Disruption of cytoskeleton does not affect Bax translocation and apoptosis on mammary epithelial cells

Introduction
　The regulation of cell number is as important as cell division and cell migration for development and normal tissue homeostasis. Apoptosis is a mechanism of cell death required for this regulation. Bcl-2 family proteins have important function in apoptotic pathway. Bax is one of proapoptotic members of Bcl-2 family proteins. Activated Bax translocates to mitochondria, whereas inactive Bax resides in cytosol.
　The physical contact of cells with extracellular matrix (ECM) is essential for survival of epithelial cells. Detachment of ECM from cell could cause anoikis, a specific form of apoptosis which has same characteristics observed in normal apoptosis, such as DNA fragmentation and change of mitochondrial membrane potential caused by Bcl-2 family proteins. Bax translocation occurs before commitment to anoikis.

Materials & Methods
　In this study, two types of established cells were used. One is FSK-7 cells which are mammary epithelial cells of mice and the other is human mammary epithelial cells, MCF-10A. These cells were treated with drugs which disrupt or stabilize cytoskeleton containing tubulin and actin filament. These drug-treated cells were stained by immunoflourescense to observe Bax translocation and DNA fragmentation. The processing status of caspase 3, which is commonly cleaved and activated during apoptosis pathway, was detected by immunoblotting.

Results
　Immunofluorescent observation revealed that disruption of cytoskeleton did not induce either Bax translocation or DNA fragmentation in both types of cells. Immunoblot analysis also indicated that there is no physical interaction between apoptosis and disruption or stabilization of cytoskeleton.

Future work
　To understand the molecular mechanism of anoikis, further examination is necessary to clarify whether cytoskeleton remains in detached cells or not. If cytoskeleton is remained in a detached cell, Anoikis could be caused by disconnection of cell to ECR, not by morphological change, such as in cytockeleton. And it is also needed to figure out how focal adhesion complexes look like in cytoskeleton disrupted cells.