Single-Cell Probe Force Studies to Identify Sox2 Overexpression-Promoted Cell Adhesion in MCF7 Breast Cancer Cells.

Jagoba Iturri,Andreas Weber,María Dm Vivanco,José L Toca-Herrera

doi:10.3390/cells9040935

Abstract

The replacement of the cantilever tip by a living cell in Atomic Force Microscopy (AFM) experiments permits the direct quantification of cell–substrate and cell–cell adhesion forces. This single-cell probe force measurement technique, when complemented by microscopy, allows controlled manipulation of the cell with defined location at the area of interest. In this work, a setup based on two glass half-slides, a non-fouling one with bacterial S-layer protein SbpA from L. sphaericus CMM 2177 and the second with a fibronectin layer, has been employed to measure the adhesion of MCF7 breast cancer cells to fibronectin films (using SbpA as control) and to other cells (symmetric vs. asymmetric systems). The measurements aimed to characterize and compare the adhesion capacities of parental cells and cells overexpressing the embryonic transcription factor Sox2, which have a higher capacity for invasion and are more resistant to endocrine therapy in vivo. Together with the use of fluorescence techniques (epifluorescence, Total Internal Fluorescence Microscopy (TIRF)), the visualization of vinculin and actin distribution in cells in contact with fibronectin surfaces is enabled, facilitating the monitoring and quantification of the formation of adhesion complexes. These findings demonstrate the strength of this combined approach to assess and compare the adhesion properties of cell lines and to illustrate the heterogeneity of adhesive strength found in breast cancer cells.

Highlights

Cancer-induced alterations at the molecular level result in changes at the cellular and tissue level, leading to defects in cell proliferation, migration, and adhesion
Strength and Work of Adhesion Differ for MCF7 and Sox2 Overexpressing Cells In Figure 2a, representative force–distance curves for control (MCF7) and Sox2-overexpressing (Sox2, see Supporting Information, Figure S2a) MCF7 cells in contact with fibronectin and SbpA
Mutations and changes in protein expression lead to distinct phenotypes in tumor cells, affecting cell adhesion and mechanics, survival, proliferation, and motility

Summary

Introduction

Cancer-induced alterations at the molecular level result in changes at the cellular and tissue level, leading to defects in cell proliferation, migration, and adhesion. The tight control of cell biochemical and mechanical microenvironment through ECM components like fibronectin, collagen, proteoglycans or laminin is disturbed during the various phases of cancer disease [1]. This leads to ECM remodeling accompanied by increased fiber alignment and matrix stiffening [2]. The process starts with cancer cell detachment, the passing of cellular barriers, transport through a fluid system, and adhesion and proliferation at a distant site [3,4]. An improved understanding of the interactions of cancer cells with ECM components is warranted

Methods

Results

Conclusion