Effects Of All-trans Retinoic Acid On Esophageal Cancer Cell EC109

Background and Objective:Esophageal cancer is a common gastrointestinal cancer. Owing to the low rate of early diagnosis, the prognosis remains poor even if surgery. About 0.3 million people died from the disease annually throughout the world. To find effective chemotherapy treatment of esophageal cancer drugs have become a hot research field. Retinoic acid is a major compounds, including vitamin A and its metabolites, with a similar structure and biological activity of natural and synthetic derivatives. Retinoic acid plays an important role in the normal physiology of human activities, such as the growth of embryo formation, visual and immunization. Numerous studies have shown that retinoic acid affects a variety of malignant tumors including induction of differentiation, inhibition of proliferation and induction of apoptosis, and so on. Squamous cell carcinoma of the skin, cervical cancer, breast cancer and other malignant tumors had entered clinical study phase, which promoted anti-tumor therapy. Retinoic acid drugs regulate cell proliferation, differentiation and apoptosis thereby inhibit tumor cell growth ,the mechanisms of which are still not fully explained. Whether ATRA may be an effective drug on the treatment of esophageal cancer or not? To answer this question, the experiments observing ATRA on esophageal cancer cell line EC109 in vitro study whether ATRA inhibit the esophageal cancer cell line EC109 and induce apoptosis or not, in the process of which the gene expression changes are detected and provide experimental evidence for further study.Materials and Methods: (1) The cell proliferation activity was detected by MTT assay after treated with ATRA The human esophageal cancer cell line EC 109 was cultured conventionally to log period and adjusted to concentration of 1×10~4/mL. EC 109 cell in adherent growth was divided into control and experimental groups 24 hours later. The control group was cultured with RPMI-1640 medium and the experimental groups was cultured with ATRA concentration of 10μmol/L,5μmoI/L,1μmol/L,0.5μmol/L and 0.1μmol/L etc, each of which had six repeated holes. The differences of cell proliferation inhibition rate between different groups was detected after cultured with ATRA 24h,48h,72h and 96h etc.(2) The cell apoptosis and cell cycle distribution was detected by flow cytometry machine The human esophageal cancer cell line EC 109 was cultured conventionally. The control group was cultured with RPMI-1640 medium and the experimental groups was cultured with 1μmol/L ATRA. Three bottles of EC109 cells was collected from the control and experimental groups respectively after replacing the new medium 24h,48h,72h and 96h. The cell apoptosis and cell cycle distribution was detected by flow cytometry machine.(3) The apoptosis-related gene mRNA levels were detected by RT-PCR The human esophageal cancer cell line EC109 was cultured conventionally. The control group was cultured with RPMI-1640 medium and the experimental groups was cultured with 1μmol/L ATRA. Three bottles of EC109 cells was collected from the control and experimental groups respectively after replacing the new medium 24h,48h,72h and 96h. The mRNA levels of apoptosis-related gene bcl-2 and bax were detected by RT-PCR technology.(4) Experimental data are disposed by SPSS 11.0 statistics soft ware, using single-factor analysis of variance , with significance of difference standard :α=0.05.RESULTS:(1) The inhibition effects of ATRA on EC109 cells were significantly dose-dependent and time-dependent. The difference of value A and inhibition rate between experimental groups and the control group and the adjacent experimental groups was statistically significant (P< 0.05) .(2) Compared with the cell of control group, there were apoptosis peak and significant cell cycle change in experimental groups after disposal by ATRA of 1.0μmol/L. With the disposal time increasing , the apoptosis rate increased ,G0/G1 cells ratio increased and S+G2/M phase cell ratio decreased. The difference of between experimental groups and the control group and the adjacent experimental groups was statistically significant (P<0.05) .(3) With the disposal time increasing ,the mRNA level of bcl-2 gene showed a significant gradual weakening trend but the mRNA level of bax gene showed a significant gradual increasing trend after EC109 cell disposal by ATRA of 1.0μmol/L. The difference of semi-quantitative results between and the control group and adjacent experimental groups were statistically significant (P<0.05) .CONCLUSION:1. ATRA inhibits the proliferation of human esophageal cancer cell line EC109 significantly.2. The apoptosis in EC109 cells could be induced by ATRA.3. The molecular mechanism of inducing apoptosis of EC109 may be related to down-regulation of bcl-2 transcription and up-regulation of bax transcription.