Telangiectasia as a presenting sign of Graves' disease

Acquired nevoid telangiectasia (ANT) is observed in several conditions including primary cutaneous disorders, systemic autoimmune disease and hyperestrogenism occurring in puberty, pregnancy and chronic liver disease. We describe a patient in whom ANT was a presenting sign of autoimmune hyperthyroidism, which improved after thyroidectomy. A 43-year-old Caucasian woman experienced an asymptomatic development of multiple widespread red skin lesions, diagnosed to be ANT. Blood tests revealed increased serum levels of free tri-iodothyronine and thyroxine and suppressed thyroid-stimulating hormone. Other causes of ANT were excluded. ANT improved but did not disappear after thyroidectomy. The possible pathogenetic factors linking ANT and Graves' disease, such as an immune-mediated process, altered estrogen metabolism or vasodilatation associated with hyperdynamic circulation, are discussed.     

Investigating the mechanism of peptide aggregation: insights from mixed monte carlo-molecular dynamics simulations

The early stages of peptide aggregation are currently not accessible by experimental techniques at atomic resolution. In this article, we address this problem through the application of a mixed simulation scheme in which a preliminary coarse-grained Monte Carlo analysis of the free-energy landscape is used to identify representative conformations of the aggregates and subsequent all-atom molecular dynamics simulations are used to analyze in detail possible pathways for the stabilization of oligomers. This protocol was applied to systems consisting of multiple copies of the model peptide GNNQQNY, whose detailed structures in the aggregated state have been recently solved in another study. The analysis of the various trajectories provides dynamical and structural insight into the details of aggregation. In particular, the simulations suggest a hierarchical mechanism characterized by the initial formation of stable parallel β-sheet dimers and identify the formation of the polar zipper motif as a fundamental feature for the stabilization of initial oligomers. Simulation results are consistent with experimentally derived observations and provide an atomically detailed view of the putative initial stages of fibril formation.     

The ubiquitin-proteasome system in cardiac dysfunction

Since proteins play crucial roles in all biological processes, the finely tuned equilibrium between their synthesis and degradation regulates cellular homeostasis. Controlling the quality of proteome informational content is essential for cell survival and function. After initial synthesis, membrane and secretory proteins are modified, folded, and assembled in the endoplasmic reticulum, whereas other proteins are synthesized and processed in the cytosol. Cells have different protein quality control systems, the molecular chaperones, which help protein folding and stabilization, and the ubiquitin-proteasome system (UPS) and lysosomes, which degrade proteins. It has generally been assumed that UPS and lysosomes are regulated independently and serve distinct functions. The UPS degrades both cytosolic, nuclear proteins, and myofibrillar proteins, whereas the lysosomes degrade most membrane and extracellular proteins by endocytosis as well as cytosolic proteins and organelles via autophagy. Over the last two decades, the UPS has been increasingly recognized as a major system in several biological processes including cell proliferation, adaptation to stress and cell death. More recently, activation or impairment of the UPS has been reported in cardiac disease and recent evidence indicate that autophagy is a key mechanism to maintain cardiac structure and function. This review mainly focuses on the UPS and its various components in healthy and diseased heart, but also summarizes recent data suggesting parallel activation of the UPS and autophagy in cardiac disease.     

Novel integrated microdialysis-amperometric system for in vitro detection of dopamine secreted from PC12 cells: design, construction, and validation

A novel dual channel in vitro apparatus, derived from a previously described design, has been coupled with dopamine (DA) microsensors for the flow-through detection of DA secreted from PC12 cells. The device, including two independent microdialysis capillaries, was loaded with a solution containing PC12 cells while a constant phosphate-buffered saline (PBS) medium perfusion was carried out using a dual channel miniaturized peristaltic pump. One capillary was perfused with normal PBS, whereas extracellular calcium was removed from extracellular fluid of the second capillary. After a first period of stabilization and DA baseline recording, KCl (75 mM) was added to the perfusion fluid of both capillaries. In this manner, a simultaneous “treatment–control” experimental design was performed to detect K⁺-evoked calcium-dependent DA secretion. For this purpose, self-referencing DA microsensors were developed, and procedures for making, testing, and calibrating them are described in detail. The electronic circuitry was derived from previously published schematics and optimized for dual sensor constant potential amperometry applications. The microdialysis system was tested and validated in vitro under different experimental conditions, and DA secretion was confirmed by high-performance liquid chromatography with electrochemical detection (HPLC–EC). PC12 cell viability was quantified before and after each experiment. The proposed apparatus serves as a reliable model for studying the effects of different drugs on DA secretion through the direct comparison of extracellular DA increase in treatment–control experiments performed on the same initial PC12 cell population.     

Chaperonins: The hunt for the Group II mechanism

Chaperonins are multi-subunit complexes that enhance the efficiency of protein-folding reactions by capturing protein substrates in their central cavities. They occur in all prokaryotic and eukaryotic cell types and, alone amongst molecular chaperones, chaperonin knockouts are always lethal. Chaperonins come in two forms; the Group I are found in bacteria, mitochondria and plastids [W.A. Fenton, A.L. Horwich, Q. Rev. Biophys. 36 (2003) 229-256, [1]] and the Group II in the eukaryotic cytoplasm and in archaea [N.J. Cowan, S.A. Lewis, Adv. Protein Chem. 59 (2001) 73-104, [2]]. Both use energy derived from ATP binding and hydrolysis to drive a series of structural rearrangements that enable them to capture, engulf and then release polypeptide chains that have either not yet acquired the native, biologically active state or have been denatured in the cell.     

Molecular characteristics of small intestinal and renal brush border thiamin transporters in rats.

The molecular characteristics of thiamin (T) transport were studied in the small intestinal and renal brush border membrane vesicles of rats, using [(3)H]T at high specific activity. The effects of various chemical modifiers (amino acid blockers) on T uptake were examined and their specificity assessed. Treatment with the carboxylic specific blockers 1-cyclohexyl-3-(2-morpholinoethyl) carbodiimide metho-p-toluene sulfonate, (1-ethyl-3-[3-(dimethylamino)propyl]-carbodiimide hydrochloride and N-ethyl-5-phenylisoaxolium-3'-sulfonate (Woodward's Reagent K) and with the sulfhydryl specific blocker p-chloromercuribenzene sulfonate inhibited T transport in both types of vesicles. Phenylglyoxal, but not ninhydrin, both reagents for arginine residues, and diethylpyrocarbonate, a reagent for histidine residues, specifically decreased T transport only in renal and small intestinal vesicles respectively. Similarly 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole reacted, but not N-acetylimidazole, both of which are reagents for tyrosine residues. However, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole inhibition was aspecific. Acetylsalicylic acid, a reagent for lysine and serine residues, decreased T transport, but the lysine effect was aspecific. Acetylsalicylic acid serine blockage also eliminated T/H(+) exchange in small intestinal vesicles. Taken together, these results suggest that for T transport carboxylic and sulfhydryl groups and serine residues are essential in both renal and small intestinal brush border membrane vesicles. In addition, arginine and histidine residues are also essential respectively for renal and small intestinal transporters. Serine was essential for the T/H(+) antiport mechanism.     

GarA is an essential regulator of metabolism in Mycobacterium tuberculosis

Alpha‐ketoglutarate is a key metabolic intermediate at the crossroads of carbon and nitrogen metabolism, whose fate is tightly regulated. In mycobacteria the protein GarA regulates the tricarboxylic acid cycle and glutamate synthesis by direct binding and regulation of three enzymes that use α‐ketoglutarate. GarA, in turn, is thought to be regulated via phosphorylation by protein kinase G and other kinases. We have investigated the requirement for GarA for metabolic regulation during growth in vitro and in macrophages. GarA was found to be essential to Mycobacterium tuberculosis, but dispensable in non‐pathogenic Mycobacterium smegmatis. Disruption of garA caused a distinctive, nutrient‐dependent phenotype, fitting with its proposed role in regulating glutamate metabolism. The data underline the importance of the TCA cycle and the balance with glutamate synthesis in M. tuberculosis and reveal vulnerability to disruption of these pathways.