Development of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21)

Nollevaux, Géraldine; Devillé, Christelle; Moualij, Benaı̈ssa El; Zorzi, Willy; Deloyer, Patricia; Schneider, Yves‐Jacques; Peulen, Olivier; Dandrifosse, Guy

doi:10.1186/1471-2121-7-20

Cited by 72 publications

(24 citation statements)

References 53 publications

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“…Zinc transport studies were performed using monocultures of Caco-2 and Caco-2/HT-29-MTX co-cultures. Co-cultures were realized with an initial cell ratio of 75% Caco-2 and 25% HT-29-MTX cells and alternated cell seeding, modified after Nollevaux et al [ 64 ]. Herein, the co-culture of Caco-2 and HT-29-MTX cells in our lab was characterized concerning the proper cellular ratio by investigating the mucin secretion and adequate differentiation of the enterocytes as reported before [ 17 ].…”

Section: Methodsmentioning

confidence: 99%

“…Prior and after the experiment, barrier integrity was monitored by measuring TEER with the epithelial volt-ohm meter Millicell ® ERS-2 (Millipore, Burlington, MA, USA). Additionally, permeability of cell monolayers during the experiment was determined using FD-20 [ 64 ] as reported before [ 17 ]. At the end of the experiment, the media of the apical and basolateral compartments were collected, and cells were harvested on ice in PBS, homogenized and centrifuged (800× g ).…”

Section: Methodsmentioning

confidence: 99%

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In Vitro Studies on Zinc Binding and Buffering by Intestinal Mucins

Maares

Keil

Koza

et al. 2018

IJMS

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The investigation of luminal factors influencing zinc availability and accessibility in the intestine is of great interest when analyzing parameters regulating intestinal zinc resorption. Of note, intestinal mucins were suggested to play a beneficial role in the luminal availability of zinc. Their exact zinc binding properties, however, remain unknown and the impact of these glycoproteins on human intestinal zinc resorption has not been investigated in detail. Thus, the aim of this study is to elucidate the impact of intestinal mucins on luminal uptake of zinc into enterocytes and its transfer into the blood. In the present study, in vitro zinc binding properties of mucins were analyzed using commercially available porcine mucins and secreted mucins of the goblet cell line HT-29-MTX. The molecular zinc binding capacity and average zinc binding affinity of these glycoproteins demonstrates that mucins contain multiple zinc-binding sites with biologically relevant affinity within one mucin molecule. Zinc uptake into the enterocyte cell line Caco-2 was impaired by zinc-depleted mucins. Yet this does not represent their form in the intestinal lumen in vivo under zinc adequate conditions. In fact, zinc-uptake studies into enterocytes in the presence of mucins with differing degree of zinc saturation revealed zinc buffering by these glycoproteins, indicating that mucin-bound zinc is still available for the cells. Finally, the impact of mucins on zinc resorption using three-dimensional cultures was studied comparing the zinc transfer of a Caco-2/HT-29-MTX co-culture and conventional Caco-2 monoculture. Here, the mucin secreting co-cultures yielded higher fractional zinc resorption and elevated zinc transport rates, suggesting that intestinal mucins facilitate the zinc uptake into enterocytes and act as a zinc delivery system for the intestinal epithelium.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

In Vitro Studies on Zinc Binding and Buffering by Intestinal Mucins

Maares

Keil

Koza

et al. 2018

IJMS

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“…HT29 cells adapted to methotrexate (MTX, 10 −5 M) were obtained from Dr. T. Lesuffleur (INSERM U505, Villejuif, France) were used between passages 30–50 with a seeding ratio Caco-2/HT29-MTX of 3:1, as previously described by Nollevaux et al (2006) [34]. Together with Caco-2 cells, they form a co-culture that reproduces the two main cellular types encountered in the human intestinal epithelium.…”

Section: Methodsmentioning

confidence: 99%

“…Therefore, with the aim to better mimic the in vivo conditions of toxinogenesis, this study assessed the enterotoxicity of 70 B. cereus strains on fully differentiated Caco-2 cells. Additionally, this work also studied the interaction between the B. cereus supernatants and the mucus layer, using a co-culture (Caco-2/HT29-MTX) cell model [34]. …”

Section: Introductionmentioning

confidence: 99%

Screening of Cytotoxic B. cereus on Differentiated Caco-2 Cells and in Co-Culture with Mucus-Secreting (HT29-MTX) Cells

Castiaux

Laloux

Schneider

et al. 2016

Toxins

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B. cereus is an opportunistic foodborne pathogen able to cause diarrhoea. However, the diarrhoeal potential of a B. cereus strain remains difficult to predict, because no simple correlation has yet been identified between the symptoms and a unique or a specific combination of virulence factors. In this study, 70 B. cereus strains with different origins (food poisonings, foods and environment) have been selected to assess their enterotoxicity. The B. cereus cell-free supernatants have been tested for their toxicity in vitro, on differentiated (21 day-old) Caco-2 cells, using their ATP content, LDH release and NR accumulation. The genetic determinants of the main potential enterotoxins and virulence factors (ces, cytK, entFM, entS, hbl, nhe, nprA, piplC and sph) have also been screened by PCR. This analysis showed that none of these genes was able to fully explain the enterotoxicity of B. cereus strains. Additionally, in order to assess a possible effect of the mucus layer in vitro, a cytotoxicity comparison between a monoculture (Caco-2 cells) and a co-culture (Caco-2 and HT29-MTX mucus-secreting cells) model has been performed with selected B. cereus supernatants. It appeared that, in these conditions, the mucus layer had no notable influence on the cytotoxicity of B. cereus supernatants.

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“… Drieschner et al, 2017 ; Vllasaliu et al, 2014 ), co-culture (e.g. Nollevaux et al, 2006 ) or through chemically induced differentiation.…”

Section: Resultsmentioning

confidence: 99%

Establishment and long-term maintenance of primary intestinal epithelial cells cultured from the rainbow trout,Oncorhynchus mykiss

2018

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A novel method for the establishment and long-term maintenance of ex vivo cultures from intestinal regions of the rainbow trout, Oncorhynchus mykiss (Walbaum), is reported. Adherence of cells was observed within hours, epithelial island formation recorded at 48 h and rapid proliferation with confluence achieved between 9-14 days. In addition to metabolic characterisation, basic morphology of growing cells was characterised using histology, immunofluorescence, transmission electron microscopy (TEM) and transepithelial electrical resistance (TEER). Regional differences in intestinal ethoxyresorufin-O-deethylase (EROD) and 7-ethoxycoumarin-O-deethylation (ECOD) activities in these primary grown enterocytes were compared following exposure to model inducers [i.e. α-NF, β-NF, B(a)P] which demonstrated significant differences. Regional differences in dietary uptake and metabolism of contaminants can therefore be studied in this in vitro system to increase our understanding of fundamental processes, while concurrently providing a means to reduce the number of fish required for biological studies in line with the principles of the 3Rs (Reduce, Refine and Replace).This article has an associated First Person interview with the first author of the paper.

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Development of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21)

Cited by 72 publications

References 53 publications

In Vitro Studies on Zinc Binding and Buffering by Intestinal Mucins

In Vitro Studies on Zinc Binding and Buffering by Intestinal Mucins

Screening of Cytotoxic B. cereus on Differentiated Caco-2 Cells and in Co-Culture with Mucus-Secreting (HT29-MTX) Cells

Establishment and long-term maintenance of primary intestinal epithelial cells cultured from the rainbow trout,Oncorhynchus mykiss

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