Isoxazoline GPIIb/IIIa antagonists bearing a phosphoramidate

Isoxazolinylacetamides bearing a phosphoramidate group alpha- to the carboxylate moiety (3) were prepared and evaluated for in vitro antiplatelet efficacy. They were found to bind GPIIb/IIIa with high affinity and were potent antagonists of ADP mediated platelet aggregation.     

Multispectroscopic studies on the interaction of a copper(ii) complex of ibuprofen drug with calf thymus DNA

The interaction of copper(II)–ibuprofenato complex with calf thymus DNA (ct-DNA) has been explored following, UV-visible spectrophotometry, fluorescence measurement, dynamic viscosity measurements, and circular dichroism spectroscopy. In spectrophotometric studies of ct-DNA it was found that [Cu(ibp)₂]₂ can form a complex with double-helical DNA. The association constant of [Cu(ibp)₂]₂ with DNA from UV-Vis study was found to be 6.19 × 10⁴ L mol^?1. The values of K_f from fluorescence measurement clearly underscore the high affinity of [Cu(ibp)₂]₂ to DNA. The experimental results showed that the conformational changes in DNA helix induced by [Cu(ibp)₂]₂ are the reason for the fluorescence quenching of the DNA-Hoechst system. In addition, the fluorescence emission spectra of intercalated methylene blue (MB) with increasing concentrations of [Cu(ibp)₂]₂ represented a significant increase of MB intensity as to release MB from MB-DNA system. The results of circular dichroism (CD) suggested that copper(II)–ibuprofenato complex can change the conformation of DNA. In addition, the results of viscosity measurements suggest that copper(II)–ibuprofenato complex may bind with non-classical intercalative mode. From spectroscopic and hydrodynamic studies, it has been found that [Cu(ibp)₂]₂ interacts with DNA by partial intercalation mode which contains intercalation and groove properties.     

DNA binding restricts the intrinsic conformational flexibility of methyl CpG binding protein 2 (MeCP2)

Mass spectrometry-based hydrogen/deuterium exchange (H/DX) has been used to define the polypeptide backbone dynamics of full-length methyl CpG binding protein 2 (MeCP2) when free in solution and when bound to unmethylated and methylated DNA. Essentially the entire MeCP2 polypeptide chain underwent H/DX at rates faster than could be measured (i.e. complete exchange in ≤10 s), with the exception of the methyl DNA binding domain (MBD). Even the H/DX of the MBD was rapid compared with that of a typical globular protein. Thus, there is no single tertiary structure of MeCP2. Rather, the full-length protein rapidly samples many different conformations when free in solution. When MeCP2 binds to unmethylated DNA, H/DX is slowed several orders of magnitude throughout the MBD. Binding of MeCP2 to methylated DNA led to additional minor H/DX protection, and only locally within the N-terminal portion of the MBD. H/DX also was used to examine the structural dynamics of the isolated MBD carrying three frequent mutations associated with Rett syndrome. The effects of the mutations ranged from very little (R106W) to a substantial increase in conformational sampling (F155S). Our H/DX results have yielded fine resolution mapping of the structure of full-length MeCP2 in the absence and presence of DNA, provided a biochemical basis for understanding MeCP2 function in normal cells, and predicted potential approaches for the treatment of a subset of RTT cases caused by point mutations that destabilize the MBD.     

Salivary Inflammatory Mediator Profiling and Correlation to Clinical Disease Markers in Asthma

Rationale

There is a need for a readily available, non-invasive source of biomarkers that predict poor asthma control.

Objectives

We sought to determine if there is an association between the salivary inflammatory profile and disease control in children and adults with asthma.

Methods

In this cross-sectional study, we collected demographic and clinical information from two independent populations at different sites, resulting in convenience samples of 58 pediatric and 122 adult urban asthmatics. Control was assessed by symptom questionnaire (children) and by Asthma Control Questionnaire and current exacerbation (adults). Saliva was collected in all subjects. We applied principal component analysis to a 10-plex panel of relevant inflammatory markers to characterize marker profiles and determined if profiles were associated with asthma control.

Results

There were similar, strong correlations amongst biologically related markers in both populations: eosinophil-related: eotaxin-1/CCL11, RANTES/CCL5, and IL-5 (p<.001); myeloid/innate: IL-1β, IL-6, MCP-1/CCL2, and IL-8/CXCL8 (p<.001). The first three principal components captured ≥74% of variability across all ten analytes in both populations. In adults, the Principal Component 1 score, broadly reflective of all markers, but with greater weight given to myeloid/innate markers, was associated with Asthma Control Questionnaire score and exacerbation. The Principal Component 3 score, reflective of IP-10/CXCL10, was associated with current exacerbation. In children, the Principal Component 1, 2, and 3 scores were associated with recent asthma symptoms. The Principal Component 2 score, reflective of higher eosinophil markers, was inversely correlated with symptoms. The Principal Component 3 score was positively associated with all symptom outcomes.

Conclusion

The salivary inflammatory profile is associated with disease control in children and adults with asthma.     

Maximal Sum of Metabolic Exchange Fluxes Outperforms Biomass Yield as a Predictor of Growth Rate of Microorganisms

Growth rate has long been considered one of the most valuable phenotypes that can be measured in cells. Aside from being highly accessible and informative in laboratory cultures, maximal growth rate is often a prime determinant of cellular fitness, and predicting phenotypes that underlie fitness is key to both understanding and manipulating life. Despite this, current methods for predicting microbial fitness typically focus on yields [e.g., predictions of biomass yield using GEnome-scale metabolic Models (GEMs)] or notably require many empirical kinetic constants or substrate uptake rates, which render these methods ineffective in cases where fitness derives most directly from growth rate. Here we present a new method for predicting cellular growth rate, termed SUMEX, which does not require any empirical variables apart from a metabolic network (i.e., a GEM) and the growth medium. SUMEX is calculated by maximizing the SUM of molar EXchange fluxes (hence SUMEX) in a genome-scale metabolic model. SUMEX successfully predicts relative microbial growth rates across species, environments, and genetic conditions, outperforming traditional cellular objectives (most notably, the convention assuming biomass maximization). The success of SUMEX suggests that the ability of a cell to catabolize substrates and produce a strong proton gradient enables fast cell growth. Easily applicable heuristics for predicting growth rate, such as what we demonstrate with SUMEX, may contribute to numerous medical and biotechnological goals, ranging from the engineering of faster-growing industrial strains, modeling of mixed ecological communities, and the inhibition of cancer growth.     

Association between Decreased Klotho Blood Levels and Organic Growth Hormone Deficiency in Children with Growth Impairment

Objective

Klotho is an aging-modulating protein expressed mainly in the kidneys and choroid plexus, which can also be shed, released into the circulation and act as a hormone. Klotho deficient mice are smaller compared to their wild-type counterparts and their somatotropes show marked atrophy and reduced number of secretory granules. Recent data also indicated an association between klotho levels and growth hormone (GH) levels in acromegaly. We aimed to study the association between klotho levels and GH deficiency (GHD) in children with growth impairment.

Design

Prospective study comprising 99 children and adolescents (aged 9.0±3.7 years, 49 male) undergoing GH stimulation tests for short stature (height-SDS = −2.1±0.6). Klotho serum levels were measured using an α-klotho ELISA kit.

Results

Klotho levels were significantly lower (p<0.001) among children with organic GHD (n = 11, 727±273 pg/ml) compared to both GH sufficient participants (n = 59, 1497±754 pg/ml) and those with idiopathic GHD (n = 29, 1645±778 pg/ml). The difference between GHS children and children with idiopathic GHD was not significant. Klotho levels positively correlated with IGF-1- standard deviation scores (SDS) (R = 0.45, p<0.001), but were not associated with gender, pubertal status, age or anthropometric measurements.

Conclusions

We have shown, for the first time, an association between low serum klotho levels and organic GHD. If validated by additional studies, serum klotho may serve as novel biomarker of organic GHD.     

Development of a mathematical model that predicts optimal muscle activation patterns by using brief trains.

Because muscles must be repetitively activated during functional electrical stimulation, it is desirable to identify the stimulation pattern that produces the most force. Previous experimental work has shown that the optimal pattern contains an initial high-frequency burst of pulses (i.e., an initial doublet or triplet) followed by a low, constant-frequency portion. Pattern optimization is particularly challenging, because a muscle's contractile characteristics and, therefore, the optimal pattern change under different physiological conditions and are different for each person. This work describes the continued development and testing of a mathematical model that predicts isometric forces from fresh and fatigued muscles in response to brief trains of electrical pulses. By use of this model and an optimization algorithm, stimulation patterns that produced maximum forces from each subject were identified.