Department of Biomedical Engineering, Fourth Military Medical University, Xi'an, Shaanxi 710032, China.
Bone. 2013 Apr;53(2):531-40. doi: 10.1016/j.bone.2013.01.008. Epub 2013 Jan 14.
Mechanical stimuli can trigger intracellular calcium (Ca(2+)) responses in osteocytes and osteoblasts. Successful construction of bone cell networks necessitates more elaborate and systematic analysis for the spatiotemporal properties of Ca(2+) signaling in the networks. In the present study, an unsupervised algorithm based on independent component analysis (ICA) was employed to extract the Ca(2+) signals of bone cells in the network. We demonstrated that the ICA-based technology could yield higher signal fidelity than the manual region of interest (ROI) method. Second, the spatiotemporal properties of Ca(2+) signaling in osteocyte-like MLO-Y4 and osteoblast-like MC3T3-E1 cell networks under laminar and steady fluid flow stimulation were systematically analyzed and compared. MLO-Y4 cells exhibited much more active Ca(2+) transients than MC3T3-E1 cells, evidenced by more Ca(2+) peaks, less time to the 1st peak and less time between the 1st and 2nd peaks. With respect to temporal properties, MLO-Y4 cells demonstrated higher spike rate and Ca(2+) oscillating frequency. The spatial intercellular synchronous activities of Ca(2+) signaling in MLO-Y4 cell networks were higher than those in MC3T3-E1 cell networks and also negatively correlated with the intercellular distance, revealing faster Ca(2+) wave propagation in MLO-Y4 cell networks. Our findings show that the unsupervised ICA-based technique results in more sensitive and quantitative signal extraction than traditional ROI analysis, with the potential to be widely employed in Ca(2+) signaling extraction in the cell networks. The present study also revealed a dramatic spatiotemporal difference in Ca(2+) signaling for osteocytic and osteoblastic cell networks in processing the mechanical stimulus. The higher intracellular Ca(2+) oscillatory behaviors and intercellular coordination of MLO-Y4 cells provided further evidences that osteocytes may behave as the major mechanical sensor in bone modeling and remodeling processes.
机械刺激可以触发骨细胞内的细胞内钙（Ca(2+)）反应。成功构建骨细胞网络需要更精细和系统地分析网络中 Ca(2+)信号的时空特性。在本研究中，采用基于独立成分分析（ICA）的无监督算法提取网络中骨细胞的 Ca(2+)信号。结果表明，基于 ICA 的技术比手动感兴趣区域（ROI）方法具有更高的信号保真度。其次，系统分析和比较了在层流和稳定流体流动刺激下，成骨细胞样 MLO-Y4 和成骨细胞样 MC3T3-E1 细胞网络中 Ca(2+)信号的时空特性。MLO-Y4 细胞比 MC3T3-E1 细胞表现出更多的 Ca(2+)瞬变，表现为更多的 Ca(2+)峰、达到第一个峰的时间更短、第一个峰和第二个峰之间的时间更短。就时间特性而言，MLO-Y4 细胞表现出更高的尖峰率和 Ca(2+)振荡频率。MLO-Y4 细胞网络中 Ca(2+)信号的细胞间同步活动比 MC3T3-E1 细胞网络更高，并且与细胞间距离呈负相关，表明 MLO-Y4 细胞网络中 Ca(2+)波的传播速度更快。本研究结果表明，与传统 ROI 分析相比，基于 ICA 的无监督技术可实现更敏感和定量的信号提取，具有在细胞网络中提取 Ca(2+)信号的广泛应用潜力。本研究还揭示了成骨细胞和破骨细胞网络在处理机械刺激时 Ca(2+)信号的显著时空差异。MLO-Y4 细胞更高的细胞内 Ca(2+)振荡行为和细胞间协调性提供了进一步的证据，表明破骨细胞可能在骨形成和重塑过程中作为主要的机械传感器。
