MiR-532 downregulation of the Wnt/β-catenin signaling via targeting Bcl-9 and induced human intervertebral disc nucleus pulposus cells apoptosis

Abstract

Objective: Intervertebral disc degeneration was characterized with aberrant intervertebral disc nucleus pulposus cells apoptosis. MiR-532 was reported to be up-regulated in the patients with intervertebral disc degeneration. However, the role of miR-532 in intervertebral disc degeneration remains unclear. Thus, current study aim to investigate the effects of miR-532 on human intervertebral disc nucleus pulposus cells.

Methods: Quantitative PCR was firstly used to evaluate the level of miR-532 expression in intervertebral disc nucleus pulposus cells. Then, luciferase reporter assays was used to confirm the target genes of miR-532. The effects of miR-532 on cell proliferation were assayed using EdU transfection. Next, apoptosis level was examined by flow cytometry utilizing Annexin V-FITC/PI double staining, and the protein expression in cells was detected with Western blotting. Finally, The expression of β-catenin protein was demonstrated by fluorescence confocal microscopy and the interaction between β-catenin and Bcl-9 was detected by co-immunoprecipitation.

Results: MiR-532 was significantly upregulated in patient with intervertebral disc degeneration. In addition, intervertebral disc nucleus pulposus cells apoptosis was significantly increased in miR-532 mimics treated group. Moreover, Bcl-9 was confirmed to be a direct target of miR-532 via luciferase reporter assays and western blot assays.

Conclusion: MiR-532 downregulation of the Wnt/β-catenin signaling via targeting Bcl-9 and induced human intervertebral disc nucleus pulposus cells apoptosis.

Keywords: Apoptosis; Intervertebral disc nucleus pulposus cells; Wnt/β-catenin; miR-532.