Liu Bi, Lei Ming, Hu Tao, Yu Fei, Xiao De-Ming, Kang Hao
Central Laboratory, Peking University Shenzhen Hospital, Shenzhen, 518000, China.
Shenzhen Luohu People's Hospital, Guangzhou Medical University, Shenzhen, 518000, China.
J Huazhong Univ Sci Technolog Med Sci. 2016 Jun;36(3):350-355. doi: 10.1007/s11596-016-1590-y. Epub 2016 Jul 5.
SRT1720, a new discovered drug, was reported to activate silent information regulator 1 (SIRT1) and inhibit the chondrocyte apoptosis. However, the underlying mechanism remains elusive. In the present study, the chondrocytes were extracted from the cartilage tissues of New Zealand white rabbits, cultured in the presence of sodium nitroprusside (SNP) (2.5 mmol/L) and divided into five groups: 1, 5, 10, and 20 μmol/L SRT1720 groups and blank control group (0 μmol/L SRT1720). MTT assay was used to detect the chondrocyte viability and proliferation, and DAPI staining and flow cytometry to measure the chondrocyte apoptosis. The expression levels of SIRT1, p53, NF-κB/p65, Bax, and peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC-1α) were detected by Western blotting and the expression levels of SIRT1, type II collagen, and aggrecan mRNA by RT-PCR. The results showed that in the SRT1720-treated groups, the nuclei of chondrocytes were morphologically intact and had uniform chromatin. In the blank control group, nuclear rupture into debris was observed in chondrocytes. With the SRT1720 concentration increasing, the chondrocyte viability increased, the apoptosis rate decreased, the protein expression levels of SIRT1 and PGC-1α and the mRNA expression levels of type II collagen and aggrecan increased ({ptP}<0.05), and the expression levels of p53, NF-κB and bax decreased (P<0.05). It was suggested that SRT1720 inhibits chondrocyte apoptosis by activating the expression of SIRT1 via p53/bax and NF-κB/PGC-1α pathways.
SRT1720是一种新发现的药物，据报道它能激活沉默信息调节因子1（SIRT1）并抑制软骨细胞凋亡。然而，其潜在机制仍不清楚。在本研究中，从新西兰白兔的软骨组织中提取软骨细胞，在硝普钠（SNP）（2.5 mmol/L）存在的情况下进行培养，并分为五组：1、5、10和20 μmol/L SRT1720组以及空白对照组（0 μmol/L SRT1720）。采用MTT法检测软骨细胞活力和增殖情况，采用DAPI染色和流式细胞术检测软骨细胞凋亡情况。通过蛋白质印迹法检测SIRT1、p53、NF-κB/p65、Bax和过氧化物酶体增殖物激活受体γ辅激活因子1-α（PGC-1α）的表达水平，通过逆转录聚合酶链反应（RT-PCR）检测SIRT1、II型胶原蛋白和聚集蛋白聚糖mRNA的表达水平。结果显示，在SRT1720处理组中，软骨细胞核形态完整，染色质均匀。在空白对照组中，观察到软骨细胞核破裂成碎片。随着SRT1720浓度的增加，软骨细胞活力增加，凋亡率降低，SIRT1和PGC-1α的蛋白表达水平以及II型胶原蛋白和聚集蛋白聚糖的mRNA表达水平升高（P<0.05），而p53、NF-κB和Bax的表达水平降低（P<0.05）。提示SRT1720可能通过p53/bax和NF-κB/PGC-1α途径激活SIRT1的表达来抑制软骨细胞凋亡。
