Radiotherapy And Sensitivity Of Tumor Necrosis Factor - α (rhTNF - α) To Human Esophageal Cancer Cells TE13, KYSE140 And SEG1

Purpose:To assess the effects of tumor necrosis factor-a (TNF-a) on the degree of differentiation of three kinds of esophageal cancer cell lines, TE 13 (poorly differentiated squamous cell carcinomas), KYSE 140 (moderately differentiated squamous cell carcinomas) and SEG 1 (adenocarcinoma), the cell growth inhibition and the radiotherapy sensitization effect in vitro.Methods:Three different differentiation degrees of esophageal cancer cell lines, TE 13, KYSE 140 and SEG 1 were cultured in the medium containing 10% fetal bovine serum RMPI-1640. rhTNF-a were respectively acted on these esophageal cancer cell lines (100 IU/ml,200 IU/ml, 400 IU/ml,1200 IU/ml,1600 IU/ml,3200 IU/ml), IC50(half maximal inhibitory concentration) were calculated according to the corresponding regression equation, which were adopted as experimental concentration. All three cell lines were divided into control group, only rhTNF-a treated group, only X-ray radiation group and combined group when they developed to the logarithmic phase. Absorbance value (OD value) were detected after 12 h,24 h,48 h,72 h. Then the cell growth inhibition curve can be drawn through MTT method. Above-mentioned grouping method still be adopted, flow cytometry was used to detect cell apoptosis after 24 h, clone formation test was aimed at radiotherapy sensitization and Western blot method was considered to detect the expression of apoptosis related gene performed by three esophageal cancer cell lines.Results:1. Cell growth inhibition curve:Apoptosis curves of three kinds of esophageal cancer cell lines those were treated by rhTNF-a with different concentrations showed that IC50 were apartly 400 IU/ml,400 IU/ml, and 1200 IU/ml. Under these concentrations, the cells’inhibition rate increased over time and slowed after 24 h. rhTNF-α has inhibitory effect among three cell lines and the strongest is combined group, which is distinctly higher than other three groups.2. Flow cytometry to detect cell apoptosis:The apoptosis rate of only rhTNF-a treated group and only X-ray radiation group were higher than the control group, the obvious phenomena was performed in the combined group, with he trend of apoptosis increasing significantly (P<0.05). rhTNF-a and X-ray irradiation have synergetic effect of all of TE13, KYSE140 and SEG1.3. Apoptosis-related protein: The results of western blotting showed that, compared to control group, the decline expression of NF-κB protein was stated between only rhTNF-a treated group and only X-ray radiation group and the maximum degree was also argued in combined group. On the contrary, Caspase-3 protein content increased, the jointed treatment group increased most. Three kinds of cells in the NF-κB protein were similar without rhTNF-a. Then SEG1 NF-κB protein content was reduced more than other two kinds with the treatment the medicine allied with X-ray radiation.Conclusion:rhTNF-a has no obvious impacts on the growth of three kinds of human esophageal cancer cells, the impacts improved greatly when we combined rhTNF-a with X ray radiation. Its mechanism may enhance Caspase-3 protein expressions and induce cell apoptosis through Caspase way, which increased the sensitivity of three kinds of human esophageal cancer cell lines, TE 13, KYSE 140 and SEG 1, to X-ray radiation therapy and enhance the effect of radiotherapy.