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Background cervical cancer (cervical cancer) is a malignant tumor that occurs in cervical intraepithelial, the world's second only to breast cancer in a serious threat to women's health and lives of the malignant tumor. The incidence rate of the first female malignancy in our According to the World Health Organization estimates, 131,000 new cases each year, accounting for 28.8% of the new cases of cervical cancer world. Women of all ages are likely to suffer from cervical cancer, however, the highest incidence and mortality of the population is concentrated in the relatively young age of 45 to 50-year-old women. In recent years, a clear upward trend in the young patients with cervical cancer, and to preserve the reproductive function and improve the quality of life is imperative. Cervical cancer occur in the low socio-economic status of women, and sexual health, HPV infection, early marriage, smoking may. The incidence of cervical cancer in developing countries is six times that of developed countries, and yet universal early screening of cervical cancer, 80% of patients at the time of diagnosis has been developed to invasive cancer. For early cervical cancer treated by surgery or radiation therapy, the use of radiation therapy combined with systemic chemotherapy treatment in late recurrence, metastatic cervical cancer patients. The clinical findings between different patients, radiotherapy and chemotherapy sensitivity differences lead to the therapeutic effect is also very different. Improve put the chemotherapy dose efficacy can be improved to a certain extent, but tolerance of different patients, and side effects increased significantly, severely reducing the quality of life of the patients. Bcl-2 is an important anti-apoptotic protein Bcl-2 family of apoptosis-related proteins in cervical cancer and other tumors with high expression of this high expression status and tumor occurrence, development, recurrence and radiotherapy and chemotherapy produce resistance is closely related. Located on the outer mitochondrial membrane Bcl-2 conformational change occurs in the early stages of apoptosis, leading to PTP channel open, thereby enabling increased mitochondrial membrane permeability, such as cytochrome C release of apoptotic protein activating factor-1 (Apaf-1) as well as apoptosis-inducing factor (AIF) and other active proteins promote apoptosis. Cytochrome C and Apaf-l, Caspases-9 composed apoptosis start complex Caspases-9 is activated, further activation of Caspase-3, ultimately leading to apoptosis. Gossypol (gossypol) is a yellow the polyphenol hydroxy bis naphthyl aldehyde compounds, there are two optical isomers, respectively L gossypol [(-)-gossypol] dextrose gossypol [()-gossypol] Currently Gossypol the preparations are mainly three kinds: gossypol acetic acid, refined of gossypol and formic gossypol. Gossypol was initially used clinically as a male contraceptive, Recent studies found that gossypol has a significant anti-tumor capacity, capable of a variety of tumor cell lines, such as prostate cancer, lymphoma and colon inhibit tumor cell proliferation and induced apoptosis. Gossypol available through a variety of ways and molecular targeting of tumor cells inhibit proliferation and (or) induction of apoptosis, including through direct mitochondrial inhibition of cell energy metabolism, reduce the expression of cell cycle regulatory proteins of Rb and cyclinDl protein , inhibition of protein kinase C activity. Large amounts of data that mitochondria induced apoptotic by gossypol role in its variety of inducing apoptosis pathway is one the most important cell death pathway. Mitochondrial-dependent apoptosis pathway is the expression of Bcl-2 down, causing cytochrome C release from the inner mitochondrial membrane start, combined with Caspase-9, Caspase-3 of Caspase-7 activation, and thus the regulation of apoptosis. Further studies confirm that gossypol is a small molecule inhibitor of anti-apoptotic proteins Bcl-2/Bcl-X L , capable of specifically binding Bcl-2/Bcl-X L BH3 locus (Bcl-2 homologydomin3), to prevent the form heterodimers with pro-apoptotic protein Bax inhibiting anti-apoptotic Bcl-2/Bcl-X L function, play a role in induction of tumor cell apoptosis. Research purposes past at home and abroad about the role of gossypol in the cervical, not yet reported, the present study intends to high expression of Bcl-2 protein in cervical carcinoma Hela study, study of gossypol acetate inhibition of proliferation and apoptosis-inducing role, and explore its possible mechanism of action, combined with X-ray study comparative gossypol acetate on human cervical cancer cell lines in vitro killing effect, provided the experimental basis for further research and clinical applications of gossypol, the theoretical basis. Experimental methods using the MTT assay of gossypol acetic acid inhibition studies in vitro proliferation of cervical cancer cell line Hela Compare gossypol acetate combined X-ray of a human cervical cancer cell lines in vitro killing effect. 2. The Hela nuclear morphology changes in the role of gossypol acetate the use of Hoechst33342/PI staining. Flow cytometry analysis of the role of gossypol acetate Hela cell apoptosis rate. Flow cytometry analysis of gossypol acetate on apoptosis related protein Bcl-2, Bax expression. 5. Colorimetric studies of gossypol acetate on Caspase-3, Caspase-8, Caspase-9 activity impact. The experimental results of gossypol acetate on Hela cells growth and proliferation inhibition, and this effect was dose-dependent effect. 10μmol / L gossypol acetic acid for 24h can significantly inhibit the growth of Hela cells 1μmol / L gossypol acetate had no significant effect on cell growth. 2. The radiotherapy group gossypol acetate the joint-ray treatment group ray dose Hela cell growth inhibition rate (7.97 ± 1.43,15.83 ± 1.16,27.96 ± 1.25)%, (42.71 ± 2.25,61.36 ± 2.33,77.45 ± 3.05 )%. The results show that the gossypol acetate the joint-ray treatment group Hela cell growth inhibition rate was significantly higher than the radiotherapy group. 3.Hoechst33342/PI staining fluorescence microscope under visible by 20μmol / L gossypol acetate after 24h was dense stain apoptotic nuclei issue a strong blue fluorescence, while the control group presented a diffuse uniform light blue normal nucleus. Flow cytometry can detect apoptotic cells in the cell cycle G 0 / G 1 before DNA hypodiploid peak, peak apoptosis. This experiment, 20 μmol / L gossypol acetate role 24h after DNA histogram hypodiploid peak, of gossypol acetate apoptosis rate of the control group, a significant difference was statistically significant (t = -19.840 P = 0.000). 5.20μmol / L gossypol acetate role after 24h, Bcl-2 protein levels decreased in the control group (78.17 ± 3.23)% to (68.97 ± 1.94)%, a significant difference was statistically significant (t = 4.239, P = 0.013 ); Bax protein expression levels increased to the control group (8.16 ± 0.67)%, (18.88 ± 1.36)%, a significant difference was statistically significant (t = -12.254, P = 0.000); the Bcl-2/Bax ratio control group, a significant difference was statistically significant (t = 17.734, P = 0.000). 6.20μmol / L gossypol acetic acid role, has appeared Caspase-3, caspase-8 expression of Caspase-9 protease activity. 12h, 24h, 48h time point Hela cells of Caspase-3 (2.32 ± 0.15,5.44 ± 0.20,4.34 ± 0.04), caspase-8 (3.70 ± 0.15,5.45 ± 0.20,2.36 ± 0.16), Caspase-9 (4.36 ± 0.33 6.40 ± 0.19,2.84 ± 0.08), the degree of activation gradually increased, peaked at 24h, followed by the activity decreased. The main conclusions of this study confirmed that gossypol acetate on cervical carcinoma Hela inhibition of proliferation and induction of apoptosis and its mechanism of action may be related to downregulation of Bcl-2, Bax protein expression rates, and Caspase-3, Caspase-8, Caspase -9 protease activation. Gossypol acetic acid-induced apoptosis of Hela cells is completed by the mitochondrial pathway and death receptor pathway. Gossypol acetate combined with X-rays can be enhanced X-rays to inhibit the proliferation of cervical carcinoma Hela synergies between.
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