crohn , Oxidasas Duales , Células Epiteliales , Íleon , Lipocalina 2 , Enfermedad de Crohn/patología , Enfermedad de Crohn/genética , Enfermedad de Crohn/inmunología , Humanos , Células Epiteliales/metabolismo , Células Epiteliales/patología , Colon/patología , Íleon/patología , Lipocalina 2/metabolismo , Lipocalina 2/genética , Oxidasas Duales/genética , Oxidasas Duales/metabolismo , Masculino , Óxido Nítrico Sintasa de Tipo II/metabolismo , Óxido Nítrico Sintasa de Tipo II/genética , Femenino , Adulto , Factor de Necrosis Tumoral alfa/metabolismo , Mucosa Intestinal/patología , Mucosa Intestinal/metabolismo , Persona de Mediana EdadRESUMEN
Undifferentiated intestinal stem cells (ISCs) are crucial for maintaining homeostasis and resolving injury. Lgr5+ cells in the crypt base constantly divide, pushing daughter cells upward along the crypt axis where they differentiate into specialized cell types. Coordinated execution of complex transcriptional programs is necessary to allow for the maintenance of undifferentiated stem cells while permitting differentiation of the wide array of intestinal cells necessary for homeostasis. Previously, members of the myeloid translocation gene (MTG) family have been identified as transcriptional co-repressors that regulate stem cell maintenance and differentiation programs in multiple organ systems, including the intestine. One MTG family member, myeloid translocation gene related 1 (MTGR1), has been recognized as a crucial regulator of secretory cell differentiation and response to injury. However, whether MTGR1 contributes to the function of ISCs has not yet been examined. Here, using Mtgr1-/- mice, we have assessed the effects of MTGR1 loss specifically in ISC biology. Interestingly, loss of MTGR1 increased the total number of cells expressing Lgr5, the canonical marker of cycling ISCs, suggesting higher overall stem cell numbers. However, expanded transcriptomic and functional analyses revealed deficiencies in Mtgr1-null ISCs, including deregulated ISC-associated transcriptional programs. Ex vivo, intestinal organoids established from Mtgr1-null mice were unable to survive and expand due to aberrant differentiation and loss of stem and proliferative cells. Together, these results indicate that the role of MTGR1 in intestinal differentiation is likely stem cell intrinsic and identify a novel role for MTGR1 in maintaining ISC function.
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Diferenciación Celular , Intestino Delgado , Células Madre , Animales , Ratones , Células Madre/metabolismo , Células Madre/citología , Intestino Delgado/citología , Intestino Delgado/metabolismo , Ratones Noqueados , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Ratones Endogámicos C57BL , Organoides/metabolismo , Organoides/citología , Proteínas Represoras/metabolismo , Proteínas Represoras/genéticaRESUMEN
BACKGROUND: A combination of proton-pump inhibitors (PPI) and topical steroids (TS) is used to treat children with eosinophilic esophagitis (EoE). However, a subset of children do not respond to this combination therapy. We aimed to identify the esophageal transcriptional, cell composition, and microbial differences between the non-responders (EoE-PPI-TSnr; n = 7) and responders (EoE-PPI-TSr; n = 7) to the combination therapy for EoE and controls (n = 9) using metatranscriptomics. METHODS: Differential gene expression analysis was used to identify transcriptional differences, validated using the EoE diagnostic panel (EDP). Deconvolution analysis was performed to identify differences in their cell type composition. Microbiome analysis was conducted from esophageal biopsies RNAseq data, and microbial abundance was correlated with esophageal gene expression. RESULTS: In all, 3164 upregulated and 3154 downregulated genes distinguished EoE-PPI-TSnr from EoE-PPI-TSr. Eosinophilic inflammatory response, cytokine signaling, and collagen formation pathways were significantly upregulated in EoE-PPI-TSnr. There was a 56% overlap in dysregulated genes between EoE-PPI-TSnr and EDP, with a perfect agreement in the directionality of modulation. Eosinophils, dendritic cells (DCs), immature DCs, megakaryocytic-erythroid progenitors, and T helper type 1 cells were significantly higher in EoE-PPI-TSnr. There was no significant difference in microbiome diversity. The relative abundance of Fusobacterium sp. and Acinetobacter sp. notably differed in EoE-PPI-TSnr and correlated with the key pathways. CONCLUSION: Our results provide critical insights into the molecular, cellular, and microbial factors associated with the lack of response to PPI and TS combination therapy in children with EoE. This study advances our understanding of the pathobiology of EoE while guiding personalized treatment strategies.
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BACKGROUND: Anaplastic thyroid carcinoma (ATC) is a highly aggressive malignancy that has consistently shown Wnt/ß-catenin (canonical) signaling activation in various study populations. There are currently no targetable treatments for BRAF-wildtype ATC and a lack of effective treatment for BRAFV600EATC. Our aim is to identify whether Wnt inhibitors could be potential therapeutic agents for ATC patients with limited treatment options. METHODS: In this Institutional Review Board-approved study, we utilize a cohort of 32 ATCs and 20 non-neoplastic multinodular goiters (MNG). We also use 4 ATC spheroid cell lines (THJ-16T, THJ-21T, THJ-29T, and THJ-11T) and two primary patient-derived ATC organoid cultures (VWL-T5 and VWL-T60). Finally, we use a murine xenograft mouse model of ATC for in vivo treatment studies. RESULTS: Using a large patient cohort, we demonstrate that this near-universal Wnt signaling activation is associated with ligand expression- rather than being mutationally-driven. We show that pyrvinium pamoate, a potent Wnt inhibitor, exhibits in vitro efficacy against both ATC cell lines and primary patient-derived ATC organoids VWL-T5 (p < 0.05) and VWL-T60 (p < 0.01) Finally, using a murine xenograft model of ATC, we show that pyrvinium significantly delays the growth of ATC tumors in THJ-16T (p < 0.005) and THJ-21T (p < 0.001). CONCLUSIONS: We tested Wnt inhibitor treatment, both in vitro and in vivo, as a potential novel therapy for this highly lethal disease. Future large-scale studies utilizing multiple Wnt inhibitors will lay the foundation for the development of these novel therapies for patients with ATC.
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The intestinal immune response is crucial in maintaining a healthy gut, but the enhanced migration of macrophages in response to pathogens is a major contributor to disease pathogenesis. Integrins are ubiquitously expressed cellular receptors that are highly involved in immune cell adhesion to endothelial cells while in the circulation and help facilitate extravasation into tissues. Here we show that specific deletion of the Tln1 gene encoding the protein talin-1, an integrin-activating scaffold protein, from cells of the myeloid lineage using the Lyz2-cre driver mouse reduces epithelial damage, attenuates colitis, downregulates the expression of macrophage markers, decreases the number of differentiated colonic mucosal macrophages, and diminishes the presence of CD68-positive cells in the colonic mucosa of mice infected with the enteric pathogen Citrobacter rodentium. Bone marrow-derived macrophages lacking expression of Tln1 did not exhibit a cell-autonomous phenotype; there was no impaired proinflammatory gene expression, nitric oxide production, phagocytic ability, or surface expression of CD11b, CD86, or major histocompatibility complex II in response to C. rodentium. Thus, we demonstrate that talin-1 plays a role in the manifestation of infectious colitis by increasing mucosal macrophages, with an effect that is independent of macrophage activation.
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Colitis , Infecciones por Enterobacteriaceae , Animales , Ratones , Citrobacter rodentium , Colitis/genética , Colitis/prevención & control , Colon/patología , Células Endoteliales/metabolismo , Infecciones por Enterobacteriaceae/metabolismo , Inflamación/patología , Mucosa Intestinal/metabolismo , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Talina/genética , Talina/metabolismoRESUMEN
OBJECTIVE: Proton pump inhibitors (PPIs) are commonly used to manage children with upper gastrointestinal symptoms and without a formal diagnosis. We investigated the effect of PPIs on esophageal mucosal transcriptome and active microbiota in children with normal esophagi. Furthermore, we examined whether the differences in host esophageal mucosal gene expression were driven by an underlying esophageal epithelial cell type composition. METHODS: Using metatranscriptomics, the host transcriptional and active microbial profiles were captured from 17 esophageal biopsy samples (PPI naïve [PPI-], n = 7; PPI exposed [PPI+], n = 10) collected from children without any endoscopic and histologic abnormalities in their esophagus (normal esophagus). Deconvolution computational analysis was performed with xCell to assess if the observed epithelial gene expression changes were related to the cell type composition in the esophageal samples. RESULTS: The median (IQR) age of our cohort was 14 years (12-16) with female (63%) preponderance. Both groups were similar in terms of their demographics and clinical features. Compared with PPI-, the PPI+ had upregulation of 27 genes including the MUC genes. The cell type composition was similar between the PPI- and PPI+ groups. Prevotella sp and Streptococcus sp were abundant in PPI+ group. CONCLUSIONS: In children with normal esophagus, PPI exposure can be associated with upregulation of esophageal mucosal homeostasis and epithelial cell function genes in a cell-type independent manner, and an altered esophageal microbiome. Additional studies are warranted to validate our findings and to investigate the causal effect of PPIs on the normal esophageal epithelium and microbial communities.
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Several probiotic-derived factors have been identified as effectors of probiotics for exerting beneficial effects on the host. However, there is a paucity of studies to elucidate mechanisms of their functions. p40, a secretory protein, is originally isolated from a probiotic bacterium, Lactobacillus rhamnosus GG. Thus, this study aimed to apply structure-functional analysis to define the functional peptide of p40 that modulates the epigenetic program in intestinal epithelial cells for sustained prevention of colitis. In silico analysis revealed that p40 is composed of a signal peptide (1-28 residues) followed by a coiled-coil domain with uncharacterized function on the N-terminus, a linker region, and a ß-sheet domain with high homology to CHAP on the C-terminus. Based on the p40 three-dimensional structure model, two recombinant p40 peptides were generated, p40N120 (28-120 residues) and p40N180 (28-180 residues) that contain first two and first three coiled coils, respectively. Compared to full-length p40 (p40F) and p40N180, p40N120 showed similar or higher effects on up-regulating expression of Setd1b (encoding a methyltransferase), promoting mono- and trimethylation of histone 3 on lysine 4 (H3K4me1/3), and enhancing Tgfb gene expression and protein production that leads to SMAD2 phosphorylation in human colonoids and a mouse colonic epithelial cell line. Furthermore, supplementation with p40F and p40N120 in early life increased H3K4me1, Tgfb expression and differentiation of regulatory T cells (Tregs) in the colon, and mitigated disruption of epithelial barrier and inflammation induced by DSS in adult mice. This study reveals the structural feature of p40 and identifies a functional peptide of p40 that could maintain intestinal homeostasis.
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Colitis , Microbioma Gastrointestinal , Probióticos , Adulto , Humanos , Animales , Ratones , Proteínas Bacterianas/genética , Péptidos , Colitis/prevención & control , Probióticos/farmacologíaRESUMEN
BACKGROUND AND AIMS: Eosinophils are present in several solid tumors and have context-dependent function. Our aim is to define the contribution of eosinophils in esophageal squamous cell carcinoma (ESCC), as their role in ESCC is unknown. METHODS: Eosinophils were enumerated in tissues from 2 ESCC cohorts. Mice were treated with 4-NQO for 8 weeks to induce precancer or 16 weeks to induce carcinoma. The eosinophil number was modified by a monoclonal antibody to interleukin-5 (IL5mAb), recombinant IL-5 (rIL-5), or genetically with eosinophil-deficient (ΔdblGATA) mice or mice deficient in eosinophil chemoattractant eotaxin-1 (Ccl11-/-). Esophageal tissue and eosinophil-specific RNA sequencing was performed to understand eosinophil function. Three-dimensional coculturing of eosinophils with precancer or cancer cells was done to ascertain direct effects of eosinophils. RESULTS: Activated eosinophils are present in higher numbers in early-stage vs late-stage ESCC. Mice treated with 4-NQO exhibit more esophageal eosinophils in precancer vs cancer. Correspondingly, epithelial cell Ccl11 expression is higher in mice with precancer. Eosinophil depletion using 3 mouse models (Ccl11-/- mice, ΔdblGATA mice, IL5mAb treatment) all display exacerbated 4-NQO tumorigenesis. Conversely, treatment with rIL-5 increases esophageal eosinophilia and protects against precancer and carcinoma. Tissue and eosinophil RNA sequencing revealed eosinophils drive oxidative stress in precancer. In vitro coculturing of eosinophils with precancer or cancer cells resulted in increased apoptosis in the presence of a degranulating agent, which is reversed with NAC, a reactive oxygen species scavenger. ΔdblGATA mice exhibited increased CD4 T cell infiltration, IL-17, and enrichment of IL-17 protumorigenic pathways. CONCLUSION: Eosinophils likely protect against ESCC through reactive oxygen species release during degranulation and suppression of IL-17.
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Carcinoma , Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Animales , Ratones , Eosinófilos , Interleucina-17 , Especies Reactivas de OxígenoRESUMEN
Background/Aims: Eosinophils are present in several solid tumors and have context-dependent function. Our aim is to define the contribution of eosinophils in esophageal squamous cell carcinoma (ESCC), since their role in ESCC is unknown. Methods: Eosinophils were enumerated in tissues from two ESCC cohorts. Mice were treated with 4-nitroquinolone-1-oxide (4-NQO) for 8 weeks to induce pre-cancer or 16 weeks to induce carcinoma. Eosinophil number was modified by monoclonal antibody to IL-5 (IL5mAb), recombinant IL-5 (rIL-5), or genetically with eosinophil-deficient (ΔdblGATA) mice or mice deficient in eosinophil chemoattractant eotaxin-1 ( Ccl11 -/- ). Esophageal tissue and eosinophil specific RNA-sequencing was performed to understand eosinophil function. 3-D co-culturing of eosinophils with pre-cancer or cancer cells was done to ascertain direct effects of eosinophils. Results: Activated eosinophils are present in higher numbers in early stage versus late stage ESCC. Mice treated with 4-NQO exhibit more esophageal eosinophils in pre-cancer versus cancer. Correspondingly, epithelial cell Ccl11 expression is higher in mice with pre-cancer. Eosinophil depletion using three mouse models ( Ccl11 -/- mice, ΔdblGATA mice, IL5mAb treatment) all display exacerbated 4-NQO tumorigenesis. Conversely, treatment with rIL-5 increases esophageal eosinophilia and protects against pre-cancer and carcinoma. Tissue and eosinophil RNA-sequencing revealed eosinophils drive oxidative stress in pre-cancer. In vitro co-culturing of eosinophils with pre-cancer or cancer cells resulted in increased apoptosis in the presence of a degranulating agent, which is reversed with N-acetylcysteine, a reactive oxygen species (ROS) scavenger. ΔdblGATA mice exhibited increased CD4 T cell infiltration, IL-17, and enrichment of IL-17 pro-tumorigenic pathways. Conclusion: Eosinophils likely protect against ESCC through ROS release during degranulation and suppression of IL-17.
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Although selenium deficiency correlates with colorectal cancer (CRC) risk, the roles of the selenium-rich antioxidant selenoprotein P (SELENOP) in CRC remain unclear. In this study, we defined SELENOP's contributions to sporadic CRC. In human single-cell cRNA-Seq (scRNA-Seq) data sets, we discovered that SELENOP expression rose as normal colon stem cells transformed into adenomas that progressed into carcinomas. We next examined the effects of Selenop KO in a mouse adenoma model that involved conditional, intestinal epithelium-specific deletion of the tumor suppressor adenomatous polyposis coli (Apc) and found that Selenop KO decreased colon tumor incidence and size. We mechanistically interrogated SELENOP-driven phenotypes in tumor organoids as well as in CRC and noncancer cell lines. Selenop-KO tumor organoids demonstrated defects in organoid formation and decreases in WNT target gene expression, which could be reversed by SELENOP restoration. Moreover, SELENOP increased canonical WNT signaling activity in noncancer and CRC cell lines. In defining the mechanism of action of SELENOP, we mapped protein-protein interactions between SELENOP and the WNT coreceptors low-density lipoprotein receptor-related proteins 5 and 6 (LRP5/6). Last, we confirmed that SELENOP-LRP5/6 interactions contributed to the effects of SELENOP on WNT activity. Overall, our results position SELENOP as a modulator of the WNT signaling pathway in sporadic CRC.
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Adenoma , Neoplasias Colorrectales , Selenio , Ratones , Animales , Humanos , Vía de Señalización Wnt , Selenoproteína P/genética , Selenoproteína P/metabolismo , Neoplasias Colorrectales/patología , Selenio/metabolismo , Carcinogénesis/genética , Adenoma/metabolismo , Regulación Neoplásica de la Expresión Génica , Proteína-5 Relacionada con Receptor de Lipoproteína de Baja Densidad/genética , Proteína-5 Relacionada con Receptor de Lipoproteína de Baja Densidad/metabolismoRESUMEN
Pathogenic enteric Escherichia coli present a significant burden to global health. Food-borne enteropathogenic E. coli (EPEC) and Shiga toxin-producing E. coli (STEC) utilize attaching and effacing (A/E) lesions and actin-dense pedestal formation to colonize the gastrointestinal tract. Talin-1 is a large structural protein that links the actin cytoskeleton to the extracellular matrix though direct influence on integrins. Here we show that mice lacking talin-1 in intestinal epithelial cells (Tln1Δepi) have heightened susceptibility to colonic disease caused by the A/E murine pathogen Citrobacter rodentium. Tln1Δepi mice exhibit decreased survival, and increased colonization, colon weight, and histologic colitis compared to littermate Tln1fl/fl controls. These findings were associated with decreased actin polymerization and increased infiltration of innate myeloperoxidase-expressing immune cells, confirmed as neutrophils by flow cytometry, but more bacterial dissemination deep into colonic crypts. Further evaluation of the immune population recruited to the mucosa in response to C. rodentium revealed that loss of Tln1 in colonic epithelial cells (CECs) results in impaired recruitment and activation of T cells. C. rodentium infection-induced colonic mucosal hyperplasia was exacerbated in Tln1Δepi mice compared to littermate controls. We demonstrate that this is associated with decreased CEC apoptosis and crowding of proliferating cells in the base of the glands. Taken together, talin-1 expression by CECs is important in the regulation of both epithelial renewal and the inflammatory T cell response in the setting of colitis caused by C. rodentium, suggesting that this protein functions in CECs to limit, rather than contribute to the pathogenesis of this enteric infection.
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Colitis , Infecciones por Enterobacteriaceae , Microbioma Gastrointestinal , Animales , Ratones , Citrobacter rodentium , Talina/genética , Escherichia coli/metabolismo , Actinas/metabolismo , Linfocitos T/metabolismo , Colitis/microbiología , Colon/microbiología , Mucosa Intestinal/metabolismo , Infecciones por Enterobacteriaceae/microbiología , Ratones Endogámicos C57BLRESUMEN
The cytokine interleukin-23 (IL-23) is involved in the pathogenesis of inflammatory and autoimmune conditions including inflammatory bowel disease (IBD). IL23R is enriched in intestinal Tregs, yet whether IL-23 modulates intestinal Tregs remains unknown. Here, investigating IL-23R signaling in Tregs specifically, we show that colonic Tregs highly express Il23r compared with Tregs from other compartments and their frequency is reduced upon IL-23 administration and impairs Treg suppressive function. Similarly, colonic Treg frequency is increased in mice lacking Il23r specifically in Tregs and exhibits a competitive advantage over IL-23R-sufficient Tregs during inflammation. Finally, IL-23 antagonizes liver X receptor pathway, cellular cholesterol transporter Abca1, and increases Treg apoptosis. Our results show that IL-23R signaling regulates intestinal Tregs by increasing cell turnover, antagonizing suppression, and decreasing cholesterol efflux. These results suggest that IL-23 negatively regulates Tregs in the intestine with potential implications for promoting chronic inflammation in patients with IBD.
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Colitis , Enfermedades Inflamatorias del Intestino , Animales , Humanos , Ratones , Colitis/patología , Factores de Transcripción Forkhead/metabolismo , Inflamación/patología , Enfermedades Inflamatorias del Intestino/patología , Interleucina-23/metabolismo , Linfocitos T ReguladoresRESUMEN
Tumor-associated macrophages (TAM) play an important role in supporting tumor growth and suppressing antitumor immune responses, and TAM infiltration has been associated with poor patient prognosis in various cancers. TAMs can be classified as pro-inflammatory, M1-like, or anti-inflammatory, M2-like. While multiple factors within the tumor microenvironment affect the recruitment, polarization, and functions of TAMs, accumulating evidence suggests that Wnt signaling represents an important, targetable driver of an immunosuppressive, M2-like TAM phenotype. TAM production of Wnt ligands mediates TAM-tumor cross-talk to support cancer cell proliferation, invasion, and metastasis. Targeting TAM polarization and the protumorigenic functions of TAMs through inhibitors of Wnt signaling may prove a beneficial treatment strategy in cancers where macrophages are prevalent in the microenvironment.
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Macrófagos Asociados a Tumores , Vía de Señalización Wnt , Macrófagos , Proliferación Celular , Fenotipo , Microambiente TumoralRESUMEN
Introduction: The pro-inflammatory cytokine interleukin-23 (IL-23) has been implicated in colorectal cancer (CRC). Yet, the cell-specific contributions of IL-23 receptor (IL-23R) signaling in CRC remain unknown. One of the cell types that highly expresses IL-23R are colonic regulatory T cells (Treg cells). The aim of this study was to define the contribution of Treg cell-specific IL-23R signaling in sporadic and inflammation-associated CRC. Methods: In mice, the role of IL-23R in Treg cells in colitis-associated cancer (CAC) was investigated using azoxymethane/dextran sodium sulphate in wild-type Treg cell reporter mice (WT, Foxp3 YFP-iCre), and mice harboring a Treg cell-specific deletion of IL-23 (Il23r ΔTreg). The role of IL-23R signaling in Treg cells in sporadic CRC was examined utilizing orthotopic injection of the syngeneic colon cancer cell line MC-38 submucosally into the colon/rectum of mice. The function of macrophages was studied using clodronate. Finally, single-cell RNA-seq of a previously published dataset in human sporadic cancer was reanalyzed to corroborate these findings. Results: In CAC, Il23r ΔTreg mice had increased tumor size and increased dysplasia compared to WT mice that was associated with decreased tumor-infiltrating macrophages. In the sporadic cancer model, Il23r ΔTreg mice had increased survival and decreased tumor size compared to WT mice. Additionally, MC-38 tumors of Il23r ΔTreg mice exhibited a higher frequency of pro-inflammatory macrophages and IL-17 producing CD4+ T cells. The decreased tumor size in Il23r ΔTreg mice was macrophage-dependent. These data suggest that loss of IL-23R signaling in Treg cells permits IL-17 production by CD4+ T cells that in turn promotes pro-inflammatory macrophages to clear tumors. Finally, analysis of TCGA data and single-cell RNA-seq analysis of a previously published dataset in human sporadic cancer, revealed that IL23R was highly expressed in CRC compared to other cancers and specifically in tumor-associated Treg cells. Conclusion: Inflammation in colorectal carcinogenesis differs with respect to the contribution of IL-23R signaling in regulatory T cells.
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BACKGROUND & AIMS: Inflammatory bowel disease (IBD) is characterized by severe gastrointestinal inflammation, but many patients experience extra-intestinal disease. Bone loss is one common extra-intestinal manifestation of IBD that occurs through dysregulated interactions between osteoclasts and osteoblasts. Systemic inflammation has been postulated to contribute to bone loss, but the specific pathologic mechanisms have not yet been fully elucidated. We hypothesized that intestinal inflammation leads to bone loss through increased abundance and altered function of osteoclast progenitors. METHODS: We used chemical, T cell driven, and infectious models of intestinal inflammation to determine the impact of intestinal inflammation on bone volume, the skeletal cytokine environment, and the cellular changes to pre-osteoclast populations within bone marrow. Additionally, we evaluated the potential for monoclonal antibody treatment against an inflammation-induced osteoclast co-receptor, myeloid DNAX activation protein 12-associating lectin-1 (MDL-1) to reduce bone loss during colitis. RESULTS: We observed significant bone loss across all models of intestinal inflammation. Bone loss was associated with an increase in pro-osteoclastogenic cytokines within the bone and an expansion of a specific Cd11b-/loLy6Chi osteoclast precursor (OCP) population. Intestinal inflammation led to altered OCP expression of surface receptors involved in osteoclast differentiation and function, including the pro-osteoclastogenic co-receptor MDL-1. OCPs isolated from mice with intestinal inflammation demonstrated enhanced osteoclast differentiation ex vivo compared to controls, which was abrogated by anti-MDL-1 antibody treatment. Importantly, in vivo anti-MDL-1 antibody treatment ameliorated bone loss during intestinal inflammation. CONCLUSIONS: Collectively, these data implicate the pathologic expansion and altered function of OCPs expressing MDL-1 in bone loss during IBD.
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Resorción Ósea , Enfermedades Inflamatorias del Intestino , Lectinas Tipo C , Osteoclastos , Osteogénesis , Receptores de Superficie Celular , Animales , Anticuerpos Monoclonales/metabolismo , Resorción Ósea/genética , Resorción Ósea/metabolismo , Resorción Ósea/patología , Diferenciación Celular/fisiología , Citocinas/metabolismo , Inflamación/genética , Inflamación/metabolismo , Inflamación/patología , Enfermedades Inflamatorias del Intestino/genética , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/patología , Intestinos/metabolismo , Lectinas/metabolismo , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Ratones , Osteoclastos/metabolismo , Osteoclastos/patología , Osteogénesis/genética , Osteogénesis/fisiología , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismoRESUMEN
Macrophages play a crucial role in the inflammatory response to the human stomach pathogen Helicobacter pylori, which infects half of the world's population and causes gastric cancer. Recent studies have highlighted the importance of macrophage immunometabolism in their activation state and function. We have demonstrated that the cysteine-producing enzyme cystathionine γ-lyase (CTH) is upregulated in humans and mice with H. pylori infection. Here, we show that induction of CTH in macrophages by H. pylori promoted persistent inflammation. Cth-/- mice had reduced macrophage and T cell activation in H. pylori-infected tissues, an altered metabolome, and decreased enrichment of immune-associated gene networks, culminating in decreased H. pylori-induced gastritis. CTH is downstream of the proposed antiinflammatory molecule, S-adenosylmethionine (SAM). Whereas Cth-/- mice exhibited gastric SAM accumulation, WT mice treated with SAM did not display protection against H. pylori-induced inflammation. Instead, we demonstrated that Cth-deficient macrophages exhibited alterations in the proteome, decreased NF-κB activation, diminished expression of macrophage activation markers, and impaired oxidative phosphorylation and glycolysis. Thus, through altering cellular respiration, CTH is a key enhancer of macrophage activation, contributing to a pathogenic inflammatory response that is the universal precursor for the development of H. pylori-induced gastric disease.
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Infecciones por Helicobacter , Helicobacter pylori , Animales , Cistationina gamma-Liasa/genética , Cistationina gamma-Liasa/metabolismo , Inflamación/metabolismo , Macrófagos/metabolismo , RatonesRESUMEN
Aberrant epithelial differentiation and regeneration contribute to colon pathologies, including inflammatory bowel disease (IBD) and colitis-associated cancer (CAC). Myeloid translocation gene 16 (MTG16, also known as CBFA2T3) is a transcriptional corepressor expressed in the colonic epithelium. MTG16 deficiency in mice exacerbates colitis and increases tumor burden in CAC, though the underlying mechanisms remain unclear. Here, we identified MTG16 as a central mediator of epithelial differentiation, promoting goblet and restraining enteroendocrine cell development in homeostasis and enabling regeneration following dextran sulfate sodium-induced (DSS-induced) colitis. Transcriptomic analyses implicated increased Ephrussi box-binding transcription factor (E protein) activity in MTG16-deficient colon crypts. Using a mouse model with a point mutation that attenuates MTG16:E protein interactions (Mtg16P209T), we showed that MTG16 exerts control over colonic epithelial differentiation and regeneration by repressing E protein-mediated transcription. Mimicking murine colitis, MTG16 expression was increased in biopsies from patients with active IBD compared with unaffected controls. Finally, uncoupling MTG16:E protein interactions partially phenocopied the enhanced tumorigenicity of Mtg16-/- colon in the azoxymethane/DSS-induced model of CAC, indicating that MTG16 protects from tumorigenesis through additional mechanisms. Collectively, our results demonstrate that MTG16, via its repression of E protein targets, is a key regulator of cell fate decisions during colon homeostasis, colitis, and cancer.
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Colitis , Enfermedades Inflamatorias del Intestino , Animales , Carcinogénesis/genética , Carcinogénesis/metabolismo , Transformación Celular Neoplásica/genética , Colitis/inducido químicamente , Colitis/genética , Colitis/metabolismo , Sulfato de Dextran/toxicidad , Humanos , Enfermedades Inflamatorias del Intestino/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factores de Transcripción/genéticaRESUMEN
IL-17R signaling is required for control of extracellular pathogens and is also implicated in development of chronic inflammatory processes. The response to the human pathogen Helicobacter pylori results in Th1 and Th17 cell activation and a chronic inflammatory process that can lead to adverse outcomes, such as gastric cancer. Previously, we identified IL-17RA as a requirement for the recruitment of neutrophils and control of H. pylori colonization in the gastric mucosa. Unexpectedly, H. pylori-infected Il17ra -/- mice had significantly more chronic inflammation than H. pylori-infected wild-type mice. In this study, human epithelial cell lines and murine models were used to investigate differential roles for IL-17A, IL-17F, and IL-17A/F during H. pylori infection. Moreover, the hypothesis that IL-17RA signaling, specifically in lymphocytes, provides an autocrine feedback loop that downregulates Th17 cytokine production was investigated. The data indicate that epithelial cells exhibit a stronger response to IL-17A and IL-17A/F than IL-17F, and that IL-17A and IL-17A/F can synergize with TNF and IL-22 to induce antimicrobial genes of gastric epithelial cells. In vivo deficiencies of IL-17A or IL-17F alone did not significantly change the immunopathological response to H. pylori, but if both cytokines were absent, a hyperinflammatory lymphocytic response developed. Using a cre/flox targeting approach for IL-17RA combined with infection, our findings demonstrate that increased chronic inflammation in Il17ra -/- mice was not attributed to a T cell-intrinsic defect. These data imply that IL-17A and IL-17F may have overlapping roles in maintenance of the gastric mucosal response to infection.
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Infecciones por Helicobacter , Helicobacter pylori , Animales , Inflamación , Interleucina-17/metabolismo , Ratones , Receptores de Interleucina-17/genéticaRESUMEN
Colorectal cancers (CRCs) arise from precursor polyps whose cellular origins, molecular heterogeneity, and immunogenic potential may reveal diagnostic and therapeutic insights when analyzed at high resolution. We present a single-cell transcriptomic and imaging atlas of the two most common human colorectal polyps, conventional adenomas and serrated polyps, and their resulting CRC counterparts. Integrative analysis of 128 datasets from 62 participants reveals adenomas arise from WNT-driven expansion of stem cells, while serrated polyps derive from differentiated cells through gastric metaplasia. Metaplasia-associated damage is coupled to a cytotoxic immune microenvironment preceding hypermutation, driven partly by antigen-presentation differences associated with tumor cell-differentiation status. Microsatellite unstable CRCs contain distinct non-metaplastic regions where tumor cells acquire stem cell properties and cytotoxic immune cells are depleted. Our multi-omic atlas provides insights into malignant progression of colorectal polyps and their microenvironment, serving as a framework for precision surveillance and prevention of CRC.
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Pólipos del Colon/patología , Neoplasias Colorrectales/patología , Microambiente Tumoral , Inmunidad Adaptativa , Adenoma/genética , Adenoma/patología , Adulto , Anciano , Animales , Carcinogénesis/genética , Carcinogénesis/patología , Muerte Celular , Diferenciación Celular , Pólipos del Colon/genética , Pólipos del Colon/inmunología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/inmunología , Progresión de la Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes , Heterogeneidad Genética , Humanos , Masculino , Ratones , Persona de Mediana Edad , Mutación/genética , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , RNA-Seq , Reproducibilidad de los Resultados , Análisis de la Célula Individual , Microambiente Tumoral/inmunologíaRESUMEN
BACKGROUND: The gut microbiome is altered in patients with inflammatory bowel disease, yet how these alterations contribute to intestinal inflammation is poorly understood. Murine models have demonstrated the importance of the microbiome in colitis since colitis fails to develop in many genetically susceptible animal models when re-derived into germ-free environments. We have previously shown that Wiskott-Aldrich syndrome protein (WASP)-deficient mice (Was-/-) develop spontaneous colitis, similar to human patients with loss-of-function mutations in WAS. Furthermore, we showed that the development of colitis in Was-/- mice is Helicobacter dependent. Here, we utilized a reductionist model coupled with multi-omics approaches to study the role of host-microbe interactions in intestinal inflammation. RESULTS: Was-/- mice colonized with both altered Schaedler flora (ASF) and Helicobacter developed colitis, while those colonized with either ASF or Helicobacter alone did not. In Was-/- mice, Helicobacter relative abundance was positively correlated with fecal lipocalin-2 (LCN2), a marker of intestinal inflammation. In contrast, WT mice colonized with ASF and Helicobacter were free of inflammation and strikingly, Helicobacter relative abundance was negatively correlated with LCN2. In Was-/- colons, bacteria breach the mucus layer, and the mucosal relative abundance of ASF457 Mucispirillum schaedleri was positively correlated with fecal LCN2. Meta-transcriptomic analyses revealed that ASF457 had higher expression of genes predicted to enhance fitness and immunogenicity in Was-/- compared to WT mice. In contrast, ASF519 Parabacteroides goldsteinii's relative abundance was negatively correlated with LCN2 in Was-/- mice, and transcriptional analyses showed lower expression of genes predicted to facilitate stress adaptation by ASF519 in Was-/-compared to WT mice. CONCLUSIONS: These studies indicate that the effect of a microbe on the immune system can be context dependent, with the same bacteria eliciting a tolerogenic response under homeostatic conditions but promoting inflammation in immune-dysregulated hosts. Furthermore, in inflamed environments, some bacteria up-regulate genes that enhance their fitness and immunogenicity, while other bacteria are less able to adapt and decrease in abundance. These findings highlight the importance of studying host-microbe interactions in different contexts and considering how the transcriptional profile and fitness of bacteria may change in different hosts when developing microbiota-based therapeutics. Video abstract.