Study Of The Controlling Effect On The Expression Of The Inhibitors Of Apoptosis Genes From DNA Damage Response Pathway And PI3K Signal Pathway Induced By Synchronious Prostate Cancer Cell DU145

Objective:To study the expressions of the genes of inhibitor of apoptosis family in the different cell cycle at the synchronous phase,the control to the genes of inhibitor of apoptosis by the DNA damage response pathway caused by the synchronization,and the control of the genes by the PI3K/AKT signal transduction pathway,and to analysis the relationship between the expression of the genes of inhibitors of apoptosis and abnormality of the construction and number of the chromosomes in which the genes of inhibitors of apoptosis are located.Method:Starved method is used to obtain synchronous G0 phase cell.Mimosine,thymidine and nocodazole are used respectively to obtain the G1 phase,S phase and G2/M phase cell causing the DNA damage response,.at the same time,PI3K inhibitor ly294002 is added to the culture in order to prevent PI3K/Akt signal transduction pathway. Semiquantitative RT-PCR method is used to detect the DNA damage checkpoint,and to detect the mRNA expression levels of inhibitors of apoptosis genes induced by PI3K/Akt signal transduction pathway at the differen cell cycle phase.The common cytogenetic G-banding technique is used to analysis the specific abnormality of chromosomes which the inhibito of apoptosis genes are located in.Result:The rate of synchronous cell at the G0/G1 phase by mimosine is 78.04%,S phase by thymidine 62.19%,G2/M phase by nocodazole 60.5%。The result of cytogenetics analyusis demonstrates those chromosomes which the investigated genes are located in present significant constructre and numerical abnormality.Chromosome 11 in which the BIRC3/API2 gene is located presents translocation,for example,t(11;12)(q;q).Chromosome 2 in which apollon/BIRC6 gene is located present deletion or duplication,for example,2p~- 2p~+.And the chromosome 20 in which the BIRC7/Livin gene is located present the haploid,for example,del(20),which affects the expression of BIRC7/Livin gene.To the fused gene API2-MALT1,the correspoding translocation of chromosome may present,t(11,18)(q21;q21).DNA damage response pathway and survival signal pathway are activated by the drugs for cell synchronization,promote the expression of BIRC3/API2 mRNA.But PI3K inhibitor ly294002 inhibit the expression of BIRC3/API2 mRNA.At the G2/M phase,ly294002 inhibit significantly the expression of apollon mRNA.Conclusion:Prostate cancer cell DU145 presents significant structure and numerical abnormality of its chromosomes,which may affect the expression of associated genes.The drugs for the cell synchronization activate the DNA damage pathway and survival signal pathway,and their signals are transmitted by PI3K pathway but not p53 signal pathway,the cell line DU145 had no functonal p53 gene or Rb gene,to regulate the expression of BIRC3 mRNA,or to regulate the expression ofBIRC6/Apollon mRNA at a certain cell cycle phase.The expression of BIRC6/Apollon mRNA is not coincident with the previous reports,we hope to continue to study this question.