title: Characterization of spontaneous and TGF-β-induced cell motility of primary human normal and neoplastic mammary cells in vitro using novel real-time technology.
abstract: The clinical complications derived from metastatic disease are responsible for the majority of all breast cancer related deaths. Since cell migration and invasion are a prerequisite for metastasis their assessment in patient cancer cells in vitro may have prognostic value for the tumor's metastatic capacity. We employed real-time cell analysis (RTCA) on the xCELLigence DP system to determine in vitro motility of patient-derived primary human breast cancer epithelial cells (HBCEC). Initially, the RTCA assay was validated using established human breast cancer cell lines with either an invasive (MDA-MB-231, MDA-MB-435s) or a non-invasive phenotype (MCF-7, MDA-MB-468), and primary NSCLC cells (Tu459). Previous standard assays of cell migration/invasion revealed that only MDA-MB-231, -435s, and Tu459 cells exhibited spontaneous and TGF-β1-stimulated migration and invasion through a Matrigel barrier. In the present study, the TGF-β1-stimulated activities could be blocked by SB431542, a potent kinase inhibitor of the TGF-β type I receptor ALK5. Application of the RTCA assay to patient-derived tumor cells showed that 4/4 primary HBCEC and primary NSCLC cells, but not normal human mammary epithelial cells (HMEC), displayed high spontaneous migratory and invasive activity which correlated with higher MMP-2 expression and uPA protein levels in HBCEC compared to HMEC. Upon treatment with TGF-β1, HBCEC exhibited morphologic and gene regulatory alterations indicative of epithelial-to-mesenchymal transition. However, exclusively the invasive but not the migratory activity of HBCEC was further enhanced by TGF-β1. This indicates the requirement for molecular, e.g. integrin interactions with Matrigel components in HBCEC in order to become responsive to pro-invasive TGF-β effects. Together, these results show for the first time that tumorigenic HBCEC but not normal HMEC possess a strong basal migratory as well as a basal and TGF-β1-inducible invasive potential. These findings qualify the RTCA assay as an in vitro migration/invasion testing system for patient-specific primary breast cancer cells.
authors: Biochemistry and Tumor Biology Laboratory, Gynecology Research Unit, Department of Gynecology and Obstetrics, Medical University, Hannover, Germany.
authors: Katharina
authors: K
authors: Mandel
authors: Daniel
authors: D
authors: Seidl
authors: Dirk
authors: D
authors: Rades
authors: Hendrik
authors: H
authors: Lehnert
authors: Frank
authors: F
authors: Gieseler
authors: Ralf
authors: R
authors: Hass
authors: Hendrik
authors: H
authors: Ungefroren
mesh_terms: D001940:Breast / Q000166:cytology*; D001943:Breast Neoplasms / Q000473:pathology*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects*; D003584:Cytological Techniques / Q000379:methods*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D020778:Matrix Metalloproteinase 2 / Q000235:genetics; D009361:Neoplasm Invasiveness; D017395:Plasminogen Activator Inhibitor 1 / Q000378:metabolism; D013997:Time Factors; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
chemical_list: D017395:Plasminogen Activator Inhibitor 1; D016212:Transforming Growth Factor beta; D014568:Urokinase-Type Plasminogen Activator; D020778:Matrix Metalloproteinase 2