Effects Of All-trans Retinoic Acid Combined With Radioactive Ray On Cell Cycle, Apoptosis And NF-κB, β-catenin, CyclinD1 Gene Expression In Esophageal Carcinoma Cell

Objective: Esophageal carcinoma is one of the most common malignant tumors in China. Its mortality rate stands the forth in all the malignant tumors, and it severely threatens people's health. Because the morbidity of most esophageal carcinoma is latent,over 50% patients were in the advanced stage when they saw a doctor and lose the opportunity of surgery. It is a hotspot to explore a more effective therapy in the EC treatment field. Therefore, chemotherapy is becoming more and more important in the therapy of esophageal carcinoma. ATRA is one of the most intensive and strongest chemotherapeutic medicines by far. If ATRA is used alone, it will induce enormously poisonous side effect and easily product continuous drug fastness. But use the chemo-treatment and radiation treatment jointly, the dose of radiation medicine could decrease and the side-effect downsize, therefore the results of cooperative synergy will be achieved. So it's an important direction in tumor research to explore the joint application of ATRA and radioactive ray.This thesis studied the effects of ATRA combined with radioactive ray on cell cycle, apoptosis and NF-κB,β-catenin, CyclinD1 gene expression in esophageal carcinoma cells to provide theoretic basis and reference index for the diagnosis and treatment of esophageal carcinoma.Methods: 1 To measure the inhibition rate of growth of the Eca109 cell lines treated at 24th hour, 48th hour and the 72nd hour with ATRA by MTT chromatometry. Chose the drug concentration that caused the inhibition rate 25%, 50% after 48h respectively as the examined concentration of flow cytometry and cell immunochemistry assays.2 Flow cytometry (FCM) was adopted to analyze the changes in cell cycle and apoptosis . Grouping: The Eca109 cell lines were divided into the following groups. The control: without ATRA and radiation interference; ATRA alone: ATRA (0.78μmol/L, 5μmol/L), respectively exposed to Eca109 cell lines; radiation alone:γ-ray radiation at a dose of 6Gy; ATRA and radiation: afterγ-ray radiation Eca109 cell lines were exposed to ATRA (0.78μmol/L, 5μmol/L) respectively. Eca109 cell lines were treated for 24 hours, 48 hours respectively.3 To survey the expression of the NF-κB,β-catenin, CyclinD1 gene before and after treated by ATRA in Eca109 cell lines by cell immunochemistry assays.4 FCM was adopted to analyze the changes in the protein product of NF-κB,β-catenin, CyclinD1 in different group. Grouping: The same as 2.5 Experimental data are disposed by SPSS 11.5 statistics software, using single-factor analysis of variance, two-sample t-test,and linear correlation, with significance of difference standard atα=0.05.Results:1 The results by MTT assay: The tendency of inhibition increased along with the dose gradual increase. There was significant difference among each group treated by ATRA with the one untreated (P<0.05).2 After induced by ATRA of 0.78μmol/L for 24 hours, the number of cells in G0/G1 phase was elevated from 35.40±1.57% to 40.47±1.96%, and 46.13±0.68% by ATRA of 5μmol/L; there was significant difference between the two groups(P<0.05). The number of cells in G0/G1 phase was elevated to 38.73±1.33% after radiation. The number of cells in G0/G1 phase was elevated to 48.13±0.90% when Eca109 cell lines were treated with ATRA of 0.78μmol/L and radiation, and 51.30±1.04% by ATRA of 5μmol/L and radiation; there was significant difference between the two groups(P<0.05). There was significant difference between the combining group and the alone one (P<0.05). There was significant difference among each group treated with the one untreated (P<0.05).The analysis of cell cycle after treated for 24 hours was the same as that treated for 48 hours.3 After induced by ATRA of 0.78μmol/L for 24 hours, the apoptosis ratio of Eca109 cells was elevated from 1.68±0.16% to 3.41±0.26%; and 5.03±0.23% by ATRA of 5μmol/L; there was significant difference between the two groups(P<0.05). The apoptosis ratio was elevated to 2.69±0.16% after radiation. The apoptosis ratio was elevated to 8.00±0.48% when Eca109 cell lines were treated with ATRA of 0.78μmol/L and radiation, and 11.44±0.51% by ATRA of 5μmol/L and radiation; there was significant difference between the two groups (P<0.05). There was significant difference between the combining group and the alone one (P<0.05). There was significant difference among each group treated with the one untreated (P<0.05).The analysis of apoptosis ratio after treated for 24 hours was the same as that treated for 48 hours.4 There was a significant down-regulation on expression of NF-κB,β-catenin, CyclinD1 in Eca109 cell lines treated with 5μmol/L ATRA after 48 hours (P<0.05).5 Decrease in NF-κB,β-catenin, CyclinD1 was demonstrated after ATRA treatment for 24 hours and 48 hours, and the decrease is obvious with the increase of ATRA concentrations(P<0.05). And decrease was observed after radiation alone. There was significant difference between the combining group and the alone one (P<0.05), and the decrease is obvious with the increase of ATRA concentrations (P<0.05). There was significant difference between each group treated and the one untreated (P<0.05). Linear correlation analysis demonstrated: NF-κB,β-catenin were in a positive correlation (P<0.05),β-catenin, CyclinD1 were in a positive correlation (P<0.05), NF-κB,β-catenin were in a positive correlation (P<0.05). Conclusions: 1 After induced by ATRA, the cell proliferation was significantly inhibited in dose-dependent manner, the tendency of inhibition increased along with the dose gradual increase.2 The transformation from G0/G1 phase cells to S phase cells was blocked by combination of ATRA with radioactive ray. At the same time, the apoptosis ratio was increased with the concentration increase.3 The combination of ATRA with radiation down-regulated the expression of NF-κB,β-catenin, CyclinD1 in Eca109 cells. With the increase of ATRA concentration, the expression of them decreased gradually, and NF-κB,β-catenin, CyclinD1 were in a positive correlation.