Dataset Information

Ulcerative Colitis-Derived Colonoid Culture: A Multi-Mineral-Approach to Improve Barrier Protein Expression.

ABSTRACT:

Background

Recent studies demonstrated that Aquamin^®, a calcium-, magnesium-rich, multi-mineral natural product, improves barrier structure and function in colonoids obtained from the tissue of healthy subjects. The goal of the present study was to determine if the colonic barrier could be improved in tissue from subjects with ulcerative colitis (UC).

Methods

Colonoid cultures were established with colon biopsies from 9 individuals with UC. The colonoids were then incubated for a 2-week period under control conditions (in culture medium with a final calcium concentration of 0.25 mM) or in the same medium supplemented with Aquamin^® to provide 1.5 - 4.5 mM calcium. Effects on differentiation and barrier protein expression were determined using several approaches: phase-contrast and scanning electron microscopy, quantitative histology and immunohistology, mass spectrometry-based proteome assessment and transmission electron microscopy.

Results

Although there were no gross changes in colonoid appearance, there was an increase in lumen diameter and wall thickness on histology and greater expression of cytokeratin 20 (CK20) along with reduced expression of Ki67 by quantitative immunohistology observed with intervention. In parallel, upregulation of several differentiation-related proteins was seen in a proteomic screen with the intervention. Aquamin^®-treated colonoids demonstrated a modest up-regulation of tight junctional proteins but stronger induction of adherens junction and desmosomal proteins. Increased desmosomes were seen at the ultrastructural level. Proteomic analysis demonstrated increased expression of several basement membrane proteins and hemidesmosomal components. Proteins expressed at the apical surface (mucins and trefoils) were also increased as were several additional proteins with anti-microbial activity or that modulate inflammation. Finally, several transporter proteins that affect electrolyte balance (and, thereby affect water resorption) were increased. At the same time, growth and cell cycle regulatory proteins (Ki67, nucleophosmin, and stathmin) were significantly down-regulated. Laminin interactions, matrix formation and extracellular matrix organization were the top three up-regulated pathways with the intervention.

Conclusion

A majority of individuals including patients with UC do not reach the recommended daily intake for calcium and other minerals. To the extent that such deficiencies might contribute to the weakening of the colonic barrier, the findings employing UC tissue-derived colonoids here suggest that adequate mineral intake might improve the colonic barrier.

SUBMITTER: Aslam MN

PROVIDER: S-EPMC7719760 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

Ulcerative Colitis-Derived Colonoid Culture: A Multi-Mineral-Approach to Improve Barrier Protein Expression.

Aslam Muhammad N MN McClintock Shannon D SD Attili Durga D Pandya Shailja S Rehman Humza H Nadeem Daniyal M DM Jawad-Makki Mohamed Ali H MAH Rizvi Areeba H AH Berner Maliha M MM Dame Michael K MK Turgeon Danielle Kim DK Varani James J

Frontiers in cell and developmental biology 20201123

<h4>Background</h4>Recent studies demonstrated that Aquamin<sup>®</sup>, a calcium-, magnesium-rich, multi-mineral natural product, improves barrier structure and function in colonoids obtained from the tissue of healthy subjects. The goal of the present study was to determine if the colonic barrier could be improved in tissue from subjects with ulcerative colitis (UC).<h4>Methods</h4>Colonoid cultures were established with colon biopsies from 9 individuals with UC. The colonoids were then incub ...[more]

PMID: 33330453

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Project description:Background. Recent studies demonstrated that Aquamin®, a calcium-, magnesium-, and multiple trace element-rich natural product, improves barrier structure and function in colonoids obtained from the tissue of healthy subjects. The goal of the present study was to determine if the colonic barrier could be improved in tissue from subjects with ulcerative colitis (UC). Methods. Colonoid cultures were established with colon biopsies from 9 individuals with UC. The colonoids were then incubated for a 2-week period under control conditions (i.e., in culture medium with a final calcium concentration of 0.25 mM) or in the same medium supplemented with Aquamin® to provide 1.5 – 4.5 mM calcium. Effects on differentiation and barrier improvement were determined using several approaches: phase-contrast & scanning electron microscopy, quantitative histology & immunohistology, mass spectrometry-based proteome assessment and transmission electron microscopy. Results. While there were no gross changes in colonoid appearance, there was evidence of increased differentiation (larger lumen diameter and wall thickness) and greater expression of cytokeratin 20 (CK20) and lesser expression of Ki67 with the intervention. Aquamin®-treated colonoids demonstrated a modest up-regulation of tight junctional proteins but stronger induction of adherens junction proteins and desmosomal proteins. Increased desmosomes were seen at the ultrastructural level. Proteomic analysis also demonstrated increased expression of basement membrane proteins and hemidesmosomal components. Proteins expressed at the apical surface (mucins and trefoils) were also increased as were several additional proteins with anti-microbial activity or that modulate inflammation. Similarly, growth and cell cycle regulatory proteins (e.g., Ki67, nucleophosmin, and stathmin) were significantly down-regulated. Laminin interactions, fibronectin matrix formation and extracellular matrix organization were the top three up-regulated pathways with the intervention. Conclusion. A majority of individuals including patients with UC do not reach the recommended daily intake for calcium and other minerals. To the extent that such deficiencies might contribute to the weakening of the colonic barrier, the findings employing UC tissue-derived colonoids here suggest that adequate mineral intake might improve the colonic barrier

Project description:Introduction: Ulcerative colitis is a chronic inflammatory condition, and continuous inflammatory stimulus may lead to barrier dysfunction. The goal of this study was to assess barrier proteomic expression by a red algae-derived multi-mineral intervention in the absence or presence of pro-inflammatory insult. Methods: Human colon organoids were maintained in a control culture medium alone or exposed to lipopolysaccharide with a combination of three pro-inflammatory cytokines [tumor necrosis factor-α, interleukin-1β and interferon-γ (LPS-cytokines)] to mimic the environment in the inflamed colon. Untreated organoids and those exposed to LPS-cytokines were concomitantly treated for 14 days with a multi-mineral product (Aquamin®) that has previously been shown to improve barrier structure/function. The colon organoids were subjected to proteomic analysis to obtain a broad view of the protein changes induced by the two interventions alone and in combination. In parallel, confocal fluorescence microscopy, tissue cohesion and transepithelial electrical resistance (TEER) measurements were used to assess barrier structure/function. Results: The LPS-cytokine mix altered the expression of multiple proteins that influence innate immunity and promote inflammation. Several of these were significantly decreased with Aquamin® alone but only a modest decrease in a subset of these proteins was detected by Aquamin® in the presence of LPS-cytokines. Among these, a subset of inflammation-related proteins including fibrinogen-β and -γ chains (FGB and FGG), phospholipase A2 (PLA2G2A) and SPARC was significantly downregulated in the presence of Aquamin® (alone and in combination with LPS-cytokines); another subset of proteins with anti-inflammatory, antioxidant or anti-microbial activity was upregulated by Aquamin® treatment. When provided alone, Aquamin® strongly upregulated proteins that contribute to barrier formation and tissue strength. Concomitant treatment with LPS-cytokines did not inhibit barrier formation in response to Aquamin®. Confocal microscopy also displayed increased expression of desmoglein-2 (DSG2) and cadherin-17 (CDH17) with Aquamin®, either alone or in the presence of the pro-inflammatory stimulus. Increased cohesion and TEER with Aquamin® (alone or in the presence of LPS-cytokines) indicates improved barrier function. Conclusion: Taken together, these findings suggest that multi-mineral intervention (Aquamin®) may provide a novel approach to combating inflammation in the colon by improving barrier structure/function as well as by directly altering the expression of certain pro-inflammatory proteins.

Dataset Information

Ulcerative Colitis-Derived Colonoid Culture: A Multi-Mineral-Approach to Improve Barrier Protein Expression.

Background

Methods

Results

Conclusion

Publications

Ulcerative Colitis-Derived Colonoid Culture: A Multi-Mineral-Approach to Improve Barrier Protein Expression.

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