Mechanically stimulated ATP release from murine bone cells is regulated by a balance of injury and repair

Abstract

Bone cells sense and actively adapt to physical perturbations to prevent critical damage. ATP release is among the earliest cellular responses to mechanical stimulation. Mechanical stimulation of a single murine osteoblast led to the release of 70 ± 24 amole ATP, which stimulated calcium responses in neighboring cells. Osteoblasts contained ATP-rich vesicles that were released upon mechanical stimulation. Surprisingly, interventions that promoted vesicular release reduced ATP release, while inhibitors of vesicular release potentiated ATP release. Searching for an alternative ATP release route, we found that mechanical stresses induced reversible cell membrane injury in vitro and in vivo. Ca2+/PLC/PKC-dependent vesicular exocytosis facilitated membrane repair, thereby minimizing cell injury and reducing ATP release. Priming cellular repair machinery prior to mechanical stimulation reduced subsequent membrane injury and ATP release, linking cellular mechanosensitivity to prior mechanical exposure. Thus, our findings position ATP release as an integrated readout of membrane injury and repair.

Transmission of mechanical information by purinergic signaling

“…We have recently reported that physiologically-relevant mechanical loading routinely injured bone cells in vitro and in vivo, resulting in release of ATP through plasma membrane disruptions, and stimulation of calcium responses in the neighbouring cells (7). These membrane disruptions in bone cells are counteracted by rapid vesicle-mediated membrane repair (7,8), which limits ATP spillage.…”

“…We have recently reported that physiologically-relevant mechanical loading routinely injured bone cells in vitro and in vivo, resulting in release of ATP through plasma membrane disruptions, and stimulation of calcium responses in the neighbouring cells (7). These membrane disruptions in bone cells are counteracted by rapid vesicle-mediated membrane repair (7,8), which limits ATP spillage. Thus, contrary to previous generalizations that ATP is released as a bolus proportional to mechanical stimulus (9), our data suggest that mechanically-stimulated ATP release contains dynamic information about both the extent of the injury and the rate of repair.…”

Vibration of osteoblastic cells using a novel motion‐control platform does not acutely alter cytosolic calcium, but desensitizes subsequent responses to extracellular ATP

“…We first used the custom motion‐control platform to investigate whether vibration induces transient changes in [Ca 2+ ] i similar to those induced by other mechanical stimuli such as fluid shear, membrane stretch and deformation (Adachi et al, ; Donahue et al, ; Hung, Pollack, Reilly, & Brighton, ; Lorusso et al, ; Mikolajewicz et al, ; Nishitani, Saif, & Wang, ; Thompson et al, ; Wu, Wong, Glogauer, Ellen, & McCulloch, ). Under the conditions tested, changes in [Ca 2+ ] i were not observed in response to mechanical vibration of UMR‐106 cells, MC3T3‐E1 cells, or primary rat osteoclasts.…”

“…The biological mechanisms underlying mechanotransduction are areas of intense study (Wall et al, 2017). In cells of the osteoblast lineage, early signaling events activated by mechanical stimuli such as fluid flow and membrane stretch include transient elevation in the concentration of cytosolic free calcium ([Ca 2+ ] i ) (Jing et al, 2014;Lorusso et al, 2016;Mikolajewicz, Zimmermann, Willie, & Komarova, 2018;Thi, Suadicani, Schaffler, Weinbaum, & Spray, 2013). Furthermore, such responses have been shown to be dependent on the release of nucleotides, such as ATP, that act in an autocrine/ paracrine manner through Ca 2+ -mobilizing P2 receptors on the cell surface (Riddle, Taylor, Rogers, & Donahue, 2007).…”

“…Furthermore, such responses have been shown to be dependent on the release of nucleotides, such as ATP, that act in an autocrine/ paracrine manner through Ca 2+ -mobilizing P2 receptors on the cell surface (Riddle, Taylor, Rogers, & Donahue, 2007). In this regard, biosensors have been used to monitor ATP release in response to the mechanical stimulation of osteoblastic cells (Hecht, Liedert, Ignatius, Mizaikoff, & Kranz, 2013;Mikolajewicz et al, 2018).…”