Search

The actin filaments of the cytoskeleton form a highly dynamic polymer scaffold which is actively involved in many essential mechanisms such as cell migration, transport, mitosis, and mechanosensitivity. We treated CHO-K1 cells with different concentrations of the actin cytoskeleton disrupting drug cytochalasin D. Then investigating the cells' elastic behaviour by scanning force microscopy-based rheology we confirmed for high cytochalasin D concentrations (≥1.5 μM) a significant decrease of mechanical stability. At lower concentrations we measured no significant softening, but flattening and a horizontal contraction was observable even at low concentrations (≥0.3 μM) of cytochalasin D. The observed changes in cell shape resulted in a lower cell volume, showing that there is compensation by volume for small decreases in cytoskeletal strength resulting from reduced numbers or lengths of actin filaments. These results suggest that the characteristic functions defining a cell's mechanical stability such as mechanosensitivity can be maintained via small changes in cell volume in order to counter fluctuations in cytoskeletal composition. Cell Motil. Cytoskeleton 2009. © 2009 Wiley-Liss, Inc.

Vegetation indices (VIs), which are derived from hyperspectral measurements, may be useful non-destructive measures to estimate crop canopy parameters. A systematic analysis of the reflectance spectrum of winter oilseed rape (OSR) for the derivation of VIs has not been conducted yet. We therefore derived in our study VIs from 61 available wavebands of the spectral range from 400 nm to 1000 nm systematically and compared the best ones to commonly used indices. Hyperspectral reflectance and destructive measurements of crop canopy parameters were therefore carried out in 2005 and 2006 in northern Germany for calibration and in 2006 for validation at the same location. For the derivation of VIs for OSR, three different approaches were tested. The approaches differed in the way of the waveband combinations by combining two wavebands in a simple ratio (SR) form λ 1 / λ 2 , a normalized difference index (NDI) form ( λ 1 − λ 2 )/( λ 1 + λ 2 ) or by using a stepwise forward multiple regression (MR), which identifies the best linear combination of all available bands in a linear combination. The derived VIs were tested for their predictive power for crop canopy parameters like green area index (GAI), shoot dry matter (DM shoot ) and total nitrogen amount in the shoot (N shoot ) and were compared to commonly used indices. Waveband combinations of two near infrared bands resulted in the best prediction of the tested crop canopy parameters for calibration and validation data sets. Correlation coefficients ( r 2 ) yielded values up to 0.82 between new indices and N shoot . Especially, NDI 750,740 was best predicting GAI, whereas either NDI or SR forms of 740 nm and 780 nm showed best results predicting DM shoot and N shoot and outperformed commonly used indices. Predicting crop canopy parameters by MR showed good results for calibration, but highest variation for validation among all newly derived indices.

Die Zeiten sind bekanntlich vorbei, in denen Kinder und Jugendliche im Schein der Taschenlampe unter der Bettdecke Abenteuerromane verschlangen, um dann als solcherart geübte und Büchern gegenüber aufgeschlossene Leserinnen und Leser die Voraussetzungen für eine in der Schule angeregte und angeleitete komplexere Lektüre mitzubringen. Wer heute Literaturunterricht planen und durchführen muß, steht vor zwei Problemen, die so schwierig wie noch nie zuvor zu lösen sind: Wie wecke ich überhaupt Bereitschaft zur Lektüre anspruchsvoller Texte, möglichst in Form eines solchen Motivationsschubs, daß dauerhaftes außerschulisches Lesen angeregt wird? Und: Wie entwickle ich die Fähigkeit zum angemessenen Umgang mit ‚schwierigen‘ Texten? Gemeint ist eine Verstehens- und Äußerungskompetenz, die auch vor dem Hintergrund moderner literaturtheoretischer Erkenntnisse vertretbar sein soll.

Biomechanical properties of tumor cells play an important role for the metastatic capacity of cancer. Cellular changes of viscoelastic features are prerequisite for cancer progression since they are essential for proliferation and metastasis. However, only little is known about the way how expression of oncogenes influences these biomechanical properties. To address this aspect we used a breast cancer cell line with inducible expression of an oncogenic version of ERBB2. ERBB2 is known to be correlated with bad prognosis in breast cancer. Cell elasticity was determined by the Optical Stretcher, where suspended cells are deformed by two slightly divergent laser beams. We found that induction of ERBB2 caused remarkable biomechanical alterations of the MCF-7 cells after 24 h: the cells actively contracted in response to mechanical stimuli, a phenomenon known as mechanoactivation. After this period, as the cells became senescent, the mechanoactivity returned to control levels. Time-resolved gene array analysis revealed that mechanoactivation was accompanied by temporal upregulation of 46 cytoskeletal genes. A possible role of these genes in tumor progression was investigated by expression analyses of 766 breast cancer patients. This showed an association of 12 out of these 46 genes with increased risk of metastasis. Our results demonstrate that overexpression of ERBB2 causes mechanoactivation of tumor cells, which may enhance tumor cell motility fostering distant metastasis.

Optical traps such as tweezers and stretchers are widely used to probe the mechanical properties of cells. Beyond their large range of applications, the use of infrared laser light in optical traps causes significant heating effects in the cell. This study investigated the effect of laser-induced heating on cell viability. Common viability assays are not very sensitive to damages caused in short periods of time or are not practicable for single cell analysis. We used cell spreading, a vital ability of cells, as a new sensitive viability marker. The optical stretcher, a two beam laser trap, was used to simulate heat shocks that cells typically experience during measurements in optical traps. The results show that about 60% of the cells survived heat shocks without vital damage at temperatures of up to 58 ± 2°C for 0.5 s. By varying the duration of the heat shocks, it was shown that 60% of the cells stayed viable when exposed to 48 ± 2°C for 5 s.

Degenerative joint diseases caused by rheumatism, joint dysplasia or traumata are particularly widespread in countries with high life expectation. Although there is no absolutely convincing cure available so far, hyaline cartilage and bone defects resulting from joint destruction can be treated today by appropriate transplantations. Recently, procedures were developed based on autologous chondrocytes from intact joint areas. The chondrocytes are expanded in cell culture and subsequently transplanted into the defect areas of the affected joints. However, these autologous chondrocytes are characterized by low expansion capacity and the synthesis of extracellular matrix of poor functionality and quality. An alternative approach is the use of adult mesenchymal stem cells (MSCs). These cells effectively expand in 2D culture and have the potential to differentiate into various cell types, including chondrocytes. Furthermore, they have the ability to synthesize extracellular matrix with properties mimicking closely the healthy hyaline joint cartilage. Beside a more general survey of the architecture of hyaline cartilage, its composition and the pathological processes of joint diseases, we will describe here which advances were achieved recently regarding the development of closed, aseptic bioreactors for the production of autologous grafts for cartilage regeneration based on MSCs. Additionally, a novel mathematical model will be presented that supports the understanding of the growth and differentiation of MSCs. It will be particularly emphasized that such models are helpful to explain the well-known fact that MSCs exhibit improved growth properties under reduced oxygen pressure and limited supply with nutrients. Finally, it will be comprehensively shown how different analytical methods can be used to characterize MSCs on different levels. Besides discussing methods for non-invasive monitoring and tracking of the cells and the determination of their elastic properties, mass spectrometric methods to evaluate the lipid compositions of cells will be highlighted.

The measurement of the mechanical properties of individual cells has received much attention in recent years. In this paper we describe the application of optically induced forces with an optical stretcher to perform step-stress experiments on individual suspended fibroblasts. The conversion from creep-compliance to frequency-dependent complex shear modulus reveals characteristic viscoelastic signatures of the underlying cytoskeleton and its dynamic molecular properties. Both normal and cancerous fibroblasts display a single stress relaxation time in the observed time and frequency space that can be related to the transient binding of actin crosslinking proteins. In addition, shear modulus and steady-state viscosity of the shell-like actin cortex as the main module resisting small deformations are extracted. These values in combination with insight into the cells’ architecture are used to explain their different deformability. This difference can then be exploited to distinguish normal from cancerous cells. The nature of the optical stretcher as an optical trap allows easy incorporation in a microfluidic system with automatic trapping and alignment of the cells, and thus a high measurement throughput. This carries the potential for using the microfluidic optical stretcher to investigate cellular processes involving the cytoskeleton and to diagnose diseases related to cytoskeletal alterations.

Isolation of individual cells from a heterogeneous cell population is an invaluable step in the analysis of single cell properties. The demands in molecular and cellular biology as well as molecular medicine are the selection, isolation, and monitoring of single cells and cell clusters of biopsy material. Of particular interest are methods which complement a passive optical or spectroscopic selection with a variety of active single cell processing techniques such as mechanical, biochemical, or genetic manipulation prior to isolation. Sophisticated laser-based cell processing systems are available which can perform single cell processing in a contact-free and sterile manner. Until now, however, these multipurpose turnkey systems offer only basic micromanipulation and are not easily modified or upgraded, whereas laboratory situations often demand simple but versatile and adaptable solutions. We built a flexible laser micromanipulation platform combining contact-free microdissection and catapulting capabilities using a pulsed ultraviolet (337nm) laser with simultaneous generation of optical tweezing forces using a continuous wave infrared (1064nm) laser. The potential of our platform is exemplified with techniques such as local laser-induced injection of biomolecules into individual living cells, laser surgery, isolation of single cells by laser catapulting, and control of neuronal growth using optical gradient forces. Arbitrary dynamic optical force patterns can be created by fast laser scanning with acousto-optical deflectors and galvanometer mirrors, allowing multibeam contact-free micromanipulation, a prerequisite for reliable handling of material in laboratory-on-a-chip applications. All common microscopy techniques can be used simultaneously with the offered palette of micromanipulation methods. Taken together, we show that advanced optical micromanipulation systems can be designed which combine quality, cost efficiency, and adaptability.