| Objective:To investigate the molecular mechanism for isatin(ISA) to induce apoptosis in human neuroblastoma cell line SH-SY5Y.Methods:Human neuroblastoma cell line SH-SY5Y cells were divided into 4 groups and treated with ISA(0,100,200,400μmol·L-1) for 48h.Fluorescent staining and DNA Ladder analysis were used to examine the changes on cell morphology,vegf and apoptosis-related proteins such as bcl-2,bax,survivin,p53 were determined by Westem blotting analysis and their mRNA expressions were detected by reverse transcriptase polymerase chain reaction(RT-PCR),and caspase-3 was tested by Flow cytometry.Results:(1) When treated with 400μmol·L-1 ISA for 48 h,SH-SY5Y cells showed the typical apoptotic morphologic changes including chromatin condensation and DNA fragment and the obvious ladder changes.(2) RT-PCR and Western blotting showed that along with the increase of ISA concentration,the expressions of bcl-2,vegf,survivin,were respectively decreased in both protein and mRNA level(P＜0.05),the expressions of bax remained unchanced(P＞0.05),while the expressions of wild type p53 was increased(P＜0.05),all these changes showed a dose-dependent manner.(3)The positive rates of activated Caspase-3 in SH-SY5Y cells were significantly higher than that in control group(18.38%,26.93%,35.66%vs.11.23%,P＜0.05).Conclusion:These data suggest that ISA can induce apoptosis in human neuroblastoma cell line SH-SY5Y cells,which maybe associated with the fact that ISA can effect several factors in apoptosis signaling pathway:1)Downregulated the expressions of bcl-2/bax,vegf,survivin.2)Upregulated the expressions of wild type p53 and Caspase-3. |
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