Dual TGFb/BMP pathway inhibition enables expansion and characterization of multiple epithelial cell types of the normal and cancerous breast

Mayuri Prasad, Brijesh Kumar, Poornima Bhat-Nakshatri, Manjushree Anjanappa, George Sandusky, Kathy Miller, Anna Maria Storniolo, Harikrishna Nakshatri

Research output: Contribution to journalArticle

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Abstract

Functional modeling of normal breast epithelial hierarchy and stromal-epithelial cell interactions have been difficult due to inability to obtain sufficient stem-progenitor-mature epithelial and stromal cells. Recently reported epithelial reprogramming assay has partially overcome this limitation, but cross-contamination of cells from the feeder layer is a concern. The purpose of this study was to develop a feeder-layer- independent and inexpensive method to propagate multiple cell types from limited tissue resources. Cells obtained after enzymatic digestion of tissues collected at surgery or by coreneedle biopsies were plated on tissue culture dishes precoated with laminin-5-rich-conditioned media from the rat bladder tumor cell line 804G and a defined growth media with inhibitors of ROCK, TGFb, and BMP signaling. Cells were characterized by flow cytometry, mammosphere assay, 3D cultures, and xenograft studies. Cells from the healthy breasts included CD10+/EpCAM- basal/myoepithelial, CD49f+/EpCAM+ luminal progenitor, CD49f-/EpCAM+ mature luminal, CD73+/EpCAM+/CD90- rare endogenous pluripotent somatic stem, CD73+/CD90+/EpCAM-, estrogen receptor alpha-expressing ALCAM (CD166)+/EpCAM+, and ALDFLUOR+ stem/luminal progenitor subpopulations. Epithelial cells were luminal (KRT19+), basal (KRT14+), or dual-positive luminal/basal hybrid cells. While breast cells derived from BRCA1, BRCA2, and PALB2 mutation carriers did not display unique characteristics, cells from women with breast cancer-protective alleles showed enhanced differentiation. Cells could also be propagated from primary tumors and metastasis of breast, ovarian, and pancreatic cancer- neuroendocrine subtype. Xenograft studies confirmed tumorigenic properties of tumor-derived cells.

Original languageEnglish (US)
Pages (from-to)1556-1570
Number of pages15
JournalMolecular Cancer Research
Volume17
Issue number7
DOIs
StatePublished - Jul 1 2019

Fingerprint

Breast
Epithelial Cells
Feeder Cells
Stromal Cells
Heterografts
Activated-Leukocyte Cell Adhesion Molecule
Breast Neoplasms
Estrogen Receptor alpha
Hybrid Cells
Conditioned Culture Medium
Tumor Cell Line
Pancreatic Neoplasms
Urinary Bladder Neoplasms
Cell Communication
Ovarian Neoplasms
Epithelial Cell Adhesion Molecule
Digestion
Neoplasms
Flow Cytometry
Alleles

ASJC Scopus subject areas

  • Molecular Biology
  • Oncology
  • Cancer Research

Cite this

Dual TGFb/BMP pathway inhibition enables expansion and characterization of multiple epithelial cell types of the normal and cancerous breast. / Prasad, Mayuri; Kumar, Brijesh; Bhat-Nakshatri, Poornima; Anjanappa, Manjushree; Sandusky, George; Miller, Kathy; Storniolo, Anna Maria; Nakshatri, Harikrishna.

In: Molecular Cancer Research, Vol. 17, No. 7, 01.07.2019, p. 1556-1570.

Research output: Contribution to journalArticle

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