Song Jin-Lian, Yue Wang, Hou Lin, Ge Yin-Lin
Department of Biochemistry and Molecular Biology, Medical College, Qingdao University, Qingdao, Shandong, 266021, P. R. China.
Ai Zheng. 2008 Mar;27(3):283-8.
BACKGROUND & OBJECTIVE: Isatin (ISA) is a natural material that exists in mammalian body fluids and tissues. ISA could inhibit the growth of tumors, but the mechanism remains unclear. This study aimed to explore the effects of ISA on the cell cycle and apoptosis of neuroblastoma cell line SH-SY5Y.
Fluorescent staining, flow cytometry, and Western blot were performed to analyze the cell cycle arrest and apoptosis of SH-SY5Y cells after treatment of ISA at different concentrations (0, 100, 200, 400 micromol/L).
When treated with 400 micromol/L ISA for 48 h, SH-SY5Y cells showed typical apoptotic morphologic changes including chromatin condensation and DNA fragment. Along with the increase of ISA concentration, Bcl-2 expression was decreased, the ratio of Bcl-2 to Bax was significantly decreased (P<0.05). When treated with ISA (100, 200, 400 micromol/L) for 48 h, the positive rates of activated Caspase-3 in SH-SY5Y cells were significantly higher than that in control SH-SY5Y cells (19.28%, 25.88%, and 33.43% vs. 10.58%, P<0.05). Moreover, inhibitor of caspase-activated DNase (ICAD), the substrate of Caspase-3, was degraded. In addition, the proportion of SH-SY5Y cells at G1 phase was significantly increased with an apparent G1 phase arrest when treated with ISA (100, 200, 400 micromol/L) for 48 h. In the progress of cell cycle arrest induced by ISA, phosphorylated ERK and CDK1 expression were down-regulated (P<0.05).
ISA can induce apoptosis and G1 phase arrest in SH-SY5Y cells, possibly by decreasing Bcl-2/Bax, activating Caspase-3 and down-regulating the expression of phosphorylated ERK and CDK1.
异吲哚酮（ISA）是一种存在于哺乳动物体液和组织中的天然物质。ISA可抑制肿瘤生长，但其机制尚不清楚。本研究旨在探讨ISA对神经母细胞瘤细胞系SH-SY5Y细胞周期及凋亡的影响。
采用荧光染色、流式细胞术及蛋白质免疫印迹法，分析不同浓度（0、100、200、400 μmol/L）ISA处理后SH-SY5Y细胞的细胞周期阻滞及凋亡情况。
400 μmol/L ISA处理48 h后，SH-SY5Y细胞呈现典型的凋亡形态学变化，包括染色质浓缩和DNA片段化。随着ISA浓度的增加，Bcl-2表达降低，Bcl-2与Bax的比值显著降低（P<0.05）。100、200、400 μmol/L ISA处理48 h后，SH-SY5Y细胞中活化的Caspase-3阳性率显著高于对照SH-SY5Y细胞（分别为19.28%、25.88%、33.43% vs. 10.58%，P<0.05）。此外，Caspase-3的底物——半胱天冬酶激活的脱氧核糖核酸酶抑制因子（ICAD）被降解。另外，100、200、400 μmol/L ISA处理48 h后，SH-SY5Y细胞G1期比例显著增加，出现明显的G1期阻滞。在ISA诱导细胞周期阻滞过程中，磷酸化的ERK和CDK1表达下调（P<0.05）。
ISA可能通过降低Bcl-2/Bax、激活Caspase-3以及下调磷酸化ERK和CDK1的表达，诱导SH-SY5Y细胞凋亡和G1期阻滞。
