EGCG On Esophageal ECA109Cell Proliferation Apoptosis And The Influence Of Survivin And VEGF Expression

Objective: human esophageal cancer cells109(ECA109) cultivatedin vitro were used to observe the influence of the cellular proliferation andapoptosis of ECA109,and the expression of mRNA of Survivin andVEGF,depending on the different concentration of Epigallaocatechin-3-gallate(EGCG),which provides the way to study the inhibition mechanism ofEGCG to ECA109,as well as the clinical significance. Method:1.MTTassay was used to measure the optical density (OD) of ECA109cultured withdifferent concentration (40、80、120μg/ml) of EGCG for24,48and72hours toanalyze the proliferation of ECA109.2. apoptosis rate of the different groupsafter coloured by Annexin V-FITC and PI was analyzed by FlowCytometry(FCM).The expressions of mRNA of Survivin and VEGF aftertreated by the drug were analyzed by Reverse Transcription-Polymerase ChainReaction(RT-PCR).Results:1.the results observed by inverted phase contrastmicroscope shows a series of morphological changes,including decrease incell volume, cell shrinkage and be more spherical, cell chromaticagglutination, vacuoles occurred in cell and so on, with the increase of EGCGconcentration.2. There was a significant difference among each group atdifferent action time（24h、48h、72h） with the same concentration(P<0.05),andthe inhibition rate of each group cultured for24h,respectively,was14.24±0.72，21.60±1.03and29.56±1.27(P<0.05, the difference wassignificant).when the co-culture time increased to48hours, the results were39.17±1.50，43.44±1.66，48.58±1.73(P<0.05, the difference wassignificant),and when the drug intervention time increased to72hours, the results were49.26±0.85，54.52±2.29，58.48±1.55(P<0.05, the difference wassignificant).while There was a significant difference among each group withdifferent concentration（40、80、120μg/ml）at the same action time(P<0.05).Andthe inhibitory effect of EGCG on the proliferationof ECA109showed in atime-dose dependent manner.3.The cell apoptosis rate of ECA109wastested by Flow cytometry using different concentration(0、40、80、120μg/m1)of EGCG.The apoptosis rate in the early cycle of each groupwas,respectively,0.0667±.03055、4.8367±0.73146、12.2533±1.22696、23.5100±1.76847,while the total apoptosis rate of that was0.5200±.09539、10.3200±0.95252、24.1267±1.44057、34.0967±1.82878.The results showed thatthere was a significant difference among each group on the apoptosis rate inearly, and was same on the total apoptosis rate(P<0.05).As well as thepromoting effect of EGCG on the both apoptosis rates of ECA109alsoshowed in a time-dose dependent manner.4.The expression quantity ofmRNA of Survivin and VEGF was tested by RT-PCR using differentconcentration(0、40、80、120μg/m1) of EGCG. The result of the expressionquantity of mRNA of Survivin of each group respectively was0.5419±0.01873、0.4844±0.01478、0.3330±0.00685、0.2155±0.00304(P<0.05,the difference was significant),while the result of that of VEGF of each grouprespectively was1.0747±0.03547、0.9312±0.03899、0.5215±0.02270、0.3846±0.01732(P<0.05, the difference was significant). And the inhibitoryeffect of EGCG on the expression quantity of mRNA of Survivin andVEGF showed that with the increasing of the concerntration of EGCG,theexpression of mRNA attended to descend.5. There was a positive correlationbetween the expression of mRNA of Survivin and VEGF(r=0.965，P<0.05). Conclusions:1. The proliferation of ECA109may be inhibited by EGCG in atime-dose dependent manner.2. EGCG may promote the apoptosis of ECA109related to time-dose dependency.3. EGCG may reduce the expression quantityof mRNA of Survivin and VEGF to induce the apoptosis.