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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

Analysis of the protein expression of tight junctions and cytoskeleton—IPEC-J2.Western blot analyses and immunofluorescence were used to examine different proteins of the tight junctions (ZO-1, occludin, claudin-3 and claudin-4) and the cytoskeleton CK18 and actin). Results comparable to IPEC-1 were found in IPEC-J2, here a weak expression was found at day 2 of culture for all proteins analysed using Western blot. The tight junctions proteins ZO-1 and occludin showed a strong expression at the cell border, however in this cell type ZO-1 had a much higher immunoreactivity as occludin. No actin-stress fibres were found in IPEC-J2. Claudin-3 and claudin-4 were present, but claudin-4 showed higher expression in the cytoplasm than at the cell border in comparison to claudin-3. (blue = DAPI; bar = 20 μm)
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pone.0132323.g005: Analysis of the protein expression of tight junctions and cytoskeleton—IPEC-J2.Western blot analyses and immunofluorescence were used to examine different proteins of the tight junctions (ZO-1, occludin, claudin-3 and claudin-4) and the cytoskeleton CK18 and actin). Results comparable to IPEC-1 were found in IPEC-J2, here a weak expression was found at day 2 of culture for all proteins analysed using Western blot. The tight junctions proteins ZO-1 and occludin showed a strong expression at the cell border, however in this cell type ZO-1 had a much higher immunoreactivity as occludin. No actin-stress fibres were found in IPEC-J2. Claudin-3 and claudin-4 were present, but claudin-4 showed higher expression in the cytoplasm than at the cell border in comparison to claudin-3. (blue = DAPI; bar = 20 μm)

Mentions: A more detailed examination of the morphology of IPEC-1 and IPEC-J2 showed the clear network of the membrane-linked proteins involved in the cell-cell-contacts. 28 genes are regulated in the microarray analyses that are involved in the network of the connected cells. Claudin-1, claudin-3, claudin-7 and occludin were chosen as marker for cell-cell contacts and tight junctions. An increased mRNA-level of claudin-3 and claudin-7 (both: p≤0.001, Table 3) was detected in IPEC-J2. In comparison to IPEC-1, a decreased mRNA-level of claudin-1 and occludin (both: p≤0.001) was noticed. These data were obtained by microarray data. Differences between the microarray and the qPCR were observed in claudin-3, which was significantly up-regulated in the qPCR but not in the microarray. IPEC-1 and IPEC-J2 were cultured on membranes and analysed at different time points: day 2, 7, 14 and 21 (Figs 4 and 5) via Western blot. ZO-1 and occludin showed opposed characteristics: ZO-1 < occludin in IPEC-1 and ZO-1 > occludin in IPEC-J2. In both cell lines, a strong expression of claudin-3 and claudin-4 was detected at day 7, 14 and 21 of culture by Western blot analyses. Claudin-3 and claudin-4 were observed in the cells. Claudin-4 showed higher expression in the cytoplasm than at the cell border in comparison to claudin-3 in IPEC-J2.


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)

Analysis of the protein expression of tight junctions and cytoskeleton—IPEC-J2.Western blot analyses and immunofluorescence were used to examine different proteins of the tight junctions (ZO-1, occludin, claudin-3 and claudin-4) and the cytoskeleton CK18 and actin). Results comparable to IPEC-1 were found in IPEC-J2, here a weak expression was found at day 2 of culture for all proteins analysed using Western blot. The tight junctions proteins ZO-1 and occludin showed a strong expression at the cell border, however in this cell type ZO-1 had a much higher immunoreactivity as occludin. No actin-stress fibres were found in IPEC-J2. Claudin-3 and claudin-4 were present, but claudin-4 showed higher expression in the cytoplasm than at the cell border in comparison to claudin-3. (blue = DAPI; bar = 20 μm)
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493080&req=5

pone.0132323.g005: Analysis of the protein expression of tight junctions and cytoskeleton—IPEC-J2.Western blot analyses and immunofluorescence were used to examine different proteins of the tight junctions (ZO-1, occludin, claudin-3 and claudin-4) and the cytoskeleton CK18 and actin). Results comparable to IPEC-1 were found in IPEC-J2, here a weak expression was found at day 2 of culture for all proteins analysed using Western blot. The tight junctions proteins ZO-1 and occludin showed a strong expression at the cell border, however in this cell type ZO-1 had a much higher immunoreactivity as occludin. No actin-stress fibres were found in IPEC-J2. Claudin-3 and claudin-4 were present, but claudin-4 showed higher expression in the cytoplasm than at the cell border in comparison to claudin-3. (blue = DAPI; bar = 20 μm)
Mentions: A more detailed examination of the morphology of IPEC-1 and IPEC-J2 showed the clear network of the membrane-linked proteins involved in the cell-cell-contacts. 28 genes are regulated in the microarray analyses that are involved in the network of the connected cells. Claudin-1, claudin-3, claudin-7 and occludin were chosen as marker for cell-cell contacts and tight junctions. An increased mRNA-level of claudin-3 and claudin-7 (both: p≤0.001, Table 3) was detected in IPEC-J2. In comparison to IPEC-1, a decreased mRNA-level of claudin-1 and occludin (both: p≤0.001) was noticed. These data were obtained by microarray data. Differences between the microarray and the qPCR were observed in claudin-3, which was significantly up-regulated in the qPCR but not in the microarray. IPEC-1 and IPEC-J2 were cultured on membranes and analysed at different time points: day 2, 7, 14 and 21 (Figs 4 and 5) via Western blot. ZO-1 and occludin showed opposed characteristics: ZO-1 < occludin in IPEC-1 and ZO-1 > occludin in IPEC-J2. In both cell lines, a strong expression of claudin-3 and claudin-4 was detected at day 7, 14 and 21 of culture by Western blot analyses. Claudin-3 and claudin-4 were observed in the cells. Claudin-4 showed higher expression in the cytoplasm than at the cell border in comparison to claudin-3 in IPEC-J2.

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