Screening and validation of differentially expressed extracellular miRNAs in acute pancreatitis

Abstract

The present study aimed to screen for differentially expressed extracellular microRNAs (miRNAs) during the development of acute pancreatitis (AP) and validate the miRNA expression in the plasma of patients with AP. The culture medium of taurolithocholic acid-3 sulfate-treated rat pancreatic acinar AR42J cells was collected to extract total RNA for miRNA microarray analysis. Compared with the miRNA test results of the AP rats in the GEO databases, the differentially expressed extracellular miRNAs were screened. The TargetScan, miRanda, and PicTar programs were used for target gene prediction of the identified miRNAs, and gene ontology-biological processes (GO-BP) functional annotation was performed. Finally, the results from the combined microarray analyses (in vitro cell line and in vivo rat samples) were validated using plasma samples from patients with mild and moderately severe AP by reverse transcription-polymerase chain reaction. The results demonstrated that extracellular MIR-24 was differentially expressed by microarray and bioinformatics analysis in both the cell line and the animal model of AP. Bioinformatics prediction analysis revealed that downstream target genes of MIR-24 included Vav2, Syk, Lhcgr, Slc9a3r1, Cacnb1, Cacna1b, Bcl10, and Fgd3. Functional enrichment analysis revealed that the main GO-BP predicted functional presentations were positive regulation of calcium-mediated signaling, activation of c-Jun N-terminal kinase activity, calcium ion transport, regulation of Rho protein signal transduction, negative regulation of the protein kinase B signaling cascade, and the T cell receptor signaling pathway. Validation analysis for the plasma MIR-24 expression in humans revealed a significant upregulation of MIR-24 in the plasma samples of AP patients compared with the healthy controls, while no significant difference was observed in the MIR-24 expression between the mild and the moderately severe AP groups. The present study confirmed the high expression of MIR-24 in peripheral blood during AP, suggesting that MIR-24 might have an intercellular communication role contributing to the AP-associated distant organ injury.