ABSTRACT
Objective To elucidate the mechanism of radiation resistant effect of LyGDI on NSCLC A549 cells.Methods A549 and H460 cells were irradiated with X-rays of 0,2,4 and 6 Gy.The clone-forming assay was used to detect cell survival and radiosensitivity.The expressions of LyGDI and COX-2 (Cyclooxygenase-2),a key radiosensitivity-related protein,were detected using Western blot.The miR-34 families were analyzed with RT-PCR.50 nmol/L mature miR-34c was transfected into A549 cells.Results The expression levels of LyGDI and COX-2 were much higher in radioresistive A549 cells than that in H460 cells.While the expression of miR-34a was quite low and miR-34b/c was hardly found in both NSCLC cells.Transfection of miR-34c into A549 cells strongly enhanced X-ray induced apoptosis by inhibiting the activations of LyGDI,COX-2,Bcl-2 and p21.Conclusions Up-regulation of LyGDI could induce COX-2 expression.The low expression of miR-34 family might be responsible for the radiation resistance of NSCLC cells.
ABSTRACT
Objective To elucidate the function and regulatory mechanism of LyGDI involved delayed cell death in the human K562 cells and HL-60 cells induced by 60Co γ-rays. Methods Erythrosine B cells staining was used to count the apoptosis rate. PI staining and flow cytometry were applied to check the cell cycle. The expression of LYGDI and Rac1 was resolved by Western blot by using monoclonal antibody of LyGDI and Racl. The distribution of Racl protein in cells was observed with immunofluorescence by using the confocal microscope. Results The K562 cells showed G2/M phase arrest and the percent age was 71.3%. The apoptosis rate was very low at early post-irradiation stage in the K562 cells. The apoptosis rate was 14% in the K562 cells at 24 h post-irradiation with 8 Gy of γ-rays, and delayed cell apoptosis was present. LyGDI was cleaved in the K562 cells irradiated by 4 Gy 60 Co γ-rays after 24 hours post-irradiation. The expression of Racl protein was not altered at all, but the distribution was changed in the irradiated cells while the Racl protein moved to cell membrane and a little in cell nucleus. The Racl was activated with the losing the binding affinity with the LyGDI. Conclusion LyGDI could promote the delayed cell apoptosis, which is through the activation of the Rac1.