The DNT-EST: a predictive embryonic stem cell-based assay for developmental neurotoxicity testing in vitro.

As the developing brain is exquisitely vulnerable to chemical disturbances, testing for developmental neurotoxicity of a substance is an important aspect of characterizing its tissue specific toxicity. Mouse embryonic stem cells (mESCs) can be differentiated toward a neural phenotype, and this can be used as a model for early brain development. We developed a new in vitro assay using mESCs to predict adverse effects of chemicals and other compounds on neural development - the so-called DNT-EST.

Neural differentiation of mouse embryonic stem cells as a tool to assess developmental neurotoxicity in vitro.

Mouse embryonic stem cells (mESCs) represent an attractive cellular system for in vitro studies in developmental biology as well as toxicology because of their potential to differentiate into all fetal cell lineages. The present study aims to establish an in vitro system for developmental neurotoxicity testing employing mESCs. We developed a robust and reproducible protocol for fast and efficient differentiation of the mESC line D3 into neural cells, optimized with regard to chemical testing.

Assaying embryotoxicity in the test tube: current limitations of the embryonic stem cell test (EST) challenging its applicability domain.

Testing for embryotoxicity in vitro is an attractive alternative to animal experimentation. The embryonic stem cell test (EST) is such a method, and it has been formally validated by the European Centre for the Validation of Alternative Methods. A number of recent studies have underscored the potential of this method.

Protein biomarkers for in vitro testing of embryotoxicity.

There are new challenges for hazard and risk assessment in the chemical industry with regard to REACH legislation in Europe and related activities in the U.S. and Japan, which require the development of novel in vitro models for the molecular characterization of drug- or chemical-related effects replacing conventional animal testing.

Unexpected common mechanistic pathways for embryotoxicity of warfarin and lovastatin.

Novel molecular content for fast in vitro strategies in the context of safety tests concerning developmental toxicity has a potential to substantially reduce animal experiments according to the "3R" concept (Reduce/Refine/Replace). Here we present and discuss data from a differential proteomic profiling of samples generated using embryonic stem cell derived in vitro models treated with a set of model substances. Among substance-dependent proteomic changes, potential surrogate markers were some isoforms of heat shock proteins and a component of the Ras pathway, present in several redundant isoforms due to posttranslational modifications.

Ponsin interacts with Nck adapter proteins: implications for a role in cytoskeletal remodelling during differentiation of skeletal muscle cells.

Skeletal muscle differentiation is a complex process: It is characterised by changes in gene expression and protein composition. Simultaneously, a dramatic remodelling of the cytoskeleton and associated cell-matrix contacts, the costameres, occurs. The expression and localisation of the protein ponsin at cell-matrix contacts marks the establishment of costameres.

Complete loss of murine Xin results in a mild cardiac phenotype with altered distribution of intercalated discs.

Aims: Xin is a striated muscle-specific F-actin binding protein that has been implicated in cardiomyopathies. In cardiomyocytes, Xin is localized at intercalated discs (IDs). Mice lacking only two of the three Xin isoforms (XinAB(-/-) mice) develop severe cardiac hypertrophy.

Beyond the sarcomere: CSRP3 mutations cause hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is a frequent genetic cardiac disease and the most common cause of sudden cardiac death in young individuals. Most of the currently known HCM disease genes encode sarcomeric proteins. Previous studies have shown an association between CSRP3 missense mutations and either dilated cardiomyopathy (DCM) or HCM, but all these studies were unable to provide comprehensive genetic evidence for a causative role of CSRP3 mutations.

Paxillin and ponsin interact in nascent costameres of muscle cells.

Muscle differentiation requires the transition from motile myoblasts to sessile myotubes and the assembly of a highly regular contractile apparatus. This striking cytoskeletal remodelling is coordinated with a transformation of focal adhesion-like cell-matrix contacts into costameres. To assess mechanisms underlying this differentiation process, we searched for muscle specific-binding partners of paxillin.

Fatigue resistance of rat extraocular muscles does not depend on creatine kinase activity.

Background: Creatine kinase (CK) links phosphocreatine, an energy storage system, to cellular ATPases. CK activity serves as a temporal and spatial buffer for ATP content, particularly in fast-twitch skeletal muscles. The extraocular muscles are notoriously fast and active, suggesting the need for efficient ATP buffering.

Dimerisation of myomesin: implications for the structure of the sarcomeric M-band.

The sarcomeric M-band is thought to provide a link between the thick and the elastic filament systems. So far, relatively little is known about its structural components and their three-dimensional organisation. Myomesin seems to be an essential component of the M-band, since it is expressed in all types of vertebrate striated muscle fibres investigated and can be found in its mature localisation pattern as soon as the first myofibrils are assembled.

Muscle-type creatine kinase interacts with central domains of the M-band proteins myomesin and M-protein.

Muscle-type creatine kinase (MM-CK) is a member of the CK isoenzyme family with key functions in cellular energetics. MM-CK interacts in an isoform-specific manner with the M-band of sarcomeric muscle, where it serves as an efficient intramyofibrillar ATP-regenerating system for the actin-activated myosin ATPase located nearby on both sides of the M-band. Four MM-CK-specific and highly conserved lysine residues are thought to be responsible for the interaction of MM-CK with the M-band.

Paradoxical absence of M lines and downregulation of creatine kinase in mouse extraocular muscle.

The M lines are structural landmarks in striated muscles, necessary for sarcomeric stability and as anchoring sites for the M isoform of creatine kinase (CK-M). These structures, especially prominent in fast skeletal muscles, are missing in rodent extraocular muscle, a particularly fast and active muscle group. In this study, we tested the hypotheses that 1).