An in vitro chronic damage model impairs inflammatory and regenerative responses in human colonoid monolayers

Rees, William D.; Telkar, Nikita; Lin, David T.S.; Wong, May Q.; Poloni, Chad; Fathi, Ayda; Kobor, Michael S.; Zachos, Nicholas C.; Steiner, Theodore S.

doi:10.1016/j.celrep.2021.110283

Cited by 13 publications

(7 citation statements)

References 103 publications

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“…Based on that, we do not recommend combining cohorts of fresh and thawed organoid cultures in projects where comparative analysis between clinical groups of patients are planned. Single cells derived from dissociated organoids are also useful for generating those primary 2D cultures that had already shown high versatility in multiple applications when seeded on conventional plastic plates ( S3A Fig ), Transwell inserts ( S3B Fig ) or microfluidic chips [ 26 , 44 – 47 ]. In this case, we observed that more aggressive organoid dissociation, like the one mediated by trypsin, is also suitable for growing 2D cultures.…”

Section: Resultsmentioning

confidence: 99%

Generation of human colon organoids from healthy and inflammatory bowel disease mucosa

Dotti

Mayorgas²,

Salas³

2022

PLoS ONE

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Ulcerative colitis and Crohn’s disease are chronic inflammatory bowel diseases (IBD) of unknown cause characterized by a relapsing-remitting behavior. Growing evidence supports the idea that the epithelial barrier plays a central role in the pathogenesis of IBD as well as in its evolution over time, thus representing a potential target for novel therapeutic options. In the last decade, the introduction of 3D epithelial cultures from ex vivo-expanded intestinal adult stem cells (ASCs) has impacted our ability to study the function of the epithelium in several gastrointestinal disorders, including IBD. Here, we describe in detail a reproducible protocol to generate Matrigel-embedded epithelial organoids from ASCs of non-IBD and IBD donors using small colonic biopsies, including steps for its optimization. A slightly modified version of this protocol is also provided in case surgical samples are used. With this method, epithelial organoids can be expanded over several passages, thereby generating a large quantity of viable cells that can be used in multiple downstream analyses including genetic, transcriptional, proteomic and/or functional studies. In addition, 3D cultures generated using our protocol are suitable for the establishment of 2D cultures, which can model relevant cell-to-cell interactions that occur in IBD mucosa.

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Section: Resultsmentioning

confidence: 99%

Generation of human colon organoids from healthy and inflammatory bowel disease mucosa

Dotti

Mayorgas²,

Salas³

2022

PLoS ONE

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“…A number of organoid-derived monolayer systems have been described recently [17][18][19][20][21][31][32][33][34][35][36][37][38] .…”

Section: Discussionmentioning

confidence: 99%

Development of a high-throughput barrier function assay using primary human gut organoids

Wright

Shaffer

et al. 2022

Preprint

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Disruptions in the gut epithelial barrier can lead to the development of chronic indications such as inflammatory bowel disease (IBD). Historically, barrier function has been assessed in cancer cell lines, which do not contain all human intestinal cell types, leading to poor translatability. To bridge this gap, we adapted human primary gut organoids grown as monolayers to assess barrier function. In this work we describe and characterize a novel 96-well human gut organoid-derived monolayer system that enables medium/high-throughput quantitative assessment of candidate therapeutics. Normal human intestine differentiation patterns and barrier function were characterized and confirmed to recapitulate key aspects of in vivo biology. Next, cellular response to TNF-α (a central driver of IBD) was determined using a diverse cadre of readouts. These outputs included transcription factor phosphorylation, target gene expression, cytokine/chemokine production, and barrier function. Additionally, we showed that TNF-α pathway antagonists rescued damage caused by TNF-α in a dose-dependent manner, indicating that this system is suitable for quantitative assessment of barrier modulating factors. Taken together, we have established a robust primary cell-based 96-well system capable of interrogating questions around mucosal response. This system is well suited to provide pivotal functional data to support translational target and drug discovery efforts.

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“…This, in turn, stimulates the expression of immune factors such as TNF‐α, interferon‐γ (IFN‐γ), and IL‐10, ultimately exacerbating the severity of IBD 22 (Figure 1). The levels of methylation modification in the ER gene were higher in patients with IBD, suggesting that methylation may serve as a potential diagnostic marker for IBD 23 . In contrast, analysis of IBD patients found no evidence of DNA methylation in the IRF5 promoter region, indicating that IRF5 is unlikely to play a role in the development of IBD 19 .…”

Section: Epigenetic Modifications In Ibdmentioning

confidence: 91%

“…The levels of methylation modification in the ER gene were higher in patients with IBD, suggesting that methylation may serve as a potential diagnostic marker for IBD. 23 In contrast, analysis of IBD patients found no evidence of DNA methylation in the IRF5 promoter region, indicating that IRF5 is unlikely to play a role in the development of IBD. 19 Additionally, the dysregulation of IFN-γ, caused by increased DNA methylation of the IFN-γ gene in T cells from peripheral blood of IBD patients, contributed to IBD's exacerbation.…”

Section: Dna Epigenetic Modifications In Ibdmentioning

confidence: 94%

Role of epigenetic modifications and aging in inflammatory bowel disease

Du,

Li,

Zhou

et al. 2023

MedComm – Future Medicine

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Inflammatory bowel disease (IBD), which encompasses ulcerative colitis (UC) and Crohn's disease (CD), refers to a chronic and recurrent nonspecific inflammatory condition affecting the mucosal and submucosal layers of the intestines. A positive family history has been identified as a risk factor for the onset of IBD, likely influenced by genetic and environmental factors. In addition to intestinal damage, patients may have extraintestinal manifestations such as inflammation of the skin, eyes, and joints, inflammation of the liver or bile ducts, kidney stones, iron‐deficiency anemia, and growth retardation in children, which may complicate diagnosis and treatment. Therefore, investigating the mechanisms of IBD and finding precise therapeutic targets provides enormous benefits to patients with IBD. Multiple studies have consistently demonstrated the influential role of dietary metabolism and aging in the development of IBD. Moreover, emerging evidence suggests that aging often coincides with alterations in epigenetic modifications, while diet metabolism mediates these epigenetic changes. Epigenetics has emerged as a prospective field for identifying novel biomarkers to facilitate the diagnosis, prognosis, and treatment of diseases. Therefore, in this perspective, we summarize the cross‐talk between epigenetic modifications, diet metabolism, and aging in the pathogenesis of IBD and attempt to identify new potential therapeutic targets and strategies.

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An in vitro chronic damage model impairs inflammatory and regenerative responses in human colonoid monolayers

Cited by 13 publications

References 103 publications

Generation of human colon organoids from healthy and inflammatory bowel disease mucosa

Generation of human colon organoids from healthy and inflammatory bowel disease mucosa

Development of a high-throughput barrier function assay using primary human gut organoids

Role of epigenetic modifications and aging in inflammatory bowel disease

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