Molecular Mechanisms of Low Intensity Pulsed Ultrasound in Human Skin Fibroblasts

Department of Chemistry, Universität Ulm, Ulm, Baden-Württemberg, Germany
Journal of Biological Chemistry (Impact Factor: 4.57). 01/2005; 279(52):54463-9. DOI: 10.1074/jbc.M404786200
Source: PubMed


Soluble factors such as polypeptide growth factors, mitogenic lipids, inflammatory cytokines, and hormones are known regulators of cell proliferation. However, the effect of mechanical stimuli on cell proliferation is less well understood. Here we examined the effect of low intensity pulsed ultrasound (US), which is used to promote wound healing, on the proliferation of primary human foreskin fibroblasts and the underlying signaling mechanisms. We show that a single 6-11-min US stimulation increases bromodeoxyuridine incorporation. In addition, an increase in the total cell number is observed after sequential US stimulation. US induced stress fiber and focal adhesion formation via activation of Rho. We further observed that US selectively induced activation of extracellular signal-regulated kinase (ERK) 1/2. Inhibition of Rho-associated coiled-coil-containing protein kinase (ROCK) prevented US-induced ERK1/2 activation, demonstrating that the Rho/ROCK pathway is an upstream regulator of ERK activation in response to US. Consequently, activation of ROCK and MEK-1 was required for US-induced DNA synthesis. Finally, an integrin beta(1) blocking antibody as well as a RGD peptide prevented US-induced DNA synthesis. In addition, US slightly increased phosphorylation of Src at Tyr(416), and Src activity was found to be required for ERK1/2 activation in response to US. In conclusion, our data demonstrate for the first time that US promotes cell proliferation via activation of integrin receptors and a Rho/ROCK/Src/ERK signaling pathway.

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    • "However, only fragmented findings have so far been reported on this topic. A general trend that has been observed is post-exposure cytoskeletal remodeling (Hauser et al. 2009; Mizrahi et al. 2012; Noriega et al. 2013; Zhang et al. 2012), and it may result in the alteration of the cell's adherence characteristics (Mahoney et al. 2009; Roper et al. 2012; Zhou et al. 2004). In line with this trend, post-exposure bio-assays have revealed that a cell's mechanotransduction pathway would be activated by low-intensity ultrasound (Louw et al. 2013; Whitney et al. 2012), and in turn, its associated molecular signaling cascades would be modulated (Favaro-Pipi et al. 2010; Ito et al. 2012; Lu et al. 2009). "
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    ABSTRACT: To establish the therapeutic potential of low-intensity ultrasound, it is important to characterize its biophysical interactions with living cells. Here, through a series of single-cell direct observations, we show that low-intensity ultrasound pulsing would give rise to a dynamic course of cytomechanical perturbations at both the membrane and nucleus levels. Our investigation was conducted using a composite platform that coupled a 1-MHz ultrasound exposure hardware to a confocal microscopy system. Short ultrasound pulses (5 cycles, 2-kHz pulse repetition frequency) with a spatial-peak time-averaged intensity of 0.24 W/cm2 (0.85-MPa peak positive acoustic pressure) were delivered over a 10-min period to adherent Neuro-2a neuroblastoma cells, and live imaging of cellular dynamics was performed before, during and after the exposure period. Bright-field imaging results revealed progressive shrinkage of cellular cross-sectional area (25%–45%, N = 7) during low-intensity ultrasound pulsing; the initial rate of size decrease was estimated to be 8%–14% per minute. This shrinkage was found to be transient, as the sonicated cells had recovered (at a rate of size increase of 0.4%–0.9% per minute) to their pre-exposure size within 30 min after the end of exposure. Three-dimensional confocal imaging results further revealed that (i) ultrasound-induced membrane contraction was volumetric in nature (21%–45% reduction), and (ii) a concomitant decrease in nucleus volume was evident (12%–25% reduction). Together, these findings indicate that low-intensity ultrasound pulsing, if applied on the order of minutes, would reversibly perturb the physical and subcellular structures of living cells.
    Ultrasound in medicine & biology 07/2014; 40(7). DOI:10.1016/j.ultrasmedbio.2014.01.003 · 2.21 Impact Factor
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    • "This is the stage at which cells migrate to the site of injury and start to divide, granulation tissue is formed, and fibroblasts begin to produce collagen. Ultrasound has been shown to enhance collagen synthesis by fibroblasts and repairing of epithelium (Zhou et al., 2004). Many laboratory-based studies have been undertaken to understand the effects of ultrasound on wound healing. "
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    ABSTRACT: Objective: To compare and investigate the effect of phonopheresis and topical salvadora persica on wound healing in rats. Design: Randomized, controlled trial. Settings: University animal laboratory. Animals: 45 adult albino male rats allocated to A, B and C groups. Interventions: After anesthesia, a wound area of 5cm2 was made on the upper dorsum of all rats. Group A was treated with phonopheresis, with frequency of 1MHz, intensity of 0.5W/cm2 and duty cycle of 40%, group B received sham phonopheresis while group C received sham ultrasonic, and treatment duration for all was six minutes/session/8 sessions. Wounds were captured by digital camera and were measured using trace method. Outcome measures: healing represents decreased in wound surface area. Results: there was a significant sequential reduction in wound surface area throughout 1st, 3rd, 5th and 8th sessions in all three groups. There was significant difference between; mean of W.S.A in group (A) and mean of W.S.A in group (B) at3rd ,5th and 8th sessions as p value (.09), (0.029) and (0.004) respectively, also there was significant difference between; mean of W.S.A in group (A) and mean of W.S.A in group (C) at 3rd, 5th and 8th sessions as p value (0.003),(0.00001) and (0.00001) respectively, while there was significant difference between; mean of W.S.A in group(B) and mean of W.S.A in group (C) at 3rd, 5th and 8th sessions as p value (0.024), (0.00001) and (0.021) respectively. Conclusion: phonophersis and topical salvadora persica gel have satisfied effect on wound healing. In addition, better healing provided by phonopheresis
    International Journal of Advanced Research 05/2014; 2(5):919-928.
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    • "Western blot analysis. Cell lysates were prepared as described before (Zhou et al, 2004). In all, 20–30 mg proteins were separated on 6% or 8% SDS-PAGE and then transferred to polyvinylidene difluoride membranes (Immobilon-P, Millipore, Bedford, MA, USA). "
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    ABSTRACT: Background: Pancreatic stellate cells (PSCs) promote metastasis as well as local growth of pancreatic cancer. However, the factors mediating the effect of PSCs on pancreatic cancer cells have not been clearly identified. Methods: We used a modified Boyden chamber assay as an in vitro model to investigate the role of PSCs in migration of Panc1 and UlaPaCa cells and to identify the underlying mechanisms. Results: PSC supernatant (PSC-SN) dose-dependently induced the trans-migration of Panc1 and UlaPaCa cells, mainly via haptokinesis and haptotaxis, respectively. In contrast to poly-L-lysine or fibronectin, collagen I resembled PSC-SN with respect to its effect on cancer cell behaviours, including polarised morphology, facilitated adhesion, accelerated motility and stimulated trans-migration. Blocking antibodies against integrin α2/β1 subunits significantly attenuated PSC-SN- or collagen I-promoted cell trans-migration and adhesion. Moreover, both PSC-SN and collagen I induced the formation of F-actin and focal adhesions in cells, which was consistent with the constantly enhanced phosphorylation of focal adhesion kinase (FAK, Tyr397). Inhibition of FAK function by an inhibitor or small interference RNAs significantly diminished the effect of PSC-SN or collagen I on haptotaxis/haptokinesis of pancreatic cancer cells. Conclusion: Collagen I is the major mediator for PSC-SN-induced haptokinesis of Panc1 and haptotaxis of UlaPaCa by activating FAK signalling via binding to integrin α2β1.
    British Journal of Cancer 11/2013; 110(2). DOI:10.1038/bjc.2013.706 · 4.84 Impact Factor
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