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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

Glucose consumption and lactate production in IPEC-1 and IPEC-J2.Apical and basolateral supernatants of cells cultured on membranes were collected and analysed using the Roche/Hitachi Cobas c system. IPEC-J2 showed a significant higher apical and basolateral glucose consumption in comparison to IPEC-1 (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 3.3 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.57 μmol/100000 cells). Comparable results were found with the focus on lactate production (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 4.1 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.98 μmol/100000 cells).
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pone.0132323.g006: Glucose consumption and lactate production in IPEC-1 and IPEC-J2.Apical and basolateral supernatants of cells cultured on membranes were collected and analysed using the Roche/Hitachi Cobas c system. IPEC-J2 showed a significant higher apical and basolateral glucose consumption in comparison to IPEC-1 (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 3.3 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.57 μmol/100000 cells). Comparable results were found with the focus on lactate production (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 4.1 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.98 μmol/100000 cells).

Mentions: To a better understanding of the metabolism of IPEC cells, glucose utilisation, lactate production, oxygen consumption and ATP-content were examined (Figs 6, 7 and 8). In Fig 6 the results for the glucose-consumption and lactate-production in both cell lines are shown. Differences were found between the cell lines. IPEC-J2 showed a higher glucose-consumption than IPEC-1 (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 3.3 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.57 μmol/100000 cells). Similar results were found with the focus on lactate-production (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 4.1 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.98 μmol/100000 cells). Furthermore, the microarray analyses showed 29 significantly regulated genes like COX5B, COX7A2, COX7C, COX8A, ATP5D, ATP5E, ATPH, NDUFA2, NDUFA4 and NDUFB2. In addition, important genes of the aerobic metabolism like CYC1, PDHB and SDHB were examined (Fig 7A). CYC 1 (cytochrome C) plays an important role in the mitochondrial respiratory chain by transferring electrons from the Rieske iron-sulfur protein to cytochrome C and is significant down-regulated in IPEC-J2 in comparison to IPEC-1 in qPCR and in the microarray. PDHB and SDHB showed opposed characteristics. Pyruvate dehydrogenase subunit B (PDHB) is a nuclear encoded mitochondrial multi-enzyme complex that catalyses the conversion of pyruvate to acetyl-CoA and carbon dioxide, it is significantly down-regulated in IPEC-J2 in comparison to IPEC-1 and. On the other hand, succinate dehydrogenase subunit B (SDHB) is significantly up-regulated and is involved in complex II and known as iron-sulfur subunit. It catalyses the oxidation of succinate. Hypoxia inducible factor (HIF1a) plays an essential role in cellular and systemic responses to hypoxia and is significantly higher expressed in IPEC-J2 in the microarray but not in qPCR. In the next step, oxygen-utilisation was examined (Fig 7B). In both cell lines, the use of a membrane growth support significantly increases the oxygen utilisation—demonstrating the membrane grown cells to be a more physiological cell culture system. In addition, IPEC-J2 (dish: 8.18 nmol/100 000 cells; membrane: 75.27 nmol/100 000 cells) showed on dishes as well as on membranes a significantly higher oxygen-consumption than IPEC-1 (dish: 3.18 nmol/100 000 cells; membrane: 20.07 nmol/100 000 cells). One important gene of the complex IV of the respiratory chain is COX5B. COX5B was analysed by microarray, qPCR and Western blot analyses. IPEC-J2 cells showed a significantly increased RNA level in the microarray but not in qPCR in comparison to IPEC-1 but a higher COX5B-protein content was observed in IPEC-J2 (Fig 8A).


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)

Glucose consumption and lactate production in IPEC-1 and IPEC-J2.Apical and basolateral supernatants of cells cultured on membranes were collected and analysed using the Roche/Hitachi Cobas c system. IPEC-J2 showed a significant higher apical and basolateral glucose consumption in comparison to IPEC-1 (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 3.3 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.57 μmol/100000 cells). Comparable results were found with the focus on lactate production (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 4.1 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.98 μmol/100000 cells).
© Copyright Policy
Related In: Results  -  Collection

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

pone.0132323.g006: Glucose consumption and lactate production in IPEC-1 and IPEC-J2.Apical and basolateral supernatants of cells cultured on membranes were collected and analysed using the Roche/Hitachi Cobas c system. IPEC-J2 showed a significant higher apical and basolateral glucose consumption in comparison to IPEC-1 (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 3.3 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.57 μmol/100000 cells). Comparable results were found with the focus on lactate production (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 4.1 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.98 μmol/100000 cells).
Mentions: To a better understanding of the metabolism of IPEC cells, glucose utilisation, lactate production, oxygen consumption and ATP-content were examined (Figs 6, 7 and 8). In Fig 6 the results for the glucose-consumption and lactate-production in both cell lines are shown. Differences were found between the cell lines. IPEC-J2 showed a higher glucose-consumption than IPEC-1 (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 3.3 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.57 μmol/100000 cells). Similar results were found with the focus on lactate-production (apical: IPEC-1: 0.48 μmol/100000 cells, IPEC-J2: 4.1 μmol/100000 cells; basolateral: IPEC-1: 0.95 μmol/100000 cells, IPEC-J2: 3.98 μmol/100000 cells). Furthermore, the microarray analyses showed 29 significantly regulated genes like COX5B, COX7A2, COX7C, COX8A, ATP5D, ATP5E, ATPH, NDUFA2, NDUFA4 and NDUFB2. In addition, important genes of the aerobic metabolism like CYC1, PDHB and SDHB were examined (Fig 7A). CYC 1 (cytochrome C) plays an important role in the mitochondrial respiratory chain by transferring electrons from the Rieske iron-sulfur protein to cytochrome C and is significant down-regulated in IPEC-J2 in comparison to IPEC-1 in qPCR and in the microarray. PDHB and SDHB showed opposed characteristics. Pyruvate dehydrogenase subunit B (PDHB) is a nuclear encoded mitochondrial multi-enzyme complex that catalyses the conversion of pyruvate to acetyl-CoA and carbon dioxide, it is significantly down-regulated in IPEC-J2 in comparison to IPEC-1 and. On the other hand, succinate dehydrogenase subunit B (SDHB) is significantly up-regulated and is involved in complex II and known as iron-sulfur subunit. It catalyses the oxidation of succinate. Hypoxia inducible factor (HIF1a) plays an essential role in cellular and systemic responses to hypoxia and is significantly higher expressed in IPEC-J2 in the microarray but not in qPCR. In the next step, oxygen-utilisation was examined (Fig 7B). In both cell lines, the use of a membrane growth support significantly increases the oxygen utilisation—demonstrating the membrane grown cells to be a more physiological cell culture system. In addition, IPEC-J2 (dish: 8.18 nmol/100 000 cells; membrane: 75.27 nmol/100 000 cells) showed on dishes as well as on membranes a significantly higher oxygen-consumption than IPEC-1 (dish: 3.18 nmol/100 000 cells; membrane: 20.07 nmol/100 000 cells). One important gene of the complex IV of the respiratory chain is COX5B. COX5B was analysed by microarray, qPCR and Western blot analyses. IPEC-J2 cells showed a significantly increased RNA level in the microarray but not in qPCR in comparison to IPEC-1 but a higher COX5B-protein content was observed in IPEC-J2 (Fig 8A).

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