Effect of Alcoholic Intoxication on the Risk of Inflammatory Bowel Disease: A Nationwide Retrospective Cohort Study

PurposeThis study investigated whether alcoholic intoxication (AI) increases the risk of inflammatory bowel disease (IBD) by using a population-based database in Taiwan.MethodsThis retrospective matched-cohort study included 57 611 inpatients with new-onset AI (AI cohort) and 230 444 randomly selected controls (non-AI cohort). Each patient was monitored for 10 years to individually identify those who were subsequently diagnosed with Crohn disease (CD) and ulcerative colitis (UC) during the follow-up period. Cox proportional hazard regression analysis was conducted to determine the risk of IBD in patients with AI compared with controls without AI.ResultsThe incidence rate of IBD during the 10-year follow-up period was 2.69 per 1 000 person-years and 0.49 per 1 000 person-years in the AI and non-AI cohorts, respectively. After adjustment for age, sex, and comorbidity, the AI cohort exhibited a 3.17-fold increased risk of IBD compared with the non-AI cohort (hazard ratio [HR] = 3.17, 95% confidence interval [CI] = 2.19–4.58). Compared with the non-AI cohort, the HRs of CD and UC were 4.40 and 2.33 for the AI cohort, respectively. After stratification for the severity of AI according to the duration of hospital stay, the adjusted HRs exhibited a significant correlation with the severity; the HRs of IBD were 1.76, 6.83, and 19.9 for patients with mild, moderate, and severe AI, respectively (p for the trend < .0001).ConclusionThe risk of IBD was higher in patients with AI and increased with the length of hospital stay.     

Structure and function of LGP2, a DEX(D/H) helicase that regulates the innate immunity response

RNA recognition receptors are important for detection of and response to viral infections. RIG-I and MDA5 are cytoplasmic DEX(D/H) helicase proteins that can induce signaling in response to RNA ligands, including those from viral infections. LGP2, a homolog of RIG-I and MDA5 without the caspase recruitment domain required for signaling, plays an important role in modulating signaling by MDA5 and RIG-I, presumably through heterocomplex formation and/or by serving as a sink for RNAs. Here we demonstrate that LGP2 can be coexpressed with RIG-I to inhibit activation of the NF-kappaB reporter expression and that LGP2 protein produced in insect cells can bind both single- and double-stranded RNA (dsRNA), with higher affinity and cooperativity for dsRNA. Electron microscopy and image reconstruction were used to determine the shape of the LGP2 monomer in the absence of dsRNA and of the dimer complexed to a 27-bp dsRNA. LGP2 has striking structural similarity to the helicase domain of the superfamily 2 DNA helicase, Hef.     

腺病毒介导犬干扰素-γ基因的表达及其体外抗犬细小病毒的作用

[目的]构建含犬干扰素-γ(c IFN-γ)基因的重组腺病毒,并在培养的犬肾细胞MDCK中分析其抗犬细小病毒的活性.[方法]首先将cIFN-γcNDA基因克隆到腺病毒穿梭质粒中,构建成含cIFN-γ基因的腺病毒穿梭质粒pShuttle3-cIFN-γ.利用特异的酶切位点,通过直接连接法将cIFN-γ表达盒插入到腺病毒基因组质粒pAdeno-X中,构建成含cIFN-γ基因的腺病毒基因组质粒pAd-cIFN-γ.pAd-cIFN-γ质粒经酶切线性化后转染人胚胎肾细胞HEK293T,在细胞中拯救出含有cIFN-γ基因的复制缺陷型重组腺病毒.然后用该重组腺病毒处理(感染)培养的犬肾细胞MDCK,再用犬细小病毒感染重组腺病毒处理的细胞,分析重组腺病毒在体外抗犬细小病毒的活性.[结果]通过连接法构建了含cIFN-γ基因的重组腺病毒,构建的重组腺病毒能够介导cIFN-γ在MDCK细胞中进行分泌表达.用含cIFN-γ基因的重组腺病毒处理MDCK细胞,可明显地抑制犬细小病毒在细胞中的增殖,表明构建的重组腺病毒具有明显的抗犬细小病毒的活性.[结论]构建了含cIFN-γ基因的重组腺病毒,并证明该重组病毒在体外具有明显的抗犬细小病毒的活性.     

Characterization of the membrane domain Nqo11 subunit of the proton-translocating NADH-quinone oxidoreductase of Paracoccus denitrificans

The proton-translocating NADH-quinone oxidoreductase (NDH-1) of Paracoccus denitrificans consists of at least 14 unlike subunits (designated Nqo1-14). The NDH-1 is composed of two segments (the peripheral and membrane segments). The membrane domain segment appears to be made up of seven subunits (Nqo7, -8, -10-14). In this report, the characterization of the Paracoccus Nqo11 subunit has been investigated. An antibody against the C-terminal 12 amino acid residues of the Paracoccus Nqo11 subunit (Nqo11c) has been raised. The Nqo11c antibody reacted with a single band (11 kDa) of the Paracoccus membranes and cross-reacted with Rhodobactor capsulatus membranes. The Nqo11 subunit was not able to be extracted from the Paracoccus membranes by NaI or alkaline treatment, unlike the peripheral subunits (Nqo1 and Nqo6). The C-terminal region of the Paracoccus Nqo11 is exposed to the cytoplasmic phase. For further characterization of the Paracoccus Nqo11 subunit, the subunit was overexpressed in Escherichia coli by using the maltose-binding protein (MBP) fusion system. The MBP-fused Nqo11 subunit was expressed in the E. coli membranes (but not in soluble phase) and was extracted by Triton X-100. The isolated MBP-fused Nqo11 subunit interacted with the phospholipid vesicles and suppressed their membrane fluidity. Topological studies of the Nqo11 subunit expressed in E. coli membranes have been performed by using cysteine mapping and immunochemical analyses. The data suggest that the Nqo11 subunit has three transmembrane segments and its C-terminus protrudes into the cytoplasmic phase.     

Epac1 mediates protein kinase A–independent mechanism of forskolin-activated intestinal chloride secretion

Intestinal Cl⁻ secretion is stimulated by cyclic AMP (cAMP) and intracellular calcium ([Ca²⁺]_i). Recent studies show that protein kinase A (PKA) and the exchange protein directly activated by cAMP (Epac) are downstream targets of cAMP. Therefore, we tested whether both PKA and Epac are involved in forskolin (FSK)/cAMP-stimulated Cl⁻ secretion. Human intestinal T84 cells and mouse small intestine were used for short circuit current (I_sc) measurement in response to agonist-stimulated Cl⁻ secretion. FSK-stimulated Cl⁻ secretion was completely inhibited by the additive effects of the PKA inhibitor, H89 (1 µM), and the [Ca²⁺]_i chelator, 1,2-bis-(o-aminophenoxy)-ethane-N,N,N’,N’-tetraacetic acid, tetraacetoxymethyl ester (BAPTA-AM; 25 µM). Both FSK and the Epac activator 8-pCPT-2’-O-Me-cAMP (50 µM) elevated [Ca²⁺]_i, activated Ras-related protein 2, and induced Cl⁻ secretion in intact or basolateral membrane–permeabilized T84 cells and mouse ileal sheets. The effects of 8-pCPT-2’-O-Me-cAMP were completely abolished by BAPTA-AM, but not by H89. In contrast, T84 cells with silenced Epac1 had a reduced I_sc response to FSK, and this response was completely inhibited by H89, but not by the phospholipase C inhibitor {type:entrez-nucleotide,attrs:{text:U73122,term_id:4098075,term_text:U73122}}U73122 or BAPTA-AM. The stimulatory effect of 8-pCPT-2’-O-Me-cAMP on Cl⁻ secretion was not abolished by cystic fibrosis transmembrane conductance (CFTR) inhibitor 172 or glibenclamide, suggesting that CFTR channels are not involved. This was confirmed by lack of effect of 8-pCPT-2’-O-Me-cAMP on whole cell patch clamp recordings of CFTR currents in Chinese hamster ovary cells transiently expressing the human CFTR channel. Furthermore, biophysical characterization of the Epac1-dependent Cl⁻ conductance of T84 cells mounted in Ussing chambers suggested that this conductance was hyperpolarization activated, inwardly rectifying, and displayed a Cl⁻>Br⁻>I⁻ permeability sequence. These results led us to conclude that the Epac-Rap-PLC-[Ca²⁺]_i signaling pathway is involved in cAMP-stimulated Cl⁻ secretion, which is carried by a novel, previously undescribed Cl⁻ channel.     

A software package for cDNA microarray data normalization and assessing confidence intervals

DNA microarray data are affected by variations from a number of sources. Before these data can be used to infer biological information, the extent of these variations must be assessed. Here we describe an open source software package, lcDNA, that provides tools for filtering, normalizing, and assessing the statistical significance of cDNA microarray data. The program employs a hierarchical Bayesian model and Markov Chain Monte Carlo simulation to estimate gene-specific confidence intervals for each gene in a cDNA microarray data set. This program is designed to perform these primary analytical operations on data from two-channel spotted, or in situ synthesized, DNA microarrays.     

MATERNAL EFFECTS,BUT NO GOOD OR COMPATIBLE GENES FOR SPERM COMPETITIVENESS IN AUSTRALIAN CRICKETS

Explanations for the evolution of polyandry often center on the idea that females garner genetic benefits for their offspring by mating multiply. Furthermore, postcopulatory processes are thought to be fundamental to enabling polyandrous females to screen for genetic quality. Much attention has focused on the potential for polyandrous females to accrue such benefits via a sexy‐ or good‐sperm mechanism, whereby additive variation exists among males in sperm competitiveness. Likewise, attention has focused on an alternative model, in which offspring quality (in this context, the sperm competitiveness of sons) hinges on an interaction between parental haplotypes (genetic compatibility). Sperm competitiveness that is contingent on parental compatibility will exhibit nonadditive genetic variation. We tested these models in the Australian cricket, Teleogryllus oceanicus, using a design that allowed us to partition additive, nonadditive genetic, and parental variance for sperm competitiveness. We found an absence of additive and nonadditive genetic variance in this species, challenging the direct relevance of either model to the evolution of sperm competitiveness in particular, and polyandry in general. Instead, we found maternal effects that were possibly sex‐linked or cytoplasmically linked. We also found effects of focal male age on sperm competitiveness, with small increments in age conferring more competitive sperm.     

Iron promotes the toxicity of amyloid beta peptide by impeding its ordered aggregation

We have previously shown that overexpressing subunits of the iron-binding protein ferritin can rescue the toxicity of the amyloid β (Aβ) peptide in our Drosophila model system. These data point to an important pathogenic role for iron in Alzheimer disease. In this study, we have used an iron-selective chelating compound and RNAi-mediated knockdown of endogenous ferritin to further manipulate iron in the brain. We confirm that chelation of iron protects the fly from the harmful effects of Aβ. To understand the pathogenic mechanisms, we have used biophysical techniques to see how iron affects Aβ aggregation. We find that iron slows the progression of the Aβ peptide from an unstructured conformation to the ordered cross-β fibrils that are characteristic of amyloid. Finally, using mammalian cell culture systems, we have shown that iron specifically enhances Aβ toxicity but only if the metal is present throughout the aggregation process. These data support the hypothesis that iron delays the formation of well ordered aggregates of Aβ and so promotes its toxicity in Alzheimer disease.