bluuebunny/crossref_metadata_2025 huggingface dataset:
Adipokines: inflammation and the pleiotropic role of white adipose tissue
Authors: Trayhurn Paul , Wood I. Stuart
Published: October 2004

White adipose tissue is now recognised to be a multifunctional organ; in addition to the central role of lipid storage, it has a major endocrine function secreting several hormones, notably leptin and adiponectin, and a diverse range of other protein factors. These various protein signals have been given the collective name ‘adipocytokines’ or ‘adipokines’. However, since most are neither ‘cytokines’ nor ‘cytokine-like’, it is recommended that the term ‘adipokine’ be universally adopted to describe a protein that is secreted from (and synthesised by) adipocytes. It is suggested that the term is restricted to proteins secreted from adipocytes, excluding signals released only by the other cell types (such as macrophages) in adipose tissue. Theadipokinome(which together with lipid moieties released, such as fatty acids and prostaglandins, constitute thesecretomeof fat cells) includes proteins involved in lipid metabolism, insulin sensitivity, the alternative complement system, vascular haemostasis, blood pressure regulation and angiogenesis, as well as the regulation of energy balance. In addition, there is a growing list of adipokines involved in inflammation (TNFα, IL-1β, IL-6, IL-8, IL-10, transforming growth factor-β, nerve growth factor) and the acute-phase response (plasminogen activator inhibitor-1, haptoglobin, serum amyloid A). Production of these proteins by adipose tissue is increased in obesity, and raised circulating levels of several acute-phase proteins and inflammatory cytokines has led to the view that the obese are characterised by a state of chronic low-grade inflammation, and that this links causally to insulin resistance and the metabolic syndrome. It is, however, unclear as to the extent to which adipose tissue contributes quantitatively to the elevated circulating levels of these factors in obesity and whether there is a generalised or local state of inflammation. The parsimonious view is that the increased production of inflammatory cytokines and acute-phase proteins by adipose tissue in obesity relates primarily to localised events within the expanding fat depots. It is suggested that these events reflect hypoxia in parts of the growing adipose tissue mass in advance of angiogenesis, and involve the key controller of the cellular response to hypoxia, the transcription factor hypoxia inducible factor-1.
Effect of a γ-aminobutyric acid-enriched dairy product on the blood pressure of spontaneously hypertensive and normotensive Wistar–Kyoto rats
Authors: Hayakawa Kazuhito , Kimura Masayuki , Kasaha Keiko , Matsumoto Keisuke , Sansawa Hiroshi , Yamori Yukio
Published: October 2004

We investigated the blood-pressure-lowering effects of γ-aminobutyric acid (GABA) and a GABA-enriched fermented milk product (FMG) by low-dose oral administration to spontaneously hypertensive (SHR/Izm) and normotensive Wistar–Kyoto (WKY/Izm) rats. FMG was a non-fat fermented milk product produced by lactic acid bacteria, and the GABA contained in FMG was made from the protein of the milk during fermentation. A single oral dose of GABA or FMG (5 ml/kg; 0·5 mg GABA/kg) significantly (P>0·05) decreased the blood pressure of SHR/Izm from 4 to 8 h after administration, but did not increase that of WKY/Izm rats. The hypotensive activity of GABA was dose-dependent from 0·05 to 5·00 mg/kg in SHR/Izm. During the chronic administration of experimental diets to SHR/Izm, a significantly slower increase in blood pressure with respect to the control group was observed at 1 or 2 weeks after the start of feeding with the GABA or FMG diet respectively (P>0·05) and this difference was maintained throughout the period of feeding. The time profile of blood-pressure change due to administration of FMG was similar to that of GABA. FMG did not inhibit angiotensin 1-converting enzyme. Furthermore, an FMG peptide-containing fraction from reverse-phase chromatography lacked a hypotensive effect in SHR/Izm rats. The present results suggest that low-dose oral GABA has a hypotensive effect in SHR/Izm and that the hypotensive effect of FMG is due to GABA.
Antioxidant vitamin status in high exposure to oxidative stress in competitive athletes
Authors: Rousseau Anne-Sophie , Hininger Isabelle , Palazzetti Stéphane , Faure Henri , Roussel Anne-Marie , Margaritis Irène
Published: October 2004

We conducted a cross-sectional study in 118 well-trained athletes to investigate 'high exposure' to sub-deficient antioxidant status, and consequently to oxidative damage, in relation to estimated daily energy expenditure (EE) and dietary antioxidant intake. Subjects completed 7 d food and activity records. Blood samples were obtained on day 8. Of the athletes 81, 60 and 43 % had intakes of vitamins E, C and β-carotene below two-thirds of the French RDA respectively, which is adjusted for EE (FRDAa). The deficit in vitamin E intake was positively correlated with EE (r0·51,P>0·0001). All the athletes had normal plasma vitamins E and C and 14 % had marginal plasma β-carotene. Plasma thiobarbituric acid-reactive substances (TBARS) did not increase with increased EE. As evidenced by ANOVA, EE-induced vitamin C intakes increased and consequently led to increased plasma ascorbic acid concentrations. In male athletes, plasma total carotenoids were negatively correlated with plasma TBARS concentrations (r–0·31,P>0·006). The relationship between vitamin C intakes and plasma concentrations was logarithmic (r0·59,P> 0·0001). To summarize, it is not clear whether vitamin E requirements are overestimated with reference to EE in the FRDAa. Daily requirements for vitamin C do not exceed 200 mg. Our present results could be interpreted as meaning that carotenoids play a protective role as exogenous antioxidants. Carotenoid intakes in athletes must be considered carefully.
Long-term almond supplementation without advice on food replacement induces favourable nutrient modifications to the habitual diets of free-living individuals
Authors: Jaceldo-Siegl Karen , Sabaté Joan , Rajaram Sujatha , Fraser Gary E.
Published: October 2004

Epidemiological and metabolic studies have shown that regular nut consumption may protect against risk of heart disease and diabetes. None has investigated the effect of adding nuts to a self-selected habitual diet (containing little or no nuts) on dietary patterns. The present study evaluated the impact of long-term almond supplementation in healthy men (n43) and women (n38) aged 25–70 years on nutrient profile and nutrient displacement. All subjects were followed for 1 year. During the first 6 months, subjects followed their habitual diets; in the second 6 months, subjects added almonds to their diets. Diets were assessed by seven random 24 h telephone diet recalls during each diet period. On average, the almond supplement was 52 g/d (about forty-two nuts) containing 1286 kJ. When subjects changed from their habitual diet to the almond-supplemented diet, the intakes of MUFA, PUFA, fibre, vegetable protein, α-tocopherol, Cu and Mg significantly (P>0·05) increased by 42, 24, 12, 19, 66, 15 and 23 % respectively; the intakes oftransfatty acids, animal protein, Na, cholesterol and sugars significantly (P>0·05) decreased by 14, 9, 21, 17 and 13 % respectively. These spontaneous nutrient changes closely match the dietary recommendations to prevent cardiovascular and other chronic diseases. Displacement estimates for total energy, total protein, total fat, SFA, MUFA, PUFA, total fibre, Ca, Fe, Mg, P, K, Zn and α-tocopherol ranged from 16 to 98 %; the estimates for total food weight, carbohydrate, sugars and Se were <245 %. A daily supplement of almonds can induce favourable nutrient modifications for chronic disease prevention to an individual's habitual diet.
Effect of plasma insulin and branched-chain amino acids on skeletal muscle protein synthesis in fasted lambs
Authors: Wester T. J. , Lobley G. E. , Birnie L. M. , Crompton L. A. , Brown S. , Buchan V. , Calder A. G. , Milne E. , Lomax M. A.
Published: October 2004

The increase in fractional rate of protein synthesis (Ks) in the skeletal muscle of growing rats during the transition from fasted to fed state has been explained by the synergistic action of a rise in plasma insulin and branched-chain amino acids (BCAA). Since growing lambs also exhibit an increase inKswith level of feed intake, the objective of the present study was to determine if this synergistic relationship between insulin and BCAA also occurs in ruminant animals. Six 30 kg fasted (72 h) lambs (8 months of age) received each of four treatments, which were based on continuous infusion into the jugular vein for 6 h of: (1) saline (155 mmol NaCl/l); (2) a mixture of BCAA (0·778 μmol leucine, 0·640 μmol isoleucine and 0·693 μmol valine/min·kg); (3) 18·7 μmol glucose/min·kg (to induce endogenous insulin secretion); (4) co-infusion of BCAA and glucose. Within each period all animals received the same isotope of phenylalanine (Phe) as follows: (1) l-[1-13C]Phe; (2) l-phenyl-[ring2H5]-alanine; (3) l-[15N]Phe; (4) l-[ring 2,6-3H]Phe. Blood was sampled serially during infusions to measure plasma concentrations of insulin, glucose and amino acids, and plasma free Phe isotopic activity; biopsies were taken 6 h after the beginning of infusions to determineKsinm. longissimus dorsiandvastusmuscle. Compared with control (saline-infused) lambs,Kswas increased by an average of 40 % at the end of glucose infusion, but this effect was not statistically significant in either of the muscles sampled. BCAA infusion, alone or in combination with glucose, also had no significant effect onKscompared with control sheep.Kswas approximately 60 % greater forvastusmuscle than form. longissimus dorsi(P>0·01), regardless of treatment. It is concluded that there are signals other than insulin and BCAA that are responsible for the feed-induced increase inKsin muscle of growing ruminant animals.
The in vivo use of the stable isotope-labelled biomarkers lactose-[15N]ureide and [2H4]tyrosine to assess the effects of pro- and prebiotics on the intestinal flora of healthy human volunteers
Authors: De Preter V. , Geboes K. , Verbrugghe K. , De Vuyst L. , Vanhoutte T. , Huys G. , Swings J. , Pot B. , Verbeke K.
Published: October 2004

Amongst the various claimed beneficial effects of pro- and prebiotics for the human host, it has been hypothesised that functional foods are able to suppress the generation and accumulation of toxic fermentation metabolites (NH3, p-cresol). Direct evidence supporting this hypothesis is lacking mainly because of the unavailability of reliable biomarkers. Preliminary data indicate that lactose-[15N]ureide and [2H4]tyrosine may be potential biomarker candidates. The aim of the present study was to evaluate the effect of pro- and prebiotics on the colonic fate of these biomarkers in a randomised, placebo-controlled, cross-over study with nineteen healthy volunteers. At the start of the study and at the end of each 2-week study period, during which they were administered either a probiotic (n 10; 6·5×109Lactobacillus casei Shirota cells twice daily) or a prebiotic (n 9; lactulose 10 g twice daily), the volunteers consumed a test meal containing the two biomarkers. Urine was collected during 48 h. Results were expressed as percentage of the administered dose. As compared with the placebo, the decrease in the percentage dose of p-[2H4]cresol in the 24–48 h urine fraction was significantly higher after probiotic intake (P=0·042). Similar changes were observed for the 15N tracer (P=0·016). After prebiotic intake, a significantly higher decrease in the percentage dose of p-[2H4]cresol (P=0·005) and 15N tracer (P=0·029) was found in the 0–24 h urine collection. The present results demonstrate that suppression of the generation and accumulation of potentially toxic fermentation metabolites by pro- and prebiotics can reliably be monitored in vivo by the use of stable isotope-labelled biomarkers.
Circulating triacylglycerol and apoE levels in response to EPA and docosahexaenoic acid supplementation in adult human subjects
Authors: Buckley Richard , Shewring Bethan , Turner Rufus , Yaqoob Parveen , Minihane Anne M.
Published: October 2004

High doses ofn–3 PUFA found in fish oils can reduce the circulating concentration of triacylglycerol (TG), which may contribute to the positive impact of these fatty acids on the risk of CVD. The present study aimed to establish the differential impact of EPA and docosahexaenoic (DHA) on plasma lipids and apo in adults. Forty-two normolipidaemic adult subjects completed a double-blind placebo controlled parallel study, receiving an EPA-rich oil (4·8 g EPA/d), DHA-rich oil (4·9 g DHA/d) or olive oil as control, for a period of 4 weeks. No effects of treatment on total cholesterol, LDL-cholesterol or HDL-cholesterol were evident. There was a significant 22 % reduction in TG level relative to the control value following the DHA treatment (P=0·032), with the 15 % decrease in the EPA group failing to reach significance (P=0·258). There were no significant inter-group differences in response to treatment for plasma apoA1, -C3 or -E levels, although a significant 15 % within-group increase in apoE was evident in the EPA (P=0·006) and DHA (P=0·003) groups. In addition, a within-group decrease in the apoA1:HDL-cholesterol ratio was observed in the DHA group, suggesting a positive impact of DHA on HDL particle size. The DHA intervention resulted in a significant increase in the proportion of EPAP=0·000 and DHAP=0·000 in plasma phospholipids, whilst significant increases in EPAP=0·000 and docosapentaenoic acidP=0·002, but not DHAP=0·193, were evident following EPA supplementation (P>0·05). Our present results indicate that DHA may be more efficacious than EPA in improving the plasma lipid profile.
Larval productivity of Fasciola gigantica in two lymnaeid snails
Authors: Dar Y. , Vignoles P. , Rondelaud D. , Dreyfuss G.
Published: October 2004

Two groups of Galba truncatula and two groups of Lymnaea natalensis were experimentally infected with Fasciola gigantica to determine if snail species had an influence on the redial burden and cercarial shedding of this trematode when snails of both species were infected with the same isolate of miracidia. In the two groups used for the study of redial burden, the total number of free rediae was significantly higher at day 49 post-exposure in L. natalensis than in G. truncatula. In the groups used for cercarial shedding, the life-span of cercaria-shedding snails and those of infected snails which died without cercarial emission, and the duration of the prepatent period were significantly longer in L. natalensis than those noted in G. truncatula. However, the mean numbers of shed cercariae did not significantly differ and showed no differences in their daily distribution throughout the shedding period. These results demonstrate that G. truncatula might be the principal intermediate host of F. gigantica in Egypt, at least in the areas where this lymnaeid species lives.
Equal partnership: two trematode species, not one, manipulate the burrowing behaviour of the New Zealand cockle, Austrovenus stutchburyi
Authors: Babirat C. , Mouritsen K.N. , Poulin R.
Published: October 2004

Metacercariae of the trematode Curtuteria australis (Echinostomatidae) accumulate in the foot of the New Zealand cockle Austrovenus stutchburyi, severely impairing the cockle's ability to burrow under the sediments. This results in increased predation by birds on cockles, and thus enhanced transmission rates of the parasite to its bird definitive hosts. This host manipulation by the trematode is costly: fish regularly crop the tip of the foot of cockles stranded on the sediment surface, killing any metacercariae they ingest. A second, previously undetected trematode species (characterized by 23 collar spines) co-existing with C. australis, has been found in the foot of cockles in the Otago Harbour, South Island, New Zealand. The relative abundance of the two species varies among localities, with the identity of the numerically dominant species also changing from one locality to the next. Both C. australis and the new species have a strong preference for encysting in the tip of the cockle's foot, where their impact on the burrowing ability of the host is greatest, and where they both face the risk of cropping by fish. Results indicate that these two species are ecological equivalents, and their combined numbers determine how the cockle population is affected.
Lesions in the liver of guinea-pigs infected with the swine lungworm, Metastrongylus apri
Authors: Yoshihara S.
Published: October 2004

Clear spot lesions were formed on the liver surface in guinea-pigs repeatedly infected with swine lungworm, Metastrongylus apri. The largest lesion, measuring 0.25 cm in diameter, was hard and yellow and showed a large granuloma in the lobule. The nematode larva was located at the centre of the lesion. This finding is likely to be an example of erratic parasitism in guinea-pigs with metastrongylidiasis.
The responses of a tropical breed of domestic rabbit, Oryctolagus cuniculus, to experimental infection with Trichostrongylus colubriformis
Authors: Musongong G.A. , Chiejina S.N. , Fakae B.B. , Ikeme M.M.
Published: October 2004

Clinical, parasitological and pathological responses of a tropical out-bred domestic rabbit to experimental Trichostrongylus colubriformis infection were used to evaluate its suitability as a laboratory host and model for studying the host–parasite relationships of T. colubriformis. In the first experiment, three groups each of 16, predominantly juvenile male, 8- to 10-week-old rabbits were given a single pulse infection with 500, 5000 or 25000 infective larvae (L3) of T. colubriformis, to represent low, medium and high levels of infection, respectively. A fourth group of 16 rabbits of similar age formed the uninfected controls. In the second experiment, two groups of 10 juvenile (8- to 10-week-old) and 10 adult (8- to 10-month-old) rabbits were similarly infected with 20000 L3, with appropriate naïve controls. Prepatency was 14 and 16 days and peak faecal egg counts occurred on days 24 and 20 after infection in young and adult rabbits respectively. Peak worm counts occurred on day 14 in both age groups and at all levels of infection. Subsequently, parasite burdens declined in a highly significantly dose- and age-dependent manner. At low and moderate levels of infection, approximately 83–98% of worms were recovered from the first 60 cm of the small intestine. Worm fecundity was also significantly influenced by host age and larval dose. Host age also had a significant effect on worm length. Infections with T. colubriformis were associated with a highly significant loss of body weight, accompanied by anorexia, diarrhoea and 25% mortality at high dose levels during the patent period of infection. There were no significant changes in packed cell volume and eosinophil counts at all ages and levels of infection but significant lymphocytosis occurred at the high dose level between days 7 and 21. Parasite-specific serum IgG responses were not related to worm burden. Overall, data showed that this miniature, docile and relatively inexpensive breed of rabbit is a potentially valuable laboratory host for studying T. colubriformis infections. The larval dose, duration of infection and host age were major determinants of host responsiveness to primary infections in this rabbit genotype.
Seasonal dynamics of Proteocephalus sagittus in the stone loach Barbatula barbatula from the Haná River, Czech Republic
Authors: Jarkovský J. , Koubková B. , Scholz T. , Prokeš M. , Baruš V.
Published: October 2004

The seasonal cycle of the cestode Proteocephalus sagittus (Cestoda: Proteocephalidae) was studied for the first time in the stone loach Barbatula barbatula from the Haná River, Czech Republic. A total of 180 loaches were examined monthly from January to December 2001. The parasite occurred in loaches throughout the year but infection parameters differed significantly among seasons, with the highest values of prevalence and abundance from the late winter to the early summer. Parasite recruitment took place in the winter and early spring and the worms sexually matured in the late spring and early summer. In contrast to P. torulosus, the gravid worms of which laid eggs only at the end of the spring/beginning of the summer, gravid worms of P. sagittus were also found, although in low numbers, in the autumn and early winter. The rate of infection of loach with P. sagittus was neither dependent on the sex nor on the size of its fish host.
Vaccination of mice with a 30 kDa Schistosoma antigen with and without human adjuvant induces high protection against S. mansoni infection
Authors: Attallah A.M. , Abdel Aziz M.M. , Abbas A.T. , Elbakry K.A. , El-Sharabasy M.M.
Published: October 2004

A 30 kDa antigen was characterized as a hydrophobic polypeptide containing 16 amino acids and evaluated as a potential candidate vaccine against infection by Schistosoma mansoni. CD1 albino mice immunized at 0, 14, and 21 days with 25 or 50 μg of the 30 kDa antigen per mouse with and without alum developed high levels of IgG antibodies (predominantly IgG2a and IgG2b isotypes). When immunized mice were infected with 200 S. mansoni cercariae, the highest protection levels (61% and 65% reduction in worm burden in two separate experiments) were obtained using the 50-μg antigen without alum adjuvant. The granuloma size decreased to 10%, a non-significant level in mice immunized using alum adjuvant. The results demonstrate the ability of the 30 kDa antigen with and without alum adjuvant to protect mice against S. mansoni infection.
Prevalence and pathology of helminths of ciconiiform birds from the Brazilian swamplands
Authors: Pinto R.M. , Barros L.A. , Tortelly L. , Teixeira R.F. , Gomes D.C.
Published: October 2004

The prevalence of helminths recovered from 108 birds representing eight species of Ciconiiformes from the Brazilian west-central region are presented. The digeneans Ascocotyle (Phagicola) longa, Clinostomum marginatum, Cotylotretus grandis, Ithyoclinostomum dimorphum, the nematodes Contracaecum multipapillatum, Desmidocercella ardeae, Eustrongylides ignotus, and the cestode Valipora mutabilis were identified. Contracaecum multipapillatum was the most prevalent species and E. ignotus the most pathogenic. Gross lesions due to infections with C. multipapillatum were characterized by ulcerative processes and hyperemia of the mucosa whereas those caused by E. ignotus consisted of perforations of the gastric mucosa and fibrotic tubular lesions in the gastric serosa. Histopathological examinations revealed necrosis and mixed leucocyte infiltrations and discrete compression of the mucosa in C. multipapillatum infections. Destruction of the mucosa and submucosa with the presence of fibrous capsules were observed in E. ignotus infections. Reports of accidental human infections, with severe clinical signs induced by these parasites, indicate the necessity of a proper evaluation of the pathogenicity of helminths of aquatic birds.
Evolutionary relationships within ‘pygmaeus’ group microphallids using genetic analysis and scanning electron microscopy
Authors: Galaktionov K.V. , Bulat S.A. , Alekhina I.A. , Saville D.H. , Fitzpatrick S.M. , Irwin S.W.B.
Published: October 2004

There are four species of ‘pygmaeus’ microphallids, namely Microphallus pygmaeus, M. piriformes, M. pseudopygmaeus and M. triangulatus (Trematoda: Microphallidae) which are parasites of marine birds and their sporocysts give rise to transmissible metacercariae inside littoral gastropods (mostly littorines). Universally primed polymerase chain reaction (UP-PCR) showed no apparent pattern between genetic diversity of the metacercariae as estimated by genomic banding profiles and their geographic region or molluscan host species. At the same time UP-PCR product cross-hybridization showed that M. pseudopygmaeus and M. triangulatus are genetically very similar, indicating that these taxa represent one species complex. In contrast, M. pygmaeus and M. piriformes are genetically well separated from each other and also from the pseudopygmaeus–triangulatus complex. Scanning electron microscopy of ventral spines, and analyses of spine angles and the number of teeth per spine, showed that all species differed significantly from one another. It was concluded that M. piriformes represents the original western member of the ‘pygmaeus’ group. Microphallus pygmaeus probably diverged from M. piriformes as it progressively specialized for sea duck final hosts. Microphallus pseudopygmaeus and M. triangulatus diverged from each other and the piriformes–pygmaeus ancestral line relatively recently. Microphallus pseudopygmaeus specialized for adoption of a wide range of gastropod host species and M. triangulatus developed morpho-functional specialization associated with final host exploitation.
Non-Hermitian Luttinger liquids and flux line pinning in planar superconductors
Authors: Ian Affleck  , Walter Hofstetter  , David R Nelson  , Ulrich Schollwöck
Published: October 2004

As a model of thermally excited flux liquids connected by a weak link, we study the effect
of a single line defect on vortex filaments oriented parallel to the surface of a thin planar
superconductor. This problem can be mapped onto the physics of a Luttinger liquid of
interacting bosons in one spatial dimension with a point impurity. When the applied
magnetic field is tilted relative to the line defect, the corresponding quantum boson
Hamiltonian is non-Hermitian. We analyse this problem using a combination of analytic
and numerical (density matrix renormalization group) methods, uncovering a delicate
interplay between enhancement of pinning due to Luttinger liquid effects and depinning
due to the tilted magnetic field. Interactions allow a single columnar defect to be very
effective in suppressing vortex tilt when the Luttinger liquid parameter .
Insulin and Hypertonicity Recruit GLUT4 to the Plasma Membrane of Muscle Cells by UsingN-Ethylmaleimide-sensitive Factor-dependent SNARE Mechanisms but Different v-SNAREs: Role of TI-VAMP
Authors: Randhawa Varinder K. , Thong Farah S.L. , Lim Dawn Y. , Li Dailin , Garg Rami R. , Rudge Rachel , Galli Thierry , Rudich Assaf , Klip Amira
Published: October 2004

Insulin and hypertonicity each increase the content of GLUT4 glucose transporters at the surface of muscle cells. Insulin enhances GLUT4 exocytosis without diminishing its endocytosis. The insulin but not the hypertonicity response is reduced by tetanus neurotoxin, which cleaves vesicle-associated membrane protein (VAMP)2 and VAMP3, and is rescued upon introducing tetanus neurotoxin-resistant VAMP2. Here, we show that hypertonicity enhances GLUT4 recycling, compounding its previously shown ability to reduce GLUT4 endocytosis. To examine whether the canonical soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) mechanism is required for the plasma membrane fusion of the tetanus neurotoxin-insensitive GLUT4 vesicles, L6 myoblasts stably expressing myc-tagged GLUT4 (GLUT4myc) were transiently transfected with dominant negative N-ethylmaleimide-sensitive factor (NSF) (DN-NSF) or small-interfering RNA to tetanus neurotoxin-insensitive VAMP (TI-VAMP siRNA). Both strategies markedly reduced the basal level of surface GLUT4myc and the surface gain of GLUT4myc in response to hypertonicity. The insulin effect was abolished by DN-NSF, but only partly reduced by TI-VAMP siRNA. We propose that insulin and hypertonicity recruit GLUT4myc from partly overlapping, but distinct sources defined by VAMP2 and TI-VAMP, respectively.
p116Rip Targets Myosin Phosphatase to the Actin Cytoskeleton and Is Essential for RhoA/ROCK-regulated Neuritogenesis
Authors: Mulder Jacqueline , Ariaens Aafke , van den Boomen Dick , Moolenaar Wouter H.
Published: October 2004

Activation of the RhoA-Rho kinase (ROCK) pathway stimulates actomyosin-driven contractility in many cell systems, largely through ROCK-mediated inhibition of myosin II light chain phosphatase. In neuronal cells, the RhoA-ROCK-actomyosin pathway signals cell rounding, growth cone collapse, and neurite retraction; conversely, inhibition of RhoA/ROCK promotes cell spreading and neurite outgrowth. The actin-binding protein p116Rip, whose N-terminal region bundles F-actin in vitro, has been implicated in Rho-dependent neurite remodeling; however, its function is largely unknown. Here, we show that p116Rip, through its C-terminal coiled-coil domain, interacts directly with the C-terminal leucine zipper of the regulatory myosin-binding subunits of myosin II phosphatase, MBS85 and MBS130. RNA interference-induced knockdown of p116Rip inhibits cell spreading and neurite outgrowth in response to extracellular cues, without interfering with the regulation of myosin light chain phosphorylation. We conclude that p116Rip is essential for neurite outgrowth and may act as a scaffold to target the myosin phosphatase complex to the actin cytoskeleton.
Short Tetracysteine Tags to β-Tubulin Demonstrate the Significance of Small Labels for Live Cell Imaging
Authors: Andresen Martin , Schmitz-Salue Rita , Jakobs Stefan
Published: October 2004

Genetically encoded tags are of fundamental importance for live cell imaging. We show that small tetracysteine (TetCys) tags can be highly advantageous for the functionality of the host protein compared with large fluorescent protein tags. One to three concatenated small TetCys tags as well as the large green fluorescent protein (GFP) were fused by integrative epitope tagging to the C terminus of β-tubulin (Tub2) in the budding yeast Saccharomyces cerevisiae. The increasing tag size correlated with functional interference to the host protein. Tub2 tagged with either 1×TetCys (10 amino acids [aa]) or 2×TetCys (20 aa) was able to substitute Tub2 in haploid cells. In contrast, C-terminal tagging of Tub2 with 3×TetCys (29 aa) or with GFP (244 aa) resulted in nonviable haploid cells. Cells expressing Tub2-1×TetCys or Tub2-2×TetCys were stained with FlAsH, which selectively binds to the TetCys-tag. The stained cells displayed dynamic FlAsH-labeled microtubules and low cellular background fluorescence. The presented approach to tag open reading frames (ORFs) at their native loci with very small TetCys-tags and the subsequent visualization of the tagged proteins in vivo can be extended in principle to any ORF in S. cerevisiae.
Enlargeosome, an Exocytic Vesicle Resistant to Nonionic Detergents, Undergoes Endocytosis via a Nonacidic Route
Authors: Cocucci Emanuele , Racchetti Gabriella , Podini Paola , Rupnik Marjan , Meldolesi Jacopo
Published: October 2004

Enlargeosomes, a new type of widely expressed cytoplasmic vesicles, undergo tetanus toxin-insensitive exocytosis in response to cytosolic Ca2+concentration ([Ca2+]i) rises. Cell biology of enlargeosomes is still largely unknown. By combining immunocytochemistry (marker desmoyokin-Ahnak, d/A) to capacitance electrophysiology in the enlargeosome-rich, neurosecretion-defective clone PC12-27, we show that 1) the two responses, cell surface enlargement and d/A surface appearance, occur with similar kinetics and in the same low micromolar [Ca2+]irange, no matter whether induced by photolysis of the caged Ca2+compound o-nitrophenyl EGTA or by the Ca2+ionophore ionomycin. Thus, enlargeosomes seem to account, at least in large part, for the exocytic processes triggered by the two stimulations. 2. The enlargeosome membranes are resistant to nonionic detergents but distinct from other resistant membranes, rich in caveolin, Thy1, and/or flotillin1. 3. Cell cholesterol depletion, which affects many membrane fusions, neither disrupts enlargeosomes nor affects their regulated exocytosis. 4. The postexocytic cell surface decline is [Ca2+]idependent. 5. Exocytized d/A-rich membranes are endocytized and trafficked along an intracellular pathway by nonacidic organelles, distinct from classical endosomes and lysosomes. Our data define specific aspects of enlargeosomes and suggest their participation, in addition to cell differentiation and repair, for which evidence already exists, to other physiological and pathological processes.
NVL2 Is a Nucleolar AAA-ATPase that Interacts with Ribosomal Protein L5 through Its Nucleolar Localization Sequence
Authors: Nagahama Masami , Hara Yoshimitsu , Seki Akihiro , Yamazoe Takeshi , Kawate Yumiko , Shinohara Takashi , Hatsuzawa Kiyotaka , Tani Katsuko , Tagaya Mitsuo
Published: October 2004

NVL (nuclear VCP-like protein), a member of the AAA-ATPase family, is known to exist in two forms with N-terminal extensions of different lengths in mammalian cells. Here, we show that they are localized differently in the nucleus; NVL2, the major species, is mainly present in the nucleolus, whereas NVL1 is nucleoplasmic. Mutational analysis demonstrated the presence of two nuclear localization signals in NVL2, one of which is shared with NVL1. In addition, a nucleolar localization signal was found to exist in the N-terminal extra region of NVL2. The nucleolar localization signal is critical for interaction with ribosomal protein L5, which was identified as a specific interaction partner of NVL2 on yeast two-hybrid screening. The interaction of NVL2 with L5 is ATP-dependent and likely contributes to the nucleolar translocation of NVL2. The physiological implication of this interaction was suggested by the finding that a dominant negative NVL2 mutant inhibits ribosome biosynthesis, which is known to take place in the nucleolus.
IC138 Is a WD-Repeat Dynein Intermediate Chain Required for Light Chain Assembly and Regulation of Flagellar Bending
Authors: Hendrickson Triscia W. , Perrone Catherine A. , Griffin Paul , Wuichet Kristin , Mueller Joshua , Yang Pinfen , Porter Mary E. , Sale Winfield S.
Published: October 2004

Increased phosphorylation of dynein IC IC138 correlates with decreases in flagellar microtubule sliding and phototaxis defects. To test the hypothesis that regulation of IC138 phosphorylation controls flagellar bending, we cloned the IC138 gene. IC138 encodes a novel protein with a calculated mass of 111 kDa and is predicted to form seven WD-repeats at the C terminus. IC138 maps near the BOP5 locus, and bop5-1 contains a point mutation resulting in a truncated IC138 lacking the C terminus, including the seventh WD-repeat. bop5-1 cells display wild-type flagellar beat frequency but swim slower than wild-type cells, suggesting that bop5-1 is altered in its ability to control flagellar waveform. Swimming speed is rescued in bop5-1 transformants containing the wild-type IC138, confirming that BOP5 encodes IC138. With the exception of the roadblock-related light chain, LC7b, all the other known components of the I1 complex, including the truncated IC138, are assembled in bop5-1 axonemes. Thus, the bop5-1 motility phenotype reveals a role for IC138 and LC7b in the control of flagellar bending. IC138 is hyperphosphorylated in paralyzed flagellar mutants lacking radial spoke and central pair components, further indicating a role for the radial spokes and central pair apparatus in control of IC138 phosphorylation and regulation of flagellar waveform.
Direct Visualization of Microtubule Flux during Metaphase and Anaphase in Crane-Fly Spermatocytes
Authors: LaFountain James R. , Cohan Christopher S. , Siegel Alan J. , LaFountain Douglas J.
Published: October 2004

Microtubule flux in spindles of insect spermatocytes, long-used models for studies on chromosome behavior during meiosis, was revealed after iontophoretic microinjection of rhodamine-conjugated (rh)-tubulin and fluorescent speckle microscopy. In time-lapse movies of crane-fly spermtocytes, fluorescent speckles generated when rh-tubulin incorporated at microtubule plus ends moved poleward through each half-spindle and then were lost from microtubule minus ends at the spindle poles. The average poleward velocity of ∼0.7 μm/min for speckles within kinetochore microtubules at metaphase increased during anaphase to ∼0.9 μm/min. Segregating half-bivalents had an average poleward velocity of ∼0.5 μm/min, about half that of speckles within shortening kinetochore fibers. When injected during anaphase, rhtubulin was incorporated at kinetochores, and kinetochore fiber fluorescence spread poleward as anaphase progressed. The results show that tubulin subunits are added to the plus end of kinetochore microtubules and are removed from their minus ends at the poles, all while attached chromosomes move poleward during anaphase A. The results cannot be explained by a Pac-man model, in which 1) kinetochore-based, minus end-directed motors generate poleward forces for anaphase A and 2) kinetochore microtubules shorten at their plus ends. Rather, in these cells, kinetochore fiber shortening during anaphase A occurs exclusively at the minus ends of kinetochore microtubules.
Prevention of Cancer Through Lifestyle Changes
Authors: Barnard R. James
Published: October 2004

Cancer is the second leading cause of death in the USA and an abundance of evidence suggests that lifestyle factors including smoking, the typical high‐fat, refined‐sugar diet and physical inactivity account for the majority of cancer. This review focuses on diet and inactivity as major factors for cancer promotion by inducing insulin resistance and hyperinsulinemia. Elevated levels of serum insulin impact on the liver primarily, increasing the production of insulin‐like growth factor I (IGF‐I) while reducing the production of insulin‐like growth factor binding protein 1 (IGFBP‐1) resulting in stimulation of tumor cell growth and inhibition of apoptosis (programmed cell death). Adopting a diet low in fat and high in fiber‐rich starch foods, which would also include an abundance of antioxidants, combined with regular aerobic exercise might control insulin resistance, reduce the resulting serum factors and thus reduce the risk for many different cancers commonly seen in the USA.
Increased adjuvant activity of minimal CD8 T cell peptides incorporated into lipid‐core‐peptides
Authors: White Karen , Kearns Philip , Toth Istvan , Hook Sarah
Published: October 2004

A problem facing the use of subunit peptide and protein vaccines is their inability to stimulate protective immune responses. Many different approaches have been utilized to overcome this inefficient immune activation. The approach we have taken is to modify the vaccine antigen so that it now has adjuvant properties. To do this, multiple copies of minimal CD8 T cell epitopes were attached to a poly lysine lipid core. These constructs are known as lipid‐core‐peptides (LCP). The research presented here examines the adjuvant activity of LCP. Using mouse models, we were able to show that LCP were indeed able to activate antigen‐presenting cells in vitro and to activate cytotoxic T‐cell responses in vivo. More importantly, LCP were able to stimulate the development of a protective antitumour immune response.
Vaccine adjuvants: Current state and future trends
Authors: Petrovsky Nikolai , Aguilar Julio César
Published: October 2004

The problem with pure recombinant or synthetic antigens used in modern day vaccines is that they are generally far less immunogenic than older style live or killed whole organism vaccines. This has created a major need for improved and more powerful adjuvants for use in these vaccines. With few exceptions, alum remains the sole adjuvant approved for human use in the majority of countries worldwide. Although alum is able to induce a good antibody (Th2) response, it has little capacity to stimulate cellular (Th1) immune responses which are so important for protection against many pathogens. In addition, alum has the potential to cause severe local and systemic side‐effects including sterile abscesses, eosinophilia and myofascitis, although fortunately most of the more serious side‐effects are relatively rare. There is also community concern regarding the possible role of aluminium in neurodegenerative diseases such as Alzheimer's disease. Consequently, there is a major unmet need for safer and more effective adjuvants suitable for human use. In particular, there is demand for safe and non‐toxic adjuvants able to stimulate cellular (Th1) immunity. Other needs in light of new vaccine technologies are adjuvants suitable for use with mucosally‐delivered vaccines, DNA vaccines, cancer and autoimmunity vaccines. Each of these areas are highly specialized with their own unique needs in respect of suitable adjuvant technology. This paper reviews the state of the art in the adjuvant field, explores future directions of adjuvant development and finally examines some of the impediments and barriers to development and registration of new human adjuvants.
The multidrug resistance gene mdr1a influences resistance to ectromelia virus infection by mechanisms other than conventional immunity
Authors: Xu Dongmei , Regner Matthias , Smith Danny , Ruby Janet , Johnstone Ricky , Müllbacher Arno
Published: October 2004

P‐glycoprotein (P‐gp), an ATP‐dependent membrane pump encoded by mdr, plays, in addition to its ability to efflux toxins, a role in the resistance to pathogens. We employed mdr1a gene knock out (mdr1a–/–) mice and ectromelia virus (EV) to elucidate the role of P‐gp in resistance to EV. Mdr1a–/– mice are more susceptible to EV infection than wild type (wt) mice, showing increased mortality and morbidity. Unexpectedly, virus titres in liver, and in vitro in macrophages and splenocytes were significantly lower in the more susceptible mdr1a–/– mice than wt littermates. Analysis of immunological mechanisms known to influence resistance to EV infection, such as NK and cytotoxic T cell responses, EV specific antibody and cytokine levels did not reveal significant differences between the two strains of mice. Only dendritic cells from mdr1a–/– mice showed impaired migration to the draining lymph nodes compared to wt mice. Our data show that P‐gp plays an important role in EV infection by as yet undefined mechanisms.
‘Proliferative’ and ‘synthetic’ airway smooth muscle cells are overlapping populations
Authors: Sukkar Maria B , Stanley Anthony J , Blake Anita E , Hodgkin Philip D , Johnson Peter R A , Armour Carol L , Hughes J Margaret
Published: October 2004

The extension of airway smooth muscle cell (ASMC) functions, from just contractile, to synthetic and/or proliferative states, is an important component of airway remodelling and inflammation in asthma. Whereas all these functions have been demonstrated in ASM, currently, it is not known whether ASMC can be differentiated on the basis of their proliferative and synthetic functions. We used flow‐cytometric techniques to determine, first, whether human ASMC are phenotypically heterogenous with regard to their secretory function, and second, the proliferative status of secretory cells. ASMC were induced to synthesize GM‐CSF by stimulation with IL‐1β and TNF‐α followed by 10% human serum. Flow‐cytometric detection of intracellular GM‐CSF revealed that only a proportion of cells in culture (∼ 20–60%) synthesize GM‐CSF. To determine the proliferative status of GM‐CSF producing cells, ASMC were pretreated with 5,6‐carboxyfluorescein diacetate succinimidyl ester (CFSE), a fluorescein based dye used to track cell division, prior to cytokine/serum stimulation. Simultaneous analysis of intracellular GM‐CSF and CFSE revealed that GM‐CSF producing cells were present in both the divided and undivided ASMC populations. Thus, cytokine production and proliferation occurred in overlapping ASMC populations and prior progression through the cell cycle was not essential for ASMC cytokine production.
Effect of incorporation of the adjuvant Quil A on structure and immune stimulatory capacity of liposomes
Authors: Demana Patrick H , Fehske Christian , White Karen , Rades Thomas , Hook Sarah
Published: October 2004

Liposomes have been widely used as drug delivery systems for many years. However, they are of limited use as delivery systems for subunit vaccines due to their low immunogenicity. Here we examine the effect of incorporating the adjuvant Quil A into liposomes on the type of particles produced, on the ability of the different particles to incorporate antigen and on the ability of the different particles to stimulate murine bone‐marrow‐derived dendritic cells (DC) and lymphocytes. The incorporation of increasing amounts of Quil A, from 20% to 70% of the total lipid into liposomes, reduces the size of the particles that form in aqueous dispersion and decreases antigen incorporation and uptake by DC. Interestingly, the particles with 20% Quil A were more toxic to cells in culture than the particles containing 70% Quil A, and the 20% particles were also more immunostimulatory.
Immunostimulatory CpG motifs induce CTL responses to HIV type I oligomeric gp140 envelope protein
Authors: Rao Mangala , Matyas Gary R , Vancott Thomas C , Birx Deborah L , Alving Carl R
Published: October 2004

In the present study we investigated the immunomodulatory effects of two adjuvants, liposomal lipid A [L(LA)] and CpG‐containing oligodeoxynucleotides (CpG ODN), to the HIV‐1 ogp140 envelope protein. Administration of each of these adjuvants separately with unencapsulated ogp140 resulted in low antibody titres. Encapsulation of ogp140 in liposomes containing lipid A resulted in a sixfold increase in anti‐ogp140 antibodies. The antibody titres were further enhanced threefold by the addition of CpG ODN. Priming and boosting BALB/c mice with unencapsulated ogp140 with L(LA) or encapsulation in liposomes containing lipid A induced a mixed Th1/Th2 type of immune response. In contrast, immunization with L(ogp140 + LA) plus CpG ODN switched the immune response to a Th‐1 response with elevated anti‐ogp140 IgG2a antibodies and IFN‐γ levels. Both adjuvants induced excellent ogp140‐specific proliferative and CTL responses. Therefore, for the induction of high titre antibodies, but not for cellular responses, the antigen and lipid A have to be present in the same liposomes. These results can have significant implications in directing the Th1 or Th2 differentiation of antigen‐specific immune responses in the context of vaccine development.
Purpose‐Based Expert Finding in a Portfolio Management System
Authors: Niu Xiaolin , McCalla Gordon , Vassileva Julita
Published: October 2004

Most of the research in the area of expert finding focuses on creating and maintaining centralized directories of experts' profiles, which users can search on demand. However, in a distributed multiagent‐based software environment, the autonomous agents are free to develop expert models or model fragments for their own purposes and from their viewpoints. Therefore, the focus of expert finding is shifting from the collection at one place as much data about a expert as possible to accessing on demand from various agents whatever user information is available at the moment and interpreting it for a particular purpose. This paper outlines purpose‐based expert modeling as an approach for finding an expert in a multiagent portfolio management system in which autonomous agents develop expert agent models independently and do not adhere to a common representation scheme. This approach aims to develop taxonomy of purposes that define a variety of context‐dependent user modeling processes, which are used by the users' personal agents to find appropriate expert agents to advise users on investing strategies.