ABSTRACT
Loss of cellular architecture and polarity of breast tissue is one of the early markers for onset of breast cancer. This loss in cellular morphology has been phenocopied using three-dimensional (3D) cultures of human mammary epithelial cells, MCF10A. MCF10A are immortalized, non-transformed and non-malignant human mammary epithelial cells that when grown in 3D matrices, exhibit a number of features of normal breast epithelium. The morphology of these acini gets disrupted in malignancy, such as an increase in size and elongation of acini. As transformation progresses, the acini loose their polarization and some may even form multi-acinar structures. Using this model system, it was observed that damage induced by N-methyl-N-nitrosourea (MNU),a SN1 type DNA methylating agent which forms the modified base O6-methylguanine,caused disruption of the well-formed polarized spheroid architectureas observed using various polarity markers such as .6-integrin, GM-130, laminin V and cell-cell junction markers such as .-catenin andE-cadherin. Interestingly, an up-regulation of Vimentin was also observed in the breast acini. Loss in the ability of the damaged cells to form tubular structures with cytoplasmic localization of Laminin V was observed in the Collagen-Matrigel invasion assays. A gain of an elongated morphology was also observed in the treated cells, an indication of attainment of a mesenchymal phenotype. The disrupted spheroids were observed to have fewer cells with larger nuclear volume but the overall spheroid volume was similar to that of the untreated. Thus, MNU has the potential to cause early transformation in MCF10A cells grown as 3D cultures. The model proposed here, thus can be further used to study early events of chemical-induced transformation.Read More…