Sirt‐1 Regulates Physiological Process and Exerts Protective Effects against Oxidative Stress
Lei Liu, Guangyuan Xia, Peifan Li, Yiming Wang, Qian Zhao
Abstract
Background . Recent studies suggest a correlation between the reduced Sirt‐1 expression with Alzheimer’s diseases (AD) and depression, respectively, suggesting a possible pathogenic role of the altered Sirt‐1 expression in neuronal degenerative diseases, such as AD and depression. However, the molecular mechanisms underlying how Sirt‐1 reduction impairs neuronal functions remain unknown. Methods . We used the SK‐N‐SH neuroblastoma cells to study the role of Sirt‐1 expression on physiological roles in neuronal cells. Gain of Sirt‐1 was achieved by transiently transfecting Sirt‐1 expression plasmid. Sirt‐1‐specific shRNA was used to elucidate the role of Sirt‐1 loss of function. CCK‐8 (Cell Counting Kit‐8) assay and flow cytometry were used to evaluate cell proliferation. Semiquantitative western blotting was used to detect relative protein levels. A further luciferase reporter gene assay was employed to examine the effect of Sirt‐1 expression on the transcriptional activity of p53. RT‐qPCR was used to determine the mRNA levels of p21, Bax, and Bcl‐2, which were the downstream target genes of p53. Results . Sirt‐1 suppressed the p53 downstream gene p21 transcription, while shRNA‐mediated Sirt‐1 knockdown resulted in a significant increase in p21 expression, implying a possibility that Sirt‐1 promotes neuron proliferation through suppressing p53 transcriptional activity. The mRNA and protein levels of p53 were not affected by the altered Sirt‐1 expression, suggesting that Sirt‐1 regulates the transcriptional regulatory activity of p53 rather than p53 expression. Indeed, we further confirmed that Sirt‐1 appeared to inhibit p53 transcriptional activity by attenuating its acetylation and resulted in a decrease of p53’s binding to the p21 promoter. Overexpressed Sirt‐1 scavenged reactive oxygen species (ROS) production in SK‐N‐SH with H 2 O 2 . Knockdown of Sirt‐1 presented opposite effect; the addition of EX527 (Sirt‐1 inhibitor) increased ROS accumulation. Conclusions . Oxidative stress induces Sirt‐1 in neuron cells, and Sirt‐1 promotes proliferation in SK‐N‐SH cells, which protects them from oxidative stress‐induced cell death, potentially via suppressing the transcriptional activity of p53. These results provide a molecular explanation underlying how the reduced Sirt‐1 potentially causes the AD and depression‐related diseases, supporting the idea that Sirt‐1 can possibly be used as a diagnostic biomarker and/or therapeutic drug target for the AD and depression‐related diseases.