A QCM-D sensing strategy for investigating the real-time effects of oxidative stress on the viscoelastic properties of pre-osteoblast cells

Abstract

Oxidative stress results from an imbalance in the cellular redox state which can cause damage to the cell cytoskeleton and morphology, thereby affecting cell viscoelasticity. In this study, we introduce a quartz crystal microbalance with dissipation (QCM-D) sensing strategy that enables the real-time assessment of the changes in the viscoelastic properties of an MC3T3 cell monolayer in response to oxidative stress. This was successfully achieved by tracking the changes in energy dissipation (ΔD-response) caused by the interaction of H2O2 with the cell monolayer adhered to a poly-d-lysine-coated polystyrene quartz sensor. While a return to baseline values of the ΔD-response was obtained at 325 min (recovery point) after the incubation of cells with 25 μM H2O2, higher concentrations (50 μM-10 mM) exhibited no recovery. We successfully validated using scanning electron, atomic force and fluorescence microscopy that at the recovery point the cell morphology recovered from 25 μM H2O2 exposure, whereas, higher concentrations (50 μM-10 mM) resulted in the shrinkage of the cytoskeleton, alteration in cell morphology, and decrease in cell density due to apoptosis/necrosis, supported by a cell viability assay. Finally, total antioxidant capacity assay (TAC) confirmed that while cells’ metabolic state recovered from 25 μM H2O2 treatment, higher H2O2 concentrations led to a decline in the TAC. Altogether, these results showed that the viscoelastic properties of cells can be used to investigate cell recovery from oxidative stress, which may allow us to distinguish oxidative eustress from distress. Overall, we have demonstrated that QCM-D is a viable tool for measuring the effects of oxidative stress on the viscoelastic properties of MC3T3 cells.