Cell stiffening is a label-free indicator of reactive oxygen species-induced intracellular acidification

Reactive oxygen species (ROS) are important secondary messengers involved in a variety of cellular processes, including activation, proliferation, and differentiation. Hydrogen peroxide (H2O2) is a major ROS typically kept in low nanomolar range that causes cell and tissue damage at supraphysiological concentrations. While ROS have been studied in detail at molecular scale, little is known about their impact on cell mechanical properties as label-free biomarker for stress response. Here, we exposed human myeloid precursor cells, T-lymphoid cells and neutrophils to varying concentrations of H2O2 and show that elevated levels of mitochondrial superoxide are accompanied by an increased Young's modulus. Mechanical alterations do not originate from global modifications in filamentous actin and microtubules but from cytosolic acidification due to lysosomal degradation. Finally, we demonstrate our findings to be independent of the presence of H2O2 and that stiffening seems to be a general response of cells to stress factors lowering cytosolic pH. Reactive oxygen species (ROS), such as hydrogen peroxide, play a key role in cellular processes but can cause damage at high concentrations. The authors demonstrate that elevated ROS levels are accompanied by an increased cell stiffness that is explained by cytosolic acidification due to lysosomal degradation and not by alterations in the cytoskeleton.