The impacts of accumulating pico-newton mechanical perturbations on the calcium signaling and ROS responses of glioma

Hsiu-Mei Wei¹, Rong-Shing Chang¹, Chi-Shuo Chen^1*

¹Biomedical Engineering and Environmental Sciences, National Tsing Hua University, Hsinchu, Taiwan

* Presenter:Chi-Shuo Chen, email:chen.chishuo@gmail.com

Mechanical forces presences in diverse physiological process in glioma, such abnormal brain pressure and tumor invasion. At the intercellular interfaces, force in tens pico-newton express in different amplitudes and periods; extracellular forces over specific threshold is known to activate intracellular signaling, but the influences of force period and applying frequency is still largely underexplored. Here, we studied the impacts of accumulating force stimuli on glioblastoma using magnetic tweezers system. Based on the data of fluorescent Fluo-4, we observed at least 2 sec pulling time was required for single force pulse (~11 pN) to activate the Ca+2 influx; no Ca+2 response was observed even under the 30 secs total pulling time of 1 Hz force stimuli. Using pharmacological inhibitor GsMTx4, we identified the transmembrane channel, Piezo-1, partially contributes to the force-induced Ca+2 influx. We speculated the transmission of low-frequency (< 0.5 Hz) mechanical force stimuli in the viscous cell membrane and evaluated the influence of high-frequency (1 Hz) force on the elastic cortical cytoskeleton. Using glioma with eliminated myosin-actin stress, we observed no significant difference in the Ca+2 peak responses to the force stimuli and the higher integrated Ca+2 alteration after the total pulling time for 30 secs, which highlighted the role of membrane deformation in force-induced signaling. We further studied the force-induced reactive oxygen species (ROS) of glioma, and observed the ROS alteration prolong for 30 min after transient force stimuli (0.5 Hz for 30 sec); the mitochondrial ROS was also interfered by the applying force. In summary, we found the duration/frequency of the force applied on cell affected the force-induced signals, such as Ca+2 and ROS; and the cellular responses correlated to the summing-up effects of force perturbations. In contrast to the critical threshold of the applied force, our results suggested the potential impacts of the accumulating effects of small force stimuli. These findings may further highlight the importance of the oscillation frequency of cellular force, and imply profound influences of the trivial force signals presented all around in organisms.

Keywords: cell mechanics, magnetic tweezers, calcium signaling, ROS