NF-κB2 signalling in enteroids modulates enterocyte responses to secreted factors from bone marrow-derived dendritic cells

Jones, Lauren G.; Vaida, Andra; Thompson, Louise; Ikuomola, Felix I.; Caamaño, Jorge H.; Burkitt, Michael D.; Miyajima, Fábio; Williams, Jonathan; Campbell, Barry J.; Pritchard, D. Mark; Duckworth, Carrie A.

doi:10.1038/s41419-019-2129-5

Cited by 24 publications

(23 citation statements)

References 49 publications

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“…At denser seeding, the enteroids gained a circular morphology with a thinned epithelial layer (Figure 2 ). This morphology indicates a condition of increased cell death without compensatory cell proliferation, as previously described in murine enteroids [ 23 ]. Both intact and fragmented enteroids could be frozen at −80 °C and brought up in culture with 80–90% recovery.…”

Section: Resultssupporting

confidence: 74%

Generation of equine enteroids and enteroid-derived 2D monolayers that are responsive to microbial mimics

Hellman

2021

Vet Res

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Enteroid cultures are three-dimensional in vitro models that reflect the cellular composition and architecture of the small intestine. One limitation with the enteroid conformation is the enclosed lumen, making it difficult to expose the apical surface of the epithelium to experimental treatments. The present study was therefore conducted to generate cultures of equine enteroids and to develop methods for culture of enteroid-derived cells on a two-dimensional plane, enabling easy access to the apical surface of the epithelium. Equine enteroids were established from small intestinal crypts within 7–9 days of culture. Transcriptional analysis of cell type markers confirmed the presence of enterocytes, stem-, Paneth-, proliferative-, enteroendocrine-, goblet- and tuft cells. This cellular composition was maintained over multiple passages, showing that the enteroids can be kept for prolonged periods. The transfer from 3D enteroids to 2D monolayers slightly modified the relative expression levels of the cell type markers, indicating a decrease of goblet- and Paneth cells in the monolayers. Stimulation with the TLR2, 3 and 4 agonists Pam3CSK4, Poly I:C and LPS, respectively, induced the pro-inflammatory cytokines TNF-α and IL-8, while the TLR5 agonist FliC only induced TNF-α. In addition, an up-regulation of TGF-β, IL-33 and IFN-β was recorded after exposure to lipofected Poly I:C that also affected the monolayer integrity. Thus, the equine enteroid-derived 2D monolayers described in the present study show both genetic and functional similarities with the equine intestine making it an interesting in vitro model for studies demanding access to the apical surface, e.g. in studies of host-microbe interactions.

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Section: Resultssupporting

confidence: 74%

Generation of equine enteroids and enteroid-derived 2D monolayers that are responsive to microbial mimics

Hellman

2021

Vet Res

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“…Sections were flushed with ice-cold sterile PBS pH 7.3, approximately 10 times until clear. Tissue sections were processed in 2 mM ethylenediaminetetraacetic acid (EDTA) chelation buffer to generate intestinal crypts, which were then resuspended in Matrigel (Corning; Loughborough, UK) and cultured over 7 days to generate enteroids as previously described ( 29 ). Cultures were passaged at day 7 by mechanically disruption through a 27G needle, resuspended in Matrigel, and plated out again on 24-well plates.…”

Section: Methodsmentioning

confidence: 99%

Impact of Interleukin 10 Deficiency on Intestinal Epithelium Responses to Inflammatory Signals

Παπουτσοπουλου¹,

Pollock²,

Walker³

et al. 2021

Front. Immunol.

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Interleukin 10 (IL-10) is a pleiotropic, anti-inflammatory cytokine that has a major protective role in the intestine. Although its production by cells of the innate and adaptive immune system has been extensively studied, its intrinsic role in intestinal epithelial cells is poorly understood. In this study, we utilised both ATAC sequencing and RNA sequencing to define the transcriptional response of murine enteroids to tumour necrosis factor (TNF). We identified that the key early phase drivers of the transcriptional response to TNF within intestinal epithelium were NFκB transcription factor dependent. Using wild-type and Il10−/− enteroid cultures, we showed an intrinsic, intestinal epithelium specific effect of IL-10 deficiency on TNF-induced gene transcription, with significant downregulation of identified NFκB target genes Tnf, Ccl20, and Cxcl10, and delayed overexpression of NFκB inhibitor encoding genes, Nfkbia and Tnfaip3. IL-10 deficiency, or immunoblockade of IL-10 receptor, impacted on TNF-induced endogenous NFκB activity and downstream NFκB target gene transcription. Intestinal epithelium-derived IL-10 appears to play a crucial role as a positive regulator of the canonical NFκB pathway, contributing to maintenance of intestinal homeostasis. This is particularly important in the context of an inflammatory environment and highlights the potential for future tissue-targeted IL-10 therapeutic intervention.

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“…In this study, cytokines secreted by macrophages such as IL-6, IL-8, IFNγ, and TGFβ1 might contribute to the potential roles of macrophages in enhancing the maturation of the intestinal epithelium and the thickening of the physical barrier ( 63 ). Similarly, Jones et al demonstrated that cytokines released by bone marrow-derived dendritic cells could modulate intestinal barrier integrity, intestinal cell proliferation, and cell death through NF-κB2 signaling ( 64 ). Despite the fact that these studies expand our knowledge about the immune cell-ISC interactions, we must realize that these networks may be far more complicated than we currently understand.…”

Section: Intestinal Stem Cell Nichementioning

confidence: 99%

“…Macrophages and dendritic cells exist throughout the lamina propria of the gut and contribute to innate and adaptive immunity, thus maintaining gut homeostasis. The co-culture of intestinal organoids with macrophages and dendritic cells has also been successfully performed to study complex networks between immune cells and ISC (62)(63)(64). Macrophages have been shown to promote mucosal repair through activation of Wntsignaling in a mouse model of IBD (83).…”

Section: Other Immune Cellsmentioning

confidence: 99%

Intestinal Stem Cells and Immune Cell Relationships: Potential Therapeutic Targets for Inflammatory Bowel Diseases

et al. 2021

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The mammalian intestine is the largest immune organ that contains the intestinal stem cells (ISC), differentiated epithelial cells (enterocytes, Paneth cells, goblet cells, tuft cells, etc.), and gut resident-immune cells (T cells, B cells, dendritic cells, innate lymphoid cell, etc.). Inflammatory bowel disease (IBD), a chronic inflammatory disease characterized by mucosa damage and inflammation, threatens the integrity of the intestine. The continuous renewal and repair of intestinal mucosal epithelium after injury depend on ISCs. Inflamed mucosa healing could be a new target for the improvement of clinical symptoms, disease recurrence, and resection-free survival in IBD treated patients. The knowledge about the connections between ISC and immune cells is expanding with the development of in vitro intestinal organoid culture and single-cell RNA sequencing technology. Recent findings implicate that immune cells such as T cells, ILCs, dendritic cells, and macrophages and cytokines secreted by these cells are critical in the regeneration of ISCs and intestinal epithelium. Transplantation of ISC to the inflamed mucosa may be a new therapeutic approach to reconstruct the epithelial barrier in IBD. Considering the links between ISC and immune cells, we predict that the integration of biological agents and ISC transplantation will revolutionize the future therapy of IBD patients.

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NF-κB2 signalling in enteroids modulates enterocyte responses to secreted factors from bone marrow-derived dendritic cells

Cited by 24 publications

References 49 publications

Generation of equine enteroids and enteroid-derived 2D monolayers that are responsive to microbial mimics

Generation of equine enteroids and enteroid-derived 2D monolayers that are responsive to microbial mimics

Impact of Interleukin 10 Deficiency on Intestinal Epithelium Responses to Inflammatory Signals

Intestinal Stem Cells and Immune Cell Relationships: Potential Therapeutic Targets for Inflammatory Bowel Diseases

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