Continuous and regulated remodelling of the cytoskeleton is crucial for many basic cell functions. In contrast to actin filaments and microtubules, it is not understood how this is accomplished for the third major cytoskeletal filament system, which consists of intermediate-filament polypeptides. Using time-lapse fluorescence microscopy of living interphase cells, in combination with photobleaching, photoactivation and quantitative fluorescence measurements, we observed that epithelial keratin intermediate filaments constantly release non-filamentous subunits, which are reused in the cell periphery for filament assembly. This cycle is independent of protein biosynthesis. The different stages of the cycle occur in defined cellular subdomains: assembly takes place in the cell periphery and newly formed filaments are constantly transported toward the perinuclear region while disassembly occurs, giving rise to diffusible subunits for another round of peripheral assembly. Remaining juxtanuclear filaments stabilize and encage the nucleus. Our data suggest that the keratin-filament cycle of assembly and disassembly is a major mechanism of intermediate-filament network plasticity, allowing rapid adaptation to specific requirements, notably in migrating cells.

lichen planus mucosae. RESULTS: The stepwise application of 2 additional approaches (morphology, DNA content, argyrophilic nucleolar organizer region counts) increased the specificity of conventional cytologic diagnosis from 92.6% to 100%. This feasibility study provided a proof of concept, demonstrating efficiency,

The paper describes the key component of the Multimodal Cell Analysis approach, a novel cytologic evaluation method for early cancer detection. The approach is based on repeated staining of a cell smear. The correlation of features and data extracted from the different stains, and related to relocated individual cells, may yield a dramatic increase of diagnostic reliability.In order to utilise the technique, fully automatic, adaptive image preprocessing techniques need to be applied, which are described in this article: coregistration of multimodal images, segmentation, and classification of cell nuclei. The presented feasibility study shows both efficiency and robustness of all steps being high regarding medical image material, and it strongly supports clinical application.

SummaryCell adhesion, a process mediated by the formation of discrete structures known as focal adhesions (FAs), is pivotal to many biological events including cell motility. Much is known about the molecular composition of FAs, although our knowledge of the spatio-temporal recruitment and the relative occupancy of the individual components present in the FAs is still incomplete. To fill this gap, an essential prerequisite is a highly reliable procedure for the recognition, segmentation and tracking of FAs. Although manual segmentation and tracking may provide some advantages when done by an expert, its performance is usually hampered by subjective judgement and the long time required in analysing large data sets. Here, we developed a model-based segmentation and tracking algorithm that overcomes these problems. In addition, we developed a dedicated computational approach to correct segmentation errors that may arise from the analysis of poorly defined FAs. Thus, by achieving accurate and consistent FA segmentation and tracking, our work establishes the basis for a comprehensive analysis of FA dynamics under various experimental regimes and the future development of mathematical models that simulate FA behaviour.

Cell stability and motility depends on a complex dynamic cytoplasmic scaffolding called the cytoskeleton. It is composed of actin filaments, intermediate filaments and microtubules, and interacts with neighbouring cells and the extracellular matrix via specialized adhesion sites -multimolecular complexes responsible for the transmission of mechanical force and regulatory signals. The dynamic behaviour of these subcellular structures in living cells can be analysed by fluorescence microscopy yielding series of 2D or 3D images. Towards a quantitative analysis, we present methods for the segmentation and motion estimation of cytoskeletal filaments as well as for the tracking of adhesion sites, allowing the quantification of cytoskeletal dynamics under different conditions.