Endothelial cells stimulate growth of normal and cancerous breast epithelial cells in 3D culture

Saevar Ingthorsson,Thorarinn Gudjonsson,Valgardur Sigurdsson,Agla Jr Fridriksdottir,Jon G Jonasson,Jens Kjartansson,Magnus K Magnusson

doi:10.1186/1756-0500-3-184

Abstract

BackgroundEpithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. However, research on the regulatory role of the endothelial component in the normal and malignant breast gland has largely been neglected. The aim of the study was to investigate the effects of endothelial cells on growth and differentiation of human breast epithelial cells in a three-dimensional (3D) co-culture assay.MethodsBreast luminal and myoepithelial cells and endothelial cells were isolated from reduction mammoplasties. Primary cells and established normal and malignant breast cell lines were embedded in reconstituted basement membrane in direct co-culture with endothelial cells and by separation of Transwell filters. Morphogenic and phenotypic profiles of co-cultures was evaluated by phase contrast microscopy, immunostaining and confocal microscopy.ResultsIn co-culture, endothelial cells stimulate proliferation of both luminal- and myoepithelial cells. Furthermore, endothelial cells induce a subpopulation of luminal epithelial cells to form large acini/ducts with a large and clear lumen. Endothelial cells also stimulate growth and cloning efficiency of normal and malignant breast epithelial cell lines. Transwell and gradient co-culture studies show that endothelial derived effects are mediated - at least partially - by soluble factors.ConclusionBreast endothelial cells - beside their role in transporting nutrients and oxygen to tissues - are vital component of the epithelial microenvironment in the breast and provide proliferative signals to the normal and malignant breast epithelium. These growth promoting effects of endothelial cells should be taken into consideration in breast cancer biology.

Highlights

Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer
The terminal duct lobular units (TDLUs) consist of an inner layer of luminal epithelial cells and an outer layer of myoepithelial cells separated from the surrounding vascular rich stroma by a basement membrane [1,2]
Breast endothelial cells cultured in rBM are quiescent but metabolically active When breast endothelial cells (BRENCs) are cultured on top of rBM they form a dense, capillary-like, network shortly after seeding (Figure 1a)

Summary

Introduction

Epithelial-stromal interaction provides regulatory signals that maintain correct histoarchitecture and homeostasis in the normal breast and facilitates tumor progression in breast cancer. Shekhar et al [17,18] showed that interaction between endothelial cells and premalignant breast epithelial cells was necessary to allow sufficient proliferation of endothelial cells as well as to induce branching ductalalveolar morphogenesis and hyperplasia of premalignant cells [17,18] In these studies, they used human umbilical vein endothelial cells (HUVEC) instead of organ-specific endothelial cells. Seeding normal and cancerous epithelial cells in rBM at clonal dilution with endothelial cells resulted in increased cloning efficacy and larger colony size This data suggests that endothelial cells in addition to providing nutrient and oxygen to tissues, might be an important microenvironmental factor for normal morphogenesis and cancerous growth in the human breast gland

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