RESUMEN
The role of the intranuclear movement of chromatin in gene expression is not well-understood. Herpes simplex virus forms replication compartments (RCs) in infected cell nuclei as sites of viral DNA replication and late gene transcription. These structures develop from small compartments that grow in size, move, and coalesce. Quantitative analysis of RC trajectories, derived from 4D images, shows that most RCs move by directed motion. Directed movement is impaired in the presence of actin and myosin inhibitors as well as a transcription inhibitor. In addition, RCs coalesce at and reorganize nuclear speckles. Lastly, distinct effects of actin and myosin inhibitors on viral gene expression suggest that RC movement is not required for transcription, but rather, movement results in the bridging of transcriptionally active RCs with nuclear speckles to form structures that enhance export of viral late mRNAs.
Asunto(s)
Transporte Activo de Núcleo Celular , Herpesviridae/fisiología , ARN Viral/metabolismo , Transcripción Genética , Replicación Viral , Animales , Núcleo Celular , Chlorocebus aethiops , Cromatina , Transfección , Células VeroRESUMEN
Invasion of malignant glioma is a highly complex phenomenon involving molecular and cellular processes at various spatio-temporal scales, whose precise interplay is still not fully understood. In order to identify the intrinsic cellular mechanisms of glioma invasion, we study an in vitro culture of glioma cells. By means of a computational approach, based on a cellular automaton model, we compare simulation results to the experimental data and deduce cellular mechanisms from microscopic and macroscopic observables (experimental data). For the first time, it is shown that the migration/proliferation dichotomy plays a central role in the invasion of glioma cells. Interestingly, we conclude that a diverging invasive zone is a consequence of this dichotomy. Additionally, we observe that radial persistence of glioma cells in the vicinity of dense areas accelerates the invasion process. We argue that this persistence results from a cell-cell repulsion mechanism. If glioma cell behavior is regulated through a migration/proliferation dichotomy and a self-repellent mechanism, our simulations faithfully reproduce all the experimental observations.
Asunto(s)
Neoplasias Encefálicas/patología , Glioma/patología , Modelos Biológicos , Algoritmos , Movimiento Celular/fisiología , Proliferación Celular , Humanos , Invasividad Neoplásica/fisiopatología , Esferoides Celulares , Células Tumorales CultivadasRESUMEN
The analysis of the pure motion of subnuclear structures without influence of the cell nucleus motion and deformation is essential in live cell imaging. In this paper, we propose a 2-D contour-based image registration approach for compensation of nucleus motion and deformation in fluorescence microscopy time-lapse sequences. The proposed approach extends our previous approach, which uses a static elasticity model to register cell images. Compared with that scheme, the new approach employs a dynamic elasticity model for the forward simulation of nucleus motion and deformation based on the motion of its contours. The contour matching process is embedded as a constraint into the system of equations describing the elastic behavior of the nucleus. This results in better performance in terms of the registration accuracy. Our approach was successfully applied to real live cell microscopy image sequences of different types of cells including image data that was specifically designed and acquired for evaluation of cell image registration methods. An experimental comparison with the existing contour-based registration methods and an intensity-based registration method has been performed. We also studied the dependence of the results on the choice of method parameters.
Asunto(s)
Núcleo Celular/fisiología , Procesamiento de Imagen Asistido por Computador/métodos , Microscopía Fluorescente/métodos , Algoritmos , Línea Celular , Elasticidad , Células HeLa , Humanos , Modelos BiológicosRESUMEN
To gain a better understanding of cellular and molecular processes, it is important to quantitatively analyze the motion of subcellular particles in live cell microscopy image sequences. Since, generally, the subcellular particles move and cell nuclei move as well as deform, it is important to decouple the movement of particles from that of the cell nuclei using non-rigid registration methods. We have developed a diffeomorphic multi-frame approach for non-rigid registration of cell nuclei in 2D and 3D live cell fluorescence microscopy images. Our non-rigid registration approach is based on local optic flow estimation, exploits information from multiple consecutive image frames, and determines diffeomorphic transformations in the log-domain, which allows efficient computation of the inverse transformations. To register single images of an image sequence to a reference image, we use a temporally weighted mean image, which is constructed based on inverse transformations and multiple consecutive frames. Using multiple consecutive frames improves the registration accuracy compared to pairwise registration, and using a temporally weighted mean image significantly reduces the computation time compared with previous work. In addition, we use a flow boundary preserving method for regularization of computed deformation vector fields, which prevents from over-smoothing compared to standard Gaussian filtering. Our approach has been successfully applied to 2D and 3D synthetic as well as real live cell microscopy image sequences, and an experimental comparison with non-rigid pairwise, multi-frame, and temporal groupwise registration has been carried out.
RESUMEN
The analysis of the motion of subcellular particles in live cell microscopy images is essential for understanding biological processes within cells. For accurate quantification of the particle motion, compensation of the motion and deformation of the cell nucleus is required. We introduce a non-rigid multi-frame registration approach for live cell fluorescence microscopy image data. Compared to existing approaches using pairwise registration, our approach exploits information from multiple consecutive images simultaneously to improve the registration accuracy. We present three intensity-based variants of the multi-frame registration approach and we investigate two different temporal weighting schemes. The approach has been successfully applied to synthetic and live cell microscopy image sequences, and an experimental comparison with non-rigid pairwise registration has been carried out.