The Expression Of BI-1 And The Mechanism Of Anti-apoptosis In Breast Cancer

Breast cancer is one of the most common malignant tumors in woman. The incidence of breast cancer has been on rise in recent 20 years,and it is seriously affecting woman's health. The regulation of programmed cell death, or apoptosis, is crucial for normal development and for the maintenance of homeostasis. Uncontrollable cellular growth resulting from the acceleration of mitosis and inhibition of cell death can promote progression of malignant tumor. Recent studies have shown that impairment of the apoptotic signalling pathway plays an important role in the initiation and progression of human breast cancers. Bax inhibitor-1 (BI-1) has been shown to represent a new type of regulator of cell death controlled by mitochondrium and endoplasmic reticulum apoptosis pathways. It was demonstrated that in mitochondrium apoptosis pathway, BI-1 can interact with Bcl-2 and Bcl-xl, but not Bax and Bad, and when over-expressed in mammalian cells, BI-1 suppressed apoptosis induced by Bax, etoposide, staurosponne and growth factor deprivation, but not by Fas (CD95) . More recently, BI-1 was shown to be over-expressed in most tumor such as prostate cancer and non-small cell lung cancer and so on.In this study we detected the expression of BI-1 in human breast cancer and their matched normal breast tissu by Fluorescence Quantative PCR,to analyse the role of BI-1 in the development of breast cancer,and we also detect the expression of ER,PR,Bcl-2,Bcl-xl and P53 in breast cancer by Immunohistochemistry. Analysing the correlation among the expression of BI-1 and ER,PR,Bcl-2,Bcl-xl ,P53 to explore the anti-apoptosis mechanism of BI-1. and provide more experimental evidence for prognosis, diagnosis and therapy of breast cancer.Methods1. Choicing patients and samplesThis study analyzed 42 case matched normal and primary cancer tissues from breast cancer patients exairesised at The First Affiliated Hospital of ZhengZhou University, from 2005 to 2006 after their informed consents. All the samples are stored in the liquid nitrogen and formalin within 5 minutes. We collected the patients clinical features such as age, tumor size , histology type and stage, and clinical grade.2.BI-1mRNA was detected in 42 case matched breast cancer and normal breast tissues by Fluorescence Quantative PCR.Totoal RNA was extracted from breast cancer specimens and FCM-7 cell lines by using TRIzol reagent (Life Technologies, Inc ) according to the standard protocal. The quality of the RNA samples was determined by electrophoresis through agarose gels and staining with ethidium bromide; Total RNA were progressed directly to cDNA by reverse transcription using AMV reverse transcriptase produced by Promega company. We also quantified, transcription ofβ-action to test the effect of reverse transcription;And the expression of BI-1 was detected by Fluorescence Quantative PCR in each specimen, at last we manufacture a standard curve from the cDNA of FCM-7 cell lines to control the detected result.3. BI-1 protein was detected in 42 matched breast cancer and normal breast tissues by western blot.We extracted the protein of kytoplasm in each specimen,and the concentration of this protein was determined by Bradford program,after the antigen were repaired, the protein was electrophoresised in polyacrylamide gel,then the protein was transfered to PVDF membrane,and the BI-1 andβ-actin were detected by special antibody,and after colorrating of the protein ,the gray scale was detect.4.ER,PR,Bcl-2,Bcl-xl and P53 protein was detected in the matched breast cancer by Immunohistochemical technique.5. SPSS11.0 was adopted to analyze the experimental data at thea=0.05 level.Results1. In 33 of 42 cases the expression of BI-1 in the breast cancer is higher than that in the normal tissues by Fluorescence Quantative PCR (P<0.05).2. There is no significant correlation between the expression of BI-1 and the age,tumor size, histology stage and clinical grade(P>0.05). but the expression of BI-1 in the group of lymph node metastasis is higher than that in the group of non-metastasis,and the difference has statistical significance (P<0.05).3. BI-1 protein was detected out in each specimen in different intension.4. The express of BI-1 mRNA and protien in breast cancer is shown to be coincident.5. The positive rate of ER,PR,Bcl-2 ,bal-xl and P53 is 59.52%, 54.76%, 61.90%, 59.52%, 47.62% respectively. Correlation analysis display:There is no correlation in the expression of BI-1 and ER or PR (P>0.05). Yet the expression of BI-1 is positive related to the Bcl-2,Bcl-xl and P53(P<0.05).6. There is no significant difference in the expression of Bcl-2 and Bcl-xl in breast cancer (P>0.05).Conclusion1. The expression of BI-1 in the breast cancer is higher than that in the normal tissues ,which indicates BI-1 can promote the development of breast cancer.2. BI-1 may take part in the developement of breast cancer in the early stage.3. BI-1 is related to the metastasis of breast cancer and contributed to lymph node metastasis.4. The united detection of BI-1 can not be used to judge the prognosis of breast cancer. 5. Both Bcl-2 and Bcl-xl take part in the mitochondrion apoptosis. BI-l,Bcl-2 and Bcl-xl will become the new targets for gene therapy of breast cancer.6. Maybe BI-1 and P53 has a relevance expression at the level of gene expression in the development of breast cancer.7. United detection of BI-l,Bd-2,Bcl-xl and P53 expression can predict the ability of invasion and metastasis of breast cancer and supply the evidence for clinical therapy and judgement of prognosis.