IgE anti Hepatitis B virus surface antigen antibodies detected in serum from inner city asthmatic and non asthmatic children

Viral Hepatitis type B (HBV) is a public health concern, but has not been linked to asthma. Immunoglobulin (Ig) G is involved in HBV immune responses; less is known about IgE antibodies (Abs) against HBV in asthma. Given the importance of HBV, we sought to determine whether HBV vaccine contributes to asthma in children, by stimulating specific IgE production. Total IgE, IgE- or IgG-anti-HBVs Abs were studied in vaccinated pediatric asthmatics and non asthmatics. We found: (1) total IgE was higher in asthmatics; (2) total IgE did not correlate with IgE anti-HBVs; (3) IgE anti-HBVs did correlate with IgG-anti-HBVs in all subjects; (4)IgE- and IgG-HBVs Abs were similar in both groups; (5) IgE- or IgG anti-HBVs Abs did not correlate with age. Our findings indicate that HBV vaccination induces IgE responses in asthmatics and non asthmatics.     

64层螺旋CT联合心肌酶检查应用于急性心源性胸痛诊断的临床价值分析

目的研究分析64层螺旋CT联合心肌酶检查应用于急性心源性胸痛诊断的临床价值。方法随机选取2008年2月~2011年3月我院收治的45例急性心源性胸痛患者作为治疗组,抽取同期来我院体检中心进行检查的45例健康者作为对照组,对照组单纯给予MSCTA(64层螺旋CT血管成像)进行检查。治疗组在此基础上联合血清心肌酶检查,观察两组患者的诊断情况。结果治疗组患者经MSCTA联合心肌酶检查诊断急性心源性胸痛特异性为100%,敏感性为95.56%,显著高于对照组93.33%、86.67%,组间差异显著,具有统计学意义(P0.05);经MSCTA分辨钙化粥样斑块CT值为(341.65±308.42)HU,非钙化粥样斑块CT值为(59.88±67.93)HU,差异显著,具有统计学意义(P0.05)。结论 64层螺旋CT联合心肌酶检查应用于诊断急性心源性胸痛效果良好,准确率较高,同时对判断冠状动脉斑块与狭窄性质具有积极的意义。     

G-protein βγ subunits are positive regulators of Kv7.4 and native vascular Kv7 channel activity

Kv7.4 channels are a crucial determinant of arterial diameter both at rest and in response to endogenous vasodilators. However, nothing is known about the factors that ensure effective activity of these channels. We report that G-protein βγ subunits increase the amplitude and activation rate of whole-cell voltage-dependent K⁺ currents sensitive to the Kv7 blocker linopirdine in HEK cells heterologously expressing Kv7.4, and in rat renal artery myocytes. In excised patch recordings, Gβγ subunits (2–250 ng /mL) enhanced the open probability of Kv7.4 channels without changing unitary conductance. Kv7 channel activity was also augmented by stimulation of G-protein–coupled receptors. Gallein, an inhibitor of Gβγ subunits, prevented these stimulatory effects. Moreover, gallein and two other structurally different Gβγ subunit inhibitors (GRK2i and a β-subunit antibody) abolished Kv7 channel currents in the absence of either Gβγ subunit enrichment or G-protein–coupled receptor stimulation. Proximity ligation assay revealed that Kv7.4 and Gβγ subunits colocalized in HEK cells and renal artery smooth muscle cells. Gallein disrupted this colocalization, contracted whole renal arteries to a similar degree as the Kv7 inhibitor linopirdine, and impaired isoproterenol-induced relaxations. Furthermore, mSIRK, which disassociates Gβγ subunits from α subunits without stimulating nucleotide exchange, relaxed precontracted arteries in a linopirdine-sensitive manner. These results reveal that Gβγ subunits are fundamental for Kv7.4 activation and crucial for vascular Kv7 channel activity, which has major consequences for the regulation of arterial tone.Increased arterial constriction and lack of responsiveness to endogenous vasodilators is a hallmark of vascular disease leading to poor health prognosis. Defining the factors that determine vascular smooth muscle (VSM) activity and modulation by vasorelaxant molecules is therefore imperative for a better understanding of vascular disease. Potassium channels are key regulators of VSM tone because they promote membrane hyperpolarization that limits the activity of voltage-dependent calcium channels known to precipitate vasoconstriction (1). The Kv7 family of voltage-dependent potassium channels and the Kv7.4 isoform, in particular, has a fundamental role in this process. There are five Kv7 isoforms (Kv7.1–Kv7.5) of which Kv7.1, Kv7.4, and Kv7.5 are consistently expressed within VSM, where the predominant molecular architecture is a Kv7.4/Kv7.5 heterotetramer (2, 3). Activation of Kv7 channels produces relaxation of numerous arteries (4–8), whereas blockade of Kv7 channels results in contraction of vessels at rest (7, 9–11) or an inhibition of endogenously derived vasorelaxations (2, 11–13). In addition, molecular reduction of Kv7.4 reduces responses to various Gs-coupled vasodilators in a number of arteries (2, 11). Crucially, Kv7.4 abundance is reduced in various arteries from hypertensive animals (6, 11, 12) where relaxant responses to endogenous vasodilators are also impaired (11, 12). Despite the key role of Kv7.4 channels in the regulation of VSM, and their involvement in mediating Gs-coupled vasodilator responses, the factors that regulate channel activity are poorly understood, and the signals linking Kv7.4 to Gs-receptor activation remain to be elucidated.G-protein–coupled receptor (GPCR) activation promotes the exchange of GDP for GTP resulting in disassociation of the heterotrimeric Gαβγ complex from the receptor into Gα-GTP and Gβγ (14). It is now established that the Gβγ complex as well as the Gα–GTP activates various intracellular signaling pathways (see refs. 15, 16 for reviews). Gβγ subunits also modulate various ion channels directly, a phenomenon of which there are only a handful of examples, with the positive regulation of an inwardly rectifying K⁺ channel in the heart the best characterized (17, 18). In this study, we explored whether Gβγ subunits modulated Kv7.4 channels and therefore function as signaling intermediates following receptor stimulation. Our results show that not only are Gβγ subunits able to enhance Kv7 channels, but also that they are a crucial requirement for the basal activity of the Kv7.4 channel.     

X-ray crystallographic and EPR spectroscopic analysis of HydG,a maturase in [FeFe]-hydrogenase H-cluster assembly

Hydrogenases use complex metal cofactors to catalyze the reversible formation of hydrogen. In [FeFe]-hydrogenases, the H-cluster cofactor includes a diiron subcluster containing azadithiolate, three CO, and two CN⁻ ligands. During the assembly of the H cluster, the radical S-adenosyl methionine (SAM) enzyme HydG lyses the substrate tyrosine to yield the diatomic ligands. These diatomic products form an enzyme-bound Fe(CO)_x(CN)_y synthon that serves as a precursor for eventual H-cluster assembly. To further elucidate the mechanism of this complex reaction, we report the crystal structure and EPR analysis of HydG. At one end of the HydG (βα)₈ triosephosphate isomerase (TIM) barrel, a canonical [4Fe-4S] cluster binds SAM in close proximity to the proposed tyrosine binding site. At the opposite end of the active-site cavity, the structure reveals the auxiliary Fe-S cluster in two states: one monomer contains a [4Fe-5S] cluster, and the other monomer contains a [5Fe-5S] cluster consisting of a [4Fe-4S] cubane bridged by a μ₂-sulfide ion to a mononuclear Fe²⁺ center. This fifth iron is held in place by a single highly conserved protein-derived ligand: histidine 265. EPR analysis confirms the presence of the [5Fe-5S] cluster, which on incubation with cyanide, undergoes loss of the labile iron to yield a [4Fe-4S] cluster. We hypothesize that the labile iron of the [5Fe-5S] cluster is the site of Fe(CO)_x(CN)_y synthon formation and that the limited bonding between this iron and HydG may facilitate transfer of the intact synthon to its cognate acceptor for subsequent H-cluster assembly.The assembly of the [FeFe]-hydrogenase diiron subcluster (1, 2) requires three maturase proteins, HydE, HydF, and HydG (3), and in vitro, they can assemble an active hydrogenase (4). The sequence and structure of the maturase HydE (5) indicates that it is a member of the radical S-adenosyl methionine (SAM) superfamily, although the biochemical function of HydE has not been experimentally determined. The GTPase HydF (6, 7) has been shown to transfer synthetic (8) or biologically derived (7, 9) diiron subclusters into apo-hydrogenase, suggesting that HydF functions as a template for diiron subcluster assembly. The tyrosine lyase HydG is also a member of the radical SAM superfamily and uses SAM and a reductant (such as dithionite) to cleave the Cα–Cβ bond of tyrosine, yielding p-cresol as the side chain-derived byproduct (10) and fragmenting the amino acid moiety into cyanide (CN⁻) (11) and carbon monoxide (CO) (12), which are ultimately incorporated as ligands in the H cluster of the [FeFe]-hydrogenase HydA (4). Two site-differentiated [4Fe-4S] clusters in HydG have been identified using a combination of spectroscopy and site-directed mutagenesis (12–16). The cluster bound close to the N terminus ([4Fe-4S]_RS) by the CX₃CX₂C cysteine triad motif (SI Appendix, Fig. S1) is typical of the radical SAM superfamily (17, 18) and has been shown to catalyze the reductive cleavage of SAM (11, 13). The resultant highly reactive 5′-deoxyadenosyl radical is thought to abstract a hydrogen atom from tyrosine, thereby inducing Cα–Cβ-bond homolysis with release of dehydroglycine (DHG) and the spectroscopically characterized 4-oxidobenzyl radical anion (16), which is quenched to yield p-cresol (Fig. 1A, step A). The second (auxiliary) Fe-S cluster is proposed to promote the conversion of DHG into CO and CN⁻ (Fig. 1A, step B) (13, 16). Two intermediates have been observed by stopped-flow IR spectroscopic analysis (19): an enzyme-bound organometallic species (complex A) (Fig. 1A, 4) that converts to a species that features an Fe(CO)₂(CN) moiety (complex B) (Fig. 1A, 5). These results, combined with ⁵⁷Fe electron-nuclear double resonance (ENDOR) studies that showed that iron from HydG is incorporated into mature hydrogenase, led to the proposal that an organometallic synthon with a minimum stoichiometry of [Fe(CO)₂CN] is synthesized at the auxiliary cluster of HydG and eventually transferred to apo-hydrogenase (19).Open in a separate windowFig. 1.Overall [FeFe]-hydrogenase H-cluster assembly and structure of TiHydG. (A) Formation of the Fe(CO)₂CN synthon is proposed to occur at the auxiliary cluster of HydG (square brackets). (B) Overall fold of HydG with an end-on view of the TIM barrel showing the radical SAM core (green), the N-terminal extension (pink), and the C-terminal extension (blue). Monomer A is shown and contains a [4Fe-4S] cluster to catalyze the formation of the 5′-deoxyadenosyl radical from SAM and a [5Fe-5S] auxiliary cluster proposed to promote the conversion of DHG into cyanide and carbon monoxide. (C) The position of the two Fe-S clusters in TiHydG. The strands of the TIM barrel are shown. The orientation is rotated 90° from B.Herein, we report the crystal structure of Thermoanaerobacter italicus HydG (TiHydG) complexed with SAM (the Protein Data Bank ID code for the structure of HydG is 4WCX). The structure, which contains two HydG monomers per asymmetric unit, reveals the auxiliary Fe-S cluster in two states: one monomer contains a [4Fe-5S] cluster, and the other monomer contains a structurally unprecedented [5Fe-5S] cluster consisting of a [4Fe-4S] cubane bridged by a μ₂-sulfide to a mononuclear Fe(II) center (which we term the labile iron). To supplement the crystallographic studies of TiHydG, we also report EPR spectroscopic studies of Shewanella oneidensis HydG (SoHydG) that provide solution-state characterization of the [5Fe-5S] cluster and show its conversion to a [4Fe-4S] cluster in the presences of exogenous cyanide. Taken together, these results support a proposed mechanism for [FeFe]-hydrogenase maturation in which the labile iron of the [5Fe-5S] cluster is the site for Fe(CO)_x(CN)_y synthon assembly.     

复合凝乳酶胶囊治疗儿童功能性消化不良临床研究

目的 观察使用复合凝乳酶胶囊治疗儿童功能性消化不良的效果及药物的安全性.方法 采用多中心、随机开放的临床试验方法,纳入2012年2月至2013年3月广东省4家三级甲等医院门诊收治的符合罗马Ⅲ标准诊断为功能性消化不良的患儿,观察并比较治疗1周及2周的总症状积分变化及药物相关不良反应,治疗效果根据总症状积分下降程度分为显效、有效及无效;并以疗效相差大于10％为优效标准对治疗1周及2周的有效性做非劣检验.结果 共有201例患儿纳入观察,196例完成2周治疗.治疗2周显效率、总有效率分别为68.88％、87.76％,明显高于治疗1周的显效率(27.04％)和总有效率(76.02％)(U=2.935,P＜0.05).治疗2周后和1周后症状积分改善率的差值、症状积分下降的差值及显效率变化差值的95％可信区间(95％ CI)下限均大于10％,治疗2周疗效优于治疗1周疗效.试验期间未见明显药物相关不良反应.结论 服用复合凝乳酶胶囊治疗消化不良对各症状改善明显.治疗2周较治疗1周的效果明显,无明显不良反应.