Topoisomerase activity assays in Neurospora

DNA topoisomerases are enzymes capable of altering the topological conformation of DNA by inducing transient single (Topoisomerase I) and double strand (Topoisomerase II) breaks

Comparison of grouper assemblages in northern areas of the wider Caribbean: A preliminary assessment

Groupers (Pisces: Serranidae) are important top-level predators in wider Caribbean, but have experienced significant exploitation, resulting in declines in abundance, size, spawning aggregations, and changes in species composition. Larger groupers are particularly vulnerable to intense fishing because of their longevity, slow growth, delayed reproduction, and aggregate spawning. Marine fishery reserves (MFR), areas permanently closed to consumptive use, offer a viable means to protect grouper resources. This study reports on fishery-independent surveys of groupers in four regions of the tropical western Atlantic during 1995 - 1997: Florida Keys, central Bahamas, southeastern Cuba, and Dominican Republic. The regions surveyed included two national parks and a national marine sanctuay, and were further categorized as: 1) intensively fished with little or no management for groupers (Cuba, Dominican Republic); 2) intensively fished with gear and effort limitations (Florida Keys); 3) lightly fished with some management (N. and S. Exuma Cays, Bahamas); and 4) a. MFR closed to fishing since 1986 (Exuma Cays Land and Sea Park, Bahamas). From 10 - 20 strip transects (20 m x 5 m) were surveyed in shallow-water (l-20 m depth) hard-bottom habitats for grouper species composition, density, and size distribution. Nine grouper species (6 Epinephelus spp., 3 Mycteroperca spp.) were documented among all regions. Areas in which groupers were partially or wholly protected from fishing had greater grouper diversity, density, and biomass, particularly for targeted species such as Nassau grouper (E. striatus). Classification of groupers by three size classes (small, intermediate, large) indicated a distinct gradient from areas with intense fishing to the MFR. In three of the regions affected by intense fishing, one of which has several grouper fishery regulations, grouper abundance and biomass were dominated by non-targeted species such as the graysby (E. cruentatus) and coney (E. fulvus). This second-order effect of fishing probably indicates competitive or predation. MFRs represent a viable means to protect grouper resources, alleviating the complications of enforcement and partially the need to gather fisheries dependent data. The ability of groupers to recover in certain regions may be deterred because of reduced larval recruitment from upstream, heavily fished sources

Norepinephrine-evoked pain in fibromyalgia. A randomized pilot study [ISRCTN70707830]

BACKGROUND: Fibromyalgia syndrome displays sympathetically maintained pain features such as frequent post-traumatic onset and stimuli-independent pain accompanied by allodynia and paresthesias. Heart rate variability studies showed that fibromyalgia patients have changes consistent with ongoing sympathetic hyperactivity. Norepinephrine-evoked pain test is used to assess sympathetically maintained pain syndromes. Our objective was to define if fibromyalgia patients have norepinephrine-evoked pain. METHODS: Prospective double blind controlled study. Participants: Twenty FM patients, and two age/sex matched control groups; 20 rheumatoid arthritis patients and 20 healthy controls. Ten micrograms of norepinephrine diluted in 0.1 ml of saline solution were injected in a forearm. The contrasting substance, 0.1 ml of saline solution alone, was injected in the opposite forearm. Maximum local pain elicited during the 5 minutes post-injection was graded on a visual analog scale (VAS). Norepinephrine-evoked pain was diagnosed when norepinephrine injection induced greater pain than placebo injection. Intensity of norepinephrine-evoked pain was calculated as the difference between norepinephrine minus placebo-induced VAS scores. RESULTS: Norepinephrine-evoked pain was seen in 80 % of FM patients (95% confidence intervals 56.3 – 94.3%), in 30 % of rheumatoid arthritis patients and in 30 % of healthy controls (95% confidence intervals 11.9 – 54.3) (p < 0.05). Intensity of norepinephrine-evoked pain was greater in FM patients (mean ± SD 2.5 ± 2.5) when compared to rheumatoid arthritis patients (0.3 ± 0.7), and healthy controls (0.3 ± 0.8) p < 0.0001. CONCLUSIONS: Fibromyalgia patients have norepinephrine-evoked pain. This finding supports the hypothesis that fibromyalgia may be a sympathetically maintained pain syndrome

A SCN9A gene-encoded dorsal root ganglia sodium channel polymorphism associated with severe fibromyalgia

<p>Abstract</p> <p>Background</p> <p>A consistent line of investigation suggests that autonomic nervous system dysfunction may explain the multi-system features of fibromyalgia (FM); and that FM is a sympathetically maintained neuropathic pain syndrome. Dorsal root ganglia (DRG) are key sympathetic-nociceptive short-circuit sites. Sodium channels located in DRG (particularly Nav1.7) act as molecular gatekeepers for pain detection. Nav1.7 is encoded in gene SCN9A of chromosome 2q24.3 and is predominantly expressed in the DRG pain-sensing neurons and sympathetic ganglia neurons. Several SCN9A sodium channelopathies have been recognized as the cause of rare painful dysautonomic syndromes such as paroxysmal extreme pain disorder and primary erythromelalgia. The aim of this study was to search for an association between fibromyalgia and several SCN9A sodium channels gene polymorphisms.</p> <p>Methods</p> <p>We studied 73 Mexican women suffering from FM and 48 age-matched women who considered themselves healthy. All participants filled out the Fibromyalgia Impact Questionnaire (FIQ). Genomic DNA from whole blood containing EDTA was extracted by standard techniques. The following SCN9A single-nucleotide polymorphisms (SNP) were determined by 5' exonuclease TaqMan assays: rs4371369; rs4387806; rs4453709; rs4597545; rs6746030; rs6754031; rs7607967; rs12620053; rs12994338; and rs13017637.</p> <p>Results</p> <p>The frequency of the rs6754031 polymorphism was significantly different in both groups (<it>P </it>= 0.036) mostly due to an absence of the GG genotype in controls. Interestingly; patients with this rs6754031 GG genotype had higher FIQ scores (median = 80; percentile 25/75 = 69/88) than patients with the GT genotype (median = 63; percentile 25/75 = 58/73; <it>P </it>= 0.002) and the TT genotype (median = 71; percentile 25/75 = 64/77; <it>P </it>= 0.001).</p> <p>Conclusion</p> <p>In this ethnic group; a disabling form of FM is associated to a particular SCN9A sodium channel gene variant. These preliminary results raise the possibility that some patients with severe FM may have a dorsal root ganglia sodium channelopathy.</p

Density, species, and size distribution of groupers (Serranidae) in three habitats at Elbow Reef, Florida Keys

We examined the density, size and species distribution of groupers in three habitats on an inshore-to-offshore transect across Elbow Reef, Florida Keys: high-relief spur-and-groove (4–9 m depth), relict spur-and-groove (10–20 m), and deep fore reef slope (21–30 m). Physical relief was greatest in the high-relief spur-and-groove (up to 3 m), lowest in the relict spur-and-groove habitat (30%). There were significant differences in the density, size, and species distribution of groupers among the three habitats. Graysby, Epinephelus cruentatus, was numerically dominant, constituting 82–91% of individual observed. Black grouper, Mycteroperca bonaci, and Nassau grouper, E. striatus, were more abundant in high to moderate relief habitats, whereas red hind, E. guttatus, was more abundant in the low-relief habitat. The size distribution was shifted towards smaller sizes in lowest relief habitat and towards larger sizes in areas with greater (\u3e0.5 m) vertical relief. We suggest that fishing pressure in the Florida Keys has resulted in an offshore grouper assemblage dominated by graysby, a small grouper species (length) which is not targeted by fishermen, and that habitat selection and biological interactions have significantly influenced the ecological structure of the grouper assemblage of this coral reef

The diversity of bioactive proteins in Australian snake venoms

Australian elapid snakes are among the most venomous in the world. Their venoms contain multiple components that target blood hemostasis, neuromuscular signaling, and the cardiovascular system. We describe here a comprehensive approach to separation and identification of the venom proteins from 18 of these snake species, representing nine genera. The venom protein components were separated by two-dimensional PAGE and identified using mass spectrometry and de novo peptide sequencing. The venoms are complex mixtures showing up to 200 protein spots varying in size from 10. These include many proteins identified previously in Australian snake venoms, homologs identified in other snake species, and some novel proteins. In many cases multiple trains of spots were typically observed in the higher molecular mass range (> 20 kDa) (indicative of post-translational modification). Venom proteins and their post-translational modifications were characterized using specific kantibodies, phosphoprotein- and glycoprotein-specific stains, enzymatic digestion, lectin binding, and antivenom reactivity. In the lower molecular weight range, several proteins were identified, but the predominant species were phospholipase A(2) and alpha-neurotoxins, both represented by different sequence variants. The higher molecular weight range contained proteases, nucleotidases, oxidases, and homologs of mammalian coagulation factors. This information together with the identification of several novel proteins (metalloproteinases, vespryns, phospholipase A(2) inhibitors, protein-disulfide isomerase, 5'-nucleotidases, cysteinerich secreted proteins, C-type lectins, and acetylcholinesterases) aids in understanding the lethal mechanisms of elapid snake venoms and represents a valuable resource for future development of novel human therapeutics

Hemps, a novel EGF-like protein, plays a central role in ascidian metamorphosis

All chordates share several characteristic features including a dorsal hollow neural tube, a notochord, a pharynx and an endostyle. Unlike other chordate taxa, ascidians have a biphasic life-history with two distinct body plans. During metamorphosis, the larval nerve cord and notochord degenerate and the pharyngeal gill slits and endostyle form. While ascidians, like other marine invertebrates, metamorphose in response to specific environmental cues, it remains unclear how these cues trigger metamorphosis. We have identified a novel gene (Hemps) which encodes a protein with a putative secretion signal sequence and four epidermal growth factor (EGF)-like repeats which is a key regulator of metamorphosis in the ascidian Herdmania curvata. Expression of Hemps increases markedly when the swimming tadpole larva becomes competent to undergo metamorphosis and then during the first 24 hours of metamorphosis. The Hemps protein is localised to the larval papillae and anterior epidermis of the larva in the region known to be required for metamorphosis. When the larva contacts an inductive cue the protein is released, spreading posteriorly and into the tunic as metamorphosis progresses. Metamorphosis is blocked by incubating larvae in anti-Hemps antibodies prior to the addition of the cue. Addition of recombinant Hemps protein to competent larvae induces metamorphosis in a concentration-dependent manner. A subgroup of genes are specifically induced during this process. These results demonstrate that the Hemps protein is a keg regulator of ascidian metamorphosis and is distinct from previously described inducers of this process in terrestrial arthropods and aquatic vertebrates

Imaging of Dysfunctional Elastogenesis in Atherosclerosis Using an Improved Gadolinium-Based Tetrameric MRI Probe Targeted to Tropoelastin

Dysfunctional elastin turnover plays a major role in the progression of atherosclerotic plaques. Failure of tropoelastin cross-linking into mature elastin leads to the accumulation of tropoelastin within the growing plaque, increasing its instability. Here we present Gd4-TESMA, an MRI contrast agent specifically designed for molecular imaging of tropoelastin within plaques. Gd4-TESMA is a tetrameric probe composed of a tropoelastin-binding peptide (the VVGS-peptide) conjugated with four Gd(III)-DOTA-monoamide chelates. It shows a relaxivity per molecule of 34.0 ± 0.8 mM-1 s-1 (20 MHz, 298 K, pH 7.2), a good binding affinity to tropoelastin (KD = 41 ± 12 μM), and a serum half-life longer than 2 h. Gd4-TESMA accumulates specifically in atherosclerotic plaques in the ApoE-/- murine model of plaque progression, with 2 h persistence of contrast enhancement. As compared to the monomeric counterpart (Gd-TESMA), the tetrameric Gd4-TESMA probe shows a clear advantage regarding both sensitivity and imaging time window, allowing for a better characterization of atherosclerotic plaques