Breast cancer (BR CA) is a major health concern for American women. The lifetime risk for developing BR CA is 1 in 8, and risks for developing and dying from BR CA increase with age. Metastatic disease kills those with BR CA. Blood-borne BR CA cells interact with endothelial cells when exiting the vasculature (extravasation), an important event in metastasis. Nothing is known about BR CA cell interactions with aged endothelial cells in vivo or in vitro. Results from our preliminary studies suggest that the addition of BR CA cells to monolayers of high passage """"""""old"""""""" endothelial cells (sequentially subcultured to passages 30 and above) causes large persistent gaps in the monolayer and endothelial cell death. In contrast, BR CA cell addition to low passage """"""""young"""""""" endothelial cells (passages I0 to 16) causes transient endothelial gaps (gaps form, then close) and no cell death. Since these differences may have relevance to metastasis and age-related increases in BR CA mortality, the Dumose of this project is to elucidate the effects of BR CA cell-induced injury on in vitro aged endothelial cells.
The specific aims are: 1. To determine if BR CA cell in addition to """"""""old"""""""" endothelial cells bases 30 to 36 causes endothelial cell apoptosis (programmed cell death). Assays to detect apoptosis include annexin-V-binding, caspase-3 activation and DNA fragmentation. The main hypotheses to be tested are that: 1, MCF-7 BR CA cell addition to """"""""old,"""""""" but not to """"""""young,"""""""" endothelial cells causes apoptosis; and 2. Addition I 2. of non-malignant mammary epithelial cells to """"""""old"""""""" endothelial cells does not induce apoptosis To determine if more ER CA cells transmigrate """"""""old"""""""" endothelial cells compared to """"""""young"""""""" endothelial ceils. A co-culture model will be used. """"""""Young"""""""" and """"""""old"""""""" endothelial cells will be grown on MatrigellM -coated, porous filters in transwell chambers, then labeled BR CA cells will added to the top chamber. BR CA cell transmigration will be measured by determining the numbers of BR CA cells that enter the bottom chamber. The hypothesis to be tested is that more MCF-7 BR CA cells will transmigrate """"""""old"""""""" endothelial cells, than """"""""young"""""""" endothelial cells. The results of these studies should provide new insight into metastasis and age-related increases in BR CA mortality. If the hypotheses are proven correct, interventions that reduce endothelial cell injury and turnover may reduce adverse clinical outcomes from BR CA, which is our long-term goal.
