Abstract
The human mammary gland is a complex and heterogeneous organ, where the interactions between mammary epithelial cells (MEC) and stromal fibroblasts are known to regulate normal biology and tumorigenesis. We aimed to longitudinally evaluate morphology and size of organoids in 3D co-cultures of normal (MCF10A) or pre-malignant (MCF10DCIS.com) MEC and hTERT-immortalized fibroblasts from reduction mammoplasty (RMF). This co-culture model, based on an isogenic panel of cell lines, can yield insights to understand breast cancer progression. However, 3D cultures pose challenges for quantitative assessment and imaging, especially when the goal is to measure the same organoid structures over time. Using optical coherence tomography (OCT) as a non-invasive method to longitudinally quantify morphological changes, we found that OCT provides excellent visualization of MEC-fibroblast co-cultures as they form ductal acini and remodel over time. Different concentrations of fibroblasts and MEC reflecting reported physiological ratios [1] were evaluated, and we found that larger, hollower, and more aspherical acini were formed only by pre-malignant MEC (MCF10DCIS.com) in the presence of fibroblasts, whereas in comparable conditions, normal MEC (MCF10A) acini remained smaller and less aspherical. The ratio of fibroblast to MEC was also influential in determining organoid phenotypes, with higher concentrations of fibroblasts producing more aspherical structures in MCF10DCIS.com. These findings suggest that stromal-epithelial interactions between fibroblasts and MEC can be modeled in vitro, with OCT imaging as a convenient means of assaying time dependent changes, with the potential for yielding important biological insights about the differences between benign and pre-malignant cells.
Highlights
The human mammary gland consists of a series of branching ducts, with each branch terminating as a hollow and spherical acinus
Interactions between mammary epithelial cells (MEC) and stromal fibroblasts are regulators of tumorigenesis [4,5], with stroma playing a vital role in the proliferation and organization of MEC, production of extracellular matrix (ECM), and regulation of cellular adhesion and migration [4]
The aim of this study was to define morphological hallmarks of stromal-epithelial interactions using optical coherence tomography (OCT) to assess 3D in vitro cultures comprised of basal-like mammary epithelial cell lines [28] and hTERT-immortalized fibroblasts from reduction mammoplasty (RMF)
Summary
The human mammary gland consists of a series of branching ducts, with each branch terminating as a hollow and spherical acinus. Chemical and physical interactions between epithelia and surrounding stroma are essential for the organ’s development and physiological functions. This intricate network of cells is a complex microenvironment [2] that maintains normal tissue architecture (homeostasis) and suppresses malignant phenotypes in healthy individuals [3], but becomes permissive or even promotes cancer during progression [4]. Fibroblasts are strongly associated with mammary epithelium, and in the vicinity of tumors, become a major cell type of the stroma [6] These cancer-associated fibroblasts appear to promote tumor growth and facilitate the progression of breast cancer [7]. By comparing normal and pre-malignant MEC co-cultured with RMF in 3D, and by modulating the ratios of the two cell types, we aimed to elucidate how stromal-epithelial interactions modulate morphological changes in acini
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