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MiR-124a Mediates the Impairment of Intestinal Epithelial Integrity by Targeting Aryl Hydrocarbon Receptor in Crohn’s Disease

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Abstract

Growing evidence suggested that microRNAs (miRNAs) contributed to the progression of Crohn’s disease (CD), but the exact physiological functions of many miRNAs in CD patients still remain illusive. In this study, we explore the potent pathogenicity of miRNAs in CD. Expressions of miRNAs and aryl hydrocarbon receptor (AHR) protein were determined in the colitic colon of 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis mice and CD patients. Colitis was induced in wild-type (WT), miR-124a overexpression (miR-124a-Nju), and AHR knockout (AHR−/−) mice. Intestinal barrier function was evaluated in colitis mice and Caco2 monolayers. There was a negative relationship between miR-124a and AHR protein in inflamed colons from CD patients. MiR-124a-Nju and AHR−/− mice treated with TNBS had more severe intestinal inflammation than WT mice. Both miR-124a-Nju mice and AHR−/− mice underwent evident intestinal barrier destruction, and anti-miR-124a administration could reverse this dysfunction in miR-124a-Nju mice but not in AHR−/− mice. In vitro studies revealed that miR-124a mimics downregulated the expression of AHR and tight junction proteins and induced hyperpermeability by increasing miR-124a expression, which was abrogated by miR-124a inhibitor and AHR antagonist FICZ. This study suggests that miR-124a can induce intestinal inflammation and cause intestinal barrier dysfunction by supressing AHR.

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Abbreviations

CD:

Crohn’s disease

AHR:

Aryl hydrocarbon receptor

bHLH:

Basic helix-loop-helix

IBD:

Inflammatory bowel disease

IL-22:

Interleukin-22

DSS:

Dextran dulfate dodium

IECs:

Intestinal epithelial cells

LP:

Lamina propria

TJ:

Tight junction

miRNAs:

MicroRNAs

TNBS:

2,4,6-Trinitrobenzenesulfonic acid

WT:

Wild type

NC:

Normal controls

SPF:

Specific pathogen free

PEI:

Polyetherimide

FBS:

Fetal bovine serum

DMEM:

Dulbecco’s modified Eagle medium

TNF-α:

Tumor necrosis factor-α

DAI:

Disease activity index

FITC:

Fluorescein isothiocyanate

TEER:

Transepithelial electrical resistance

SEM:

Standard error of the mean

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Acknowledgments

The authors wish to thank Xuerui Zhang for his excellent technical assistance.

Funding

This study was supported by grants from the National Natural Science Foundation of China (81770553 and 81470827), Jiangsu Province’s Key Medical Talents Program (ZDRCB2016001), and Postgraduate Research & Practice Innovation Program of Jiangsu Province (KYCX18_1484).

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Author notes

  1. Xiaojing Zhao and Jiajia Li contributed equally to this work.

Authors and Affiliations

  1. Department of Gastroenterology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, 210029, Jiangsu, China

    Xiaojing Zhao, Jiajia Li, Jingjing Ma, Chunhua Jiao, Xinyun Qiu, Xiufang Cui, Di Wang & Hongjie Zhang

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Previous presentations

An abstract of the study’s results was invited as an oral presentation at the 2019 United European Gastroenterology Week in Barcelona. The abstract was also chosen as a postpresentation at the 6th Annual Meeting of Asian Organization for Crohn’s & Colitis, which was published in Journal of Digestive Diseases banner.

Electronic Supplementary Material

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ESM 2

Mice were enema treatment using anti-miR-124a 12 h after induction of colitis. Frozen colon sections were obtained from mice with Cy3 anti-miR-124a enema treatment and stained with markers of intestinal epithelia cell [red, Cy3 anti-miR-124a; green, pan cytokeratin [PCK]; blue, DAPI nuclear staining]. Scale bar = 50 μm (PNG 932 kb)

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ESM 3

Effect of increasing concentrations of TNF-α (0, 10, 25, 50, and 100 ng/ml) on Caco-2 epithelial resistance [Ω•cm2] for increasing time periods (0-48 h). Results are presented as mean ± SEM. *P < 0.05 (PNG 123 kb)

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ESM 4

The examination of the level of miR-124a in Caco-2 cells after transfected with miR-124a mimic, pre-scramble, miR-124a inhibitor and anti-scramble as well as stimulated with TNF-α. (A) qRT-PCR analysis of miR-124a expression in Caco-2 cells in different groups. (B) AHR mRNA expression was determined by qRT-PCR analysis in different groups. Results are presented as mean ± SEM. *P < 0.05; *** P < 0.001 (PNG 145 kb)

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Zhao, X., Li, J., Ma, J. et al. MiR-124a Mediates the Impairment of Intestinal Epithelial Integrity by Targeting Aryl Hydrocarbon Receptor in Crohn’s Disease. Inflammation 43, 1862–1875 (2020). https://doi.org/10.1007/s10753-020-01259-0

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