Estradiol increases VEGF in human breast studied by whole-tissue culture

Sex steroid exposure constitutes a risk factor for breast cancer, but little is known about the effects of sex steroids on the normal breast, largely because of the lack of convenient models. We have developed a method of culturing normal breast tissue ex vivo. We have applied this method to investigate the effects of estradiol and progesterone on the key angiogenic mediator, vascular endothelial growth factor (VEGF), in the breast. Whole breast tissue was obtained from routine reduction mammoplasty. Tissue biopsies were cultured in vitro for 1-3 weeks, and the expression of luminal cytokeratin 18 was determined by immunohistochemistry. As an application, tissue biopsies were treated in vitro for 1 week with or without estradiol or estradiol and progesterone. Estrogen receptor, progesterone receptor, and Ki-67 were analyzed, and VEGF levels were examined by quantitative immunoassay and immunohistochemistry. Whole breast tissue was cultured ex vivo for 1 week with preserved morphology. Increased detachment of the luminal epithelium was observed after 2 weeks. Estradiol increased extracellular levels of VEGF in normal breast tissue biopsy medium. The addition of progesterone had neither stimulatory nor inhibitory effects on secreted VEGF. The method of whole breast tissue culturing thus provide a means by which to explore the biology of normal breast tissue. Our results suggest that estradiol exerts pro-angiogenic effects in normal breast by increasing levels of biologically active VEGF.