Serum resistin and polymorphisms of androgen receptor GAGn and GGNn and aromatase TTTAn

There is evidence that androgens are regulators of insulin resistance (IR), and may be involved in the regulation of resistin, a cytokine that has been related with IR. Earlier studies found that androgen receptor length polymorphisms CAGn and GGNn and the aromatase polymorphism TTTAn may influence receptor or enzyme activity and serum concentrations of androgens. This study was designed to determine whether polymorphism length was related to serum resistin concentration and to other variables related with IR. In 1,580 persons chosen randomly from the general population of the Canary Islands (Spain), we measured polymorphism length, waist circumference, waist/hip ratio, BMI, and serum glucose concentration. In smaller subgroups, we also measured C-peptide (n = 677), resistin (n = 583), and leptin concentration (n = 754) and estimated IR (homeostasis model assessment-IR (HOMA2-IR)). In men, polymorphism length correlated with resistin concentration (CAGn, r = 0.13, P = 0.031; TTTAn, r = 0.15, P = 0.005; GGNn, r = -0.15, P = 0.026), and the correlations were confirmed in multivariate regression models. The length of CAGn and TTTAn correlated inversely with C-peptide (r = -0.13, P = 0.016 and r = -0.21, P < 0.001, respectively) and with estimated IR (r = -0.12, P = 0.032 and r = -0.19, P = 0.001, respectively). In men, length of the CAGn, GGNn, and TTTAn was associated with serum resistin concentration. These results support the hypothesis that androgens may be involved in the regulation of resistin. Resistin may be a link between IR and androgens.     

Accuracy of immunodiagnostic tests for active tuberculosis using single and combined results: a multicenter TBNET-Study

Background

The clinical application of IFN-γ release assays (IGRAs) has recently improved the diagnosis of latent tuberculosis infection. In a multicenter study of the Tuberculosis Network European Trialsgroup (TBNET) we aimed to ascertain in routine clinical practice the accuracy of a novel assay using selected peptides encoded in the mycobacterial genomic region of difference (RD) 1 for the diagnosis of active tuberculosis in comparison with tuberculin skin test (TST), QuantiFERON-TB GOLD In-Tube (Cellestis Ltd., Carnegie, Australia) and T-SPOT.TB (Oxfordimmunotec, Abingdon, UK).

Principal Findings

425 individuals from 6 different European centres were prospectively enrolled. We found that sensitivity of the novel test, TST, QuantiFERON-TB GOLD In-Tube and T-SPOT.TB was respectively 73.1%, 85.3%, 78.1%, and 85.2%; specificity was respectively 70.6%, 48.0%, 61.9% and 44.3%; positive likelihood ratios were respectively 2.48, 1.64, 2.05, and 1.53; negative likelihood ratios were respectively 0.38, 0.31, 0.35, 0.33. Sensitivity of TST combined with the novel test, QuantiFERON-TB GOLD In-Tube and T-SPOT.TB increased up to 92.4%, 97.7% and 97.1%, respectively. The likelihood ratios of combined negative results of TST with, respectively, the novel test, QuantiFERON-TB GOLD In-Tube and T-SPOT.TB were 0.19, 0.07 and 0.10.

Conclusions

The assay based on RD1 selected peptides has similar accuracy for active tuberculosis compared with TST and commercial IGRAs. Then, independently of the spectrum of antigens used in the assays to elicit mycobacterial specific immune responses, the novel test, IGRAs, and the TST do not allow an accurate identification of active tuberculosis in clinical practice. However, the combined use of the novel assay or commercial IGRAs with TST may allow exclusion of tuberculosis.     

Centrosome dysfunction in Drosophila neural stem cells causes tumors that are not due to genome instability

Genome instability (GI) and centrosomal alterations are common traits in human cancer [1, 2]. It is suspected that centrosome dysfunction may cause tumors by bringing about GI, but direct experimental proof is still lacking [3]. To explore the possible functional link between centrosome function and overgrowth, we have assayed the tumorigenic potential of a series of mutants that affect different centrosomal proteins in Drosophila. We have found that a significant number of such mutant conditions are tumorigenic in larval brain tissue, where self-renewing asymmetric division of neural stem cells is frequent, but not in symmetrically dividing epithelial cells. We have also found that mutations that increase GI without causing centrosome dysfunction are not tumorigenic in our assay. From these observations, we conclude that the tumors caused by centrosome dysfunction cannot be explained solely by the resulting genome instability. We propose that such tumors might be caused by impaired asymmetric division of neural stem cells [4]. These results show that centrosome loss, far from being innocuous, is a potentially dangerous condition in flies.     

Mucin secretion by the human colon cell line LS174T is regulated by bile salts

We recently reported that bile salts play a role in the regulation of mucin secretion by cultured dog gallbladder epithelial cells. In this study we have examined whether bile salts also influence mucin secretion by the human epithelial colon cell line LS174T. Solutions of bile salts were applied to monolayers of LS174T cells. Mucin secretion was quantified by measuring the secretion of [3H]GlcNAc labeled glycoproteins. Both unconjugated bile salts as well as taurine conjugated bile salts stimulated mucin secretion by the colon cells in a dose-dependent fashion. Hydrophobic bile salts were more potent stimulators than hydrophilic bile salts. Free (unconjugated) bile salts were more stimulatory compared with their taurine conjugated counterparts. Stimulation of mucin secretion by LS174T cells was found to occur at much lower bile salt concentrations than in the experiments with the dog gallbladder epithelial cells. The protein kinase C activators PMA and PDB had no stimulatory effect on mucin secretion. We conclude that mucin secretion by the human colon epithelial cell line LS174T is regulated by bile salts. We suggest that regulation of mucin secretion by bile salts might be a common mechanism, by which different epithelia protect themselves against the detergent action of bile salts, to which they are exposed throughout the gastrointestinal tract.      

Dynamics and metal exchange properties of C4C4 RING domains from CNOT4 and the p44 subunit of TFIIH

Zinc fingers are small structured protein domains that require the coordination of zinc for a stable tertiary fold. Together with FYVE and PHD, the RING domain forms a distinct class of zinc-binding domains, where two zinc ions are ligated in a cross-braced manner, with the first and third pairs of ligands coordinating one zinc ion, while the second and fourth pairs ligate the other zinc ion. To investigate the relationship between the stability and dynamic behaviour of the domains and the stability of the metal-binding site, we studied metal exchange for the C4C4 RING domains of CNOT4 and the p44 subunit of TFIIH. We found that Zn(2+)-Cd(2+) exchange is different between the two metal-binding sites in the C4C4 RING domains of the two proteins. In order to understand the origins of these distinct exchange rates, we studied the backbone dynamics of both domains in the presence of zinc and of cadmium by NMR spectroscopy. The differential stability of the two metal-binding sites in the RING domains, as reflected by the different metal exchange rates, can be explained by a combination of accessibility and an electrostatic ion interaction model. A greater backbone flexibility for the p44 RING domain as compared to CNOT4 may be related to the distinct types of protein-protein interactions in which the two C4C4 RING domains are involved.     

Crystal structure of monomeric actin in the ATP state. Structural basis of nucleotide-dependent actin dynamics

A nucleotide-dependent conformational change regulates actin filament dynamics. Yet, the structural basis of this mechanism remains controversial. The x-ray crystal structure of tetramethylrhodamine-5-maleimide-actin with bound AMPPNP, a non-hydrolyzable ATP analog, was determined to 1.85-A resolution. A comparison of this structure to that of tetramethylrhodamine-5-maleimide-actin with bound ADP, determined previously under similar conditions, reveals how the release of the nucleotide gamma-phosphate sets in motion a sequence of events leading to a conformational change in subdomain 2. The side chain of Ser-14 in the catalytic site rotates upon Pi release, triggering the rearrangement of the loop containing the methylated His-73, referred to as the sensor loop. This in turn causes a transition in the DNase I-binding loop in subdomain 2 from a disordered loop in ATP-actin to an ordered alpha-helix in ADP-actin. Despite this conformational change, the nucleotide cleft remains closed in ADP-actin, similar to ATP-actin. An analysis of the existing structures of members of the actin superfamily suggests that the cleft is open in the nucleotide-free state.     

Circadian pattern of blood pressure,heart rate,and double product in liver glycogen storage disease

The objective of this study was to determine systolic, diastolic, and mean arterial blood pressure (SBP, DBP, and MAP), heart rate (HR), double-product (DP: SBP x HR), and activity levels and their 24h pattern in liver glycogen storage disease (LGSD) patients. A case series of 12 (11 pediatric and one adult) diurnally active LGSD (seven type I, three type III, and two type IX) subjects were simultaneously assessed by 24h ambulatory blood pressure monitoring and wrist actigraphy. Nine subjects were judged to be hypertensive based on the criterion of an elevated 24h mean SBP and/or DBP being elevated beyond reference standards or the SBP and/or DBP load (percentage of time BP exceeds normal values) being greater than 25%. Two of the three other subjects, not viewed as hypertensive based on their 24h average SBP or DBP, exhibited daytime or nighttime SBP and/or DBP load hypertension. Each study variables displayed statistically significant (p < 0.001) group circadian rhythmicity. The SBP, DBP, and MAP displayed comparable 24h patterns of appreciable amplitude (total peak-trough variation equal to 17.7, 23.6, and 19.6%, respectively, of the 24h mean) with highest values (orthophase) occurring approximately 11 h after the commencement of daytime activity. The sleep-time trough (bathyphase) occurred approximately 4.5 h before morning awakening. The statistically significant (p < 0.006) circadian rhythms of HR (amplitude equal to 33.2% of the 24h mean) and DP (amplitude equal to 49.4% of the 24h mean) peaked earlier, approximately 7.4 h into the daytime activity span. The sleep-time trough occurred approximately 3 h before morning awakening. The 24h pattern in the cardiovascular variables was correlated with the 24h pattern of activity, with r ranging from 0.50 for DBP to 0.39 for HR.     

Cytotoxicity mechanisms of sodium hypochlorite in cultured human dermal fibroblasts and its bactericidal effectiveness

We investigated the therapeutic efficacy of the topical antiseptic sodium hypochlorite (NaOCl) for antibacterial activity and in parallel the cytotoxicity mechanisms by which hypochlorite and the chloramines generated therefrom induce oxidative tissue damage, which further influences the wound-healing process. Human dermal fibroblasts were exposed to increasing concentrations of reagent NaOCl (0.00005-0.1%) at exposure times varying between 2 and 24 h and the protective effects of fetal calf serum (FCS) determined. Antibacterial power was studied by testing a wide range of hypochlorite concentrations (0.00025-0.5%) against four isolated bacterial species. Total bactericidal effects were observed only for 0.5%; concentration range 0.25-0.025% produced partial antimicrobial activity. The early NaOCl-produced cytotoxic action on cultured fibroblasts was cell ATP depletion which occurred at 0.00005% (with FCS 2%) followed by dose- and time-dependent decreases, reaching levels below 5% of control values. Using the 3'-[1-(phenylamino-carbonyl)-3,4-tetrazolium]-bis(4-methoxy-6-nitro)benzene sulfonic acid metabolic assay to evaluate cell death, we observed that NaOCl concentrations greater than 0.05% provoked null fibroblast survival at all exposure times assayed. Hypochlorous acid proved to exert a rapid inhibitory effect on DNA synthesis, consistent with its primary role in bacterial killing by phagocytes. Cytotoxicity produced by increasing NaOCl concentrations and assessed by measuring both mitochondrial function and cell DNA synthesis was reduced with the greatest presence of FCS (10%) in culture media.