| Objective: In order to investigate the different radiosensitivities, we isolated the radioresistant cells from a human esophageal cancer cell line by repeated irradiation. Further, the aim of the study was to observe the effect of a human monoclonal antibody to EGFR (hR3) combined with recombinant adenoviruses-p53 anticancer injection (SBN-1) on esophageal cancer cells with different radiosensitivities.Methods: 1. Isolation and characterization of radioresistant human esophageal cancer cells in vitro. A radioresistant variant cell line, TE13R120, was isolated by repeated cobalt-60 gamma-radiation exposure of a radiosensitive esophageal cancer cell line, TE13. TE13 and TE13R120 cells were observed by phase-contrast microscopy and HE staining. The relative radiosensitivities of tumor cells were assessed by standard colony formation assays. The time-course of cell cycle distribution after irradiation was investigated by flow cytometry (FCM). 2. To examine the effect of a human monoclonal antibody to EGFR (hR3) combined with recombinant adenoviruses-p53 anticancer injection on esophageal cancer cells with different radiosensitivities. The expressions of epidermal growth factor receptor (EGFR) were detected by immunohistochemistry (IHC). The inhibition effect of cell growth and the influence by hR3 and SBN-1 interval 0,6,12,24h were measured with MTT method. Changes of cell cycle and expressions of Bax and Bcl-2 after treatment with hR3 and SBN-1 were detected by FCM.Results: 1. We isolated a radioresistant variant cell line, designated TE13R120, from the TE13 cell line by repeated gamma-radiation exposure and this variant cell line has retained its radioresistance. 1.1 Morphological features of this radioresistant TE13R120 and parental TE13 cell line. TE13R120 were somewhat larger cells with a non-uniform size and spindle shapes as compared to TE13. TE13 cells showed a more uniform shape and with more cytoplasm than TE13R120 cells by HE staining. 1.2 The difference of cell growth kinetics of the two kinds of cells. The population doubling time of TE13R120 was approximately 39.93h, while that of TE13 was 33.94h. 1.3 The plating efficiency (PE) for TE13R120 cells was 47.67±5.84%. PE for TE13 cells was 38.17±7.65%. For TE13R120 cells, D0 for radiation sensitivity was 2.31Gy while for TE13 cells D0 was 1.29Gy. The surviving fractions at 2 Gy (SF2) were 0.63 for TE13R120 cells and 0.50 for TE13 cells. SF2 of TE13R120 cells was greater than that of the parent cells, TE13. 1.4 Time-courses of cell cycle distributions after irradiation. The fractions of G0/G1, S and G2/M cells were 57.89±11.56%,23.97±6.31%,18.14±5.25% for TE13 respectively. Corresponding fractions were 71.77±6.54%,13.22±2.18%,15.12±4.54% for TE13R120 cells. The proliferation index (PI) for TE13R120 was smaller than that of TE13 cells (28.35±6.72% versus 42.11±11.56%). A marked accumulation of cells in the G1 phase was observed after irradiation with a dose of 4Gy for TE13 cells. The accumulation of cells in G1 phase reached its peak at 12h after irradiation. At 48h after irradiation, the cell cycle distribution for TE13 cell lines returned to that for the non-irradiated control cells. However, the apparent change of cell cycle distribution for TE13R120 cells was not observed after irradiation and only a slight accumulation of cells in G2/M phase was observed at 12h to 24h after irradiation.2. To examine the effect of a human monoclonal antibody to EGFR (hR3) combined with recombinant adenoviruses-p53 anticancer injection (SBN-1) on esophageal cancer cells with different radiosensitivity. 2.1 EGFR is overexpressed in TE13 cell lines and the radioresistant variant cell lines TE13R120 detected by immunohistochemistry. 2.2 hR3, SBN-1 and every experimental group all showed the antiproliferative effect in vitro growth profile of each esophageal cell line and they were significant (P<0.05). Inhibition rate of the simultaneous administration on TE13 cells and TE13R120 cells was 77.93±3.81% and 86.09±4.35% respectively, which w... |
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