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MicroRNA MiR-27a-5p Alleviates the Cerulein-Induced Cell Apoptosis and Inflammatory Injury of AR42J Cells by Targeting Traf3 in Acute Pancreatitis

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Abstract

Acute pancreatitis (AP), a sudden inflammatory process of pancreas, is painful and may contribute to death. The aberrant expression of miR-27a-5p has been reported in many types of cancers and diseases including AP. Thus, it is urgent to manifest the functions and mechanism of miR-27a-5p in AP. The levels of miR-27a-5p, tumor necrosis factor (TNF) receptor–associated factor 3 (Traf3) in serum of AP patient, or cerulein-treated AR42J cells were detected by qRT-PCR. Functionally, the apoptotic rate, the protein levels of Bcl-2 and Bax, the caspase-3 activity, and the levels of IL-1β, IL-6, and TNF-α in cerulein-treated AR42J cells were measured by flow cytometry, Western blot, caspase-3 activity assay, and qRT-PCR and ELISA assay, respectively. In addition, the putative target of miR-27a-5p was predicted by TargetScan online database, and the dual luciferase reporter assay and RNA immunoprecipitation (RIP) assay were conducted to verify this interaction. Cerulein-treated mouse AP model was established to explore the role of miR-27a-5p in AP in vivo. The level of miR-27a-5p was notably downregulated in AP patients and cerulein-treated AR42J cells. The functional experiments indicated that miR-27a-5p mimics attenuated the promotion effects on cell apoptosis and the inflammatory response in AR42J cells caused by cerulein. The interaction between miR-27a-5p and Traf3 was predicted by TargetScan online database and validated by dual luciferase reporter assay and RIP assay. Following qRT-PCR results exhibited that Traf3 was apparently enhanced in cerulein-treated AR42J cells. The further functional experiments disclosed that Traf3 overexpression relieved the inhibitory effects on cell apoptosis and the inflammatory response induced by miR-27a-5p mimics in cerulein-treated AR42J cells. Moreover, miR-27a-5p alleviated cerulein-induced injury in vivo. In this study, we established the cerulein-treated AR42J cells as AP model in vitro. We validated that miR-27a-5p was significantly downregulated, and Traf3 was strikingly upregulated in AP patient and/or cerulein-treated AR42J cells. The further mechanistical and functional experiments unraveled that miR-27a-5p regulated Traf3 to relieve the cerulein-induced cell apoptosis and inflammatory injury of AR42J cells. Therefore, this novel regulatory network may provide therapeutic target for AP patients.

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Authors and Affiliations

  1. Department of Emergency, Shan County Dongda Hospital, No. 1 Yanshi East Road, Shan County, Heze, 274300, Shandong, China

    Yi Zhu

  2. Department of General Surgery, Shan County Dongda Hospital, Heze, 274300, Shandong, China

    Shuguang Liu & Feng Wang

Authors
  1. Yi Zhu
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  2. Shuguang Liu
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  3. Feng Wang
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Correspondence to Yi Zhu.

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The study was permitted by the Ethics Committee of Shan County Dongda Hospital and performed according to the Declaration of Helsinki Principles. Informed consent was provided for all AP patients. The animal experiment was authorized by the Animal Ethical Committee of Shan County Dongda Hospital and conducted according to the National Guide for Care and Use of Laboratory Animals.

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Zhu, Y., Liu, S. & Wang, F. MicroRNA MiR-27a-5p Alleviates the Cerulein-Induced Cell Apoptosis and Inflammatory Injury of AR42J Cells by Targeting Traf3 in Acute Pancreatitis. Inflammation 43, 1988–1998 (2020). https://doi.org/10.1007/s10753-020-01272-3

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