Abstract: Objective  To investigate the protective role and mechanism of berberine(Ber) against hypoxia/reoxygenation(H/R)-induced injury in H9c2 cells via miR-29b.
Methods  The H9c2 cells cultured in vitro were divided into control group(normal culture), H/R group(H/R treatment) and H/R+Ber group(treated with 150 μmol/L Ber before H/R treatment). Cell viability was detected by MTT assay, and LDH release from cell supernatant was detected by LDH kit. Apoptosis was detected by flow cytometry, Caspase-3 activity was detected by Caspase-3 kit, and the expression level of miR-29b was detected by RT-PCR. The H9c2 cell line with low expression of miR-29b was established, and the effects of Ber and miR-29blow expression on H/R-induced injury in H9c2 cells were observed by the above methods. The bioinformatics prediction and dual luciferase reporter gene were used to verify the targeting relationship between miR-29b and PTEN, and the effect of miR-29b on PTEN protein expression was detected by Western blot.
Results  Compared with the control group, the cell survival rate and miR-29b expression level of H9c2 were significantly decreased in H/R group, while the LDH release in the cells upernatant, apoptosis rate and Caspase-3 activity were significantly increased(P＜0.05). Compared with H/R group, the above changes caused by H/R were significantly inhibited after Ber treatment(P＜0.05). miR-29b down-regulation significantly alleviated the protective effect of Ber on H/R-induced H9c2 cell injury(P＜0.05). The dual luciferase reporter gene assay confirmed that PTEN was a target gene of miR-29b. Western blot analysis confirmed that miR-29b could negatively regulate the expression of PTEN protein.
Conclusion  miR-29b plays an active role in the protective effect of Ber on H/R-induced H9c2 cell injury, and its mechanism may be related to the targeted regulation of PTEN expression.

Key words: myocytes, cardiac； berberine, apoptosis