A proteome resource of ovarian cancer ascites: integrated proteomic and bioinformatic analyses to identify putative biomarkers

Epithelial ovarian cancer is the most lethal gynecological malignancy, and disease-specific biomarkers are urgently needed to improve diagnosis, prognosis, and to predict and monitor treatment efficiency. We present an in-depth proteomic analysis of selected biochemical fractions of human ovarian cancer ascites, resulting in the stringent and confident identification of over 2500 proteins. Rigorous filter schemes were applied to objectively minimize the number of false-positive identifications, and we only report proteins with substantial peptide evidence. Integrated computational analysis of the ascites proteome combined with several recently published proteomic data sets of human plasma, urine, 59 ovarian cancer related microarray data sets, and protein-protein interactions from the Interologous Interaction Database I (2)D ( http://ophid.utoronto.ca/i2d) resulted in a short-list of 80 putative biomarkers. The presented proteomics analysis provides a significant resource for ovarian cancer research, and a framework for biomarker discovery.     

Impact of simulated moose densities on abundance and richness of vegetation,herbivorous and predatory arthropods along a productivity gradient

Large herbivores can affect vegetation structure and species composition as well as material and energy flows in the ecosystem through their selective feeding, defecation, urination and trampling. These changes have a large potential to indirectly affect other trophic levels, but the mechanisms are poorly known. We studied the impacts of moose Alces alces browsing along a gradient of site productivity by experimentally simulating four different moose densities. Here we show that moose can affect the richness and abundance of three trophic levels in Swedish boreal forests through complex direct and indirect impacts, but in qualitatively different ways depending on how the physical habitat or food resources of a trophic level are affected. Vegetation richness had a hump‐shaped (unimodal) response to increased moose density. Leaf litter production decreased when browsing increased, which in turn depressed the abundance of flying prey for spiders. Consequently, spider abundance and richness declined monotonically. The responses of spider richness to moose density were further conditioned by site productivity: the response was positive at productive and negative at unproductive sites. In contrast, herbivorous Hemiptera were not affected by moose, most likely because the abundance of their food plants was not affected. The highest simulated moose density had an impact on all variables responding to moose even after a few years of treatment and can be considered as overabundance. We also show that the impacts of low or moderate moose density can be positive to some of the organisms negatively affected by high density. The level of herbivore population density that leads to substantial community impacts also depends on site factors, such as productivity.     

Analysis of the mechanism by which tryptophan analogs inhibit human myeloperoxidase

Myeloperoxidase (MPO) catalyzes the formation of oxidants that have been implicated in the pathogenesis of various diseases, including cardiovascular and pulmonary diseases and cancer. Inhibition of MPO oxidant-generating activity represents an attractive target for preventing the progression of inflammatory conditions. Peroxidation and chlorination catalytic activity were utilized to screen for the most effective tryptophan analog that inhibits MPO. Rapid kinetic measurements were performed to determine the mechanisms through which these compounds inhibit the catalytic activity of MPO. Substituents on the amino and carboxyl termini of tryptophan enhance its affinity toward MPO compared to a substituent on the indole ring. Hydrogen-bond donor capabilities and a positive charge on the amino group are not required for MPO inhibition. Hydroxyl-containing indoles did not inhibit MPO H₂O₂-consumption activity. Elimination of the negative charge from the carboxyl terminus by introducing a hydrophobic character significantly enhanced tryptophan analog affinity for MPO and improved its inhibitory properties. Further mechanistic studies indicated that indole compounds inhibited MPO activity through the accumulation of compound II, an inactive MPO intermediate. Our results show that specific structural features of tryptophan analogs are involved in increasing the affinity for MPO and providing a significantly greater inhibition of MPO's catalytic activities.     

5-Aryl-4-(5-substituted-2,4-dihydroxyphenyl)-1,2,3-thiadiazoles as inhibitors of Hsp90 chaperone

A series of 5-aryl-4-(5-substituted-2,4-dihydroxyphenyl)-1,2,3-thiadiazoles were synthesized and their binding to several constructs of human Hsp90 chaperone measured by isothermal titration calorimetry (ITC). The most potent compound bound Hsp90 with the dissociation constant of about 5 nM.     

Preparation and use of cross-linked enzyme aggregates (CLEAs) of laccases

Cross-linked enzyme aggregates (CLEA^®) were prepared from laccases from three different sources: Trametes versicolor, Trametes villosa and Agaricus bisporus. The effect of the various parameters – nature of the precipitant, pH, temperature, glutaraldehyde concentration and cross-linking time – on the activity recovery and storage and operational stability of the resulting CLEAs was different. The laccase CLEAs exhibited the expected increased stability compared to the free enzyme but there was no direct correlation with the number of surface lysine residues in the latter. It is clearly not the only parameter influencing the properties of the CLEA. Co-aggregation with albumin did not improve the stability. The laccase CLEAs, in combination with the stable N-oxy radical, TEMPO, were shown to be active and stable catalysts for the aerobic oxidation of linear C₅–C₁₀ aliphatic alcohols, to the corresponding aldehydes, in aqueous buffer (pH 4). Rates were an order of magnitude higher than those observed with the corresponding free enzyme and the CLEAs could be recycled several times without appreciable loss of activity. The addition of water immiscible or water miscible solvents showed no further improvement in rate compared with reactions in aqueous buffer alone.     

Plantago lanceolata and Plantago rugelii Extracts are Toxic to Meloidogyne incognita but not to Certain Microbes

Extracts from the plants Plantago lanceolata and P. rugelii were evaluated for toxicity to the root-knot nematode Meloidogyne incognita, the beneficial microbes Enterobacter cloacae, Pseudomonas fluorescens and Trichoderma virens, and the plant-pathogenic fungi Fusarium oxysporum f. sp. gladioli, Phytophthora capsici, Pythium ultimum, and Rhizoctonia solani. Wild plants were collected, roots were excised from shoots, and the plant parts were dried and ground to a powder. One set of extracts (10% w/v) was prepared in water and another in methanol. Treatments included extract concentrations of 25%, 50%, 75% and 100%, and water controls. Meloidogyne incognita egg hatch was recorded after 7-day exposure to the treatments, and second-stage juvenile (J2) activity after 48 hours. All extracts were toxic to eggs and J2, with P. lanceolata shoot extract tending to have the most activity against M. incognita. Numbers of active J2 remained the same or decreased in a 24-hour water rinse following the 48-hour extract treatment, indicating that the extracts were lethal. When data from water- and methanol-extracted roots and shoots of both plant species were combined for analysis, J2 tended to be more sensitive than eggs to the toxic compounds at lower concentrations, while the higher concentrations (75% and 100%) were equally toxic to both life stages. The effective concentrations causing 50% reduction (EC₅₀) in egg hatch and in J2 viability were 44.4% and 43.7%, respectively. No extract was toxic to any of the bacteria or fungi in our assays.     

No response of plasma substance P, but delayed increase of interleukin-1 receptor antagonist to acute psychosocial stress

Psychosocial stress has been shown to induce inflammatory reactions, followed by the release of immunosuppressive glucocorticoids. This may be mediated by catecholamines or other stress reactive substances such as neuropeptides or cytokines. We here set out to explore the effects of acute psychosocial stress on plasma levels of substance P (SP), a possible mediator of stress-induced inflammatory reactions, and interleukin-1 receptor antagonist (IL-1ra). Twelve healthy male subjects (mean age 27 yrs.) were subjected to the psychosocial stress test "Trier Social Stress Test" (TSST) and a resting control condition. Blood and saliva samples were taken before, as well as 1, 20, 45, and 90 min after TSST or rest, respectively. Salivary cortisol and plasma SP and IL-1ra were measured using immunoassays, salivary alpha-amylase (sAA) was measured by an enzyme kinetic method, and plasma epinephrine (E) and norepinephrine (NE) were measured by HPLC. The TSST induced immediate increases of E, NE, and sAA, and a delayed increase of free cortisol. Plasma IL-1ra showed an even further delayed peak at 90 min after stress. Plasma levels of SP did not respond to stress. No significant associations between changes of stress hormones and IL-1ra or SP were found. We conclude that substance P, epinephrine, and norepinephrine are probably not involved in mediating peripheral inflammation following psychosocial stress, at least with respect to IL-1ra. Further studies have to reveal the mechanisms involved in the stress-induced up regulation of IL-1ra.