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Vascular Endothelial-Breast Epithelial Cell Coculture Model Created from 3D Cell Structures.

ABSTRACT: Endothelial cell interactions with normal and cancerous breast epithelial cells have been widely studied in tissue growth and development, as well as in angiogenesis and metastasis. Despite the understanding that 3D multicellular architecture is critical to the cell phenotype, 3D vascular structures have not yet been cocultured with 3D breast spheroids in vitro. The objective of this study was therefore to create a hierarchical, multiscale model of vascular endothelial-breast epithelial cell interactions in which both cell types were assembled into their 3D architectures. The model was successfully fabricated by adding preformed breast spheroids onto preformed endothelial tube-like networks. Through this model, we observed that breast spheroids maintain vascular tube-like networks. Over time, breast epithelial cells migrate out of the spheroid structure along the endothelial networks. This research shows that 3D cell structures serve as an important building block for creating multicellular coculture models to study physiologically relevant cell-cell interactions.

SUBMITTER: Swaminathan S

PROVIDER: S-EPMC8670894 | biostudies-literature | 2017 Nov

REPOSITORIES: biostudies-literature

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Vascular Endothelial-Breast Epithelial Cell Coculture Model Created from 3D Cell Structures.

Swaminathan Swathi S Ngo Olivia O Basehore Sarah S Clyne Alisa Morss AM

ACS biomaterials science & engineering 20170110 11

Endothelial cell interactions with normal and cancerous breast epithelial cells have been widely studied in tissue growth and development, as well as in angiogenesis and metastasis. Despite the understanding that 3D multicellular architecture is critical to the cell phenotype, 3D vascular structures have not yet been cocultured with 3D breast spheroids <i>in vitro</i>. The objective of this study was therefore to create a hierarchical, multiscale model of vascular endothelial-breast epithelial c ...[more]

PMID: 33418720

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Project description:IntroductionAbout 70% of breast cancers express oestrogen receptor alpha (ESR1/ERalpha) and are oestrogen-dependent for growth. In contrast with the highly proliferative nature of ERalpha-positive tumour cells, ERalpha-positive cells in normal breast tissue rarely proliferate. Because ERalpha expression is rapidly lost when normal human mammary epithelial cells (HMECs) are grown in vitro, breast cancer models derived from HMECs are ERalpha-negative. Currently only tumour cell lines are available to model ERalpha-positive disease. To create an ERalpha-positive breast cancer model, we have forced normal HMECs derived from reduction mammoplasty tissue to express ERalpha in combination with other relevant breast cancer genes.MethodsCandidate genes were selected based on breast cancer microarray data and cloned into lentiviral vectors. Primary HMECs prepared from reduction mammoplasty tissue were infected with lentiviral particles. Infected HMECs were characterised by Western blotting, immunofluorescence microscopy, microarray analysis, growth curves, karyotyping and SNP chip analysis. The tumorigenicity of the modified HMECs was tested after orthotopic injection into the inguinal mammary glands of NOD/SCID mice. Cells were marked with a fluorescent protein to allow visualisation in the fat pad. The growth of the graft was analysed by fluorescence microscopy of the mammary glands and pathological analysis of stained tissue sections. Oestrogen dependence of tumour growth was assessed by treatment with the oestrogen antagonist fulvestrant.ResultsMicroarray analysis of ERalpha-positive tumours reveals that they commonly overexpress the Polycomb-group gene BMI1. Lentiviral transduction with ERalpha, BMI1, TERT and MYC allows primary HMECs to be expanded in vitro in an oestrogen-dependent manner. Orthotopic xenografting of these cells into the mammary glands of NOD/SCID mice results in the formation of ERalpha-positive tumours that metastasise to multiple organs. The cells remain wild type for TP53, diploid and genetically stable. In vivo tumour growth and in vitro proliferation of cells explanted from tumours are dependent on oestrogen.ConclusionWe have created a genetically defined model of ERalpha-positive human breast cancer based on normal HMECs that has the potential to model human oestrogen-dependent breast cancer in a mouse and enables the study of mechanisms involved in tumorigenesis and metastasis.

Dataset Information

Vascular Endothelial-Breast Epithelial Cell Coculture Model Created from 3D Cell Structures.

Publications

Vascular Endothelial-Breast Epithelial Cell Coculture Model Created from 3D Cell Structures.

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