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Establishment of immortalized mouse intestinal epithelial cells line and study of effects of Arg-Arg on inflammatory response

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Abstract

Primary mouse intestinal epithelial cells (MIEs) are not ideal models for long-term culture in vitro and a limited amount of approximate three generations. In addition, the mechanism that arginine-arginine dipeptide (Arg-Arg) regulates mouse intestinal inflammatory response remains unknown. Therefore, the aim of this study was to establish immortal MIEs and study the effects of Arg-Arg on inflammatory response after challenging the MIEs with lipopolysaccharide (LPS) or staphylococcal enterotoxin C (rSEC). Our data showed that immortalized MIEs could be cultured over 100 generations. The immortalized MIEs showed positive reaction against cytokeratine 18 antigen, E-cadherin, and peptide transporters (Pept1) using indirect immunofluorescence. Cytokeratine 18 and Pept1 can be expressed in immortalized MIEs by immunoblotting. Fatty acid-binding proteins (FABPs) and villin known as intestinal epithelial cell functional protein were constitutively expressed in immortalized MIEs. For inflammatory response, these results showed that Arg-Arg can decrease the LPS-induced expression of IL-1β and the rSEC-induced expression of TNF-α; however, it can upregulate the LPS-induced expression of IL-6 and TNF-α and the rSEC-induced expression level of IL-1β. In addition, in the MAPK signaling pathway, pSAPK/JNK and p-Erk1/2 in LPS with Arg-Arg treatment were upregulated than that in LPS treatment. p-p38 in LPS with Arg-Arg treatment was attenuated than that in LPS treatment. pSAPK/JNK and p-p38 in rSEC with Arg-Arg treatment were enhanced than that in rSEC treatment. Conversely, p-Erk1/2 in rSEC with Arg-Arg treatment was attenuated than that in rSEC treatment. These novel findings suggest that Arg-Arg dipeptide plays an important role for regulation of the immunologic balance in mouse intestinal inflammatory response.

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Acknowledgements

This study was supported partly by the National Natural Science Foundation of China (No. 31572430).

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Authors and Affiliations

  1. College of Animal Science and Technology, Yangzhou University, Yangzhou, 225009, People’s Republic of China

    Kang Zhan, Maocheng Jiang, Yannan Sui, Kang Yan, Miao Lin & Guoqi Zhao

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  1. Kang Zhan
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  2. Maocheng Jiang
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  3. Yannan Sui
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  4. Kang Yan
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  5. Miao Lin
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  6. Guoqi Zhao
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Correspondence to Guoqi Zhao.

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Kang Zhan and Maocheng Jiang authors contributed equally to this work.

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Zhan, K., Jiang, M., Sui, Y. et al. Establishment of immortalized mouse intestinal epithelial cells line and study of effects of Arg-Arg on inflammatory response. In Vitro Cell.Dev.Biol.-Animal 53, 538–546 (2017). https://doi.org/10.1007/s11626-017-0143-4

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  • DOI: https://doi.org/10.1007/s11626-017-0143-4

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