The Study On Relationship Between Methylation, Apoptosis And Carcinogenesis In Esophageal Cancer

Objective:To investigate the relationship between methyltransferase(5-Aza-2' -Deoxycytidine,5-Aza-CdR),apoptosis associated genes(Survivin;Caspase-3)and carcinogenesis in esophageal cancer.Methods:Human esophageal squamous cell cancer(ESCC)Eca109 Line were treated by 5-Aza-CdR with 10^-7,10^-6,10^-5,10^-4,0 mol/L, respectively.Consequently,the growth rate of the cells was detected by MTT assay and morphological structure observation;Meanwhile,cell apoptosis and cell cycle were analyzed by flow cytometry(FCM)method.The transcription of Survivin mRNA and expression of Caspase-3 were examined in 81 ESCC and 58 non-cancer esophageal mucosa tissues by in situ hybridization(ISH)and Immunohistochemistry(IHC)technique. Results:The proliferation of Eca109 cells was inhibited by 5-Aza-CdR from the concentration of 10^-7to 10^-4mol/L,and the inhibitive rate showed time-and-concentration-dependent manner(1～4 d,10^-7～10^-4mol/L,P＜0.05),inhibitory curves were achieved peak at 96 hours in each concentration group.An apoptosis peak appeared before diploid peak,and the proportion of G₀/G₁ phase cells was significantly increased;however,the proportion of S phase cells was decreased and apoptotic morphology was observed by invert-microscope.In additional,the positive frequence of Survivin mRNA transcription was 79.01%(64/81)in ESCC group and 22.41%(13/58)in non-cancer esophageal mucosa groups.There was a significant difference between ESCC and non-cancer group (P＜0.05),whereas there was no significant difference between carcinogenesis of ESCC and either of parameters,shch as gender,age,cell differentiation,lymph node metastasis (P＞0.05).The positive frequence of Caspase-3 protein expression was 27.16%(22/81) in ESCC and 82.76%(48/58)in non-cancer group.There was a statistically difference between ESCC and non-cancer group(P＜0.05).But there was no statistically difference between ESCC and either of parameters,shch as gender,age,cell differentiation,lymph node metastasis(P＞0.05).The level of Survivin transctrption was reversely related to the expression of Caspase-3 protein in ESCC group(P＜0.05).Conclusions:The growth of Eca109 cells were inhibited by 5-Aza-CdR via reactivating tumor suppressive genes silenced by promoter methylation in vitro,which suggested hypermethylation was involved in the carcinogenesis in ESCC.Survivin and Caspase-3 genes may play a significant role in the ESCC.