Impaired protein stability of 11beta-hydroxysteroid dehydrogenase type 2: a novel mechanism of apparent mineralocorticoid excess

Apparent mineralocorticoid excess (AME) is a severe form of hypertension that is caused by impaired activity of 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD2), which converts biologically active cortisol into inactive cortisone. Mutations in HSD11B2 result in cortisol-induced activation of mineralocorticoid receptors and cause hypertension with hypokalemia, metabolic alkalosis, and suppressed circulating renin and aldosterone concentrations. This study uncovered the first patient with AME who was described in the literature, identified the genetic defect in HSD11B2, and provided evidence for a novel mechanism of reduced 11beta-HSD2 activity. This study identified a cluster of amino acids (335 to 339) in the C-terminus of 11beta-HSD2 that are essential for protein stability. The cluster includes Tyr(338), which is mutated in the index patient, and Arg(335) and Arg(337), previously reported to be mutated in hypertensive patients. It was found that wild-type 11beta-HSD2 is a relatively stable enzyme with a half-life of 21 h, whereas that of Tyr(338)His and Arg(337)His was 3 and 4 h, respectively. Enzymatic activity of Tyr(338)His was partially retained at 26 degrees C or in the presence of the chemical chaperones glycerol and dexamethasone, indicating thermodynamic instability and misfolding. The results provide evidence that the degradation of both misfolded mutant Tyr(338)His and wild-type 11beta-HSD2 occurs through the proteasome pathway. Therefore, impaired 11beta-HSD2 protein stability rather than reduced gene expression or loss of catalytic activity seems to be responsible for the development of hypertension in some individuals with AME.

Conformational preferences of natural and C3-modified epothilones in aqueous solution

The conformational properties of the microtubule-stabilizing agent epothilone A ( 1a) and its 3-deoxy and 3-deoxy-2,3-didehydro derivatives 2 and 3 have been investigated in aqueous solution by a combination of NMR spectroscopic methods, Monte Carlo conformational searches, and NAMFIS calculations. The tubulin-bound conformation of epothilone A ( 1a), as previously proposed on the basis of solution NMR data, was found to represent a significant fraction of the ensemble of conformations present for the free ligands in aqueous solution.

Liquid–Vapor Interface of Formic Acid Solutions in Salt Water: A Comparison of Macroscopic Surface Tension and Microscopic in Situ X-ray Photoelectron Spectroscopy Measurements

The liquid–vapor interface is difficult to access experimentally but is of interest from a theoretical and applied point of view and has particular importance in atmospheric aerosol chemistry. Here we examine the liquid–vapor interface for mixtures of water, sodium chloride, and formic acid, an abundant chemical in the atmosphere. We compare the results of surface tension and X-ray photoelectron spectroscopy (XPS) measurements over a wide range of formic acid concentrations. Surface tension measurements provide a macroscopic characterization of solutions ranging from 0 to 3 M sodium chloride and from 0 to over 0.5 mole fraction formic acid. Sodium chloride was found to be a weak salting out agent for formic acid with surface excess depending only slightly on salt concentration. In situ XPS provides a complementary molecular level description about the liquid–vapor interface. XPS measurements over an experimental probe depth of 51 Å gave the C 1s to O 1s ratio for both total oxygen and oxygen from water. XPS also provides detailed electronic structure information that is inaccessible by surface tension. Density functional theory calculations were performed to understand the observed shift in C 1s binding energies to lower values with increasing formic acid concentration. Part of the experimental −0.2 eV shift can be assigned to the solution composition changing from predominantly monomers of formic acid to a combination of monomers and dimers; however, the lack of an appropriate reference to calibrate the absolute BE scale at high formic acid mole fraction complicates the interpretation. Our data are consistent with surface tension measurements yielding a significantly more surface sensitive measurement than XPS due to the relatively weak propensity of formic acid for the interface. A simple model allowed us to replicate the XPS results under the assumption that the surface excess was contained in the top four angstroms of solution.

Protein synthesis in jejunum and liver of neonatal calves fed vitamin A and lactoferrin

Rates of protein synthesis (PS) and turnover are more rapid during the neonatal period than during any other stage of postnatal life. Vitamin A and lactoferrin (Lf) can stimulate PS in neonates. However, newborn calves are vitamin A deficient and have a low Lf status, but plasma vitamin A and Lf levels increase rapidly after ingestion of colostrum. Neonatal calves (n = 6 per group) were fed colostrum or a milk-based formula without or with vitamin A, Lf, or vitamin A plus Lf to study PS in the jejunum and liver. l-[(13)C]Valine was intravenously administered to determine isotopic enrichment of free (nonprotein-bound) Val (AP(Free)) in the protein precursor pool, atom percentage excess (APE) of protein-bound Val, fractional protein synthesis rate (FSR) in the jejunum and liver, and isotopic enrichment of Val in plasma (APE(Pla)) and in the CO(2) of exhaled air (APE(Ex)). The APE, AP(Free), and FSR in the jejunum and liver did not differ significantly among groups. The APE(Ex) increased, whereas APE(Pla) decreased over time, but there were no group differences. Correlations were calculated between FSR(Jej) and histomorphometrical and histochemical data of the jejunum, and between FSR(Liv) and blood metabolites. There were negative correlations between FSR(Liv) and plasma albumin concentrations and between FSR(Jej) and the ratio of villus height:crypt depth, and there was a positive correlation between FSR(Jej) and small intestinal cell proliferation in crypts. Hence, there were no effects of vitamin A and Lf and no interactions between vitamin A and Lf on intestinal and hepatic PS. However, FSR(Jej) was correlated with histomorphometrical traits of the jejunum and FSR(Liv) was correlated with plasma albumin concentrations.

Mass spectrometric study of molecular and ionic sublimation of lanthanum triiodide

The molecular and ionic composition of saturated vapor over lanthanum triiodide was studied by Knudsen effusion mass spectrometry. The (LaI3)n molecules (n = 1–3) and the [I(LaI3)n]− ions (n = 0–4) were observed. The partial pressures of the molecules were determined and the enthalpies of sublimation, ΔsH° (298.15 K) in kJ mol−1, in the form of monomers (304 ± 7), dimers (428 ± 25), and trimers (455 ± 50) were obtained by the second and third laws of thermodynamics. The enthalpy of formation, ΔfH° (298.15 K) in kJ mol−1, of the LaI3 (−376 ± 10), La2I6 (−932 ± 25), La3I9 (−1585 ± 50) molecules and the LaI4− (−841 ± 24), La2I7− (−1486 ± 32) ions were determined. The electron work function, φe = 3.5 ± 0.3 eV, for the LaI3 crystal was calculated from the thermochemical cycle.

Rebound-associated vertebral fractures after discontinuation of denosumab-from clinic and biomechanics.

Rebound-associated vertebral fractures may follow treatment discontinuation of highly potent reversible bone antiresorptives, resulting from the synergy of rapid bone resorption and accelerated microdamage accumulation in trabecular bone. INTRODUCTION The purposes of this study are to characterize rebound-associated vertebral fractures following the discontinuation of a highly potent reversible antiresorptive therapy based on clinical observation and propose a pathophysiological rationale. METHODS This study is a case report of multiple vertebral fractures early after discontinuation of denosumab therapy in a patient with hormone receptor-positive non-metastatic breast cancer treated with an aromatase inhibitor. RESULTS Discontinuation of highly potent reversible bone antiresorptives such as denosumab may expose patients to an increased fracture risk due to the joined effects of absent microdamage repair during therapy followed by synchronous excess activation of multiple bone remodelling units at the time of loss-of-effect. We suggest the term rebound-associated vertebral fractures (RVF) for this phenomenon characterized by the presence of multiple new clinical vertebral fractures, associated with either no or low trauma, in a context consistent with the presence of high bone turnover and rapid loss of lumbar spine bone mineral density (BMD) occurring within 3 to 12 months after discontinuation (loss-of-effect) of a reversible antiresorptive therapy in the absence of secondary causes of bone loss or fractures. Unlike atypical femoral fractures that emerge from failure of microdamage repair in cortical bone with long-term antiresorptive treatment, RVF originate from the synergy of rapid bone resorption and accelerated microdamage accumulation in trabecular bone triggered by the discontinuation of highly potent reversible antiresorptives. CONCLUSIONS Studies are urgently needed to i) prove the underlying pathophysiological processes suggested above, ii) establish the predictive criteria exposing patients to an increased risk of RVF, and iii) determine appropriate treatment regimens to be applied in such patients.

The Contribution of the Activation Entropy to the Gas-Phase Stability of Modified Nucleic Acid Duplexes

Tricyclo-DNA (tcDNA) is a sugar-modified analogue of DNA currently tested for the treatment of Duchenne muscular dystrophy in an antisense approach. Tandem mass spectrometry plays a key role in modern medical diagnostics and has become a widespread technique for the structure elucidation and quantification of antisense oligonucleotides. Herein, mechanistic aspects of the fragmentation of tcDNA are discussed, which lay the basis for reliable sequencing and quantification of the antisense oligonucleotide. Excellent selectivity of tcDNA for complementary RNA is demonstrated in direct competition experiments. Moreover, the kinetic stability and fragmentation pattern of matched and mismatched tcDNA heteroduplexes were investigated and compared with non-modified DNA and RNA duplexes. Although the separation of the constituting strands is the entropy-favored fragmentation pathway of all nucleic acid duplexes, it was found to be only a minor pathway of tcDNA duplexes. The modified hybrid duplexes preferentially undergo neutral base loss and backbone cleavage. This difference is due to the low activation entropy for the strand dissociation of modified duplexes that arises from the conformational constraint of the tc-sugar-moiety. The low activation entropy results in a relatively high free activation enthalpy for the dissociation comparable to the free activation enthalpy of the alternative reaction pathway, the release of a nucleobase. The gas-phase behavior of tcDNA duplexes illustrates the impact of the activation entropy on the fragmentation kinetics and suggests that tandem mass spectrometric experiments are not suited to determine the relative stability of different types of nucleic acid duplexes.

Human monocytoid cells as a model to study Toll-like receptor-mediated activation

THP-1 2A9, a subclone of the monocytoid cell line THP-1 and known to be exquisitely sensitive to LPS, was tested for TNF production following triggering by excess doses of TLR ligands. TLR2, TLR4 and TLR5 agonists, but neither TLR3 nor TLR9 agonists, induced TNF production. When used at lower concentrations, priming by calcitriol strongly influenced the sensitivity of cells to LPS and different TLR2 triggers (lipoteichoic acid (LTA), trispalmitoyl-cysteyl-seryl-lysyl-lysyl-lysyl-lysine (Pam3Cys) and peptidoglycan (PGN)). Priming by calcitriol failed to modulate TLR2 and TLR4 mRNA and cell surface expression of these receptors. TNF signals elicited by TLR2 agonists were blocked by the TLR-specific antibody 2392. CD14-specific antibodies showed variable effects. CD14-specific antibodies inhibited TNF induction by LTA. High concentrations partially inhibited TNF induction by Pam3Cys. The same antibodies failed to inhibit TNF induction by PGN. Thus, THP-1 2A9 cells respond by TNF production to some, but not all TLR agonists, and the wide variety of putative TLR2 agonists interact to variable degrees also with other cell-surface-expressed binding sites such as CD14. THP-1 2A9 cells might provide a model by which to investigate in more detail the interaction of pathogen-associated molecular patterns and monocytoid cell-surface-expressed pattern recognition receptors.

[Urinary hormone analysis]

Urinary hormone analysis is applied to detect an altered steroid hormone metabolism, an elevated production of biogenic amines and to non-invasively determine the protein hormone human beta-choriogonadotropin indicating a pregnancy. Occasionally, these determinations need to be complemented by plasma- or serum hormone analysis. Clinical data including current drug therapy and urinary creatinine as reference are required to interpret any urine analysis. Diseases to be investigated by steroid hormone analysis are excess production of a typical or atypical mineralocorticoid active steroid hormones, the hormonal activity of adrenal or ovarian tumors, acne of unknown origin, hirsutism, a PCO-, an adrenogenital or a suspected Cushing syndrome. Biogenic amines should be determined in suspected secondary or refractory arterial hypertension, in case of pheochromocytoma- or paraganglioma-associated symptoms or if a serotonin-producing tumor is suspected. In children genetically determined diseases are the primary background to perform an analysis.

Introduction XXIX European Seminar in Ethnomusicology: Cultural Mapping and Musical Diversity

«Cultural mapping» has become a central keyword in the UNESCO strategy to protect world cultural and natural heritage. It can be described as a tool to increase the awareness of cultural diversity. As Crawhall (2009) pointed out, cultural mapping was initially considered to represent the «landscapes in two or three dimensions from the perspectives of indigenous and local peoples». It thus transforms the intangible cultural heritage to visible items by establishing profiles of cultures and communities, including music traditions. Cultural mapping is used as a resource for a variety of purposes as broad as peace building, adaptation to climate change, sustainability management, heritage debate and management, but can also become highly useful in the analysis of conflict points. Music plays a significant role in each of these aspects. This year’s symposium invites to highlight, yet also to critically reassess this topic from the following ethnomusicological perspectives: - The method of cultural mapping in ethnomusicology What approaches and research techniques have been used so far to establish musical maps in this context? What kinds of maps have been developed (and, for example, how far do these relate to indigenous mental maps that have only been transmitted orally)? How far do these modern approaches deviate from the earlier cultural mapping approaches of the cultural area approaches that were still evident with Alan P. Merriam and in Alan Lomax` Cantometrics? In how far are the methods of cultural mapping and of ethnomusicological fieldwork different and how can they benefit from each other? - Intangible cultural heritage and musical diversity As the 2003 UNESCO Convention for the Safeguarding of the Intangible Cultural Heritage pointed out in Article 12, each state signing the declaration «shall draw up, in a manner geared to its own situation, one or more inventories of the intangible cultural heritage, present in its territory and monitor these.» This symposium calls for a critical re-assessment of the hitherto established UNESCO intangible cultural heritage lists. The idea is to highlight the sensitive nature and the effects of the various heritage representations. «Heritage» is understood here as a selection from a selection – a small subset of history that relates to a given group of people in a particular place, at a specific time (Dann and Seaton 2001:26). This can include presentations of case studies, yet also a critical re-analysis of the selection process, e.g. who was included – or even excluded (and why)? Who were the decision makers? How can the role of ethnomusicology be described here? Where are the (existent and possible) conflict points (politically, socially, legally, etc.)? What kinds of solution strategies are available to us? How is the issue of diversity – that has been so strongly emphasized in the UNESCO declarations – reflected in the approaches? How might diversity be represented in future approaches? How does the selection process affect musical canonization (and exclusion)? What is the role of archives in this process? - Cultural landscape and music As defined by the World Heritage Committee, cultural landscapes can be understood as a distinct geographical area representing the «combined work of nature and man» (http://whc.unesco.org/en/culturallandscape/). This sub-topic calls for a more detailed – and general – exploration of the exact relation between nature/landscape (and definition of such) and music/sound. How exactly is landscape interrelated with music – and identified (and vice versa)? How is this interrelation being applied and exploited in a (inter-)national context?