miR-15b and miR-16 induce the apoptosis of rat activated pancreatic stellate cells by targeting Bcl-2 in vitro.

OBJECTIVES

Activated pancreatic stellate cells (PSCs) play a pivotal role in the development of pancreatic diseases, especially chronic pancreatitis and pancreatic cancer. MicroRNAs have become a focal point of interest as post-transcriptional regulators of gene expression via their interaction with the 3' untranslated region of target mRNAs, which results in gene silencing. We examined the relative expression of microRNAs (miR-15b and miR-16) and their target gene, Bcl-2, during activation of rat PSCs, and determined their effects on apoptosis of rat PSCs in vitro.

METHODS

miR-15b and miR-16 expression levels were analyzed in quiescent and activated PSCs by stem-loop RT-PCR. In addition, the effects of miR-15b and miR-16 on apoptosis of activated PSCs were investigated by immunofluorescence microscopy with Hoechst 33342 staining, and flow cytometry with annexin-V/propidium (PI) co-labeling. Bcl-2 and Bcl-xl were also analyzed by real-time RT-PCR and Western blotting.

RESULTS

During activation of PSCs, from the quiescent stage to activated stage, miR-15b and miR_16 were downregulated, while Bcl-2 expression was upregulated. Restoring intracellular miRNA levels by miR-15b and miR-16 administration greatly reduced Bcl-2 protein levels, and significantly induced apoptosis in activated PSCs.

CONCLUSIONS

miR-15b and miR-16 could induce apoptosis of rat PSCs by targeting Bcl-2.