Study On Anti - Breast Cancer Of "Fritillaria Thunbergii - Fritillaria

BackgroundBreast cancer is now one of the most common cancers for women around the world. It has high morbidity and mortality, which hazards women’s physical and mental health seriously. Surgery, radiotherapy, chemotherapy, endocrine therapy and molecular target therapies comprehensively were given priority to breast cancer treatment, having cured or alleviated most of the patients. But there are still parts of the patients having been found in the advanced stage, loss of surgery opportunity, strong side effects of chemo-radiotherapy, varying degrees of drug resistance, recurrence or metastasis, etc. Such problems impact on life quality and survival period of those patients. More and more scholars have focused attention on the traditional Chinese medicine, have carried out a large number of clinical observations and basic researches, and make a great progress. The advantage of traditional Chinese medicine therapies to enhance body immunity, improve effect and decrease toxicity of chemo-radiotherapy, reverse the drug-resistant, perfect survival quality, lower the risks of recurrence and metastasis, extend the lifetime, etc. Bolbostemma Paniculatum and Peimine are used commonly in traditional Chinese medicine therapy for breast cancer, especially in long-term maintenance course after surgery, chemotherapy or radiotherapy. The modern pharmacology research confirms anticancer effect of Bolbostemma Paniculatum, reversal of multidrug-resistance and increased sensitivity effects of Peimine. However, the cooperation effect of Bolbostemma Paniculatum and Peimine for the treatment of breast cancer seldom reported. Professor Hu Kaiwen prefers to apply Bolbostemma Paniculatumand Peimine to treat breast cancer patients after surgery, which effectiveness is still lack of systematic reviews.ObjectiveIn Vitro and In Vivo experiments, to study the cooperation effect of Bolbostemma Paniculatum and Peimine for the treatment of breast cancer by clinical and basic research, which supply experimental evidence for compatible application in traditional Chinese medicine.MethodsDesign in vitro and in vivo experiments:1. In vitro:By using MTT, observe the cooperation effect of Bolbostemma Paniculatumand Peimine for the treatment of four breast cancer cells. By transfection and filtering, build four green and red fluorescent protein transferred human breast cancer models:MDA-MB-231 DUAL, BT-549 DUAL, MCF-7 DUAL, and MCF-7/ADR-RES DUAL. Using inverted real-time fluorescence microscopy imaging, observe morphological change and death modes of the cell models on the cooperation effect of Bolbostemma Paniculatum and Peimine.2. In vivo:Using orthotopic transplantation method, build green fluorescent protein markers in human breast cancer model. Through the comparison of the negative control group(CON), Bolbostemma Paniculatum group(T20), low dose of Bolbostemma Paniculatum and Peimine (T20+Z15), high dose of Bolbostemma Paniculatum and Peimine (T50+Z20) of the tumor inhibition rate, evaluate the effects on tumor growth,by fluorescence microscopy(the FluorVivo Model 100). In the end, apply the flow cytometry on tumor tissue for detection of apoptosis and cell cycle, to research the mechanism of anti-breast cancer effect of the two medicines.Results1.In vitro:The Bolbostemma Paniculatum attract has growth-inhibitory for the four human breast cancer cell models. At the same concentrations of Bolbostemma Paniculatum attract, combined application of Peimine can improve the growth-inhibitory rate of the other three human breast cancer cells (P <0.05), except MDA-MB-231 cell. However, Peimine alone has no significant impact on four human breast cancer cell models.2. The Bolbostemma Paniculatum attract has stronger inhibition of triple negative breast cancer BT-549,MDA-MB-231 than that of MCF-7, and MCF-7/ADR-RES. Peimine alone has no significant impact on four human breast cancer cell models. Combined application of Peimine can improve the growth-inhibitory rate of triple negative breast cancer BT-549 and receptor positive breast cancer MCF-7, and can reverse of multidrug-resistance breast cancer MCF-7/ADR-RES. But they are no effect of triple negative and high metastasis breast cancer MDA-MB-231.3. In morphological observation, four green and red fluorescent protein transferred human breast cancer models:MDA-MB-231 DUAL, BT-549 DUAL, MCF-7 DUAL, and MCF-7/ADR-RES DUAL were built successfully. There were both apoptosis and oncosis after the treatment of Bolbostemma Paniculatum and Peimine.4. In vivo:Green fluorescent protein markers in human breast cancer MDA-MB-231 GFP model were built. The experimental results show that tumor size and weight in the three treatment groups(T20,T20+Z15,T50+Z20) were decreased comparing the control group, but the changes were no statistically significant difference (P>0.05). There were lung and lymph metastasis in the four group, and no statistically significant difference (P>0.05). Flow cytometry results indicate that apoptosis in the early and late stage in T50+Z20 and T20+Z15 is more than other that of other groups significantly (P<0.05). The cell cycle were no significant difference in the four groups (P>0.05)5. At the end of experiment, the overall weight of mice were increased, with good general condition. Anatomy of the important organ, there were no significant bleeding, degeneration, necrosis and other symptoms. But before and after treatment T20+Z15 increases mice body weight were significantly lower than the Con group. Combination of Bolbostemma Paniculatum and Peimine has a certain toxicity.Conclusion1. In vitro, Bolbostemma Paniculatum and Peimine have the cooperation effect for breast cancer, and Peimine can improve the effect of Bolbostemma Paniculatum. The modes of inducing cell death are apoptosis and oncosis.2. Bolbostemma Paniculatum and Peimine are different sensitivity to different biological characteristics of breast cancer cells. Highly metastatic and triple-negative cell line MDA-MB-231 is of the lowest sensitivity, and multidrug resistance cell line MCF-7/ADR-RES is of the best effect.3. There was possibility of anti-breast cancer growth in Green Fluorescent Protein Gene Transferred Mouse Model of Human Breast Cancer MDA-MB-231 GFP, which has a certain toxicity in vivo studies.