The Study On The Effects Of5-aza-2’-deoxycytidine,in Combination With Valproic Acid On The Growth Inhibition In Bile Duct Cancer Cells

Objective To investigate the effects of5-aza-2’-deoxycytidine,incombi-nation with Valproic acid on the growth inhibition in bile duct cancer cell TFK-1and preliminary discusses its antitumor mechanismMethods1. Applicating Cell Proliferation Inhibiting Methods（MTT Method）to detect the survival rate of bile duct cancer cell TFK-1exposure to differentconcentration of5-Aza-CdR and VPA,and in combination with each other.2. Applicating Flow Cytometry (FCM)to detect cell cycle and apoptosis of TFK-1exposure to5-Aza-CdR in combination with VPA.3.Applicating Werstern-Blot technology to detect cell cycle and apoptosis relatedprotein expression of TFK-1exposure to5-Aza-CdR in combination with VPA.Results1. Both5-Aza-CdR and VPA have growth inhibition effect,and showingtime and dose effect relations; Compared to the single group, at the same time thecombined group shows significantly higher inhibition on cell vitality.this differenceis of statistical significance in comparision with the single group(P<0.05).2.the control group shows cells mainly distributed in G0/G1phase and S phase, G2/M phase is less.TFK-1cell exposure to5-Aza-CdR or VPA alone after24h,G0/G1phase cells drop and S phase cells increase significantly(P<0.05); showingS phase block in dominant; the combined group shows G0/G1phase block andG2/M phase block, this difference is of statistical significance in comparision withthe single group(P<0.05);BothVPA and5-Aza-CdR group all shows cell apoptosis,showing time and dose effect relation;compared to the single group, the combinedgroup shows significantly higher rate.(P<0.05)3. Compared with the control group, TFK-1cell exposure to5-Aza-CdR or VPAalone after24h, HOXA5, P53, P21, Bax expression was increased(P<0.05);exposure to5-Aza-CdR alone,bcl-2expression was reduced(P<0.05),but VPAgroup wasn’t obvious(P>0.05).the combined group shows P53,P21, Bax, HOXA5protein expression is much increased (P<0.01)and Bcl-2expression is reduced(P<0.05).Conclusions1.5-Aza-CdR in combination with VPA collaborative increasecell growth inhibition effect。2. Through raising the expression of p53, p21, bax and reducing bcl-2,5-Aza-CdRin combination with VPA regulates cycle arrest and cell apoptosis, inhibiting thegrowth of bile duct cancer cell TFK-1.3. HOXA5may induce TFK-1cell apoptosis by P53depending apoptosis way;Hoxa5and P53gene silencing may be one of the mechanism of early oncogenesisin cholangiocarcinoma.