| Objective: To study the inhitory effects and its mechanisms of Oridonin on human lung adenocarcinoma SPC-A-1 cells.Methods: Inhibitory effect of cell proliferation on SPC-A-1 cell caused by Oridonin was determined by MTT assay. The morphological changes were observed by phase contrast microscope, Hoechst33258 fluorochrome staining and electron microscope. Cell cycle and apoptosis rate were analyzed by flow cytometry. Apoptosis was also detected by TUNEL. The activities of Caspase-3 and Caspase-9 in SPC-A-1 cells treated with Oridonin were detected by colorimetric assay kit. The mechanisms involved in the effects of Oridonin on SPC-A-1 cells were studied by Real-time RT-PCR and Western blot.Results: MTT(3-(4,5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide,MTT) assay showed that Oridonin had anti-proliferation effect on SPC-A-1 cells in a time-dependant and dose-dependant manner. Morphologically apoptotic changes about chromatic aggregation and nuclear condensation were surveyed by phase contrast microscope, Hoechst 33258 fluorochrome staining and electron microscope. The results of FCM represented the apoptosis rates of SPC-A-1 cells treated with Oridonin were 3.2%(20μmo/L), 12.0%(30μmo/L), 34.1%(40μmo/L)and 4.1%(12h),11.4%(24h),34.1%(48h) respectively, and revealed that the cell cycle were arrested in G2/M phase. Oridonin demonstrated a moderate apoptosis and cell arrest effects on SPC-A-1 cells in a time-dependant and dose-dependant manner. TUNEL analysis showed that Oridonin had apoptosis effect on SPC-A-1 cells. The activity of Caspase-3 and Caspase-9 were increased also. The mRNA levels of p53, p21, bax were increased, meanwhile bcl-2 and c-Myc at mRNA level were reduced. The protein of P53, P21, Bax were up-regulated, and Bcl-2 was down-regulated. Conclusion: Oridonin shows obvious inhibitory effects on SPC-A-1 cells. The effects may be related to induce apoptosis via mitochondrial pathways and arrest cell cycle via up regulating P53 and P21, down regulating c-Myc. |
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