Eosinophils have emerged as multifaceted cells that contribute to tissue homeostasis. However, the impact of the microbiota on their frequency and function at mucosal sites remains unclear. Here, we ...investigated the role of the microbiota in the regulation of enteric eosinophils. We found that small intestinal (SI) eosinophilia was significantly greater in germ-free (GF) mice compared to specific pathogen free (SPF) controls. This was associated with changes in the production of enteric signals that regulate eosinophil attraction and survival, and was fully reversed by complex colonization. Additionally, SI eosinophils of GF mice exhibited more cytoplasmic protrusions and less granule content than SPF controls. Lastly, we generated a novel strain of eosinophil-deficient GF mice. These mice displayed intestinal fibrosis and were less prone to allergic sensitization as compared to GF controls. Overall, our study demonstrates that commensal microbes regulate intestinal eosinophil frequency and function, which impacts tissue repair and allergic sensitization to food antigens. These data support a critical interplay between the commensal microbiota and intestinal eosinophils in shaping homeostatic, innate, and adaptive immune processes in health and disease.
Chronic intestinal inflammation contributes to pathologies such as inflammatory bowel disease (IBD) and colon cancer. While the precise etiology remains controversial, IBD is believed to manifest as ...a result of various factors. We previously reported that intestinal-specific overexpression of the transcription factor Kaiso results in an intestinal inflammatory response; however, the cause of this inflammation is unknown. To elucidate the underlying mechanism(s) of the Kaiso-mediated intestinal inflammatory phenotype, we evaluated two independent transgenic mouse lines that express varying levels of Kaiso (KaisoTg). Histological analyses of KaisoTg mice revealed intestinal damage including thickening of the mucosa, intestinal "lesions" and crypt abscesses, which are reminiscent of IBD pathology. Additionally, higher Kaiso levels induced intestinal neutrophilia as early as 12 weeks, which worsened as the mice aged. Notably, the Kaiso-induced intestinal inflammation correlated with a leaky intestinal barrier and mis-regulation of E-cadherin expression and localization. Interestingly, Kaiso overexpression resulted in reduced proliferation but enhanced migration of intestinal epithelial cells prior to the onset of inflammation. Collectively, these data suggest that Kaiso plays a role in regulating intestinal epithelial cell integrity and function, dysregulation of which contributes to a chronic inflammatory phenotype as mice age.
The function and structure of the mammalian epithelial cell layer is maintained by distinct intercellular adhesion complexes including
adherens
junctions (AJs), tight junctions, and desmosomes. The ...AJ is most integral for stabilizing cell-cell adhesion and conserving the structural integrity of epithelial tissues. AJs are comprised of the transmembrane protein E-cadherin and cytoplasmic catenin cofactors (α, β, γ, and p120-catenin). One organ where malfunction of AJ is a major contributor to disease states is the mammalian intestine. In the intestine, cell-cell adhesion complexes work synergistically to maintain structural integrity and homeostasis of the epithelium and prevent its malfunction. Consequently, when AJ integrity is compromised in the intestinal epithelium, the ensuing homeostatic disruption leads to diseases such as inflammatory bowel disease and colorectal carcinoma. In addition to their function at the plasma membrane, protein components of AJs also have nuclear functions and are thus implicated in regulating gene expression and intracellular signaling. Within the nucleus, AJ proteins have been shown to interact with transcription factors such as TCF/LEF and Kaiso (
ZBTB33
), which converge on the canonical Wnt signaling pathway. The multifaceted nature of AJ proteins highlights their complexity in modulating homeostasis and emphasizes the importance of their subcellular localization and expression in the mammalian intestine. In this review, we summarize the nuclear roles of AJ proteins in intestinal tissues; their interactions with transcription factors and how this leads to crosstalk with canonical Wnt signaling; and how nuclear AJ proteins are implicated in intestinal homeostasis and disease.
Since its discovery, several studies have implicated the POZ-ZF protein Kaiso in both developmental and tumorigenic processes. However, most of the information regarding Kaiso's function to date has ...been gleaned from studies in Xenopus laevis embryos and mammalian cultured cells. To examine Kaiso's role in a relevant, mammalian organ-specific context, we generated and characterized a Kaiso transgenic mouse expressing a murine Kaiso transgene under the control of the intestine-specific villin promoter. Kaiso transgenic mice were viable and fertile but pathological examination of the small intestine revealed distinct morphological changes. Kaiso transgenics (Kaiso(Tg/+)) exhibited a crypt expansion phenotype that was accompanied by increased differentiation of epithelial progenitor cells into secretory cell lineages; this was evidenced by increased cell populations expressing Goblet, Paneth and enteroendocrine markers. Paradoxically however, enhanced differentiation in Kaiso(Tg/+) was accompanied by reduced proliferation, a phenotype reminiscent of Notch inhibition. Indeed, expression of the Notch signalling target HES-1 was decreased in Kaiso(Tg/+) animals. Finally, our Kaiso transgenics exhibited several hallmarks of inflammation, including increased neutrophil infiltration and activation, villi fusion and crypt hyperplasia. Interestingly, the Kaiso binding partner and emerging anti-inflammatory mediator p120(ctn) is recruited to the nucleus in Kaiso(Tg/+) mice intestinal cells suggesting that Kaiso may elicit inflammation by antagonizing p120(ctn) function.
The relationship between autophagic cell death and apoptosis is a poorly understood aspect of programmed cell death (PCD). We have examined this relationship by studying the elimination of an ...extra-embryonic tissue, known as the amnioserosa (AS), during Drosophila development. The AS becomes autophagic during the final stages of embryogenesis; ultimately, however, the elimination of the AS involves caspase-dependent nuclear fragmentation, tissue dissociation, and engulfment by phagocytic macrophages. Mutants that are defective in the activation or execution of caspase-dependent PCD fail to degrade and eliminate the AS but show no abatement in AS autophagy. Sustained autophagy does not, therefore, necessarily result in cell death. Surprisingly, the down-regulation of autophagy also results in a persistent AS phenotype and reduced cell death. Conversely, up-regulation of autophagy results in caspase-dependent premature AS dissociation. These observations are consistent with the interpretation that autophagy is a prerequisite for caspase-dependent cell death in the AS.
A number of food allergies (eg, fish, shellfish, and nuts) are lifelong, without any disease-transforming therapies, and unclear in their underlying immunology. Clinical manifestations of food ...allergy are largely mediated by IgE. Although persistent IgE titers have been attributed conventionally to long-lived IgE
plasma cells (PCs), this has not been directly and comprehensively tested.
We sought to evaluate mechanisms underlying persistent IgE and allergic responses to food allergens.
We used a model of peanut allergy and anaphylaxis, various knockout mice, adoptive transfer experiments, and in vitro assays to identify mechanisms underlying persistent IgE humoral immunity over almost the entire lifespan of the mouse (18-20 months).
Contrary to conventional paradigms, our data show that clinically relevant lifelong IgE titers are not sustained by long-lived IgE
PCs. Instead, lifelong reactivity is conferred by allergen-specific long-lived memory B cells that replenish the IgE
PC compartment. B-cell reactivation requires allergen re-exposure and IL-4 production by CD4 T cells. We define the half-lives of antigen-specific germinal centers (23.3 days), IgE
and IgG
PCs (60 and 234.4 days, respectively), and clinically relevant cell-bound IgE (67.3 days).
These findings can explain lifelong food allergies observed in human subjects as the consequence of allergen exposures that recurrently activate memory B cells and identify these as a therapeutic target with disease-transforming potential.
A number of food allergies (eg, fish, shellfish, and nuts) are lifelong, without any disease-transforming therapies, and unclear in their underlying immunology. Clinical manifestations of food ...allergy are largely mediated by IgE. Although persistent IgE titers have been attributed conventionally to long-lived IgE+ plasma cells (PCs), this has not been directly and comprehensively tested.
We sought to evaluate mechanisms underlying persistent IgE and allergic responses to food allergens.
We used a model of peanut allergy and anaphylaxis, various knockout mice, adoptive transfer experiments, and in vitro assays to identify mechanisms underlying persistent IgE humoral immunity over almost the entire lifespan of the mouse (18-20 months).
Contrary to conventional paradigms, our data show that clinically relevant lifelong IgE titers are not sustained by long-lived IgE+ PCs. Instead, lifelong reactivity is conferred by allergen-specific long-lived memory B cells that replenish the IgE+ PC compartment. B-cell reactivation requires allergen re-exposure and IL-4 production by CD4 T cells. We define the half-lives of antigen-specific germinal centers (23.3 days), IgE+ and IgG1+ PCs (60 and 234.4 days, respectively), and clinically relevant cell-bound IgE (67.3 days).
These findings can explain lifelong food allergies observed in human subjects as the consequence of allergen exposures that recurrently activate memory B cells and identify these as a therapeutic target with disease-transforming potential.
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Constitutive Wnt/β-catenin signaling is a key contributor to colorectal cancer (CRC). Although inactivation of the tumor suppressor adenomatous polyposis coli (APC) is recognized as an early event in ...CRC development, it is the accumulation of multiple subsequent oncogenic insults facilitates malignant transformation. One potential contributor to colorectal carcinogenesis is the POZ-ZF transcription factor Kaiso, whose depletion extends lifespan and delays polyp onset in the widely used ApcMin/+ mouse model of intestinal cancer. These findings suggested that Kaiso potentiates intestinal tumorigenesis, but this was paradoxical as Kaiso was previously implicated as a negative regulator of Wnt/β-catenin signaling. To resolve Kaiso's role in intestinal tumorigenesis and canonical Wnt signaling, we generated a transgenic mouse model (KaisoTg/+) expressing an intestinal-specific myc-tagged Kaiso transgene. We then mated KaisoTg/+ and ApcMin/+ mice to generate KaisoTg/+:ApcMin/+ mice for further characterization. KaisoTg/+:ApcMin/+ mice exhibited reduced lifespan and increased polyp multiplicity compared to ApcMin/+ mice. Consistent with this murine phenotype, we found increased Kaiso expression in human CRC tissue, supporting a role for Kaiso in human CRC. Interestingly, Wnt target gene expression was increased in KaisoTg/+:ApcMin/+ mice, suggesting that Kaiso's function as a negative regulator of canonical Wnt signaling, as seen in Xenopus, is not maintained in this context. Notably, KaisoTg/+:ApcMin/+ mice exhibited increased inflammation and activation of NFκB signaling compared to their ApcMin/+ counterparts. This phenotype was consistent with our previous report that KaisoTg/+ mice exhibit chronic intestinal inflammation. Together our findings highlight a role for Kaiso in promoting Wnt signaling, inflammation and tumorigenesis in the mammalian intestine.
•Kaiso expression is increased in CRC compared to normal intestinal tissue.•Kaiso is highly expressed in polyps of the ApcMin/+ mouse model of colon cancer.•Kaiso overexpression increases polyp burden and shortens lifespan of ApcMin/+ mice.•KaisoTg/+ mice exhibit increased Wnt signaling activity.•KaisoTg/+ and KaisoTg/+:ApcMin/+ mice display increased NFκB signaling and inflammation.
Since the discovery of the p120ctn binding partner, Kaiso, a BTB/POZ transcription factor, several studies have implicated the protein in both development and tumourigenesis. Most information about ...Kaiso’s function in vertebrate development has been gleaned from studies in Xenopus laevis embryos where Kaiso negatively regulates the Wnt signalling pathway. Since the Wnt signalling pathway is crucial in intestinal development, intestinal-specific Kaiso overexpressing mice were generated and characterized to elucidate Kaiso’s role in a mammalian context. Kaiso transgenic (KaisoTg/+) mice were viable and fertile but developed gross histopathological changes in the small intestine. The KaisoTg/+ mice exhibited enlarged crypts accompanied by increased secretory cell differentiation reminiscent of inhibition of the Notch pathway. Indeed, the Notch effector protein, HES1, is decreased in KaisoTg/+ mice. Additionally, KaisoTg/+ mice display a neutrophil-specific intestinal inflammation reminiscent of the knockdown of p120ctn. Interestingly, the KaisoTg/+ mice display decreased p120ctn localization at the membranes and an increase in the neutrophil adhesion molecule, ICAM-1, both of which induce neutrophilia. Notably, the KaisoTg/+ mice developed multiple crypt abscesses over time due to massive neutrophil infiltration of the epithelial cell layers. This is the first study to examine the in vivo roles of Kaiso in a mammalian context and our findings suggest a regulatory role for Kaiso in the inflammatory and Notch signalling pathways.
Thesis
Candidate in Philosophy
Background A number of food allergies (eg, fish, shellfish, and nuts) are lifelong, without any disease-transforming therapies, and unclear in their underlying immunology. Clinical manifestations of ...food allergy are largely mediated by IgE. Although persistent IgE titers have been attributed conventionally to long-lived IgE+ plasma cells (PCs), this has not been directly and comprehensively tested. Objective We sought to evaluate mechanisms underlying persistent IgE and allergic responses to food allergens. Methods We used a model of peanut allergy and anaphylaxis, various knockout mice, adoptive transfer experiments, and in vitro assays to identify mechanisms underlying persistent IgE humoral immunity over almost the entire lifespan of the mouse (18-20 months). Results Contrary to conventional paradigms, our data show that clinically relevant lifelong IgE titers are not sustained by long-lived IgE+ PCs. Instead, lifelong reactivity is conferred by allergen-specific long-lived memory B cells that replenish the IgE+ PC compartment. B-cell reactivation requires allergen re-exposure and IL-4 production by CD4 T cells. We define the half-lives of antigen-specific germinal centers (23.3 days), IgE+ and IgG1+ PCs (60 and 234.4 days, respectively), and clinically relevant cell-bound IgE (67.3 days). Conclusions These findings can explain lifelong food allergies observed in human subjects as the consequence of allergen exposures that recurrently activate memory B cells and identify these as a therapeutic target with disease-transforming potential.
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