Diamond Blackfan anemia is mediated by hyperactive Nemo-like kinase.

Diamond Blackfan Anemia (DBA) is a congenital bone marrow failure syndrome associated with ribosomal gene mutations that lead to ribosomal insufficiency. DBA is characterized by anemia, congenital anomalies, and cancer predisposition. Treatment for DBA is associated with significant morbidity. Here, we report the identification of Nemo-like kinase (NLK) as a potential target for DBA therapy. To identify new DBA targets, we screen for small molecules that increase erythroid expansion in mouse models of DBA. This screen identified a compound that inhibits NLK. Chemical and genetic inhibition of NLK increases erythroid expansion in mouse and human progenitors, including bone marrow cells from DBA patients. In DBA models and patient samples, aberrant NLK activation is initiated at the Megakaryocyte/Erythroid Progenitor (MEP) stage of differentiation and is not observed in non-erythroid hematopoietic lineages or healthy erythroblasts. We propose that NLK mediates aberrant erythropoiesis in DBA and is a potential target for therapy.

Novel ranking of protective and risk factors for adolescent adiposity in US females.

OBJECTIVE: Rank the importance of potentially modifiable psychosocial, dietary and environmental risk and protective factors for female adolescent obesity in order to target and inform public health prevention efforts. Utilizing the largest dataset available that captures the onset of the adolescent obesity surge in the USA, the study provides a more robust understanding of paediatric obesity risk factors. METHODS: Data were obtained from an observational, longitudinal study conducted between 1989 and 2001, the NHLBI Growth and Health Study. This study includes girls aged 9-19 years from three urban US locations, with Black and White girls generally represented equally. Data were analysed using multiple regression, random forest and propensity score matching to determine the strongest adiposity risk and protective factors during ages 9-12 predicting adiposity at age 19 with multiple methods to maximize the ability to identify possible public health interventions. Multiple linear regression and random forest analysis identified the strongest associations among 288 risk and protective factors selected from the study's literature review. For the 190 factors associated with follow-up adiposity from the data, propensity score matching was used to control for confounding of these factors. RESULTS: Findings suggest that highest priority interventional targets across the domains surveyed are lowering specific nutrients; eating meals with others or during activities without skipping; parents fixing evening snacks; improving perceptions of non-extremes as the healthy weight; improving self-worth, physical activity and social competence; and limiting any negative impact of dieting relatives. Similar associations were observed for Black and White girls. CONCLUSION: The clinical implications of these findings allow health practitioners to target behavioural change efforts and address social and environmental factors that have demonstrated higher prioritization value for early obesity interventional efforts for adolescents.

Deterministic Remote Entanglement of Superconducting Circuits through Microwave Two-Photon Transitions.

Large-scale quantum information processing networks will most probably require the entanglement of distant systems that do not interact directly. This can be done by performing entangling gates between standing information carriers, used as memories or local computational resources, and flying ones, acting as quantum buses. We report the deterministic entanglement of two remote transmon qubits by Raman stimulated emission and absorption of a traveling photon wave packet. We achieve a Bell state fidelity of 73%, well explained by losses in the transmission line and decoherence of each qubit.

Continuous Quantum Nondemolition Measurement of the Transverse Component of a Qubit.

Quantum jumps of a qubit are usually observed between its energy eigenstates, also known as its longitudinal pseudospin component. Is it possible, instead, to observe quantum jumps between the transverse superpositions of these eigenstates? We answer positively by presenting the first continuous quantum nondemolition measurement of the transverse component of an individual qubit. In a circuit QED system irradiated by two pump tones, we engineer an effective Hamiltonian whose eigenstates are the transverse qubit states, and a dispersive measurement of the corresponding operator. Such transverse component measurements are a useful tool in the driven-dissipative operation engineering toolbox, which is central to quantum simulation and quantum error correction.

Coordinate regulation of residual bone marrow function by paracrine trafficking of AML exosomes.

We recently demonstrated that acute myeloid leukemia (AML) cell lines and patient-derived blasts release exosomes that carry RNA and protein; following an in vitro transfer, AML exosomes produce proangiogenic changes in bystander cells. We reasoned that paracrine exosome trafficking may have a broader role in shaping the leukemic niche. In a series of in vitro studies and murine xenografts, we demonstrate that AML exosomes downregulate critical retention factors (Scf, Cxcl12) in stromal cells, leading to hematopoietic stem and progenitor cell (HSPC) mobilization from the bone marrow. Exosome trafficking also regulates HSPC directly, and we demonstrate declining clonogenicity, loss of CXCR4 and c-Kit expression, and the consistent repression of several hematopoietic transcription factors, including c-Myb, Cebp-ß and Hoxa-9. Additional experiments using a model of extramedullary AML or direct intrafemoral injection of purified exosomes reveal that the erosion of HSPC function can occur independent of direct cell-cell contact with leukemia cells. Finally, using a novel multiplex proteomics technique, we identified candidate pathways involved in the direct exosome-mediated modulation of HSPC function. In aggregate, this work suggests that AML exosomes participate in the suppression of residual hematopoietic function that precedes widespread leukemic invasion of the bone marrow directly and indirectly via stromal components.

Quantum engineering. Confining the state of light to a quantum manifold by engineered two-photon loss.

Physical systems usually exhibit quantum behavior, such as superpositions and entanglement, only when they are sufficiently decoupled from a lossy environment. Paradoxically, a specially engineered interaction with the environment can become a resource for the generation and protection of quantum states. This notion can be generalized to the confinement of a system into a manifold of quantum states, consisting of all coherent superpositions of multiple stable steady states. We have confined the state of a superconducting resonator to the quantum manifold spanned by two coherent states of opposite phases and have observed a Schrödinger cat state spontaneously squeeze out of vacuum before decaying into a classical mixture. This experiment points toward robustly encoding quantum information in multidimensional steady-state manifolds.

Tracking photon jumps with repeated quantum non-demolition parity measurements.

Quantum error correction is required for a practical quantum computer because of the fragile nature of quantum information. In quantum error correction, information is redundantly stored in a large quantum state space and one or more observables must be monitored to reveal the occurrence of an error, without disturbing the information encoded in an unknown quantum state. Such observables, typically multi-quantum-bit parities, must correspond to a special symmetry property inherent in the encoding scheme. Measurements of these observables, or error syndromes, must also be performed in a quantum non-demolition way (projecting without further perturbing the state) and more quickly than errors occur. Previously, quantum non-demolition measurements of quantum jumps between states of well-defined energy have been performed in systems such as trapped ions, electrons, cavity quantum electrodynamics, nitrogen-vacancy centres and superconducting quantum bits. So far, however, no fast and repeated monitoring of an error syndrome has been achieved. Here we track the quantum jumps of a possible error syndrome, namely the photon number parity of a microwave cavity, by mapping this property onto an ancilla quantum bit, whose only role is to facilitate quantum state manipulation and measurement. This quantity is just the error syndrome required in a recently proposed scheme for a hardware-efficient protected quantum memory using Schrödinger cat states (quantum superpositions of different coherent states of light) in a harmonic oscillator. We demonstrate the projective nature of this measurement onto a region of state space with well-defined parity by observing the collapse of a coherent state onto even or odd cat states. The measurement is fast compared with the cavity lifetime, has a high single-shot fidelity and has a 99.8 per cent probability per single measurement of leaving the parity unchanged. In combination with the deterministic encoding of quantum information in cat states realized earlier, the quantum non-demolition parity tracking that we demonstrate represents an important step towards implementing an active system that extends the lifetime of a quantum bit.

Autonomously stabilized entanglement between two superconducting quantum bits.

Quantum error correction codes are designed to protect an arbitrary state of a multi-qubit register from decoherence-induced errors, but their implementation is an outstanding challenge in the development of large-scale quantum computers. The first step is to stabilize a non-equilibrium state of a simple quantum system, such as a quantum bit (qubit) or a cavity mode, in the presence of decoherence. This has recently been accomplished using measurement-based feedback schemes. The next step is to prepare and stabilize a state of a composite system. Here we demonstrate the stabilization of an entangled Bell state of a quantum register of two superconducting qubits for an arbitrary time. Our result is achieved using an autonomous feedback scheme that combines continuous drives along with a specifically engineered coupling between the two-qubit register and a dissipative reservoir. Similar autonomous feedback techniques have been used for qubit reset, single-qubit state stabilization, and the creation and stabilization of states of multipartite quantum systems. Unlike conventional, measurement-based schemes, the autonomous approach uses engineered dissipation to counteract decoherence, obviating the need for a complicated external feedback loop to correct errors. Instead, the feedback loop is built into the Hamiltonian such that the steady state of the system in the presence of drives and dissipation is a Bell state, an essential building block for quantum information processing. Such autonomous schemes, which are broadly applicable to a variety of physical systems, as demonstrated by the accompanying paper on trapped ion qubits, will be an essential tool for the implementation of quantum error correction.

Opioid, cannabinoid and vanilloid receptor localization on porcine cultured myenteric neurons.

Opioids and cannabinoids have profound inhibitory actions on intestinal motility which are mediated in part by their cognate receptors in the enteric nervous system. In the present study, we examined the expression of immunoreactivity for delta- and kappa-opioid receptors, CB(1)-cannabinoid receptors and type 1 vanilloid receptors by immunocytochemistry and confocal laser scanning microscopy on ileal myenteric neurons, isolated from juvenile pigs, that were <70 microm diameter in either axis and maintained for 1-2 weeks in primary culture. Immunoreactivities for delta-opioid and cannabinoid receptors were present in neurons immunoreactive for the cholinergic marker, choline acetyltransferase. Some neurons with delta-opioid receptor-like immunoreactivity were also immunoreactive for kappa-opioid, cannabinoid or vanilloid receptors. These observations indicate that receptors for cannabinoids or vanilloids are co-localized in opioid receptor-expressing myenteric neurons which modulate intestinal sensorimotor function.

Opioid receptors on bone marrow neutrophils modulate chemotaxis and CD11b/CD18 expression.

Opiates impair neutrophil-mediated host defense, but the involvement of kappa-opioid receptors in this action has not been defined. The selective kappa-opioid receptor agonist [trans-(+)3,4-dichloro-N-methyl-N[2-(1-pyrrolidinyl)-cyclohexyl]benzeneacetamide methanesulfonate inhibited macrophage inflammatory protein-2-induced chemotaxis of bone marrow neutrophils from C57BL/6 mice. Its effects were concentration-dependent (pIC(50)=10.40+/-0.61) and inhibited by naloxone (K(e)=0.27 nM). The kappa-opioid receptor agonists bremazocine and ICI-204, 488 also inhibited chemotaxis, as did the respective mu- and delta-opioid receptor agonists [D-Ala(2), N-methyl-Phe(4), Gly(5)-ol]enkephalin and [D-Pen(2,5)]enkephalin albeit with lower potencies. U-50,488H also decreased neutrophil expression of the beta(2) integrin CD11b/CD18 (Mac-1) and adhesion to plastic in a naloxone-reversible manner. The results indicate that kappa-opioid receptors expressed by neutrophils rapidly modulate chemotaxis and adhesion in vitro.

Localization of CB1-cannabinoid receptor immunoreactivity in the porcine enteric nervous system.

Cannabis has been used for centuries in the medicinal treatment of gastrointestinal disorders. Endogenous cannabinimimetic substances such as 2-arachidonylglycerol have been isolated from gut homogenates and CB1-cannabinoid binding sites have been identified in small intestine. In this study, CB1-cannabinoid receptors (CB1-R) were immunohistochemically localized within the enteric nervous system of the pig, an omnivorous species whose digestive tract is functionally similar to humans. Two anti-CB1-R antisera, raised against N-terminal epitopes in the human CB1-R, were employed to localize receptor immunoreactivity by secondary immunofluorescence. CB1-R immunoreactivity was observed in the myenteric and submucosal ganglionated plexuses of porcine ileum and colon. In the ileum, all CB1-R-immunoreactive neurons coexpressed immunoreactivity to the cholinergic marker, choline acetyltransferase (ChAT). CB1-R/ChAT-immunoreactive neurons appeared to be in close apposition to ileal Peyer's patches, submucosal blood vessels, and intestinal crypts. In the distal colon, CB1-R-immunoreactive neurons also expressed immunoreactivity to ChAT, albeit less frequently than in ileum. Immunoreactivity to vasoactive intestinal peptide or nitric oxide synthase was not colocalized in ileal or colonic CB1-R-immunoreactive neurons. These studies indicate that CB1-R are present in cholinergic neurons in the porcine enteric nervous system. The potential roles of these receptors in intestinal motility and epithelial transport, host defense and visceral pain transmission are discussed.

Intestinal type 2 proteinase-activated receptors: expression in opioid-sensitive secretomotor neural circuits that mediate epithelial ion transport.

Trypsin and mast cell tryptase cleave within the extracellular N terminus of proteinase-activated receptor-2 (PAR-2), exposing a tethered ligand (SLIGRL) that binds and activates the cleaved receptor. We examined the neuronal expression of PAR-2 and its role in intestinal ion transport. Short-circuit current elevations in response to trypsin or the receptor-activating peptide SLIGRL-NH(2) were measured in sheets of mucosa-submucosa from porcine ileum. SLIGRL-NH(2) or trypsin rapidly elevated short-circuit current after their contraluminal application with respective 50% effective concentrations of 184 and 769 nM. Their actions were attenuated after contraluminal administration of the neuronal conduction blocker saxitoxin (0.1 microM); the cyclooxygenase inhibitor indomethacin (10 microM); or the Na(+)/K(+)/Cl(-) cotransport inhibitor furosemide (10 microM), but not by atropine (0.1 microM), a muscarinic cholinergic antagonist. In addition, soybean trypsin inhibitor (5 microgram/ml) reduced mucosal responses to trypsin. The delta-opioid agonist [D-Pen(2,5)]-enkephalin (0.1 microM) inhibited trypsin action, an effect that was prevented by naltrindole (0.1 microM), a delta-opioid antagonist. PAR-2 immunofluorescence was localized in the mucosa using a receptor-specific antibody. PAR-2-like immunoreactivity was detected in myenteric and submucosal neurons, nerve fibers innervating ileal smooth muscle and mucosa, and in enteroendocrine cells. Some neurons coexpressed PAR-2- and choline acetyltransferase-like immunoreactivity. These results indicate that PAR-2 is expressed on cholinergic and noncholinergic submucosal neurons in porcine ileum. PAR-2 agonists stimulate active anion secretion by a neurogenic mechanism that is modulated by prostanoids and opioids. These receptors may have a potentially important role in intestinal neuroimmunomodulation.

Catecholaminergic, cholinergic and peptidergic innervation of gut-associated lymphoid tissue in porcine jejunum and ileum.

With its abundance of neurons and immunocytes, the gut is a potentially important site for the study of the interaction between the nervous and immune systems. Using immunohistochemical techniques, we tested the hypothesis that gut-associated lymphoid tissue in the porcine small intestine might receive catecholaminergic, cholinergic and peptidergic innervation. Antibodies against protein gene product (PGP) 9.5 were employed to detect neuronal membranes; antibodies against tyrosine hydroxylase (TH), type 2 vesicular monoamine transporter (VMAT-2) and choline acetyltransferase (ChAT) were used to detect catecholaminergic and cholinergic neurons; and antibodies to neuromedin U-8 (NMU-8), substance P (SP) and vasoactive intestinal peptide (VIP) were also used. PGP9.5-immunoreactive nerve fibers were observed between jejunal Peyer's patch (PP) follicles and in submucosal ganglia localized at the base of continuous ileal PP. Many ChAT-positive and a few TH-/VMAT-2-immunoreactive neurons or axons adjacent to jejunal and ileal PP were observed. Neurons and fibers from ganglia situated between or at the base of PP follicles manifested robust immunoreactivities to VIP and NMU-8; relatively less SP immunoreactivity was observed at these locations. All neuromedin-U 8-positive neurons observed exhibited immunoreactivity to ChAT as did some VIP-positive neurons. The specific chemical coding of enteric neurons in close apposition to jejunal and ileal PP and the differential localization of neuropeptides within the jejunal and ileal PP are indicative of neuroimmunomodulation at these sites.

Differential expression of manganese and copper-zinc superoxide dismutases in the olfactory and vomeronasal receptor neurons of rats during ontogeny.

Superoxide dismutases (SODs) protect cells from damage by oxygen free radicals. Manganese (Mn) SOD is preferentially induced in terminally differentiating cells; induction of copper-zinc (CuZn) SOD is more closely associated with postnatal exposure to environmental sources of oxygen free radicals. The purpose of this study was to investigate ontogenetic changes in immunoreactivity for MnSOD and CuZnSOD relative to the expression of markers of neuronal and chemosensory differentiation in olfactory and vomeronasal receptor neurons (ORNs and VRNs, respectively), which mature with different time courses. Immunoreactivity for both SODs was detected in rat ORNs at embryonic day (E) 14, the earliest time point investigated, but not until E16 in vomeronasal neuroblasts. ORNs also expressed the neuronal marker protein gene product (PGP) 9.5 and the chemosensory cell marker olfactory marker protein (OMP) at E14; vomeronasal neuroblasts expressed PGP 9.5 at E16 but were not immunoreactive for OMP until postnatal day (P) 2. Immunoreactivity for MnSOD in ORNs and VRNs generally increased pre- and postnatally to a maximum at P11. Immunoreactivity for CuZnSOD did not increase markedly until after birth, reaching maximal levels at P11-P24. Within ORNs and VRNs, the most intense immunoreactivity was localized in the dendritic and supranuclear regions. The results indicate that in ORNs and VRNs, increases in MnSOD immunoreactivity during ontogeny parallel the ongoing differentiation and maturation of chemosensory receptor neurons; in contrast, the induction of immunoreactivity for CuZnSOD is associated with postnatal exposure to the ambient oxygen and xenobiotic environment.

Manganese and copper-zinc superoxide dismutases in the human olfactory mucosa: increased immunoreactivity in Alzheimer's disease.

Superoxide dismutases are the cell's major enzymatic defenses against cytotoxic reactive oxygen species and oxidative stress. Reactive oxygen species, which induce the expression of these enzymes, have been implicated in the neurodegeneration associated with Alzheimer's disease (AD), and individuals with AD exhibit early, severe deficits in olfactory ability. We used immunohistochemistry to examine the cellular localization of managanese and copper-zinc superoxide dismutases in the olfactory mucosae of nondemented young/middle-aged and old subjects as well as age-and postmortem-interval matched nondemented elderly individuals and those with AD. Tissues were obtained at autopsy from individuals ranging in age from 19 to 98 years old. Immunoreactivity for both enzymes was localized in olfactory receptor neurons, sustentacular and basal cells in the olfactory epithelium, and in olfactory and extrinsic nerves, Bowman's glands, and vascular endothelium in the lamina propria. Computer-assisted quantitative analysis demonstrated that very intense immunoreactivity for both managanese and copper-zinc superoxide dismutases occupied significantly more area, particularly near the surface and in the basal region, of the olfactory epithelium from subjects with AD than from the age-and postmortem interval-matched nondemented elderly subjects. The pronounced increase in superoxide dismutase immunoreactivity in the olfactory epithelium of AD subjects suggests that oxidative stress may be responsible, at least in part, for the olfactory deficits in subjects with AD.

Age-dependent phenotypic switching of mast cells in NGF-transgenic mice.

Effects of overexpression of nerve growth factor (NGF) on mast cell phenotype and numbers were investigated in nasal and oral mucosae and skin of 3- and 6-week-old transgenic mice in which NGF expression in epithelial basal cells was driven by the keratin-14 promoter. Mast cell phenotypes were identified by Alcian blue/safranin and berberine sulfate histochemistry. In the 3-week-old transgenic mice, NGF overexpression had no effect on phenotype except in tongue, where mast cells exhibited mixed or connective tissue phenotypes compared with the mucosal phenotype in the non-transgenic. In 6-week-old transgenic animals, NGF overexpression resulted in the mucosal phenotype in tissues which contained connective tissue or mixed mast cells in non-transgenics. Mast cell hyperplasia occurred at both ages. NGF effects on mast cell phenotype were age-dependent and involve complex microenvironmental interactions.