Identification of Potential Muscle Biomarkers in McArdle Disease: Insights from Muscle Proteome Analysis

Glycogen storage disease type V (GSDV, McArdle disease) is a rare genetic myopathy caused by deficiency of the muscle isoform of glycogen phosphorylase (PYGM). This results in a block in the use of muscle glycogen as an energetic substrate, with subsequent exercise intolerance. The pathobiology of GSDV is still not fully understood, especially with regard to some features such as persistent muscle damage (i.e., even without prior exercise). We aimed at identifying potential muscle protein biomarkers of GSDV by analyzing the muscle proteome and the molecular networks associated with muscle dysfunction in these patients. Muscle biopsies from eight patients and eight healthy controls showing none of the features of McArdle disease, such as frequent contractures and persistent muscle damage, were studied by quantitative protein expression using isobaric tags for relative and absolute quantitation (iTRAQ) followed by artificial neuronal networks (ANNs) and topology analysis. Protein candidate validation was performed by Western blot. Several proteins predominantly involved in the process of muscle contraction and/or calcium homeostasis, such as myosin, sarcoplasmic/endoplasmic reticulum calcium ATPase 1, tropomyosin alpha-1 chain, troponin isoforms, and alpha-actinin-3, showed significantly lower expression levels in the muscle of GSDV patients. These proteins could be potential biomarkers of the persistent muscle damage in the absence of prior exertion reported in GSDV patients. Further studies are needed to elucidate the molecular mechanisms by which PYGM controls the expression of these proteins.     

Accumulation of silver nanoparticles by cultured primary brain astrocytes

Silver nanoparticles (AgNP) are components of various food industry products and are frequently used for medical equipment and materials. Although such particles enter the vertebrate brain, little is known on their biocompatibility for brain cells. To study the consequences of an AgNP exposure of brain cells we have treated astrocyte-rich primary cultures with polyvinylpyrrolidone (PVP)-coated AgNP. The incubation of cultured astrocytes with micromolar concentrations of AgNP for up to 24 h resulted in a time- and concentration-dependent accumulation of silver, but did not compromise the cell viability nor lower the cellular glutathione content. In contrast, the incubation of astrocytes for 4 h with identical amounts of silver as AgNO(3) already severely compromised the cell viability and completely deprived the cells of glutathione. The accumulation of AgNP by astrocytes was proportional to the concentration of AgNP applied and significantly lowered by about 30% in the presence of the endocytosis inhibitors chloroquine or amiloride. Incubation at 4?°C reduced the accumulation of AgNP by 80% compared to the values obtained for cells that had been exposed to AgNP at 37?°C. These data demonstrate that viable cultured brain astrocytes efficiently accumulate PVP-coated AgNP in a temperature-dependent process that most likely involves endocytotic pathways.     

Spray drying formulation of albendazole microspheres by experimental design. In vitro–in vivo studies

Both an experimental design and optimization techniques were carried out for the development of chitosan–pectin–carboxymethylcellulose microspheres to improve the oral absorption of albendazole as a model drug. The effect of three different factors (chitosan, pectin and carboxy methyl cellulose concentrations) was studied on five responses: yield, morphology, dissolution rate at 30 and 60?min, and encapsulation efficiency of the microspheres. During the screening phase, the factors were evaluated in order to identify those which exert a significant effect. Simultaneous multiple response optimizations were then used to find out experimental conditions where the system shows the most adequate results. The optimal conditions were found to be: chitosan concentration, 1.00% w/v, pectin concentration 0.10% w/v and carboxymethylcellulose concentration 0.20% w/v. The bioavailability of the loaded drug in the optimized microspheres was evaluated in Wistar rats which showed an area under curve (AUC) almost 10 times higher than the pure drug.     

Further utilisation of 2-naphthol esters: polarographic determination of phosphatase and arylsulphatase activities

Summary The Polarographic method of the determination of alkaline and acid phosphatases and arylsulphatases using esters of 2-naphthol has been elaborated. The enzymatically released 2-naphthol is nitrosated and the nitrosation product (1-nitroso-2-naphthol) is reduced on the mercury drop-electrode with the production of a well-developed four-electron wave. The method was applied for the determination of the enzymatic activity of tissue homogenates and commercial enzyme preparations.

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Weitere Anwendbarkeit von 2-Naphthol-Estern : Polarographische Bestimmung der Aktivität von Phosphatase und Arvlsulphatase

Zusammenfassung Es wurde eine Methode zur polarographischen Bestimmung der Aktivität der alkalischen und sauren Phosphatasen und Arylsulphatasen bei Benützung der 2-Naphthol-Ester ausgearbeitet. Das durch enzymatische Hydrolyse freigesetzte 2-Naphthol wird nitrosiert und das Nitrosierungsprodukt (1-Nitroso-2-Naphthol) auf der Quecksilber-Tropfelektrode bei Entstehung einer gut entwickelten Welle reduziert. Diese Methode wurde bei Bestimmung der enzymatischen Aktivität in Gewebehomogenaten wie auch in enzymatischen Handelspräparaten verwendet.

     

Targeting the Surface of the Protein 14-3-3 by Ultrasmall (1.5 nm) Gold Nanoparticles Carrying the Specific Peptide CRaf

The surface of ultrasmall gold nanoparticles with an average diameter of 1.55 nm was conjugated with a 14-3-3 protein-binding peptide derived from CRaf. Each particle carries 18 CRaf peptides, leading to an overall stoichiometry of Au(115)Craf(18). The binding to the protein 14-3-3 was probed by isothermal titration calorimetry (ITC) and fluorescence polarization spectroscopy (FP). The dissociation constant (K_D) was measured as 5.0 μM by ITC and 0.9 μM by FP, which was close to the affinity of dissolved CRaf to 14-3-3σ. In contrast to dissolved CRaf, which alone did not enter HeLa cells, CRAF-conjugated gold nanoparticles were well taken up by HeLa cells, opening the opportunity to target the protein inside a cell.     

Integrated capacity planning over highly volatile horizons

Today production planning has to deal with highly dynamic markets and increasing uncertainties. Moreover, it has to take into account possibilities of the surrounding production network. By combining a queueing theory model with a stochastic, dynamic optimization approach, a method to support decision making in production planning was developed. Hereby, a Markovian Decision Process is solved to find cost minimal policies as reactions to volatile market demands for minimizing costs due to capacity adaptations, changes in process steps, and locations. The method was applied at an automotive supplier to find suitable system configurations and investment decisions for an uncertain future.     

Simulation of personnel requirements during production ramp-up

Nowadays the number of product models constantly increases, due to a strong competition to innovate and the reduction of product life cycles, resulting in a more frequent occurrence of production ramp-ups. The production ramp-up combines the product development with the series production and has an impact on the market entry timing. In order to ensure an ideal ramp-up with new developed products being placed on the market as quickly as possible, there has to be an efficient use of existing resources, especially of human resources. Nevertheless, the planning of these resources turns out to be the most important challenge because it is directly related to a great uncertainty. The following article represents an approach that is being developed at the Institute of Production Science (wbk) of the Karlsruhe Institute of Technology (KIT) to optimize the forecast of the personnel requirements during ramp-up by taking into account the dynamic planning variables and organizational basic conditions. Therefore, a method will be developed that provides support to the responsible persons of the respective production ramp-up to calculate the necessary manpower for every single ramp-up phase and to realize the economic optimum. This method focuses on a simulation of the ramp-up process that allows an economically more efficient use of human resources.