Attenuation of the cytoprotection induced by hypoxic preconditioning upon transfection with BNIP3-siRNA in human neuroblastoma SH-SY5Y cells

Abstract

Purpose

The aim of this study was to investigate the functional role of hypoxic preconditioning (HPC) in human neuroblastoma cells.

Methods

BNIP3 small-interfering RNA (BNIP3-siRNA) sequence was synthesized and used to transfect human neuroblastoma SH-SY5Y cell lines. Thereafter, BNIP3 expression at mRNA and protein levels and its effects on the cell proliferation were analyzed. The most effective pair of siRNA was selected to knockdown the expression level of BNIP3. Moreover, the effects of HPC on oxygen-glucose deprivation/reperfusion (OGD/R)-induced apoptosis and autophagy in SH-SY5Y cells were explored to further reveal the possible mechanisms underlying HPC.

Results

BNIP3-siRNA attenuated the protective effects of HPC by decreasing the cell viability, increasing the enzymatic activity of caspase-3 and 9, increasing the rate of apoptosis, and increasing the protein expression level of activated caspase-3. Additionally, BNIP3-siRNA had no significant influence on the expression level of HIF-1α induced by HPC, while it substantially inhibited HPC-induced BNIP3/Beclin1 and autophagy.

Conclusions

HPC promoted autophagy through regulating BNIP3 to reduce OGD/R.

Keywords:

• SH-SY5Y
• BNIP3
• Beclin1
• hypoxic preconditioning
• oxygen glucose deprivation/reperfusion

Authors’ contributions

Lu Na designed and supervised the project. Lu Na and Bai Ruiying drafted the manuscript, designed the experiments, and interpreted the data. Lu Na and Liu Bo were jointly responsible for funding acquisition. Lu Na, Liu Bo, and Weijia Cheng performed the experiments and acquired the data. Weijia Cheng and Wu Zekun carried out statistical analysis. All the authors read and approved the final manuscript.

Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Ethics statement

The Ethics Committee of the Xinxiang Medical University (Xinxiang, China; Approval No. XYLL-B2013001) approved the study protocol.

Additional information

Funding

This research was financially supported by the National Natural Science Foundation of China (Grant Nos. 81771517 and 81571205) and the Key Research Project for Science and Technology of the Education Department of Henan Province, China (Grant No. 14A310009).