Trace amines: identification of a family of mammalian G protein-coupled receptors.

Tyramine, beta-phenylethylamine, tryptamine, and octopamine are biogenic amines present in trace levels in mammalian nervous systems. Although some "trace amines" have clearly defined roles as neurotransmitters in invertebrates, the extent to which they function as true neurotransmitters in vertebrates has remained speculative. Using a degenerate PCR approach, we have identified 15 G protein-coupled receptors (GPCR) from human and rodent tissues. Together with the orphan receptor PNR, these receptors form a subfamily of rhodopsin GPCRs distinct from, but related to the classical biogenic amine receptors. We have demonstrated that two of these receptors bind and/or are activated by trace amines. The cloning of mammalian GPCRs for trace amines supports a role for trace amines as neurotransmitters in vertebrates. Three of the four human receptors from this family are present in the amygdala, possibly linking trace amine receptors to affective disorders. The identification of this family of receptors should rekindle the investigation of the roles of trace amines in mammalian nervous systems and may potentially lead to the development of novel therapeutics for a variety of indications.     

Effects of aripiprazole on circadian prolactin secretion related to pharmacogenetics in healthy volunteers

Aripiprazole treatment in schizophrenic patients was previously associated with lower or normalized prolactin levels. Genetic variants in cytochrome P450 (CYP) (CYP2D6), dopamine receptor (DRD2, DRD3) and serotonin receptor (HTR2A, HTR2C) genes were previously associated with antipsychotic‐induced hyperprolactinaemia. Our aim was to evaluate whether aripiprazole affects prolactin secretion and its relationship with pharmacogenetics. Thirty‐one healthy volunteers receiving a 10‐mg single oral dose of aripiprazole were genotyped for 12 polymorphisms in CYP2D6, DRD2, DRD3, HTR2A and HTR2C genes by qPCR. Aripiprazole and dehydro‐aripiprazole plasma concentrations were measured by HPLC‐MS/MS. Prolactin concentrations of the 31 volunteers taking aripiprazole and 12 volunteers receiving ibuprofen were determined by ELISA. Prolactin concentrations after ibuprofen intake were considered as control, since it is known to cause no effect. Prolactin concentrations were slightly higher in the aripiprazole group compared to the ibuprofen group. All prolactin pharmacokinetic parameters were higher in females than in males. CYP2D6 poor and intermediate metabolizers had notably higher prolactin C_max and AUC_0‐12 than normal and ultrarapid metabolizers. The DRD3 rs6280 polymorphism affected prolactin levels: volunteers carrying Ser/Ser genotype had significantly lower prolactin levels than volunteers carrying the Gly allele. Furthermore, HTR2C rs3813929 C/C homozygotes had significantly lower prolactin levels than T allele carriers. Nevertheless, aripiprazole did increase prolactin levels compared to ibuprofen.     

Differential Interleukin-8 thresholds for chemotaxis and netosis in human neutrophils

In humans, IL-8 (CXCL8) is a key chemokine for chemotaxis of polymorphonuclear leukocytes and monocytes/macrophages when acting on CXCR1 and CXCR2. CXCL8 activity on neutrophils includes chemotaxis and eliciting the extrusion of neutrophil extracellular traps (NETs). In this study, we show that concentrations of IL-8 that induce NETosis surpass in at least one order of magnitude those required to elicit chemoattraction in human neutrophils. IL-8-induced NETosis was less dependent on G-proteins than migration, while extracellular Ca⁺² chelation similarly inhibited both processes. Reactive oxygen species (ROS) were more important for NETosis than for chemotaxis as evidenced by neutralization with N-acetyl -cysteine. Interestingly, selective blockade with anti-CXCR1 mAb inhibited NETosis much more readily than chemotaxis, while pharmacological inhibition of both CXCR1 and CXCR2, or selective inhibition for CXCR2 alone, similarly inhibited both functions. Together, these results propose a model according to which low concentrations of IL-8 in a gradient attract neutrophils to the inflammatory foci, while high receptor-saturating concentrations of IL-8 give rise to NETosis once leukocytes reach the core of the inflammatory insult.     

Clinical features and outcome of T-cell acute lymphoblastic leukemia in childhood with respect to alterations at the TAL1 locus: a Pediatric Oncology Group study

Alteration of the TAL1 locus is the most common nonrandom genetic defect in childhood T-cell acute lymphoblastic leukemia (T-ALL). To determine if rearrangements of the TAL1 proto-oncogene confer a distinct leukemic phenotype, we studied leukemic peripheral blood or bone marrow samples from 182 children with newly diagnosed T-ALL enrolled on Pediatric Oncology Group treatment protocols. Forty-eight (26%) of the samples had a local rearrangement of the TAL1 locus. Demographic and clinical features were compared for patient subgroups with and without TAL1 rearrangements. The only clinical correlates that were significantly associated with TAL1 gene rearrangements were higher white blood cell count (P = .017) and higher hemoglobin (P = .007) at diagnosis. Immunophenotypically, samples with altered TAL1 were more likely to be CD2+ (P = .001) and lack CD10 (cALLa) expression (P = .007) than those without the rearrangement. There was a trend toward improved event-free survival (EFS) in patients with TAL1 rearrangements (4-year EFS was 44% +/- 7% for patients without the rearrangements v 59% +/- 11% for those with rearrangements), but the difference was not significant (P = .34). The role of TAL1 in leukemogenesis has yet to be clearly defined, and the prognostic significance of TAL1 gene rearrangements in T-ALL deserves further study.     

Minor histocompatibility antigens HA-1-, -2-, and -4-, and HY-specific cytotoxic T-cell clones inhibit human hematopoietic progenitor cell growth by a mechanism that is dependent on direct cell-cell contact

HLA-identical bone marrow transplantation (BMT) may be complicated by graft-versus-host disease or graft rejection. Both complications are thought to be initiated by recognition of minor histocompatibility (mH) antigens by HLA-restricted mH-antigen-specific T lymphocytes. Using HLA- A2-restricted mH antigens HA-1-, -2-, and -4-, and HY-specific cytotoxic T lymphocyte (CTL) clones, we studied the recognition by these CTL clones of interleukin-2 (IL-2)-stimulated T cells (IL-2 blasts), BM mononuclear cells (BMMNCs), and hematopoietic progenitor cells (HPCs). We showed that, when IL-2 blasts from the BM donors who were investigated were recognized by the HA-1-, -2-, and -4-, and HY- specific CTL clones, their BMMNCs and HPCs were recognized as well by these CTL clones, resulting in antigen-specific growth inhibition of erythrocyte burst-forming units (BFU-E), colony-forming units- granulocyte (CFU-G), and CFU-macrophage (CFU-M). the HA-2-specific CTL clone, however, inhibited BFU-E and CFU-G growth from four donors to a lesser extent than from two other donors. We further investigated whether inhibitory cytokines released into the culture medium by the antigen-specific stimulated CTLs or by stimulated BMMNCs were responsible for suppression of HPC growth or whether this effect was caused by direct cell-cell contact between CTLs and HPCs. HPC growth inhibition was only observed after preincubation of BMMNCs and CTLs together for 4 hours before plating the cells in semisolid HPC culture medium. When no cell-cell contact was permitted before plating, neither antigen-stimulated CTL nor antigen-nonstimulated CTLs provoked HPC growth inhibition. Culturing BMMNCs in the presence of supernatants harvested after incubation of BMMNCs and CTL clones together for 4 or 72 hours did also not result in HPC growth inhibition. Both suppression of HPC growth and lysis of IL-2 blasts and BMMNCs in the 51Cr-release assay appeared to be dependent on direct cell-cell contact between target cells and CTLs and were not caused by the release of inhibitory cytokines into the culture medium by antigen-specific stimulated CTLs or by stimulated BMMNCs. Our results show that mH-antigen-specific CTLs can inhibit HPC growth by a direct cytolytic effect and may therefore be responsible for BM graft rejection after HLA-identical BMT.     

Foam film stratification studies probe intermicellar interactions

Ultrathin foam films containing supramolecular structures like micelles in bulk and adsorbed surfactant at the liquid–air interface undergo drainage via stratification. At a fixed surfactant concentration, the stepwise decrease in the average film thickness of a stratifying micellar film yields a characteristic step size that also describes the quantized thickness difference between coexisting thick–thin flat regions. Even though many published studies claim that step size equals intermicellar distance obtained using scattering from bulk solutions, we found no reports of a direct comparison between the two length scales. It is well established that step size is inversely proportional to the cubic root of surfactant concentration but cannot be estimated by adding micelle size to Debye length, as the latter is inversely proportional to the square root of surfactant concentration. In this contribution, we contrast the step size obtained from analysis of nanoscopic thickness variations and transitions in stratifying foam films using Interferometry Digital Imaging Optical Microscopy (IDIOM) protocols, that we developed, with the intermicellar distance obtained using small-angle X-ray scattering. We find that stratification driven by the confinement-induced layering of micelles within the liquid–air interfaces of a foam film provides a sensitive probe of non-DLVO (Derjaguin–Landau–Verwey–Overbeek) supramolecular oscillatory structural forces and micellar interactions.

Molecules in simple liquids and supramolecular structures in complex fluids can stratify or undergo confinement-induced layering induced by symmetry breaking at a solid–liquid or a fluid–fluid interface (1–8). In freestanding or foam films, the confinement-induced layering of supramolecular structures including micelles (9–17), lipid layers (18, 19), polyelectrolyte–surfactant complexes (20, 21), nanoparticles (9, 22), and liquid crystalline assemblies (23) can result in drainage via stratification. Due to thin film interference, foam films visualized under white light illumination display iridescent colors for thick films (h > 100 nm) (24–28), but ultrathin films (h < 100 nm) exhibit shades of gray that get progressively darker as the film gets thinner (9–21). In reflected light microscopy, micellar foam films exhibit coexisting thick−thin regions with distinct shades of gray. Interferometry-based measurement of the average film thickness over time decreases in a stepwise fashion yielding a step size, Δh (9–17). Many published studies argue (9–12, 22, 29–34) that foam films containing charged micelles or latex particles stratify analogously due to the formation of “ordered colloidal crystals” (OCCs) and step size, Δh, equals the intermicellar distance, d, in bulk solutions. However, a comparison of concentration-dependent Δh obtained from the dynamic foam stratification studies (influenced by confinement effects) with d measured using small-angle X-ray or neutron scattering (SAXS or SANS) or other direct measurements of static equilibrium structure, and related evidence for or against the formation of OCCs in micellar foam films, are lacking in the literature. Thus, the motivations of this contribution are threefold: 1) contrast the step size, Δh, obtained via stratification studies with the intermicellar distance, d, and micelle dimensions determined using SAXS; 2) examine the SAXS data for any evidence of OCCs; and 3) elucidate the influence of ionic micelles on foam film stability and topography, as well as on colloidal forces, in multicomponent complex fluids.Micelles, formed by self-assembly of soaps and detergents and ever present in typical household foams, facilitate cleaning and detergent action by solubilizing oils and oil-soluble dirt within their hydrophobic core (2, 34, 35). Micelles formed by biosurfactants like bile salt and rhamnolipids can be used for delivering nonpolar, bio-active polyunsaturated oils, flavonoids, vitamins, and hydrophobic drugs (36–38). Therefore, understanding the stability and lifetime of micellar foams is essential toward molecular engineering of formulations, controlling foams in industrial reactors, rivers, and lakes and developing bio-surfactants (36–38). Foam film drainage involves interfacial flows that are influenced both by bulk rheology and interfacial rheology as well as Laplace or capillary pressure, Pc=σC (set by surface tension, σ and curvature, C) (27, 28, 39–41). Additionally, thickness transitions and variations in ultrathin (h < 100 nm) freestanding as well as supported (containing one or two solid boundaries) films (41–43) depend on disjoining pressure, Π(h)=−(∂G/∂h)P,T,A,Ni, defined as the free energy required to change unit thickness at constant temperature, T, pressure, P, surface area, A, and mole number, N_i (1, 34, 40–42). Intermolecular and surface forces determine the strength and range of disjoining pressure, Π(h), as well as of colloidal interaction forces, F(h) (1–3, 35, 40–42). Physical properties of surfactant solutions like surface tension and conductivity show distinct change around a critical micelle concentration (CMC), beyond which spheroidal micelles can form (2, 34, 35), and rod-like micelles, lamellar phases, etc., emerge at higher concentrations (44–46). In foam films formed with ionic surfactant at c < CMC, drainage below h < 30 nm often leads to the formation of relatively long-lived common black (CB) film attributed to counterbalancing of P_c by ΠDLVO(h), the disjoining pressure due to DLVO (Derjaguin–Landau–Verwey–Overbeek) forces contributed by van der Waals and electrostatic double-layer interactions (1–3, 35, 39, 40). Even thinner Newton black (NB) films attest to the role of shorter-range, non-DLVO surface forces (14, 25–27, 40, 41). In contrast, in micellar foam films (c > CMC), a non-DLVO, oscillatory structural force, ΠOS(h), counterbalances P_c at multiple flat thicknesses, manifested as distinct shades of gray in reflected light microscopy (9–17, 21, 40, 47–50).For micellar fluids containing charged micelles, the step size, Δh, obtained using thickness–time plots from stratification experiments, and periodicity, λ, of ΠOS(h) directly measured using thin-film balance (47, 48) show that both periodicity and step size exceed micelle size, a, implying λ>a and Δh>a. In 1971, Bruil and Lyklema (51) were the first to report that the concentration-dependent decrease in step size measured for sodium dodecyl sulfate (SDS) solutions followed a power law of the form Δh∝csoap−1/3 and wrote that step size values “seem to be related to intermolecular distance in the (unmicellized) bulk solution.” In 1988, Nikolov et al. (9) reported that foam films containing latex particles stratified in a fashion similar to micellar foam films and argued that diffusion-driven, layer-by-layer removal of micelles or particles from an ordered colloidal crystal (OCC) structure drives stratification. In their OCC or “micelle-vacancy diffusion” mechanism, they proposed that the effective film viscosity increases with decrease in stratified film thickness (9, 10, 29–31, 33). Contrastingly, in the “hydrodynamic” mechanism, Bergeron and Radke (13, 47) described stratification using a thin-film equation, by incorporating ΠOS(h) and bulk solution viscosity. Nikolov et al. (9, 10, 29–31) suggested that the step size, Δh, was equal to an effective diameter, deff=2lSDS+κ−1, computed by adding the fixed length of SDS molecules, l_SDS, to the Debye length, κ−1, that captures the range of screened electrostatic interactions. However, the step size Δh∼c−1/3 and the Debye length κ−1∼c−1/2 display distinct power laws, and the measured step size exceeds the micelle size, a, as well as the computed effective diameter, d_eff, for ionic micellar systems, or typically Δh>a and Δh>deff.Studies on charged nanoparticle dispersions find that the periodicity, λ, of the oscillatory structural force, F(h), measured directly with surface force apparatus (SFA), or colloidal probe atomic force microscopy (CP-AFM), correlates well with the interparticle distance, d, obtained using scattering and simulations (4, 5, 52–55). Furthermore, the periodicity, λ≈d≫a, is primarily set by the particle number density, ρ, and is relatively independent of added salt, charge at solid surfaces, and particle size, a (4, 5, 53–55). Assuming that analogy between λ∝ρ−1/3 in the nanoparticle studies and Δh∝c−1/3 in stratified foam studies arises due to similar underlying physics, Danov et al. (32) and Anachkov et al. (11) argued that Δh≈d or step size equals the intermicellar distance, d, in bulk solutions and hypothesized that step size from stratification studies could be used for determining aggregation number as Nagg≈(c−CMC)(Δh)3. However, Yilixiati et al. (17) showed that on salt addition, the measured Δh values for micellar SDS solutions do not collapse onto a single curve even if plotted against micellar number density, ρ, as micelle number and dimensions can change on the addition of salt (or surfactant) (2), whereas nanoparticle dimensions remain constant. Furthermore, solid boundaries that can impact SFA and AFM measurements are absent in stratifying foam films. However, the thickness of stratifying films is rather heterogeneous, and the average thickness changes in a stepwise fashion. Thus, the analogy between stratifying micelles in foam films and stratifying nanoparticle dispersions under confinement between solid surfaces requires further investigation. In particular, a comparison between multiple length-scales including micelle dimensions, Debye length, intermicellar distance, d and step-size, Δh, and the consequences of thickness heterogeneities within foam films are warranted.In this study, we contrast the concentration-dependent changes in step-size measured in foam stratification studies with micellar dimensions and intermicellar distances in bulk solutions obtained using SAXS for aqueous solutions of SDS. For the range of concentrations (25 mM ≤ c_SDS ≤ 250 mM) explored here, bulk rheology, interfacial tension, micelle shape and size, and interfacial charge (or potential) are nearly constant. Hence, the observed concentration-dependent changes in step-size and nanoscopic topography in stratifying films are dictated by the corresponding changes in intermicellar interactions and the resulting disjoining pressure, ΠOS(h). We visualize and analyze nanoscopic thickness variations and transitions in stratifying foam films using IDIOM (Interferometry Digital Imaging Optical Microscopy) protocols (16) (Fig. 1A) that provide requisite spatiotemporal resolution (thickness ∼1 nm, in-plane < 1 μm, time < 1 ms). We analyze SAXS data to compute micelle dimensions, volume fraction, and microstructure (order) in bulk solutions and obtain the intermicellar distance from structure factor peak in SAXS data. Finally, we discuss the ramifications of the close comparison between step size from the foam film stratification studies and micellar dimensions and intermicellar distance determined using SAXS analysis on the intermicellar interactions and the mechanistic basis of stratification.Open in a separate windowFig. 1.Schematic of the setup used for examining stratification using IDIOM protocols and illustrative examples of stepwise thinning. (A) The Scheludko-like cell contains a plane-parallel film and surrounding meniscus that emulates a single foam film and its Plateau border. The cell is placed in a closed container and stratification is visualized using reflected light microscopy. A finite volume of fluid is inserted into the cell using the side-arm connected to a syringe. No liquid is added or withdrawn during the stratification experiment, and drainage from the film into the meniscus occurs freely and spontaneously. (B) Spatiotemporal variation in interference intensity I(x, y, t; λ) is used for computing thickness transitions and variations in stratifying films. (C) Average film thickness plotted as a function of time shows stepwise thinning for foam films made with aqueous SDS solutions. The spikes and dips in thickness plots appear when mesas or domains emerge in the region selected for computing average thickness. The data are shifted horizontally for clarity.     

Rotavirus Seasonal Distribution and Prevalence Before and After the Introduction of Rotavirus Vaccine in a Peri-Urban Community of Lima,Peru

We evaluated the monthly distribution of rotavirus diarrhea in a cohort of children 12–24 months of age followed as part of a diarrhea clinical trial in a peri-urban community of Lima. We observed a peak of rotavirus diarrhea in the winter months and a decrease in rotavirus prevalence after the introduction of the rotavirus vaccine in Peru.