Solid-phase enzyme-linked immunosorbent assay for hepatitis E virus IgG and IgM antibodies utilizing recombinant antigens and synthetic peptides.

Four recombinant antigens representing two distinct antigenic domains from two different strains of hepatitis E virus (HEV), were used individually to develop four ELISAs designed to detect antibodies to HEV. Both IgG and IgM class antibodies to HEV were detected in 7 of 8 pedigreed serum/plasma from known outbreaks of HEV in Mexico, Burma, Somalia and Pakistan. In addition, specific HEV-antibodies were detected in cynomolgus macaques following inoculation with various HEV strains. Anti-HEV was also detected in 8 of 386 (2.1%) randomly selected American blood donors. Supplemental tests utilizing both synthetic peptides and specific blocking assays provided additional serologic data confirming the presence of anti-HEV. Similar prevalence studies on a limited number of available sera from other geographical regions (Alaska, Japan, Germany, New Zealand, Thailand and Mexico) confirmed the presence of anti-HEV in at least 1.1 to 7.6% of the specimens.     

Association of DJ-1 and parkin mediated by pathogenic DJ-1 mutations and oxidative stress

The identification of rare monogenic forms of Parkinson's disease (PD) has provided tremendous insight into the molecular pathogenesis of this disorder. Heritable mutations in alpha-synuclein, parkin, DJ-1 and PINK1 cause familial forms of PD. In the more common sporadic form of PD, oxidative stress and derangements in mitochondrial complex-I function are considered to play a prominent role in disease pathogenesis. However, the relationship of DJ-1 with other PD-linked genes and oxidative stress has not been explored. Here, we show that pathogenic mutant forms of DJ-1 specifically but differentially associate with parkin, an E3 ubiquitin ligase. Chemical cross-linking shows that pathogenic DJ-1 mutants exhibit impairments in homo-dimer formation, suggesting that parkin may bind to monomeric DJ-1. Parkin fails to specifically ubiquitinate and enhance the degradation of L166P and M26I mutant DJ-1, but instead promotes their stability in cultured cells. The interaction of parkin with L166P DJ-1 may involve a larger protein complex that contains CHIP and Hsp70, perhaps accounting for the lack of parkin-mediated ubiquitination. Oxidative stress also promotes an interaction between DJ-1 and parkin, but this does not result in the ubiquitination or degradation of DJ-1. Parkin-mediated alterations in DJ-1 protein stability may be pathogenically relevant as DJ-1 levels are dramatically increased in the detergent-insoluble fraction from sporadic PD/DLB brains, but are reduced in the insoluble fraction from parkin-linked autosomal recessive juvenile-onset PD brains. These data potentially link DJ-1 and parkin in a common molecular pathway at multiple levels that may have important implications for understanding the pathogenesis of inherited and sporadic PD.     

Contextual modulation of synchronization to random dots in the cat visual cortex

Synchronization of neuronal activity has been proposed as a binding mechanism for integration of image properties into one coherent percept. In the present study, we investigated the contextual modulation of synchronization to random dot patterns. Coherent motion of random dots evoked well synchronized responses in area 17 of anaesthetized cats when the stimulus was presented in the compound receptive field of recorded sites. Gradually changing the directional coherence of random dots in the surround while maintaining fully coherent motion of the stimulus in the receptive field significantly suppressed synchronization of neuronal activity for some stimulus conditions. However, usually one or two peaks of increased synchronization were found in the surround coherence tuning curves with low (8–12%) and/or moderate (25–50%) coherence in the surround. At the population level, synchronization was significantly depressed with incoherent motion in the receptive field and when both the surround and the receptive field were jointly stimulated with 0% coherence. The intriguing finding was the discovery of two distinct groups of cells with opposite synchronization changes dependent on the presence or absence of significant synchronization in their spontaneous activity. The latter group of neurons showed peaks of increased synchronization with lower surround coherence, thus probably being more sensitive to the direction of the surround motion. Overall, our findings support the notion that binding of stimulus properties can be achieved by synchronized activity of cortical cells. However, our findings go further than the original hypothesis of feature binding by synchrony to show that synchronization of cortical activity may be directly related to the decision making processes, which in turn are related to the threshold of perception of coherent motion.     

Brief intense exercise followed by passive recovery modifies the pattern of fuel use in humans during subsequent sustained intermittent exercise

The role of work period duration as the principal factor influencing carbohydrate metabolism during intermittent exercise has been investigated. Fuel oxidation rates and muscle glycogen and free carnitine content were compared between two protocols of sustained intermittent intense exercise with identical treadmill speed and total work duration. In the first experiment subjects (n=6) completed 40 min of intermittent treadmill running involving a work : recovery cycle of 6 : 9 s or 24 : 36 s on separate days. With 24 : 36 s exercise a higher rate of carbohydrate oxidation approached significance (P=0.057), whilst fat oxidation rate was lower (P ≤ 0.01) and plasma lactate concentration higher (P ≤ 0.01). Muscle glycogen was lower post‐exercise with 24 : 36 s (P ≤ 0.05). Muscle free carnitine decreased (P ≤ 0.05), but there was no difference between protocols. In the second experiment a separate group of subjects (n=5) repeated the intermittent exercise protocols with the addition of a 10‐min bout of intense exercise, followed by 43 ± 5 min passive recovery, prior to sustained (40 min) intermittent exercise. For this experiment the difference in fuel use observed previously between 6 : 9 s and 24 : 36 s was abolished. Carbohydrate and fat oxidation, plasma lactate and muscle glycogen levels were similar in 6 : 9 s and 24 : 36 s. When compared with the first experiment, this result was because of reduced carbohydrate oxidation in 24 : 36 s (P ≤ 0.05). There was no difference, and no change, in muscle free carnitine between protocols. A 10‐min bout of intense exercise, followed by 43 ± 5 min of passive recovery, substantially modifies fuel use during subsequent intermittent intense exercise.     

Sequence relations and coding properties of a subgenomic RNA isolated from barley stripe mosaic virus

A subgenomic (SG) RNA ( approximately 800 nucleotides) is a minor component of barley stripe mosaic virus RNAs. The SG-RNAs isolated from the Type and North Dakota 18 (ND18) strains of BSMV have sequence homology with RNA 2 of the ("pseudo-two component") Type strain, which has two electrophoretic components, but only limited homology is evident with RNA 2 of the ND18 and Norwich strains, which have three electrophoretic components ("three component" strain). Instead, eDNAs from SG-RNA hybridize most efficiently with RNA 3 of the ND18 and Norwich strains. In wheat germ extracts the SG-RNAs direct the synthesis of two polypeptides with apparent molecular weights of 20 to 21 x 10(3). However, these two polypeptides were difficult to detect by polyacrylamide gel electrophoresis of extracts from Type- or ND18-infected barley and so appear not to accumulate during infection.     

Mutations in Emery-Dreifuss muscular dystrophy and their effects on emerin protein expression

Seventeen families with Emery-Dreifuss muscular dystrophy (EDMD) have been studied both by DNA sequencing and by emerin protein expression. Fourteen had mutations in the X-linked emerin gene, while three showed evidence of autosomal inheritance. Twelve of the 14 emerin mutations caused early termination of translation. An in-frame deletion of six amino acids from the C-terminal transmembrane helix caused almost complete absence of emerin from muscle with no localization to the nuclear membrane, although mRNA levels were normal. This shows that mutant emerin proteins are unstable if they are unable to integrate into a membrane. A 22 bp deletion in the promoter region was expected to result in reduced emerin production, but normal amounts of emerin of normal size were found in leucocytes and lymphoblastoid cell lines. This shows that DNA analysis is necessary to exclude emerin mutations in suspected X-linked EDMD. Emerin levels in female carriers often deviated from the expected 50% and this was due, in at least two families, to skewed emerin mRNA expression from the normal and mutated alleles. In one family with a novel deletion of the last three exons of the emerin gene, a carrier had a cardiomyopathy and very low emerin levels (<5% of normal) due to skewed X-inactivation. In the three autosomal cases of EDMD, emerin was normal on western blots of blood cells, which suggests that autosomal EDMD is not caused by indirect reduction of emerin levels.      

Anxiety during pregnancy and fetal attachment after in-vitro fertilization conception

The aim of this study was to compare 70 couples who had conceived by in- vitro fertilization (IVF) with 63 matched controls for the prevalence of anxiety and quality of attachment to the baby during pregnancy. Results for mothers showed no group differences using a global measure of anxiety, the Spielberger State-Trait Anxiety Inventory. However, pregnancy-specific measures revealed significantly higher levels of anxiety in IVF mothers about the survival and normality of their unborn babies, about damage to their babies during childbirth and about separating from their babies after birth. When IVF mothers were differentiated according to the number of treatment cycles, more differences in anxiety level were revealed, with most increases occurring in mothers who had experienced two or more treatment cycles. IVF fathers did not differ from controls on the global anxiety measure. No data on pregnancy-specific anxiety were available for fathers. Neither IVF mothers nor IVF fathers differed from controls on measures of attachment to the baby during pregnancy. Results are discussed in the context of the need for researchers to employ differentiated and issue-specific measures to identify concerns that may be unique to IVF couples. Clinical implications regarding the need for psychological support during pregnancy are also discussed.