The Study For The Roles Of CPLA 2α In Breast Cancer Cells Chemotaxis And Invasion

Objectives:To investigate the role of cPLA2αin breast cancer cell invasion and illustrate the mechanisms of cPLA2αsignaling pathway in breast cancer cell invasion.Methods:Breast cancer cells were transfected with small RNA interference plasmids to disrupt cPLA2αexpression. cPLA2αprotein levels and mRNA levels were detected by Western blotting and RT-PCR. Scratch assay and chemotaxis assay were examined to detect the migration and chemotaxis ability of breast cancer cells. The in vitro invasion ability of breast cancer cells was examined using matrigel invasion assay. F-actin content of breast cancer cells was analyzed with F-actin polymirazation assay. Western Blotting was used to check the level of phosphorylated cofilin, integrinβ1 and FAK, and check the rescue expression level of phosphorylated FAK by adding arachidonic acid to sicPLA2α/MDA231 cell, and meanwhile detect the migration and chemotaxis ability of the cells. The sicPLA2α/MDA231 cell was injected or hypodermic inoculated into SCID rat in orde to examine the in vivo invasion ability of breast cancer cells.Results:Western blotting results showed significant decrease in cPLA2αprotein levels, consistent with a decrease in mRNA levels. The cPLA2α-reduced MDA231 cells showed decreased chemotaxis ability compared with the control cells (P<0.01). When a scratch was created in the fluent monolayer cells, it took the cPLA2α-reduced MDA231 cells a longer time to fill the gap (P<0.01). Adhesion assay showed the number of the adhering cells was decreased for reduction of cPLA2αwithin 5 and 15 minutes (P<0.01). The invasion assay showed prominent differences between the cPLA2α-reduced MDA231 cells and the control cells (P<0.01). In the cPLA2α-reduced MDA231 cells, the actin polymerization in response to the EGF stimulation was significantly reduced. Western blotting results showed that the level of phosphorylated integrinβ1 and phosphorylated FAK was inhibited for the cPLA2αreduction. The phosphorylated cofilin and Akt expression in cPLA2α-reduced MDA231 cells was at a level comparable to control cells. The invasion and metastasis assay of human breast cancer cells in the SCID rat showed that the number of tumors in the surface of lung has been clearly decreased in the cPLA2α-impaired MDA-MB-231 cells group (p<0.05).Conclusions:(1) The expression of cPLA2αin MDA-MB-231 cell was associated with tumor metastasis.(2) cPLA2a participated in EGF-induced chemotaxis and invasion of human breast cancer cells.(3) cPLA2a modulate the EGF-induced expression of phosphorylated FAK,which regulated chemotaxis and actin polymerization.(3) cPLA2a regulates cell adhesion signal pathway directly through phosphorylation of integrinβ1.(4) Arachidonic acid and its metabolite were associated with chemotaxis and migration of breast cancer cells. (5) cPLA2a plays an important role in invasion and metastasis of human breast cancer cells in vivo.