Silencing Of THBS1 Gene Inhibits The Biological Function Of Breast Cancer Cells

Background:Breast cancer is one of the most common malignant tumors that endanger women’s health,and its incidence is increasing year by year globally.In clinical treatment of breast cancer,surgical resection and postoperative chemotherapy are mainly used.However,due to the characteristics of strong invasiveness and high recurrence rate,the fatality rate of breast cancer ranks first among female malignant tumors.Platelet thrombin protein-1(thrombospondin-1,THBS1)is a multifunctional glycoprotein synthesized and secreted by a variety of cells.Abnormal expression in tumors has carcinogenic or tumor suppressor functions.It plays a role in the development of tumors.The complex regulation and its role in the occurrence and development of breast cancer is not yet clear.Objective:1.To analyze the expression level of THBS1 m RNA in breast cancer and its correlation with clinicopathological characteristics such as age,gender,TNM staging,stage and survival.2.To explore the effect of THBS1 gene silencing mediated by small interfering RNA(si RNA)on the proliferation,invasion and apoptosis of human breast cancer cells MCF-7 and MDA-MB-231.Methods:1.Bioinformatics analysis: Use TCGA database and GEPIA database to analyze the expression of THBS1 m RNA in breast cancer,and analyze whether the expression of THBS1 is correlated with age,gender,TNM staging and other clinicopathological characteristics.2.To study the effect of si RNA-mediated THBS1 gene silencing on the biological behavior of breast cancer cells MCF-7 and MDA-MB-231:(1)Three different sequences were specifically targeted at THBS1 gene through liposome transfection method Si RNA(si RNA-THBS1 for short)was transfected into human breast cancer MCF-7 and MDA-MB-231 cells.(2)Three different sequences of si RNA-THBS1 versus THBS1 were detected by real-time fluorescent quantitative PCR(q RT-PCR)The sequence with the best silencing effect was screened out.Subsequent experiments were all used to transfect the sequence for THBS1 gene silencing.(3)Cell Counting Kit-8(CCK-8)proliferation experiment to detect THBS1 gene silencing on breast cancer cells The effect of proliferation.(4)Transwell cell matrix invasion test to detect the effect of THBS1 gene silencing on breast cancer cell invasion.(5)Flow cytometry technique Annexin V-FITC/PI double staining method to detect THBS1 gene silencing on breast cancer The effect of apoptosis.All experiments were repeated three times,and the experimental results were statistically analyzed using SPSS23.0 statistical software,P <0.05 considered the difference to be statistically significant.Results :1.Bioinformatics analysis: The expression of THBS1 in breast cancer tissues was higher than that in normal tissues(P<0.05);in breast cancer,the expression of THBS1 was related to age,gender,stage,T(the size),and M(Distant metastasis)are not related,(P >0.05);It is related to N(lymph node metastasis),(P <0.05).2.Cell experiment:(1)q RT-PCR: Three sequences of si RNA-THBS1 can silence THBS1,among which sequence-1(si RNA-THBS1-1)has the highest silencing efficiency.This sequence of si RNA can affect MDA-MB-231 cells The silencing efficiency of MCF-7 cells reached 62.9±2.9%,and the silencing efficiency of MCF-7 cells reached 94.98±0.37%.(2)CCK-8 cell proliferation experiment: After silencing the THBS1 gene,the proliferation ability of MCF-7 cells was better The transfection reagent group,the blank control group,and the si RNA-NC group were significantly reduced(P<0.05).The proliferation ability of MDA-MB-231 cells was not significantly different from other controls(P>0.05).(3)Transwell cell matrix Invasion experiment: After counting by hematoxylin staining,it was found that the number of invaded cells in the si RNA-THBS1 group was less than that in the si RNA-NC group(P<0.05),and the difference was statistically significant.(4)Flow cytometry technology to detect cell apoptosis: The apoptosis rate of the two kinds of cells in the si RNA-THBS1 group was higher than that in the si RNA-NC group(P<0.05),and the difference was statistically significant.Conclusion : 1.The expression of THBS1 in breast cancer is higher than that in normal tissues,and the expression of THBS1 is related to lymph node metastasis.2.THBS1 gene promotes the proliferation of breast cancer cells MCF-7 cells,promotes the invasion of MDA-MB-231 and MCF-7cells,and inhibits the apoptosis of both cells.