Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region

Cation and anion transport in the colonocyte apical membrane is highly spatially organized along the cryptal axis. Because of lack of experimental accessibility, information about the functionality of ion transporters in the colonocyte apical membrane in the lower part of the crypt is scarce. The ai...

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Autores principales: Salari, Azam, Zhou, Kunyan, Nikolovska, Katerina, Seidler, Ursula, Amiri, Mahdi
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2023
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Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10001859/
https://www.ncbi.nlm.nih.gov/pubmed/36901695
http://dx.doi.org/10.3390/ijms24054266
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author Salari, Azam
Zhou, Kunyan
Nikolovska, Katerina
Seidler, Ursula
Amiri, Mahdi
author_facet Salari, Azam
Zhou, Kunyan
Nikolovska, Katerina
Seidler, Ursula
Amiri, Mahdi
author_sort Salari, Azam
collection PubMed
description Cation and anion transport in the colonocyte apical membrane is highly spatially organized along the cryptal axis. Because of lack of experimental accessibility, information about the functionality of ion transporters in the colonocyte apical membrane in the lower part of the crypt is scarce. The aim of this study was to establish an in vitro model of the colonic lower crypt compartment, which expresses the transit amplifying/progenitor (TA/PE) cells, with accessibility of the apical membrane for functional study of lower crypt-expressed Na(+)/H(+) exchangers (NHEs). Colonic crypts and myofibroblasts were isolated from human transverse colonic biopsies, expanded as three-dimensional (3D) colonoids and myofibroblast monolayers, and characterized. Filter-grown colonic myofibroblast–colonic epithelial cell (CM-CE) cocultures (myofibroblasts on the bottom of the transwell and colonocytes on the filter) were established. The expression pattern for ion transport/junctional/stem cell markers of the CM-CE monolayers was compared with that of nondifferentiated (EM) and differentiated (DM) colonoid monolayers. Fluorometric pH(i) measurements were performed to characterize apical NHEs. CM-CE cocultures displayed a rapid increase in transepithelial electrical resistance (TEER), paralleled by downregulation of claudin-2. They maintained proliferative activity and an expression pattern resembling TA/PE cells. The CM-CE monolayers displayed high apical Na(+)/H(+) exchange activity, mediated to >80% by NHE2. Human colonoid–myofibroblast cocultures allow the study of ion transporters that are expressed in the apical membrane of the nondifferentiated colonocytes of the cryptal neck region. The NHE2 isoform is the predominant apical Na(+)/H(+) exchanger in this epithelial compartment.
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spelling pubmed-100018592023-03-11 Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region Salari, Azam Zhou, Kunyan Nikolovska, Katerina Seidler, Ursula Amiri, Mahdi Int J Mol Sci Article Cation and anion transport in the colonocyte apical membrane is highly spatially organized along the cryptal axis. Because of lack of experimental accessibility, information about the functionality of ion transporters in the colonocyte apical membrane in the lower part of the crypt is scarce. The aim of this study was to establish an in vitro model of the colonic lower crypt compartment, which expresses the transit amplifying/progenitor (TA/PE) cells, with accessibility of the apical membrane for functional study of lower crypt-expressed Na(+)/H(+) exchangers (NHEs). Colonic crypts and myofibroblasts were isolated from human transverse colonic biopsies, expanded as three-dimensional (3D) colonoids and myofibroblast monolayers, and characterized. Filter-grown colonic myofibroblast–colonic epithelial cell (CM-CE) cocultures (myofibroblasts on the bottom of the transwell and colonocytes on the filter) were established. The expression pattern for ion transport/junctional/stem cell markers of the CM-CE monolayers was compared with that of nondifferentiated (EM) and differentiated (DM) colonoid monolayers. Fluorometric pH(i) measurements were performed to characterize apical NHEs. CM-CE cocultures displayed a rapid increase in transepithelial electrical resistance (TEER), paralleled by downregulation of claudin-2. They maintained proliferative activity and an expression pattern resembling TA/PE cells. The CM-CE monolayers displayed high apical Na(+)/H(+) exchange activity, mediated to >80% by NHE2. Human colonoid–myofibroblast cocultures allow the study of ion transporters that are expressed in the apical membrane of the nondifferentiated colonocytes of the cryptal neck region. The NHE2 isoform is the predominant apical Na(+)/H(+) exchanger in this epithelial compartment. MDPI 2023-02-21 /pmc/articles/PMC10001859/ /pubmed/36901695 http://dx.doi.org/10.3390/ijms24054266 Text en © 2023 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Salari, Azam
Zhou, Kunyan
Nikolovska, Katerina
Seidler, Ursula
Amiri, Mahdi
Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region
title Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region
title_full Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region
title_fullStr Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region
title_full_unstemmed Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region
title_short Human Colonoid–Myofibroblast Coculture for Study of Apical Na(+)/H(+) Exchangers of the Lower Cryptal Neck Region
title_sort human colonoid–myofibroblast coculture for study of apical na(+)/h(+) exchangers of the lower cryptal neck region
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10001859/
https://www.ncbi.nlm.nih.gov/pubmed/36901695
http://dx.doi.org/10.3390/ijms24054266
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