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Published on: January 7, 2019
miR-30d Inhibition Protects IPEC-J2 Cells Against Clostridium perfringens Beta2 Toxin-Induced Inflammatory Injury
Kaihui Xie1, Qiaoli Yang1, Zunqiang Yan1
1College of Animal Science and Technology, Gansu Agricultural University, Lanzhou, China.
Abstract:
Clostridium perfringens beta2 (CPB2) toxin, one of the virulence factors of Clostridium perfringens (C. perfringens), can cause necrotizing enterocolitis in piglets. Accumulating pieces of evidence indicate that microRNAs (miRNAs) refer to the regulation of inflammatory processes. Previously, we have discovered that miR-30d was differentially expressed between the ileum of normal piglets and C. perfringens type C-infected diarrheal piglets. Here, we found that miR-30d expression was lowered in CPB2 toxin-treated intestinal porcine epithelial cells (IPEC-J2) at different time points. Subsequently, we determined that miR-30d inhibitor attenuated CPB2 toxin revulsive inflammatory damage in IPEC-J2 cells and promoted cell proliferation and cell cycle progression, whereas miR-30d mimic had opposite results. In addition, we confirmed that Proteasome activator subunit 3 (PSME3) was a downstream target gene of miR-30d via a dual luciferase reporter assay, qPCR, and western blot. We also found that overexpression of PSME3 suppressed CPB2 toxin-induced inflammatory damage and promoted cell proliferation and cycle progression. Our results demonstrate that miR-30d aggravates CPB2 toxin revulsive IPEC-J2 cells inflammatory injury via targeting PSME3, thereby providing a novel perspective for the prevention and treatment of piglet diarrhea at the molecular level.
Insights
MicroRNA-30d exacerbates Clostridium perfringens beta2 toxin-induced intestinal injury by targeting PSME3. This finding offers new molecular insights for preventing and treating piglet diarrhea.
Area of Science:
- Molecular Biology
- Veterinary Medicine
- Microbiology
Background:
- Clostridium perfringens beta2 (CPB2) toxin is a virulence factor causing necrotizing enterocolitis in piglets.
- MicroRNAs (miRNAs) are key regulators of inflammatory processes.
- Previous studies identified differential expression of miR-30d in piglets infected with C. perfringens type C.
Purpose of the Study:
- To investigate the role of miR-30d in CPB2 toxin-induced intestinal injury in porcine epithelial cells (IPEC-J2).
- To identify the downstream target of miR-30d involved in CPB2 toxin-induced inflammation.
- To elucidate the molecular mechanism of miR-30d in regulating CPB2 toxin-induced intestinal damage.
Main Methods:
- Treatment of IPEC-J2 cells with CPB2 toxin and manipulation of miR-30d expression (inhibitor and mimic).
- Assessment of cell proliferation, cell cycle progression, and inflammatory damage.
- Dual luciferase reporter assay, qPCR, and western blot to identify and validate the target gene PSME3.
Main Results:
- miR-30d expression was downregulated in CPB2 toxin-treated IPEC-J2 cells.
- miR-30d inhibition attenuated CPB2 toxin-induced inflammation and promoted cell proliferation and cycle progression.
- PSME3 was confirmed as a direct target of miR-30d, and its overexpression suppressed CPB2 toxin-induced injury.
Conclusions:
- miR-30d aggravates CPB2 toxin-induced inflammatory injury in IPEC-J2 cells by targeting PSME3.
- The miR-30d/PSME3 axis plays a critical role in the pathogenesis of CPB2 toxin-induced intestinal damage.
- This study provides a novel molecular perspective for preventing and treating piglet diarrhea.
