Investigation of changes in optical attenuation of bone and neuronal cells in organ culture or three-dimensional constructs in vitro with optical coherence tomography: relevance to cytochrome oxidase monitoring
Investigation of changes in optical attenuation of bone and neuronal cells in organ culture or three-dimensional constructs in vitro with optical coherence tomography: relevance to cytochrome oxidase monitoring
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DOI：
10.1007/s00249-003-0285-z
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发表时间：
2003-03
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影响因子：
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通讯作者：
Xiangqun Xu;Ruikang K. Wang;A. E. El Haj
中科院分区：
文献类型：
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作者：
Xiangqun Xu;Ruikang K. Wang;A. E. El Haj
利用光学相干断层扫描（OCT）监测了外植培养的大鼠骨膜组织和三维琼脂糖构建的人神经细胞的光衰减变化，这些变化与细胞色素氧化酶有关，在组织工程和诊断中具有潜在的应用前景。采用超发光二极管（SLD），其峰值发射波长（λ=820 nm）为氧化型CytOx的近红外吸收波段。衰减系数通过液体模型（含脂内萘酚绿B）、外植体培养（大鼠颅骨骨膜）和三维琼脂糖构建的细胞的深度分辨反射率曲线获得。利用衰减系数与浓度的线性关系，可以准确地定量测定萘酚绿B的吸收系数。减少CytOx前后星形细胞瘤细胞在琼脂糖中的衰减系数的差异为0.26±0.10 mm−1(n=9)，而琼脂糖对照未观察到衰减。骨膜组织中酶的减少导致衰减系数变化0.43±0.24 mm−1（n=7）。为了进行比较，使用生化测定，在星形细胞瘤细胞和大鼠骨膜中，氧化还原形式的CytOx的吸收系数分别在820 nm处约8.3±1.5×10−3mm−1（n=4）和8.7±2.5×10−3mm−1（n=4）处测量。与OCT相比，生化测量的CytOx浓度较低可能是由于散射曲线的变化和多种血红素氧化酶的放大影响，这表明常规OCT不足以监测细胞色素氧化酶的氧化还原变化。然而，使用该技术，氧化态的定性变化是明显的。我们的研究结果表明，OCT在提供器官培养组织和体外三维结构细胞的高分辨率断层成像方面具有潜在的应用前景。
Changes in optical attenuation, relevant to cytochrome oxidase, of the rat bone periosteal tissue in explanted culture and human neuronal cells in three-dimensional agarose constructs have been monitored by the use of optical coherence tomography (OCT), with potential applications in tissue engineering and diagnosis. A superluminescent diode (SLD) with a peak emission wavelength (λ=820 nm) that is the near-infrared absorption band of the oxidized form of CytOx was employed. The attenuation coefficient was obtained from the depth-resolved reflectance profiles of liquid phantoms (naphthol green B with intralipid), explant culture (periosteum of calvaria from rats) and cells in 3D agarose constructs. The absorption coefficient of naphthol green B can be accurately quantified by the linear relationship between attenuation coefficients and the concentration. The difference in the attenuation coefficient of astrocytoma cells in agarose before and after reduction of CytOx is 0.26±0.10 mm−1(n=9), whereas no attenuation is observed with the agarose control. Reduction of the enzyme in periosteal tissue leads to a change in attenuation coefficient of 0.43±0.24 mm−1(n=7). For comparison, using a biochemical assay, the absorption coefficient of the oxidized-reduced form of CytOx is measured at approximately 8.3±1.5×10−3mm−1(n=4) and 8.7±2.5×10−3mm−1(n=4) at 820 nm for astrocytoma cells and rat periosteum, respectively. The lower value of CytOx concentration using biochemical versus OCT measurements may result from shifts in the scattering profile and the amplifying influences of multiple heme-based oxidases, indicating that conventional OCT is not specific enough to monitor redox changes in cytochrome oxidase. However, qualitative shifts in oxidation state are apparent using the technique. Our results suggest the potential application of OCT in providing high-resolution tomographic imaging of tissues in organ culture and cells grown in three-dimensional constructs in vitro.