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Comparing Two Intestinal Porcine Epithelial Cell Lines (IPECs): Morphological Differentiation, Function and Metabolism.

Nossol C, Barta-Böszörményi A, Kahlert S, Zuschratter W, Faber-Zuschratter H, Reinhardt N, Ponsuksili S, Wimmers K, Diesing AK, Rothkötter HJ - PLoS ONE (2015)

Bottom Line: On the other hand, "spliceosome", "ribosome", "RNA-degradation" and "tight junction" are significantly down-regulated pathways in IPEC-J2 in comparison to IPEC-1.These cells seem to be more active in their metabolism than IPEC-1 cells due to a significant higher ATP-content as well as a higher O2- and glucose-consumption.In addition, IPEC-J2 cells are a preferential tool for in vitro studies with the focus on metabolism.

View Article: PubMed Central - PubMed

Affiliation: Institute of Anatomy, Otto-von-Guericke University Magdeburg, 39120, Magdeburg, Germany.

ABSTRACT
The pig shows genetical and physiological resemblance to human, which predestines it as an experimental animal model especially for mucosal physiology. Therefore, the intestinal epithelial cell lines 1 and J2 (IPEC-1, IPEC-J2)--spontaneously immortalised cell lines from the porcine intestine--are important tools for studying intestinal function. A microarray (GeneChip Porcine Genome Array) was performed to compare the genome wide gene expression of IPECs. Different significantly up-regulated pathways were identified, like "lysosome", "pathways in cancer", "regulation of actin cytoskeleton" and "oxidative phosphorylation" in IPEC-J2 in comparison to IPEC-1. On the other hand, "spliceosome", "ribosome", "RNA-degradation" and "tight junction" are significantly down-regulated pathways in IPEC-J2 in comparison to IPEC-1. Examined pathways were followed up by functional analyses. ATP-, oxygen, glucose and lactate-measurement provide evidence for up-regulation of oxidative phosphorylation in IPEC-J2. These cells seem to be more active in their metabolism than IPEC-1 cells due to a significant higher ATP-content as well as a higher O2- and glucose-consumption. The down-regulated pathway "ribosome" was followed up by measurement of RNA- and protein content. In summary, IPEC-J2 is a morphologically and functionally more differentiated cell line in comparison to IPEC-1. In addition, IPEC-J2 cells are a preferential tool for in vitro studies with the focus on metabolism.

No MeSH data available.


Related in: MedlinePlus

Polarisation of IPEC-1 and IPEC-J2, confocal microscopy and transmission electron microscopy.(A) Ten days old IPEC-1 and IPEC-J2 cells were fixed and stained for ZO-1 (Zonula occludens, green) and beta-catenin (Zonula adherens, red). Both cells lines showed a polarised structure with ZO-1 immunoreactivity at the apical pole and the underlying beta-catenin expression. (B) Important genes of a polarised brush border were analysed using microarray and qPCR. Here, significant differences were found between the cell lines. VIL1 (villin-1), VIL2 (villin-2 = ezrin), TLR4 (toll like receptor 4), MUC4 (mucin 4) and ESPN (espin) were significant up-regulated in IPEC-J2 in comparison to IPEC-1 in microarray as well as in qPCR. Villin1 and villin2 (= ezrin) were followed up by Western blot analyses. No villin-1 was detected in IPEC-1 but a strong expression in IPEC-J2 over time. Villin-2 was expressed in both cell lines. (C) Cells were cultured for 10 or 31 days and analysed using transmission electronmicroscopy. Both cell lines showed well-developed tight junctions at day 10 and 31 (red arrows). Desmosomes (blue arrows) and interdigitation (yellow arrow) were also observed. No differences were detected concerning microvilli length between both cell lines and at both time points. In contrast, the number of microvilli differed between the cell lines (IPEC-J2>IPEC-1).
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pone.0132323.g002: Polarisation of IPEC-1 and IPEC-J2, confocal microscopy and transmission electron microscopy.(A) Ten days old IPEC-1 and IPEC-J2 cells were fixed and stained for ZO-1 (Zonula occludens, green) and beta-catenin (Zonula adherens, red). Both cells lines showed a polarised structure with ZO-1 immunoreactivity at the apical pole and the underlying beta-catenin expression. (B) Important genes of a polarised brush border were analysed using microarray and qPCR. Here, significant differences were found between the cell lines. VIL1 (villin-1), VIL2 (villin-2 = ezrin), TLR4 (toll like receptor 4), MUC4 (mucin 4) and ESPN (espin) were significant up-regulated in IPEC-J2 in comparison to IPEC-1 in microarray as well as in qPCR. Villin1 and villin2 (= ezrin) were followed up by Western blot analyses. No villin-1 was detected in IPEC-1 but a strong expression in IPEC-J2 over time. Villin-2 was expressed in both cell lines. (C) Cells were cultured for 10 or 31 days and analysed using transmission electronmicroscopy. Both cell lines showed well-developed tight junctions at day 10 and 31 (red arrows). Desmosomes (blue arrows) and interdigitation (yellow arrow) were also observed. No differences were detected concerning microvilli length between both cell lines and at both time points. In contrast, the number of microvilli differed between the cell lines (IPEC-J2>IPEC-1).

Mentions: IPEC-1 and IPEC-J2 form a continuous monolayer characterised by polarised growth, transepithelial resistance (TEER) and the expression of actin, villin-1 and villin-2 (= ezrin) (Fig 2). The polarised structure was confirmed by immunofluorescence-based detection of ZO-1 and β-catenin (Fig 2A). Three-dimensional reconstruction of optical sections of confocal microscopy showed an apical localisation of ZO-1 (green) and a lateral localisation of β-catenin (red). Both cell lines showed increasing TEER-values with increasing duration of cultivation. On day 10, IPEC-1 and IPEC-J2 exhibited a TEER about 7 kOhm*cm2 (IPEC-1: 7.0±1.2 kOhm*cm2; IPEC-J2: 7.8±1.0 kOhm*cm2).


Comparing Two Intestinal Porcine Epithelial Cell Lines (IPECs): Morphological Differentiation, Function and Metabolism.

Nossol C, Barta-Böszörményi A, Kahlert S, Zuschratter W, Faber-Zuschratter H, Reinhardt N, Ponsuksili S, Wimmers K, Diesing AK, Rothkötter HJ - PLoS ONE (2015)

Polarisation of IPEC-1 and IPEC-J2, confocal microscopy and transmission electron microscopy.(A) Ten days old IPEC-1 and IPEC-J2 cells were fixed and stained for ZO-1 (Zonula occludens, green) and beta-catenin (Zonula adherens, red). Both cells lines showed a polarised structure with ZO-1 immunoreactivity at the apical pole and the underlying beta-catenin expression. (B) Important genes of a polarised brush border were analysed using microarray and qPCR. Here, significant differences were found between the cell lines. VIL1 (villin-1), VIL2 (villin-2 = ezrin), TLR4 (toll like receptor 4), MUC4 (mucin 4) and ESPN (espin) were significant up-regulated in IPEC-J2 in comparison to IPEC-1 in microarray as well as in qPCR. Villin1 and villin2 (= ezrin) were followed up by Western blot analyses. No villin-1 was detected in IPEC-1 but a strong expression in IPEC-J2 over time. Villin-2 was expressed in both cell lines. (C) Cells were cultured for 10 or 31 days and analysed using transmission electronmicroscopy. Both cell lines showed well-developed tight junctions at day 10 and 31 (red arrows). Desmosomes (blue arrows) and interdigitation (yellow arrow) were also observed. No differences were detected concerning microvilli length between both cell lines and at both time points. In contrast, the number of microvilli differed between the cell lines (IPEC-J2>IPEC-1).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4493080&req=5

pone.0132323.g002: Polarisation of IPEC-1 and IPEC-J2, confocal microscopy and transmission electron microscopy.(A) Ten days old IPEC-1 and IPEC-J2 cells were fixed and stained for ZO-1 (Zonula occludens, green) and beta-catenin (Zonula adherens, red). Both cells lines showed a polarised structure with ZO-1 immunoreactivity at the apical pole and the underlying beta-catenin expression. (B) Important genes of a polarised brush border were analysed using microarray and qPCR. Here, significant differences were found between the cell lines. VIL1 (villin-1), VIL2 (villin-2 = ezrin), TLR4 (toll like receptor 4), MUC4 (mucin 4) and ESPN (espin) were significant up-regulated in IPEC-J2 in comparison to IPEC-1 in microarray as well as in qPCR. Villin1 and villin2 (= ezrin) were followed up by Western blot analyses. No villin-1 was detected in IPEC-1 but a strong expression in IPEC-J2 over time. Villin-2 was expressed in both cell lines. (C) Cells were cultured for 10 or 31 days and analysed using transmission electronmicroscopy. Both cell lines showed well-developed tight junctions at day 10 and 31 (red arrows). Desmosomes (blue arrows) and interdigitation (yellow arrow) were also observed. No differences were detected concerning microvilli length between both cell lines and at both time points. In contrast, the number of microvilli differed between the cell lines (IPEC-J2>IPEC-1).
Mentions: IPEC-1 and IPEC-J2 form a continuous monolayer characterised by polarised growth, transepithelial resistance (TEER) and the expression of actin, villin-1 and villin-2 (= ezrin) (Fig 2). The polarised structure was confirmed by immunofluorescence-based detection of ZO-1 and β-catenin (Fig 2A). Three-dimensional reconstruction of optical sections of confocal microscopy showed an apical localisation of ZO-1 (green) and a lateral localisation of β-catenin (red). Both cell lines showed increasing TEER-values with increasing duration of cultivation. On day 10, IPEC-1 and IPEC-J2 exhibited a TEER about 7 kOhm*cm2 (IPEC-1: 7.0±1.2 kOhm*cm2; IPEC-J2: 7.8±1.0 kOhm*cm2).

Bottom Line: On the other hand, "spliceosome", "ribosome", "RNA-degradation" and "tight junction" are significantly down-regulated pathways in IPEC-J2 in comparison to IPEC-1.These cells seem to be more active in their metabolism than IPEC-1 cells due to a significant higher ATP-content as well as a higher O2- and glucose-consumption.In addition, IPEC-J2 cells are a preferential tool for in vitro studies with the focus on metabolism.

View Article: PubMed Central - PubMed

Affiliation: Institute of Anatomy, Otto-von-Guericke University Magdeburg, 39120, Magdeburg, Germany.

ABSTRACT
The pig shows genetical and physiological resemblance to human, which predestines it as an experimental animal model especially for mucosal physiology. Therefore, the intestinal epithelial cell lines 1 and J2 (IPEC-1, IPEC-J2)--spontaneously immortalised cell lines from the porcine intestine--are important tools for studying intestinal function. A microarray (GeneChip Porcine Genome Array) was performed to compare the genome wide gene expression of IPECs. Different significantly up-regulated pathways were identified, like "lysosome", "pathways in cancer", "regulation of actin cytoskeleton" and "oxidative phosphorylation" in IPEC-J2 in comparison to IPEC-1. On the other hand, "spliceosome", "ribosome", "RNA-degradation" and "tight junction" are significantly down-regulated pathways in IPEC-J2 in comparison to IPEC-1. Examined pathways were followed up by functional analyses. ATP-, oxygen, glucose and lactate-measurement provide evidence for up-regulation of oxidative phosphorylation in IPEC-J2. These cells seem to be more active in their metabolism than IPEC-1 cells due to a significant higher ATP-content as well as a higher O2- and glucose-consumption. The down-regulated pathway "ribosome" was followed up by measurement of RNA- and protein content. In summary, IPEC-J2 is a morphologically and functionally more differentiated cell line in comparison to IPEC-1. In addition, IPEC-J2 cells are a preferential tool for in vitro studies with the focus on metabolism.

No MeSH data available.


Related in: MedlinePlus