Milt characteristics and plasma levels of gonadal steroids in greenback flounder Rhombosolea tapirina following treatment with exogenous hormones

Fish were treated with exogenous hormones, and milt and blood samples were collected for up to 96 h post‐treatment. Blood plasma samples were assayed for the gonadal steroids testosterone (T), 11‐ketotestosterone (11KT) and 17,20ß‐dihydroxy‐4‐pregnen‐3‐one (17.20ßP). Milt volume, spermatocrit and sperm motility were measured from milt samples. Non‐spermiated fish showed increased plasma T and 11KT in response to human chorionic gonadotropin (hCG) but not luteinising hormone releasing hormone analogue (LHRHa). Fish did not become spermiated in response to treatment with hCG, LHRHa, 11KT, 17‐hydroxyprogesterone (17P) or 17,20ßP. Spermiated fish showed an increase in milt volume in response to hCG and LHRHa but not exogenous steroids. Sperm motility declined to zero over 120 s and was not affected by hormone treatment or sampling time. Increased milt volume was accompanied by increased plasma T and 11KT, but not 17.20ßP levels. In a separate experiment, LHRHa delivered by injection or pellet was equally effective at increasing milt volume but had no effect on plasma steroid levels. Spermatocrit declined with stripping but was not affected by hormone treatment, nor was sperm motility. Co‐treatment of fish with 17P plus LHRHa had no additive effect on plasma steroid concentrations or milt volume. The results suggest that as in other teleosts, gonadotropin mediates milt production in greenback flounder.     

Temporal and spatial changes in the microbial bioaerosol communities in green-waste composting

In this study, the microbial community within compost, emitted into the airstream, downwind and upwind from a composting facility was characterized and compared through phospholipid fatty acid analysis and 16S rRNA gene analysis using denaturing gradient gel electrophoresis and bar-coded pyrosequencing techniques. All methods used suggested that green-waste composting had a significant impact upon bioaerosol community composition. Daily variations of the on-site airborne community showed how specific site parameters such as compost process activity and meteorological conditions affect bioaerosol communities, although more data are required to qualify and quantify the causes for these variations. A notable feature was the dominance of Pseudomonas in downwind samples, suggesting that this genus can disperse downwind in elevated abundances. Thirty-nine phylotypes were homologous to plant or human phylotypes containing pathogens and were found within compost, on-site and downwind microbial communities. Although the significance of this finding in terms of potential health impact was beyond the scope of this study, it clearly illustrated the potential of molecular techniques to improve our understanding of the impact that green-waste composting emissions may have on the human health.     

Safety Implications of High-Field MRI: Actuation of Endogenous Magnetic Iron Oxides in the Human Body

Background

Magnetic Resonance Imaging scanners have become ubiquitous in hospitals and high-field systems (greater than 3 Tesla) are becoming increasingly common. In light of recent European Union moves to limit high-field exposure for those working with MRI scanners, we have evaluated the potential for detrimental cellular effects via nanomagnetic actuation of endogenous iron oxides in the body.

Methodology

Theoretical models and experimental data on the composition and magnetic properties of endogenous iron oxides in human tissue were used to analyze the forces on iron oxide particles.

Principal Finding and Conclusions

Results show that, even at 9.4 Tesla, forces on these particles are unlikely to disrupt normal cellular function via nanomagnetic actuation.     

Characterization of specifically oxidized apolipoproteins in mildly oxidized high density lipoprotein

Atherosclerosis is a state of heightened oxidative stress. Oxidized LDL is present in atherosclerotic lesions and used as marker for coronary artery disease, although in human lesions lipids associated with HDL are as oxidized as those of LDL. Here we investigated specific changes occurring to apolipoprotein A-I (apoA-I) and apoA-II, as isolated HDL and human plasma undergo mild, chemically induced oxidation, or autoxidation. During such oxidation, Met residues in apoA-I and apoA-II become selectively and consecutively oxidized to their respective Met sulfoxide (MetO) forms that can be separated by HPLC. Placing plasma at -20 degrees C prevents autoxidation, whereas metal chelators and butylated hydroxytoluene offer partial protection. Independent of the oxidation conditions, apoA-I and apoA-II (dimer) with two MetO residues accumulate as relatively stable oxidation products. Compared to controls, serum samples from subjects with the endothelial cell nitric oxide synthase a/b genotype that is associated with increased coronary artery disease contain increased concentrations of apoA-I with two MetO residues. Our results show that during the early stages, oxidation of HDL gives rise to specifically oxidized forms of apoA-I and apoA-II, some of which may be useful markers of in vivo HDL oxidation, and hence potentially atherosclerosis.     

Preliminary evaluation of nanoscale biogenic magnetite in Alzheimer's disease brain tissue

Elevated iron levels are associated with many types of neurodegenerative disease, such as Alzheimer's, Parkinson's and Huntington's diseases. However, these elevated iron levels do not necessarily correlate with elevated levels of the iron storage or transport proteins, ferritin and transferrin. As such, little is known about the form of this excess iron. It has recently been proposed that some of the excess iron in neurodegenerative tissue may be in the form of the magnetic iron oxide magnetite (Fe(3)O(4)). We demonstrate, for the first time to our knowledge, using highly sensitive superconducting quantum interference device (SQUID) magnetometry, that the concentrations of magnetite are found to be significantly higher in three samples of Alzheimer's disease tissue than in three age- and sex-matched controls. These results have implications, not only for disease progression, but also for possible early diagnosis.     

The propeptide of the transforming growth factor-beta superfamily member, macrophage inhibitory cytokine-1 (MIC-1), is a multifunctional domain that can facilitate protein folding and secretion

Macrophage inhibitory cytokine-1 (MIC-1) is a divergent member of the transforming growth factor-beta (TGF-beta) superfamily. While it is synthesized in a pre-pro form, it is unique among superfamily members because it does not require its propeptide for correct folding or secretion of the mature peptide. To investigate factors that enable these propeptide independent events to occur, we constructed MIC-1/TGF-beta1 chimeras, both with and without a propeptide. All chimeras without a propeptide secreted less efficiently compared with the corresponding constructs with propeptide. Folding and secretion were most affected after replacement of the predicted major alpha-helix in the mature protein, residues 56-68. Exchanging the human propeptide in this chimera with either the murine MIC-1 or TGF-beta1 propeptide resulted in secretion of the unprocessed, monomeric chimera, suggesting a specific interaction between the human MIC-1 propeptide and mature peptide. Propeptide deletion mutants enabled identification of a region between residues 56 and 78, which is important for the interaction between the propeptide and the mature peptide. Cotransfection experiments demonstrated that the propeptide must be in cis with the mature peptide for this phenomenon to occur. These results suggest a model for TGF-beta superfamily protein folding.     

Differential effects of temperature and maturity stage on hepatic estrogen receptor characteristics of Atlantic salmon

In order to determine if elevated temperature during vitellogenesis had a detrimental effect on hepatic estrogen receptors of Atlantic salmon (Salmo salar), 3H-estradiol saturation binding analysis, using one- and two-site binding models, was carried out on extracts of hepatic cytosols from fish held at 14, 18 or 22 degrees C over the austral period of peak vitellogenesis (February to April). With one-site binding analysis, no temperature related difference in either receptor affinity (Kd) or number (Bmax) was found at each sampling point, but there was an apparent decrease in both affinity and number at each temperature over the period of the study. However, some analyses, notably at 22 degrees C during February, were best described using a two-site binding model. At this temperature and time, there was a clear separation of binding affinity into high and low components (Kd = 0.67+/-S.E. 0.05 and 20+/-S.E. 5.6 nM, respectively) (n = 4), which suggests that February was a critical time of temperature related hepatic sensitivity to estrogen. These results support those of other studies where we found that February was also a sensitive time with respect to temperature impairment of in vitro follicular estrogen synthesis, and the greatest period of in vivo temperature sensitivity.     

Engineering water to act as an active site acid catalyst in a soluble fumarate reductase

The ability of an arginine residue to function as the active site acid catalyst in the fumarate reductase family of enzymes is now well-established. Recently, a dual role for the arginine during fumarate reduction has been proposed [Mowat, C. G., Moysey, R., Miles, C. S., Leys, D., Doherty, M. K., Taylor, P., Walkinshaw, M. D., Reid, G. A., and Chapman, S. K. (2001) Biochemistry 40, 12292-12298] in which it acts both as a Lewis acid in transition-state stabilization and as a Br?nsted acid in proton delivery. This proposal has led to the prediction that, if appropriately positioned, a water molecule would be capable of functioning as the active site Br?nsted acid. In this paper, we describe the construction and kinetic and crystallographic analysis of the Q363F single mutant and Q363F/R402A double mutant forms of flavocytochrome c(3), the soluble fumarate reductase from Shewanella frigidimarina. Although replacement of the active site acid, Arg402, with alanine has been shown to eliminate fumarate reductase activity, this phenomenon is partially reversed by the additional substitution of Gln363 with phenylalanine. This Gln --> Phe substitution in the inactive R402A mutant enzyme was designed to "push" a water molecule close enough to the substrate C3 atom to allow it to act as a Br?nsted acid. The 2.0 A resolution crystal structure of the Q363F/R402A mutant enzyme does indeed reveal the introduction of a water molecule at the correct position in the active site to allow it to act as the catalytic proton donor. The 1.8 A resolution crystal structure of the Q363F mutant enzyme shows a water molecule similarly positioned, which can account for its measured fumarate reductase activity. However, in this mutant enzyme Michaelis complex formation is impaired due to significant and unpredicted structural changes at the active site.     

Effect of density on the reproductive behaviour of the territorial male demoiselle Chromis dispilus (Pisces: Pomacentridae)

Synopsis Characteristics of nest sites and reproductive behaviour of territorial male demoiselles Chromis dispilus were examined at sites of low, medium and high population density in northeastern New Zealand, by use of SCUBA diving and a remote underwater video system. Nest territories were closer together at high density (relative to areas of low population density) and this was accompanied by an increase in the frequency of spawning sequences and territorial defence against conspecifics. Territorial males were more aggressive during periods of display and spawning than during egg guarding. Fish from areas of low density spent more time on courtship display (signal jumps) and territorial defence against fish of other species, than demoiselles from areas of high population density. Territories and nest sizes tended to be largest in areas of medium population density. As in other damselfishes, population density is a major determinant of the frequency and intensity of reproductive behaviours.     

Immunological similarities between specific chloroplast ribosomal proteins from Chlamydomonas reinhardtii and ribosomal proteins from Escherichia coli

Polyclonal antibodies were elicited against seven of the 33 different proteins of the large subunit of the chloroplast ribosome from Chlamydomonas reinhardtii. Three of these proteins are synthesized in the chloroplast and four are made in the cytoplasm and imported. In western blots, six of the seven antisera are monospecific for their respective large subunit ribosomal proteins, and none of these antisera cross-reacted with any chloroplast small subunit proteins from C. reinhardtii. Antisera to the three chloroplast-synthesized ribosomal proteins cross-reacted with specific Escherichia coli large subunit proteins of comparable charge and molecular weight. Only one of the four antisera to the chloroplast ribosomal proteins synthesized in the cytoplasm cross-reacted with an E. coli large subunit protein. None of the antisera cross-reacted with any E. coli small subunit proteins. On the assumption of a procaryotic, endosymbiotic origin for the chloroplast, those chloroplast ribosomal proteins still synthesized within the organelle appear to have retained more antigenic sites in common with E. coli ribosomal proteins than have those which are now the products of cytoplasmic protein synthesis. Antisera to this cytoplasmically synthesized group of chloroplast ribosomal proteins did not recognize any antigenic sites among C. reinhardtii cytoplasmic ribosomal proteins, suggesting that the genes for the cytoplasmically synthesized chloroplast ribosomal proteins either are not derived from the cytoplasmic ribosomal protein genes or have evolved to a point where no antigenic similarities remain.