Hauser Joerg, Hauser Manfred, Muhr Gert, Esenwein Stefan
Department of Surgery, BG-Kliniken Bergmannsheil, Ruhr-University-Bochum, Buerkle-de-la-Camp-Platz 1, 44789 Bochum Germany.
J Orthop Res. 2009 Mar;27(3):286-94. doi: 10.1002/jor.20741.
In clinical and experimental studies an acceleration of fracture healing and increased callus formation induced by low-intensity pulsed ultrasound (LIPUS) has been demonstrated. The exact molecular mechanisms of ultrasound treatment are still unclear. In this study ultrasound transmitted cytoskeletal and growth rate changes of SAOS-2 cells were examined. Osteoblast-like cell lines (SAOS-2) were treated using low-intensity pulsed ultrasound. Cytoskeletal changes were analyzed using rhodamine phalloidine for f-actin staining and indirect immunofluorescence techniques with different monoclonal antibodies against several tubulin modifications. To examine changes of cell number after ultrasound treatment cell counts were done. Significant changes in cytoskeleton structure were detected compared to controls, including an enhancement of stress fiber formation combined with a loss of cell migration after ultrasound application. We further observed that sonication altered the proportion of the more stable microtubules to the more labile microtubule subclass. The labile tyrosinated microtubules appeared highly enhanced, whereas the amount of the more stable acetylated microtubules was remarkably diminished. All these observations were quantified by fluorometric measurements. The centrosomal gamma-tubulin was frequently scattered throughout the cell's cytoplasm, giving rise to additional polyglu-positive microtubular asters, which induced multipolar spindles, leading either to aneuploid mini-or giant cells. Moreover, a significant increase of cell number was noticed in the sonicated group. These experiments demonstrate that ultrasound treatment increases cell number and leads to significant changes of the cytoskeletal structure and composition in vitro.
在临床和实验研究中，已证实低强度脉冲超声（LIPUS）可加速骨折愈合并增加骨痂形成。超声治疗的确切分子机制仍不清楚。在本研究中，检测了超声对SAOS-2细胞细胞骨架和生长速率变化的影响。使用低强度脉冲超声处理成骨样细胞系（SAOS-2）。使用罗丹明鬼笔环肽进行f-肌动蛋白染色，并采用针对几种微管蛋白修饰的不同单克隆抗体的间接免疫荧光技术分析细胞骨架变化。为了检测超声处理后细胞数量的变化，进行了细胞计数。与对照组相比，检测到细胞骨架结构有显著变化，包括超声处理后应力纤维形成增强以及细胞迁移能力丧失。我们进一步观察到，超声处理改变了更稳定的微管与更不稳定的微管子类的比例。不稳定的酪氨酸化微管明显增加，而更稳定的乙酰化微管数量显著减少。所有这些观察结果均通过荧光测量进行量化。中心体γ-微管蛋白经常分散在整个细胞质中，产生额外的多聚谷氨酸阳性微管星状体，进而诱导多极纺锤体，导致非整倍体小细胞或巨细胞。此外，在超声处理组中观察到细胞数量显著增加。这些实验表明，超声处理可增加细胞数量，并在体外导致细胞骨架结构和组成发生显著变化。
