Data mining-based statistical analysis of biological data uncovers hidden significance: clustering Hashimoto’s thyroiditis patients based on the response of their PBMC with IL-2 and IFN-γ secretion to stimulation with Hsp60

The pathogenesis of Hashimoto’s thyroiditis includes autoimmunity involving thyroid antigens, autoantibodies, and possibly cytokines. It is unclear what role plays Hsp60, but our recent data indicate that it may contribute to pathogenesis as an autoantigen. Its role in the induction of cytokine production, pro- or anti-inflammatory, was not elucidated, except that we found that peripheral blood mononucleated cells (PBMC) from patients or from healthy controls did not respond with cytokine production upon stimulation by Hsp60 in vitro with patterns that would differentiate patients from controls with statistical significance. This “negative” outcome appeared when the data were pooled and analyzed with conventional statistical methods. We re-analyzed our data with non-conventional statistical methods based on data mining using the classification and regression tree learning algorithm and clustering methodology. The results indicate that by focusing on IFN-γ and IL-2 levels before and after Hsp60 stimulation of PBMC in each patient, it is possible to differentiate patients from controls. A major general conclusion is that when trying to identify disease markers such as levels of cytokines and Hsp60, reference to standards obtained from pooled data from many patients may be misleading. The chosen biomarker, e.g., production of IFN-γ and IL-2 by PBMC upon stimulation with Hsp60, must be assessed before and after stimulation and the results compared within each patient and analyzed with conventional and data mining statistical methods.     

Clozapine impairs insulin action by up-regulating Akt phosphorylation and Ped/Pea-15 protein abundance

Clinical and experimental evidence indicates that atypical antipsychotics impair glucose metabolism. We investigated whether clozapine may directly affect insulin action by analyzing insulin signaling in vitro and in vivo. Clozapine reduced insulin-stimulated glucose uptake in PC12 and in L6 cells, representative models of neuron and skeletal muscle, respectively. Consistently, clozapine reduced insulin effect on insulin receptor (IR) by 40% and on IR substrate-1 (IRS1) tyrosine phosphorylation by 60%. Insulin-stimulated Akt phosphorylation was also reduced by about 40%. Moreover, insulin-dependent phosphorylation of protein kinase C-ζ (PKC-ζ) was completely blunted in clozapine-treated cells. Interestingly, clozapine treatment was accompanied by an insulin-independent increase of Akt phosphorylation, with no change of IR, IRS1, and PKC-ζ basal phosphorylation. The cellular abundance of Ped/Pea-15, an Akt substrate and inducer of insulin resistance, was also increased following clozapine exposure, both in the absence and in the presence of cyclohexymide, a protein synthesis inhibitor. Similar as in cellular models, in the caudate-putamen and in the tibialis muscle of clozapine-treated C57/BL/KsJ mice, Akt phosphorylation and Ped/Pea-15 protein levels were increased and PKC-ζ phosphorylation was decreased. Thus, in these experimental models, clozapine deranged Akt function and up-regulated Ped/Pea-15, thereby inhibiting insulin stimulation of PKC-ζ and of glucose uptake.     

Wireless GPS-based phase-locked synchronization system for outdoor environment

Synchronization of data coming from different sources is of high importance in biomechanics to ensure reliable analyses. This synchronization can either be performed through hardware to obtain perfect matching of data, or post-processed digitally. Hardware synchronization can be achieved using trigger cables connecting different devices in many situations; however, this is often impractical, and sometimes impossible in outdoors situations. The aim of this paper is to describe a wireless system for outdoor use, allowing synchronization of different types of - potentially embedded and moving - devices. In this system, each synchronization device is composed of: (i) a GPS receiver (used as time reference), (ii) a radio transmitter, and (iii) a microcontroller. These components are used to provide synchronized trigger signals at the desired frequency to the measurement device connected. The synchronization devices communicate wirelessly, are very lightweight, battery-operated and thus very easy to set up. They are adaptable to every measurement device equipped with either trigger input or recording channel. The accuracy of the system was validated using an oscilloscope. The mean synchronization error was found to be 0.39 μs and pulses are generated with an accuracy of <2 μs. The system provides synchronization accuracy about two orders of magnitude better than commonly used post-processing methods, and does not suffer from any drift in trigger generation.     

Lead optimization of 2-(piperidin-3-yl)-1H-benzimidazoles: identification of 2-morpholin- and 2-thiomorpholin-2-yl-1H-benzimidazoles as selective and CNS penetrating H₁-antihistamines for insomnia

The structure-activity relationships of 2-(piperidin-3-yl)-1H-benzimidazoles, 2-morpholine and 2-thiomorpholin-2-yl-1H-benzimidazoles are described. In the lead optimization process, the pK(a) and/or logP of benzimidazole analogs were reduced either by attachment of polar substituents to the piperidine nitrogen or incorporation of heteroatoms into the piperidine heterocycle. Compounds 9a and 9b in the morpholine series and 10g in the thiomorpholine series demonstrated improved selectivity and CNS profiles compared to lead compound 2 and these are potential candidates for evaluation as sedative hypnotics.     

Might the observed α2A-adrenoreceptor agonism or antagonism of allyphenyline analogues be ascribed to different molecular conformations?

We recently reported that the α(2)-adrenoreceptor (AR) ligand allyphenyline (9) significantly enhanced morphine analgesia (due to its α(2C)-AR agonism), was devoid of sedative side effects (due to its α(2A)-AR antagonism), prevented and reversed morphine tolerance and dependence. To highlight the molecular characteristics compatible with this behaviour and to obtain novel agents potentially useful in chronic pain and opioid addiction management, the allyl group of 9 was replaced by substituents of moderate steric bulk (MR) and positive or negative lipophilic (π) and electronic (σ) contributions in all the possible combinations. Effective novel α(2C)-agonists/α(2A)-antagonists (2, 3, 10, 12, and 17) were obtained. This study also demonstrated that contradictory combinations of the physicochemical parameters were similarly able to induce the α(2A)-activation. Since we had previously observed that the absolute configuration affected only the potency, but not the functional profile of the ligands, we hypothesized that the α(2A)-activation was governed by a ligand preferred conformation. From a structural overlay investigation it emerged that an extended conformation appeared to be associated with dual α(2C)-agonism/α(2A)-antagonism, whereas a folded conformation associated with α(2C)-/α(2A)-agonism.     

Stability,oviposition attraction,and larvicidal activity of binary toxin from <Emphasis Type="Italic">Bacillus sphaericus</Emphasis> expressed in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Bacillus sphaericus produces a two-chain binary toxin composed of BinA (42 kDa) and BinB (51 kDa), which are deposited as parasporal crystals during sporulation. The toxin is highly active against Culex larvae and Aedes and Anopheles mosquitoes, which are the principal vectors for the transmission of malaria, yellow fever, encephalitis, and dengue. The use of B. sphaericus and Bacillus thuringiensis in mosquito control programs is limited by their sedimentation in still water. In this study, the binA and binB genes were cloned and the recombinant BinAB protein was expressed in three strains of Escherichia coli. These recombinant strains were used in a toxicity assay against Culex quinquefasciatus larvae. The highest expression level was achieved when both proteins were expressed in a single operon construct. The BinAB protein expressed in the E. coli Arctic strain showed higher larvicidal activity than either of the recombinant proteins from the E. coli Ril or pLysS strains. Furthermore, it had the highest oviposition attraction (49.1%, P?相似文献   

Virulence factor prediction in <Emphasis Type="Italic">Streptococcus pyogenes</Emphasis> using classification and clustering based on microarray data

Interesting biological information as, for example, gene expression data (microarrays), can be extracted from publicly available genomic data. As a starting point in order to narrow down the great possibilities of wet lab experiments, global high throughput data and available knowledge should be used to infer biological knowledge and emit biological hypothesis. Here, based on microarray data, we propose the use of cluster and classification methods that have become very popular and are implemented in freely available software in order to predict the participation in virulence mechanisms of different proteins coded by genes of the pathogen Streptococcus pyogenes. Confidence of predictions is based on classification errors of known genes and repetitive prediction by more than three methods. A special emphasis is done on the nonlinear kernel classification methods used. We propose a list of interesting candidates that could be virulence factors or that participate in the virulence process of S. pyogenes. Biological validations should start using this list of candidates as they show similar behavior to known virulence factors.     

The molecular anatomy of human Hsp60 and its similarity with that of bacterial orthologs and acetylcholine receptor reveal a potential pathogenetic role of anti-chaperonin immunity in myasthenia gravis

Heat-shock protein 60 (Hsp60) is ubiquitous and highly conserved being present in eukaryotes and prokaryotes, including pathogens. This chaperonin, although typically a mitochondrial protein, can also be found in other intracellular sites, extracellularly, and in circulation. Thus, it can signal the immune system and participate in the development of inflammation and immune reactions. Both phenomena can be elicited by human and foreign Hsp60 (e.g., bacterial GroEL), when released into the blood by infectious agents. Consequently, all these Hsp60 proteins become part of a complex autoimmune response characterized by multiple cross reactions because of their structural similarities. In this study, we demonstrate that Hsp60 proteins from humans and two common pathogens, Chlamydia trachomatis and Chlamydia pneumoniae, share various sequence segments of potentially highly immunogenic epitopes with acetylcholine receptor α1 subunit (AChRα1). The structural data indicate that AChRα1 antibodies, implicated in the pathogenesis of myasthenia gravis, could very well be elicited and/or maintained by self- and/or bacterial Hsp60.     

In vitro characteristics of 1-phenyl-3-methyl-4-acylpyrazol-5-ones iron chelators

Iron chelators represent a group of structurally different compounds sharing the ability of iron binding. The group has been evolving in recent years mainly due to novel experimental indications associated with variable requirements for iron chelators. A group of synthetic 1-phenyl-3-methyl-4-acyl-pyrazol-5-ones has been known for many years but data on their potential biological activity are rather limited.In this study, we analysed a series of these compounds for their iron-chelating properties as well as for their effects on iron based Fenton chemistry. For the former ferrozine spectrophotometric method and for the latter HPLC method with salicylic acid were used.All of the tested compounds were very efficient ferric chelators but their ferrous-chelating effects differed according to the acyl substitution. Notwithstanding various ferrous chelation activities, the individual Fe²⁺-affinities were not significantly different through pathophysiologically relevant pH conditions and some of the tested substances were more potent ferrous chelators at pH 4.5 than clinically used standard deferoxamine. Of particular interest is H₂QpyQ /2,6-bis[4(1-phenyl-3-methylpyrazol-5-one)carbonyl]pyridine/ which iron-chelating affinity increased when pH was decreasing. In spite of ferrous chelation differences, most of the tested acylpyrazolones were similarly active powerful inhibitors of Fenton chemistry as deferoxamine.Conclusively, acylpyrazolones are efficient iron chelators and H₂QpyQ may represent a prototype of novel specific chelators designated particularly for chelation at acidic conditions.