Guo Ang, Song Long-Sheng
Division of Cardiovascular Medicine, Department of Internal Medicine and Francois M. Abboud Cardiovascular Research Center, Carver College of Medicine, University of Iowa, Iowa City, Iowa.
Division of Cardiovascular Medicine, Department of Internal Medicine and Francois M. Abboud Cardiovascular Research Center, Carver College of Medicine, University of Iowa, Iowa City, Iowa.
Biophys J. 2014 Jun 17;106(12):2729-36. doi: 10.1016/j.bpj.2014.05.013.
Cardiac transverse (T)-tubules provide a specialized structure for synchronization and stabilization of sarcoplasmic reticulum Ca(2+) release in healthy cardiomyocytes. The application of laser scanning confocal microscopy and the use of fluorescent lipophilic membrane dyes have boosted the discoveries that T-tubule remodeling is a significant factor contributing to cardiac contractile dysfunction. However, the analysis and quantification of the remodeling of T-tubules have been a challenge and remain inconsistent among different research laboratories. Fast Fourier transformation (FFT) is the major analysis method applied to calculate the spatial frequency spectrum, which is used to represent the regularity of T-tubule systems. However, this approach is flawed because the density of T-tubules as well as non-T-tubule signals in the images influence the spectrum power generated by FFT. Preprocessing of images and topological architecture extracting is necessary to remove non-T-tubule noise from the analysis. In addition, manual analysis of images is time consuming and prone to errors and investigator bias. Therefore, we developed AutoTT, an automated analysis program that incorporates image processing, morphological feature extraction, and FFT analysis of spectrum power. The underlying algorithm is implemented in MATLAB (The MathWorks, Natick, MA). The program outputs the densities of transversely oriented T-tubules and longitudinally oriented T-tubules, power spectrum of the overall T-tubule systems, and averaged spacing of T-tubules. We also combined the density and regularity of T-tubules to give an index of T-tubule integrity (TTint), which provides a global evaluation of T-tubule alterations. In summary, AutoTT provides a reliable, easy to use, and fast approach for analyzing myocyte T-tubules. This program can also be applied to measure the density and integrity of other cellular structures.
心脏横管（T管）为健康心肌细胞中肌浆网Ca(2+)释放的同步化和稳定提供了一种特殊结构。激光扫描共聚焦显微镜的应用以及亲脂性荧光膜染料的使用推动了相关发现，即T管重塑是导致心脏收缩功能障碍的一个重要因素。然而，T管重塑的分析和量化一直是一个挑战，不同研究实验室之间的结果仍不一致。快速傅里叶变换（FFT）是用于计算空间频谱的主要分析方法，该频谱用于表示T管系统的规律性。然而，这种方法存在缺陷，因为图像中T管的密度以及非T管信号会影响FFT产生的频谱功率。图像预处理和拓扑结构提取对于从分析中去除非T管噪声是必要的。此外，图像的人工分析既耗时又容易出错，还存在研究者偏差。因此，我们开发了AutoTT，这是一个自动化分析程序，它结合了图像处理、形态特征提取和频谱功率的FFT分析。底层算法在MATLAB（The MathWorks，Natick，MA）中实现。该程序输出横向T管和纵向T管的密度、整个T管系统的功率谱以及T管的平均间距。我们还结合T管的密度和规律性给出了一个T管完整性指数（TTint），它提供了对T管改变的整体评估。总之，AutoTT为分析心肌细胞T管提供了一种可靠、易用且快速的方法。该程序也可用于测量其他细胞结构的密度和完整性。
