Expression Of CXCL16 In Breast Cancer And Activation Of NF-kappa B Signaling Pathway In Regulation Of Breast Cancer

Objective(s):In this study,the expression levels of chemokine ligand 16(CXCL16)mRNA in carcinoma and adjacent tissues were detected in human breast cancer clinical samples and human breast cancer cell lines,and its correlation between the expression level and the clinical characteristics of invasive breast cancer patients was analyzed.To detect the expression level of CXCL16 mRNA and protein in triple negative breast cancer cells(MDA-MB-231)and estrogen receptor positive breast cancer cells(MCF-7).The effect of CXCL16 on the proliferation,migration and invasion ability of breast cancer cells,and CXCL16 regulates the expression levels of downstream gene u-Pa(Uridylyl Phosphate Adenosine),MMP9(Matrix Metallo Proteinase-9)and VEGF(Vascular endothelial growth factor).The expression levels of p65,phosphorylated p65,I?Ba,phosphorylated I?Ba mRNA and protein in this pathway.Furthermore,to explore the mechanism of CXCL16 regulating the proliferation,migration and invasion of breast cancer cells is of great significance for finding new diagnostic markers and therapeutic targets of breast cancer.Methods:1.Collect 43 cases of breast cancer diagnosed as invasive breast cancer at the First Affiliated Hospital of Kunming Medical University and Chenggong Hospital of Kunming Medical University from March to August 2019.The expression level of CXCL16 mRNA in breast cancer and adjacent tissues was detected by real-time fluorescent quantitative reverse transcription polymerase chain reaction(qRT-PCR).The correlation between the expression level of CXCL16 mRNA in breast cancer and the clinical characteristics of axillary lymph node metastasis,TNM stage,pathological molecular typing and postoperative recurrence and metastasis risk was analyzed.2.Cultivate MDA-MB-231 and MCF-7 cells.The expression of CXCL16 mRNA and protein in different invasive breast cancer cell lines was detected by qRT-PCR and western blot test method.And the correlation between CXCL16 and different invasive breast cancer cells was analyzed and compared.3.MDA-MB-231 and MCF-7 cells overexpressed CXCL6 plasmids by instantaneous transfection,use qRT-PCR and western blot test method to detect the transfection of MDA-MB-231 and MCF-7 cells overexpressing CXCL16.4.Using the CCK8 cell proliferation test method to detect the proliferation of MDA-MB-231 and MCF-7 cells overexpressing CXCL16.5.Detect the migration and invasion ability of MAD-MB-231 and MCF-7 cells overexpressing CXCL16 by scratch and transwell test method.6.Detect the expression level of u-PA,MMP9 and VEGF mRNA by qRT-PCR test method,and detect the expression level of t-p65,p-p65,t-I?B?,p-I?B? by western blot test method in NF-?B signaling pathway downstream MDA-MB-231 and MCF-7 cells overexpressing CXCL16.Results:1.CXCL16 mRNA is expressed in both breast cancer tissues and adjacent tissues,the expression in cancer tissues is significantly higher than that in adjacent tissues.(P<0.001,the difference was statistically significant)2.The expression level of CXCL16 mRNA in cancer tissues is correlated with the axillary lymph node metastasis,TNM stage,pathological molecular classification and postoperative risk of recurrence and metastasis.And the expression is higher in invasive breast cancer with axillary lymph node metastasis,higher TNM stage,and higher risk of postoperative recurrence and metastasis.(P<0.05,the difference was statistically significant)3.CXCL16 mRNA and protein are expressed in both MDA-MB-231 and MCF-7,and in MDA-MB-231 cells are expressed higher than MCF-7 cells.(P<0.05,the difference was statistically significant)4.MDA-MB-231 and MCF-7 cells overexpressing CXCL16 have enhanced proliferation,migration and invasion ability.(P<0.05,the difference was statistically significant)5.The expression of u-PA,MMP9,VEGF mRNA and the related molecules t-p65,p-p65,t-I?B?,p-I?B? in MDA-MB-231 and MCF-7 cells overexpressing CXCL16 were up-regulated.Under the action of NF-?B pathway inhibitors,the expression of these genes and molecules were down regulated.(P<0.05,the difference was statistically significant)Conclusion(s):1.The expression level of CXCL16 mRNA in breast cancer is higher than that in adjacent tissues,which suggests that CXCL16 may play a role in promoting breast cancer development.2.The expression of CXCL16 mRNA was correlated with axillary lymph node metastasis,TNM stage,pathological molecular typing and postoperative recurrence and metastasis risk.The higher the expression of CXCL16 mRNA in invasive breast cancer with axillary lymph node metastasis,higher TNM stage,worse pathological molecular typing and higher risk of recurrence and metastasis after operation,suggesting that the expression level of CXCL16 mRNA is positively correlated with the malignant behavior of invasive breast cancer.3.The expression level of CXCL16 mRNA and protein in high invasive breast cancer cells(MDA-MB-231)was higher than that in low invasive breast cancer cells(MCF-7),suggesting that the expression level of CXCL16 mRNA was positively correlated with the invasion of breast cancer.4.The proliferation,migration and invasion ability of breast cancer cells overexpressing CXCL16 are increased,suggesting that CXCL16 may have the role of promoting breast cancer cell proliferation,migration and invasion.5.In overexpressing CXCL16 breast cancer cells,the downstream genes of the NF-?B pathway u-PA,MMP9 and VEGF mRNA and the related molecules t-p65,p-p65,t-I?B? and p-I?B? protein expression were up-regulated.When using the NF-?B pathway inhibitor(BAY 11-7082),the expression of its downstream genes and related molecules were down-regulated.It is suggested that CXCL16 may regulate downstream genes and relted molecules by activating NF-?B signaling pathway,thereby promoting the development of breast cancer.