Expression Of TRAF1 And Its Relationship With TRAF2 In The Different Metastasis Breast Cancer Cell Lines

ObjectiveTRAFs (Tumor necrosis factor receptor-associated factors), is an important intracellular connection protein. It can participate in many signal transducers pathways to active genes such as NF-κB and JNK. It can regulate reproduction, survival, apoptosis of normal cells and tumor cells and it can lead to sprouting of endothelial cells. Seven TRAFs members have already been found. TRAF1 is the most special. Several model systems have been used to examine TRAF1 function and the results obtained have led to a variety of often contradictory conclusions about whether TRAF1 is a positive or negative regulator of TNFR signaling pathways and in which pathway it is important. TRAF1 can bind receptors either directly or by heterotrimerizing with other TRAFs. Among all TRAFs members the relationship between TRAF1 and TRAF2 is the most closed. Up to now studies on the expression and function of TRAF1 in breast cancer cell lines are rare.This study investigated the expression of TRAF1 in the breast cancer cell lines through immunohistochemistry staining and western blot. Co-immunoprecipitation is used to study the relationship of TRAF1 and TRAF2.Materials and Methods1. MaterialsThe normal human breast cell line MCF-10A, the low metastasis human breast cancer cell line MCF-7, the moderate metastasis human breast cancer cell line MDA-MB-231 and the high metastasis human breast cancer cell line MDA-MB-435s.2. Methods(1) The expressions of TRAF1 in the above-mentioned cultured cell lines were examined through immunohistochemistry staining.(2) The expressions of TRAF1 in the above-mentioned cultured cell lines were also examined with Western blot.(3) The relationships between TRAF1 and TRAF2 with co-immunoprecipitation in the above-mentioned cultured cell lines except MDA-MB-231 were also examined(4) A statistical software Statistical Product and Services Solutions (SPSS) (version 12.0) was utilized for data analysis. Analysis of variance was done to compare the expressions of TRAF1 in different cell lines. A P-value less than 0.05 was considered valid in terms of statictics and Values less than 0.01 were considered predominantly significant.Results1. Expressions of TRAF1 through immunohistochemistry staining innormal breast and breast cancer cell lines.(1) It's expression in MCF-10A is lower than those in MCF-7 (P<0.05), MDA-MB-231 (P<0.01),and MDA-MB-435s (P<0.01).(2) It's expression in MCF-7 is lower than those in MDA-MB-231 (P<0.01), ,and MDA-MB-435s (P<0.01). It's expression in MDA-MB-231 is lower than those in MDA-MB-435s (P<0.05).As the metastasis of breast cancer cell lines increase, the expression of TRAF1 would be stronger.2. Expression of TRAF1 through western blot staining in normalbreast and breast cancer cell lines.(1) It's expression in MCF-10A is lower than those in MCF-7 (P<0.05), MDA-MB-231 (P<0.01), and MDA-MB-435s (P<0.01).(2) It's expression in MCF-7 is lower than those in MDA-MB-231 (P<0.05), and MDA-MB-435s (P<0.01). It's expression in MDA-MB-231 is lower than those in MDA-MB-435s (P<0.05).As the metastasis of breast cancer cell lines increase, the expression of TRAF1 would be stronger.3. Relationships between TRAF1 and TRAF2 in normal breast andbreast cancer cell lines.(1) It is found through co-immunoprecipitation that TRAF1 and TRAF2 combined in normal human breast cell line MCF-10A, human breast cancer cell line MCF-7 and MDA-MB-435s.(2) The quantity of TRAF1 in combination with TRAF2 is in MCF-10A is larger than the quantities in MCF-7 (P<0.05) and MDA-MB-435s (P<0.01), and its quantity in MCF-7 is larger in that in MDA-MB-435s (P<0.05).As the metastasis of breast cancer cell lines increases, the quantity of TRAF1 in combination with TRAF2 would be smaller.Conclusions1. The expression of TRAF1 in breast cancer cell lines are stronger than that in normal human breast cell line. As the metastasis of breast cancer cell lines increase, the expression of TRAF1 would be stronger.2. TRAF1 would combine with TRAF2 in both the normal human breast cell line and different types of human breast cancer cell lines.3. The quantity of TRAF1 in combination with TRAF2 in normal breast cell line was larger than those in breast cancer cell lines. As the metastasis of breast cancer cell lines increases, the quantity would be smaller.It suggests that TRAF1 is a negative regulator of TRAF2 signaling pathways through the combination with TRAF2.