Quantitative in situ time-series evaluation of osteoblastic collagen synthesis under cyclic strain using second-harmonic-generation microscopy
Quantitative in situ time-series evaluation of osteoblastic collagen synthesis under cyclic strain using second-harmonic-generation microscopy
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使用二次谐波显微镜对循环应变下成骨细胞胶原合成进行定量原位时间序列评估
DOI：
10.1117/12.2316607
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发表时间：
2019
影响因子：
3.5
通讯作者：
Takeshi Yasui
中科院分区：
文献类型：
--
作者：
Katsuya Sato;Oki Matsubara;Eiji Hase;Takeo Minamikawa;Takeshi Yasui
本研究的目的是使用二次谐波发生（SHG）显微镜评估机械刺激下成骨细胞胶原蛋白的合成。我们应用 SHG 显微镜来监测由成骨细胞样细胞 (MC3T3-E1) 合成的胶原纤维，无需固定和染色。为了定量评估机械刺激对成骨细胞胶原蛋白合成的影响，我们比较了使用实验室制造的拉伸装置施加和不施加循环拉伸刺激的成骨细胞合成的胶原纤维的 SHG 图像。我们在不同的刺激条件下每 7 天获取一次 SHG 图像，持续 3 周（5 分钟/天和 3 小时/天，应变幅度为 5%，频率为 0.5 Hz）。平均SHG强度的图像分析表明，与非拉伸条件相比，循环拉伸使成骨细胞胶原合成量显着增强，而两种机械刺激条件之间没有显着差异。此外，成骨早期胶原纤维的成熟度不受机械刺激的影响。研究结果可用于骨再生医学，通过人工刺激对胶原蛋白合成进行反馈控制。
The aim of this study is to evaluate the osteoblastic collagen synthesis under mechanical stimulation using second-harmonic-generation (SHG) microscopy. We apply SHG microscopy to monitor the collagen fibers synthesized by osteoblast-like cells (MC3T3-E1) without the need for fixation and staining. To quantitatively evaluate the influence of mechanical stimulation on osteoblastic collagen synthesis, we compare SHG images of osteoblast-synthesized collagen fibers with and without a cyclic stretch stimulus applied using a lab-made stretching device. We acquire SHG images every 7 days for 3 weeks at different stimulus conditions (5 min/day and 3 h/day with a strain magnitude of 5% and a frequency of 0.5 Hz). Image analysis of the average SHG intensity indicates that the amount of osteoblastic collagen synthesis is significantly enhanced by the cyclic stretch compared with the nonstretched condition, while there is no significant difference between the two mechanical stimulation conditions. Furthermore, the maturity of the collagen fibers in the early stage of bone formation is not affected by the mechanical stimulation. The results can be used in bone regenerative medicine to apply feedback control of collagen synthesis by artificial stimulation.