Smurova K M, Alieva I B, Vorob'ev I A
Tsitologiia. 2007;49(4):270-9.
In the internal cytoplasm of interphase cells the density of microtubules is the highest in the centrosome area and decreases to the cell periphery. As a rule, the quantity of fluorescent microtubules cannot be counted up in the internal cytoplasm, but it is possible to estimate microtubules quantity using measuring of their optical density. In living 3T3 and CHO cells the microtubules optical density decreased according to different mathematical dependences that apparently reflected the differences of their microtubule system organization. To determine appropriateness that circumscribe the reduction of microtubules optical density from the centrosome region to the direction of cell margin, we modeled cell contours with the certain ratio and interposition of centrosome-attached and free microtubules in vector schedules CorelDraw program. The decrease of optical density was analyzed in MetaMorph program as it was described earlier (Smurova et al., 2002). It was shown that fluorescent microtubules optical density decreased exponentially (y = ae(-bx)) if the system joined only microtubules growing from the centrosome up to the cell margin. The curve became smoother in the case of not all radial centrosome-attached microtubules reached the margin, and adding of free microtubules into the system led to the sharp fall in optical density in the centrosome area and to its gradual decrease at the cell periphery. The increase in free microtubules quantity changed the character of the curve describing the reduction of optical density microtubule system which included free and centrosome-attached microtubules in proportions of 5 : 1 was described by the equation of linear regression (f= k . x + b). Thus, the mathematical dependence describing the microtubules distribution from the centrosome to the cell periphery, depends on the ratio of microtubules and their relative positioning in the cell volume. The data obtained using model systems have coincided with the results of experiments. The graphs which described the increase in microtubules optical density during microtubule repolymerization after nocodazole treatment, corresponded to the graphs for model cells. Thus, the method we used allows to analyze the microtubule system in the cases when the direct observation of individual microtubules is difficult.
在间期细胞的内部细胞质中,微管密度在中心体区域最高,并向细胞周边递减。通常,无法在内部细胞质中对荧光微管数量进行计数,但可以通过测量其光密度来估算微管数量。在活的3T3和CHO细胞中,微管光密度根据不同的数学关系下降,这显然反映了它们微管系统组织的差异。为了确定从中心体区域到细胞边缘方向微管光密度降低的适当性,我们在CorelDraw程序的矢量图中,以特定比例和中心体附着微管与游离微管的位置关系对细胞轮廓进行建模。如前所述(Smurova等人,2002年),在MetaMorph程序中分析光密度的降低情况。结果表明,如果系统仅包含从中心体生长到细胞边缘的微管,荧光微管光密度呈指数下降(y = ae(-bx))。当并非所有与中心体相连的径向微管都到达边缘时,曲线会变得更平滑,并且向系统中添加游离微管会导致中心体区域光密度急剧下降,而在细胞周边则逐渐下降。游离微管数量的增加改变了描述微管系统光密度降低的曲线特征,其中游离微管与中心体附着微管比例为5:1的微管系统光密度降低情况由线性回归方程(f = k·x + b)描述。因此,描述从中心体到细胞周边微管分布的数学关系取决于微管的比例及其在细胞体积中的相对位置。使用模型系统获得的数据与实验结果一致。描述诺考达唑处理后微管再聚合过程中微管光密度增加的图表,与模型细胞的图表相符。因此,我们使用的方法能够在难以直接观察单个微管的情况下分析微管系统。