Integrin targeting of glyphosate and its cell adhesion modulation effects on osteoblastic MC3T3-E1 cells revealed by label-free optical biosensing

Abstract

This study is a discovery of interesting and far reaching properties of the world leading herbicide active ingredient glyphosate. Here we demonstrate the cell adhesion-modifying characteristics of glyphosate affecting cellular interactions via Arg-Gly-Asp (RGD)-dependent integrins. This conclusion was supported by the observations that a glyphosate surface coating induced integrin-specific cell adhesion, while glyphosate in solution inhibited cell adhesion on an RGD-displaying surface. A sensitive, real-time, label-free, whole cell approach was used to monitor the cell adhesion kinetic processes with excellent data quality. The half maximal inhibitory concentration (IC50) for glyphosate was determined to be 0.47 ± 0.07% (20.6 mM) in serum-free conditions. A three-dimensional dissociation constant of 0.352 mM was calculated for the binding between RGD-specific integrins in intact MC3T3-E1 cells and soluble glyphosate by measuring its competition for RGD-motifs binding, while the affinity of those RGD-specific integrins to the RGD-motifs was 5.97 µM. The integrin-targeted affinity of glyphosate was proven using competitive binding assays to recombinant receptor αvβ3. The present study shows not only ligand-binding properties of glyphosate, but also illustrates its remarkable biomimetic power in the case of cell adhesion.

Single-cell adhesion force kinetics of cell populations from combined label-free optical biosensor and robotic fluidic force microscopy

“…Over the years, numerous methods have been introduced to examine and quantify cell adhesion, from simple observations in an optical microscope to increasingly elaborate atomic force microscopy (AFM) techniques [15][16][17][18][19] . These techniques either measure cell-surface interactions and cell adhesion kinetics 10,[20][21][22][23][24] , or they are based on applying a force that can lead to cell detachment (termed adhesion strength measurements) on single-cells (e.g., micropipette aspiration, AFM, optical tweezer techniques) or on cell populations (e.g., centrifugation assay, spinning disk, flow chamber) 25 . One critical parameter of cell adhesion measurement methods is the throughput, describing the number of cells that can be detached in a certain period of time 26 .…”

“…This novel method has been used to examine the behavior of live cells on biofunctionalized surfaces with extremely high sensitivity and throughput 23,[46][47][48] . The possibility to chemically modify the sensing surface allows for the examination of specific molecular interactions, such as Arg-Gly-Asp tripeptide (RGD motif) mediated activation and clustering of integrin proteins.…”

Biophysical and biomechanical properties of neural progenitor cells as indicators of developmental neurotoxicity

“…Exposure to heavy metals (e.g., lead, methylmercury) and organic sulfates led to altered expression and function of cell adhesion molecules such as cadherins and IgCAMs (Grunwald 2018). Upon exposure of glyphosate (a herbicide), a concentration-dependent reduction in cell adhesion was observed in MC3T3-E1 cells, via blocking of RGD-specific integrin (Szekacs et al 2018). Inhibiting fibronectin-integrin binding leads to a strong decrease in adhesion force of primary gastrulating cells as detected by AFM analysis (Puech et al 2005).…”

Surface Functionalization by Hydrophobin-EPSPS Fusion Protein Allows for the Fast and Simple Detection of Glyphosate

“…However, these findings could not be confirmed by some other studies [11,12,13]. Furthermore, there are studies showing an effect of glyphosate or its formulations for example on human placental cells and aromatase [14] or on cell adhesion properties [15]. Different results in several studies on glyphosate and its assessment of the International Agency for Research on Cancer (IARC) [16] as a probably carcinogenic compound raised the debate about the hazardous risks and the approval of glyphosate in the European Union.…”