Detection And Analysis Of The Twist Gene Which Was Activated By Wnt Signal Pathway In Caski Cell Lines

ObjectiveWnt signal pathway in cervical cancer caski cell line was activated by lithiumchloride,then detected the different expression of Twist gene in mRNA and proteinlevel. To investigate the relationship between Wnt signal pathway and Twist gene incervical cancer caski cell line, provided the experimental basis of the pathogenesisfor cervical cancer.MethodsAfter cultivated cervical cancer caski cell line,the Wnt signal pathway wasactivated by different concentration of lithium chloride,then selected the suitableactivator concentration by MTT.This experiment was divided into seven groups:control group (without lithium chloride) and0.8、0.4、0.2、0.1、0.05、0.025mol/L sixconcentration of lithium chloride groups. After fostered12hours、24hours,the cellgrowth inhibition rate in different concentration and time was detected by MTT,thenscreened out the minimum inhibition concentration and second low inhibitionconcentration of lithium chloride.Frist, the expression of beta-catenin which was themark that represented the activation of Wnt signal pathway was detected by Westernblot, then detected the expression of Twist gene in mRNA and protein level byRT-PCR and Western blot when Wnt signal pathway was activated in caski celllines.The experiment was divided into five groups:(1)control group (without lithiumchloride);(2) the minimum inhibition concentration of lithium chloride fostered12hours;(3)the minimum inhibition concentration of lithium chloride fostered24hours;(4) the second low inhibition concentration of lithium chloride fostered12hours;(5) the second low inhibition concentration of lithium chloride fostered24hours.The expression of Twist gene in five groups of cells were detected by RT-PCRand Western blot in mRNA and protein level.At last, analysed the expression of Twist gene in mRNA and protein level when the Wnt signal pathway was activated.Results1.The caski cells was dealed with different concentration of lithium chlorideand fostered12hours and24hours respectively,then calculated the minimuminhibition concentration and the second low inhibition concentration of lithiumchloride were0.05mol/L、0.1mol/L, the two concentration of lithium chloridecorresponded to cell inhibition rates in12hours and24hours were6.600±0.027，4.200±0.063and21.800±0.021，46.200±0.073.2.The Wnt signal pathway was activated by lithium chloride,then the expressionof beta-catenin was increased（P<0.01).3.The expression of Twist gene in experimental groups that dealed with the0.05mol/L、0.1mol/L of lithium chloride and fostered12hours、24hours respectivelywere significantly increased than the control group in mRNA and protein level(P<0.05).This result showed that the activation of Wnt signal pathway could lead to theexpression of Twist gene increased in caski cell line.Conclusion1.The expression of beta-catenin was increased after the lithium chloride dealedwith the caski cell line, it indicated that Wnt signal pathway was activated.2.The Wnt signal pathway was activated by lithium chloride,then promoted theexpression of Twist gene in caski cell lines.