[ZGDHu-1-inducing apoptosis of SHI-1 leukemia cells and its molecular mechanism].
The aim of study was to investigate the mechanism of N, N'-di-(m-methylphenyl)-3, 6-dimethyl-1, 4-dihydro-1, 2, 4, 5-tetrazine-1, 4-dicarboamide (ZGDHu-1) inducing apoptosis in SHI-1 human leukemia cell line. Different concentrations of ZGDHu-1 and different times of culture were used to treat SHI-1 cells; the apoptosis of SHI-1 cells was analyzed by morphology, DNA agarose gel electrophoresis, DNA content detection, Annexin-V/PI and Hoechst33258 labeling method, the mitochondrial transmembrane potential (Delta Psi m) were measured by dihydrorhodamin 123, and expressions of bcl-2, bax, Fas, p53 and mitochondrial membrane protein were analyzed by flow cytometry, while the bcl-2, bax and p53 gene were analyzed by RT-PCR. The transcriptional level of hTERT-mRNA was measured by real-time fluorescence quantitative RT-PCR. The results showed that after exposure to ZGDHu-1, SHI-1 cells were induced to apoptosis in a time-and does-dependent manner. SHI-1 cell apoptosis was confirmed by typical cell morphology, DNA fragmentation, sub-G(1) phase, Hoechst33258 and Annexin-V/PI labeling etc. The expression of bax, bax/bcl-2, p53 and Fas gene significantly increased and bcl-2 slightly decreased. ZGDHu-1 could increased the expression of mitochondrial membrane protein in a dose-dependent manner while Delta Psi m reduced. The expression of hTERT-mRNA significantly decreased. It is concluded that ZGDHu-1 can up-regulate the expression of p53, bax and bax/bcl-2. The mitochondrial pathway mediated by descent of mitochondrial transmembrane potential may be one of the mechanisms inducing apoptosis by ZGDHu-1, in which Fas gene also participates. Telomerase may be an effective gene target for anti-tumour effect of ZGDHu-1.
Zhou
Yong-Lie
YL
Central Laboratory, Zhejiang Provincial People Hospital, Hangzhou 310014 China. ZYL@126.com
Lü
Ya-Ping
YP
Hu
Wei-Xiao
WX
Qiu
Lian-Nü
LN
Wang
Wen-Song
WS
Liu
Jian-Dong
JD
Wu
Jian-Guo
JG
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D006573:Heterocyclic Compounds, 1-Ring / Q000494:pharmacology*; D006801:Humans; D007948:Leukemia, Monocytic, Acute / Q000473:pathology*; D051379:Mice; D008819:Mice, Nude; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D019098:Telomerase / Q000378:metabolism*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D000970:Antineoplastic Agents; D006573:Heterocyclic Compounds, 1-Ring; C522668:N,N-di-(m-methylphenyl)-3,6-dimethyl-1,4-dihydro-1,2,4,5-tetrazine-1,4-dicarboxamide; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D051028:bcl-2-Associated X Protein; C509186:TERT protein, human; D019098:Telomerase