AUTHOR=Lan Tao , Zheng Yu-chen , Li Ning-dao , Chen Xiao-sheng , Shen Zhe , Yan Bin 

TITLE=CRISPR/dCas9-Mediated Parkin Inhibition Impairs Mitophagy and Aggravates Apoptosis of Rat Nucleus Pulposus Cells Under Oxidative Stress

JOURNAL=Frontiers in Molecular Biosciences

VOLUME=8

YEAR=2021

URL=https://www.frontiersin.org/journals/molecular-biosciences/articles/10.3389/fmolb.2021.674632

DOI=10.3389/fmolb.2021.674632

ISSN=2296-889X

ABSTRACT=<sec><title>Objective</title><p>The aim of this study is to explore the role of Parkin in intervertebral disk degeneration (IDD) and its mitophagy regulation mechanism.</p></sec><sec><title>Study design and methods</title><p>Rat nucleus pulposus (NP) cells were stimulated with hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) to a mimic pathological condition. Apoptosis and mitophagy were assessed by Western blot, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, and immunofluorescence staining. The CRISPR–dCas9–KRAB system was used to silence the expression of Parkin.</p></sec><sec><title>Result</title><p>In this study, we found that Parkin was downregulated in rat NP cells under oxidative stress. In addition, treatment with H<sub>2</sub>O<sub>2</sub> resulted in mitochondrial dysfunction, autophagy inhibition, and a significant increase in the rate of apoptosis of NP cells. Meanwhile, mitophagy inhibition enhanced H<sub>2</sub>O<sub>2</sub>-induced apoptosis. Furthermore, repression of Parkin significantly attenuated mitophagy and exacerbated apoptosis.</p></sec><sec><title>Conclusion</title><p>These results suggested that Parkin may play a protective role in alleviating the apoptosis of NP cells <italic>via</italic> mitophagy, and that targeting Parkin may provide a promising therapeutic strategy for the prevention of IDD.</p></sec>