Vibration induced osteogenic commitment of mesenchymal stem cells is enhanced by cytoskeletal remodeling but not fluid shear

Abstract

Consistent across studies in humans, animals and cells, the application of vibrations can be anabolic and/or anti-catabolic to bone. The physical mechanisms modulating the vibration-induced response have not been identified. Recently, we developed an in vitro model in which candidate parameters including acceleration magnitude and fluid shear can be controlled independently during vibrations. Here, we hypothesized that vibration induced fluid shear does not modulate mesenchymal stem cell (MSC) proliferation and mineralization and that cell’s sensitivity to vibrations can be promoted via actin stress fiber formation. Adipose derived human MSCs were subjected to vibration frequencies and acceleration magnitudes that induced fluid shear stress ranging from 0.04Pa to 5Pa. Vibrations were applied at magnitudes of 0.15g, 1g, and 2g using frequencies of both 100Hz and 30Hz. After 14d and under low fluid shear conditions associated with 100Hz oscillations, mineralization was greater in all vibrated groups than in controls. Greater levels of fluid shear produced by 30Hz vibrations enhanced mineralization only in the 2g group. Over 3d, vibrations led to the greatest increase in total cell number with the frequency/acceleration combination that induced the smallest level of fluid shear. Acute experiments showed that actin remodeling was necessary for early mechanical up-regulation of RUNX-2 mRNA levels. During osteogenic differentiation, mechanically induced up-regulation of actin remodeling genes including Wiskott-Aldrich syndrome (WAS) protein, a critical regulator of Arp2/3 complex, was related to the magnitude of the applied acceleration but not to fluid shear. These data demonstrate that fluid shear does not regulate vibration induced proliferation and mineralization and that cytoskeletal remodeling activity may play a role in MSC mechanosensitivity.

Bone marrow stem cells adapt to low-magnitude vibrations by altering theircytoskeleton during quiescence and osteogenesis

“…Therefore, our results regarding cell morphology and roughness should be treated as relative rather than absolute (Qian et al, 2010). Mechanical vibrations were shown to be anabolic for osteoprogenitor cell pools in vitro, whether the application of stimulus was vertical (Kim et al, 2012) or horizontal (Uzer et al, 2013). Our results were consistent with the increase in cell proliferation for quiescent stem cells, but we did not observe an increase in cell numbers of stem cells during osteogenic commitment, even though our results suggested increased viability.…”

“…Consistent with the idea that cytoskeletal elements are strong determinants of mechanotransductive pathways, chemical blocking of actin polymerization inhibits the response of bone cells to mechanical stimulation (Rosenberg, 2003). Similarly, molecular response to mechanical stimuli is augmented in progenitor cells during osteogenesis, when the treatment is combined with lysophosphatidic acid, an agent that induces rapid actin stress fiber formation (Uzer et al, 2013). Though cytoskeletal elements can guide the mechanical sensitivity of bone-forming cells during lowintensity vibrations, it is not clear whether cytoskeletal elements can adapt to this oscillatory stimulus, similar to adaptations observed in response to fluid shear (Malone et al, 2007;Ponik et al, 2007).…”

“…Another alternative hypothesis for the effectiveness of vibratory signals is the "oscillatory motion" component of these signals that prescribes accelerations on the cell and subcellular structures (Garman et al, 2007a(Garman et al, , 2007bOzcivici et al, 2007). Recent in vitro evidence suggests that during oscillatory fluid shear applications, a mechanical response is modulated by accelerations occurring during fluid flow rather than drag forces acting on the cellular membrane (Uzer et al, 2012(Uzer et al, , 2013.…”

Vibrational stimulation induces osteoblast differentiation and the upregulation of osteogenic gene expression in vitro

“…Because the vertical displacement of the vibrator at 60 Hz and 5 m/s 2 was 35 lm, this was too small to induce a horizontal flow. Therefore, it appears that the vibration-induced fluid shear stress acting on the cell layer in this study was less than that measured in Uzer's report (Uzer et al 2012(Uzer et al , 2013; (4) the maximum strain increased proportionally with peak acceleration (Fig. 4).…”

“…We confirmed that there was no standing wave on the fluid surface within the range of accelerations and frequencies used in this study. Finite element methods revealed that the range of fluid shear stress placed upon the cell layer was 0.04-0.94 Pa (Uzer et al 2013) when the plate was horizontally vibrated (LMHF; 30-100 Hz, 0.98-9.81 m/s 2 ). Because the vertical displacement of the vibrator at 60 Hz and 5 m/s 2 was 35 lm, this was too small to induce a horizontal flow.…”