Influence of electrical stimulation on bone remodelling

电刺激（ES）方法在骨科中用于促进骨再生。成骨细胞和破骨细胞相互作用导致的骨重塑是一种成功的再生治疗。生物物理和电刺激对成骨细胞或破骨细胞的影响已有文献报道，然而，交流电场对这两种细胞类型的影响以及在类似刺激条件下在一个ES装置中两者的相互作用以前没有描述过。为了研究ES对骨重塑的影响及其对骨重塑的影响，我们将利用已经建立的ES体外系统刺激羟基磷灰石（HA）涂层玻璃和钛上的骨细胞。由于透明质酸是骨的主要无机成分，因此它是培养成骨细胞和破骨细胞两种细胞类型的合适底物。HA涂层将通过体液模拟来实现，涂层将通过不同的分析和成像方法来控制。在实验初期，ES对成骨细胞和破骨细胞活力的影响将在28天内分别进行实验研究。分析的时间点是根据细胞特异性分化阶段选择的。到目前为止，尚不清楚胚胎干细胞是否会影响细胞分化。此外，尚不清楚分化相是强烈分离还是流畅过渡。因此，ES诱导的两种细胞类型的分化将通过分析所选标记的基因表达来检验，并将在蛋白质水平上得到证实。然后用相反细胞类型的ES产生的细胞培养上清培养成骨细胞和破骨细胞。介质交换允许通过相互独立的可溶性因子来研究两种细胞类型之间的相互作用。细胞培养上清含有体内骨细胞相互作用所需的所有分泌因子的总和。目前尚不清楚可溶性因子的表达是否受ES的调节，这必须在提出的建议中进行研究。因此，将分析上清液的组成和骨重塑特异性蛋白质的调节。利用条件培养基培养细胞的知识将用于建立成骨细胞和破骨细胞的共同培养，以在随后的资助期内同时进行ES。在这个项目中，将获得关于电刺激下成骨细胞和破骨细胞相互作用的基本知识，从而改善骨组织的再生。

Electrical stimulation (ES) approaches are used in orthopaedics to enhance bone regeneration. A successful regenerative therapy is assumed by bone remodeling resulting from the interaction of osteoblasts and osteoclasts. Effects of bio-physical and electrical stimulation on either osteoblasts or osteoclasts have been reported in literature, however, the influence of alternating current fields on both cell types and the interaction of both in one ES set-up under similar stimulation conditions have not been described before. To investigate the influence of ES on bone remodeling and their effects on it, an already established in vitro system for ES will be adapted to stimulate the bone cells on hydroxyapatite (HA) coated glass and titanium. As HA is the main anorganic component of bone, it is a suitable substrate for the cultivation of both cell types, osteoblasts and osteoclasts. The HA coating will be achieved by body fluid simulation and the coating will be controlled by different analytic and imaging methods. At the beginning, the influence of ES on the viability of osteoblasts and osteoclasts will be investigated in separate experiments over 28 days in maximum. The time points for analyses are chosen due to cell-specific differentiation phases. By now, it is not known whether ES effects cellular differentiation. Moreover, it is unknown whether the differentiation phases are strongly separated or in fluent transitions. Therefore, the differentiation of both cell types induced by ES will be examined via analysis of the gene expression of selected markers and will be confirmed on protein level. Furthermore, the osteoblasts and osteoclasts will be cultivated with the cell culture supernatant generated by ES of the opposite cell type. The medium exchange allows the investigation of the interplay between both cell types through soluble factors independently from each other. The cell culture supernatants contain the total of all secreted factors necessary for bone cell interaction in vivo. It is already unknown whether the expression of soluble factors is regulated by ES which has to be examined in the presented proposal. Thus, supernatants will be analysed for composition and regulation of proteins specific for bone remodeling. The knowledge of cell cultivation with the conditioned medium will be used for the establishment of a co-culture of osteoblasts and osteoclasts for a simultaneous ES in the subsequent funding period. Within this project, fundamental knowledge about interactions between osteoblasts and osteoclasts under electrical stimulation will be acquired, allowing improved regeneration of bone tissue.