Fungal biomass and decomposition in Spartina maritima leaves in the Mondego salt marsh (Portugal)

Spartina maritima (Curtis) Fernald is a dominant species in the Mondego salt marsh on the western coast of Portugal, and it plays a significant role in estuarine productivity. In this work, leaf litter production dynamics and fungal importance for leaf decomposition processes in Spartina maritima were studied. Leaf fall was highly seasonal, being significantly higher during dry months. It ranged from 42 g m^-2 in June to less than 6 g m^-2 during the winter. Fungal biomass, measured as ergosterol content, did not differ significantly between standing-decaying leaves and naturally detached leaves. Fungal biomass increased in wet months, with a maximum of 614 g g^-1 of ergosterol in January in standing-decaying leaves, and 1077 g g^-1 in December, in naturally detached leaves, decreasing greatly in summer. Seasonal pattern of fungal colonization was similar in leaves placed in litterbags on the marsh-sediment surface. However, ergosterol concentrations associated with standing-decaying and naturally detached leaves were always much higher than in litterbagged leaves, suggesting that fungal activity was more important before leaf fall. Dry mass of litterbagged leaves declined rapidly after 1 month (about 50%), mostly due to leaching of soluble organic compounds. After 13 months, Spartina leaves had lost 88% of their original dry weight. The decomposition rate constant (k) for Spartina maritima leaves was 0.151 month^-1.     

Monoclonal anti-HMGB1 (high mobility group box chromosomal protein 1) antibody protection in two experimental arthritis models

High mobility group box chromosomal protein 1 (HMGB1) is a DNA-binding nuclear protein that can be released from dying cells and activated myeloid cells. Extracellularly, HMGB1 promotes inflammation. Experimental studies demonstrate HMGB1 to be a pathogenic factor in many inflammatory conditions including arthritis. HMGB1-blocking therapies in arthritis models alleviate disease and confer significant protection against cartilage and bone destruction. So far, the most successful HMGB1-targeted therapies have been demonstrated with HMGB1-specific polyclonal antibodies and with recombinant A box protein, a fragment of HMGB1. The present study is the first to evaluate the potential of a monoclonal anti-HMGB1 antibody (2G7, mouse IgG2b) to ameliorate arthritis. Effects of repeated injections of this antibody have now been studied in two conceptually different models of arthritis: collagen type II-induced arthritis (CIA) in DBA/1 mice and in a spontaneous arthritis disease in mice with combined deficiencies for genes encoding for the enzyme DNase type II and interferon type I receptors. These mice are unable to degrade phagocytozed DNA in macrophages and develop chronic, destructive polyarthritis. Therapeutic intervention in CIA and prophylactic administration of anti-HMGB1 monoclonal antibody (mAb) in the spontaneous arthritis model significantly ameliorated the clinical courses. Anti-HMGB1 mAb therapy also partially prevented joint destruction, as demonstrated by histological examination. The beneficial antiarthritic effects by the anti-HMGB1 mAb in two diverse models of arthritis represent additional proof-of-concept, indicating that HMGB1 may be a valid target molecule to consider for development of future clinical therapy.     

Direct analysis of nitrocatechol-type glucuronides in urine by capillary electrophoresis–electrospray ionisation mass spectrometry and tandem mass spectrometry

Direct, quantitative capillary electrophoresis–electrospray ionisation mass spectrometric (CE–ESI-MS) and tandem mass spectrometric (CE–ESI-MS–MS) methods are described for the quantitation of 3-O-glucuronides of E- and Z-entacapone isomers (EEG and EZG) and tolcapone (TG) in urine. 3-O-Glucuronide of nitecapone was used as internal standard. Good separation of glucuronides was achieved with 20 mM ammonium acetate as separation solution at pH 6.84. Stacking was used to increase the sensitivity of the method by introducing samples in 5 mM ammonium acetate. CE–ESI-MS and CE–ESI-MS–MS methods are linear with correlation coefficients better than 0.9983 and 0.9982, and repeatable with relative standard deviations below 9 and 14%, respectively. The limit of detection (LOD) in CE–ESI-MS at signal-to-noise ratio 3 is 100 ng/ml for EEG and EZG and 250 ng/ml for TG. The CE–ESI-MS–MS method was the more sensitive; LOD was 7 ng/ml for all compounds, without any concentration of the sample.     

Nox4 NADPH oxidase-derived reactive oxygen species, via endogenous carbon monoxide, promote survival of brain endothelial cells during TNF-α-induced apoptosis

We investigated the role of reactive oxygen species (ROS) in promoting cell survival during oxidative stress induced by the inflammatory mediator tumor necrosis factor-α (TNF-α) in cerebral microvascular endothelial cells (CMVEC) from newborn piglets. Nox4 is the major isoform of NADPH oxidase responsible for TNF-α-induced oxidative stress and apoptosis in CMVEC. We present novel data that Nox4 NADPH oxidase-derived ROS also initiate a cell survival mechanism by increasing production of a gaseous antioxidant mediator carbon monoxide (CO) by constitutive heme oxygenase-2 (HO-2). TNF-α rapidly enhanced endogenous CO production in a superoxide- and NADPH oxidase-dependent manner in CMVEC with innate, but not with small interfering RNA (siRNA)-downregulated Nox4 activity. CORM-A1, a CO-releasing compound, inhibited Nox4-mediated ROS production and enhanced cell survival in TNF-α-challenged CMVEC. The ROS-induced CO-mediated survival mechanism requires functional interactions between the protein kinase B/Akt and extracellular signal-related kinase (ERK)/p38 MAPK signaling pathways activated by TNF-α. In Akt siRNA-transfected CMVEC and in cells with pharmacologically inhibited Akt, Erk1/2, and p38 mitogen-activated protein kinase (MAPK) activities, CORM-A1 was no longer capable of blocking Nox4 activation and apoptosis caused by TNF-α. Overall, Nox4 NADPH oxidase-derived ROS initiate both death and survival pathways in TNF-α-challenged CMVEC. The ROS-dependent cell survival pathway is mediated by an endogenous antioxidant CO, which inhibits Nox4 activation via a mechanism that includes Akt, ERK1/2, and p38 MAPK signaling pathways. The ability of CO to inhibit TNF-α-induced ERK1/2 and p38 MAPK activities in an Akt-dependent manner appears to be the key element in ROS-dependent survival of endothelial cells during TNF-α-mediated brain inflammatory disease.     

Susceptibility of different populations of ticks to entomopathogenic fungi

This study aimed to evaluate the in vitro effect of the entomopathogenic fungi Metarhizium anisopliae sensu lato (s.l.) and Beauveria bassiana sensu lato (s.l.) on two distinct populations of Rhipicephalus microplus, from two different experimental farms. Bioassays were performed with engorged females, eggs and larvae. Fungal infection was evaluated based on biological parameters of treated engorged females, percentage of hatch from treated eggs, and percentage of mortality and mean lethal time (LT90) of treated larvae. When the treatments were compared between the two populations, there were significant differences in the following parameters: pre-oviposition period, hatching period and egg production index. Moreover, the results showed that B. bassiana s.l., isolate Bb 986, was more virulent than M. anisopliae s.l., isolate 959, for engorged females, showing a control percentage of 49%. In the bioassay with eggs, the hatching percentage ranged from 3.1% to 49.5% in one population and from 3.4% to 42.7% in the other, with no significant difference between the two populations. In the bioassay with unfed larvae, the mortality percentage ranged from 91.8% to 98.7% in one population and from 71.0% to 94.0% in the other. The LT(90) varied from 19.52 to 27.51 days in one of the populations and 22.89 to 37.31 days in the other. These results suggest that populations of R. microplus show distinct variation in their susceptibility to B. bassiana s.l. and M. anisopliae s.l.     

Advanced glycation end-product pentosidine is not a relevant marker of disease activity in patients with rheumatoid arthritis

Advanced glycation end-product (AGE) pentosidine has previously been demonstrated in different tissues and body fluids. It was suggested as a novel marker for evaluating the pathologic activity in rheumatoid arthritis (RA). In this study we analyzed the relation between pentosidine and markers of inflammation, cartilage turnover, immune response, and disease status of RA. Using HPLC, we analyzed pentosidine in serum and synovial fluid from 39 patients with RA and in serum from 38 healthy controls. Cartilage oligomeric matrix protein (COMP) and antibodies to CCP (anti-CCP) were measured by ELISA. Clinical disease status was assessed by Disease Activity Score 28 (DAS 28) and functional status by Health Assessment Questionnaire (HAQ). We demonstrated significantly higher serum levels of pentosidine in RA patients in comparison with controls. Pentosidine in serum significantly correlated with pentosidine in synovial fluid. Serum pentosidine levels were associated with erythrocyte sedimentation rate (p<0.03) but not with CRP, COMP, anti-CCP antibodies, DAS 28, or HAQ. In contrast to previous studies, we could not show any correlation of pentosidine levels with inflammatory status, clinical disease activity, markers of immune response, or cartilage breakdown. However, AGEs can be suggested as important players participating in joint destruction rather than markers of disease activity.     

Factors potentially affecting fertility of lactating dairy cow recipients

Objectives of this study were to evaluate factors that could affect pregnancy rate after embryo transfer (ET) in lactating dairy cow recipients. The trial was conducted at a dairy farm located in Descalvado, SP, Brazil from October 2003 to September 2004. From 1037 cows with CL that were treated with an injection of PGF2alpha, 43.3% were detected in heat; 263 were previously assigned at day of PGF2alpha injection for AI and 186 for ET. Ovulation rate was 85.7% (385/449). Pregnancy rate for cows with CL for AI and embryo transfer recipients were 36.5% (84/230) and 58.7% (91/155) at day 25 and 33.0% (76/230) and 45.8% (71/155) at day 46, respectively. Embryonic loss were 9.5% (8/84) for the AI group and 21.9% (20/91) for the ET group. Average milk production was 31.4 L/day/cow. Average daily milk production from 7 days before PGF2alpha injection to 7 days after ET tended (P < 0.10) to influence pregnancy rate on days 25 and 46. Average daily milk production from the day of embryo transfer to 7 days after influenced embryonic loss (P < 0.05). Cows with higher milk production had lower probability of pregnancy and higher probability of embryonic loss. Cows with higher days in milk had higher probability of pregnancy. Cows with higher rectal body temperature had lower probability of pregnancy and higher probability of embryonic loss. The influence of high milk yield and body temperature on fertility in lactating dairy cow recipients suggests that these effects can occur also after embryo reaches the blastocyst stage.     

1.5-Mb YAC Contig in Xq28 Formatted with Sequence-Tagged Sites and Including a Region Unstable in the Clones

A contig of 20 yeast artificial clones (YACs) has been assembled across 1.5 Mb of Xq28 and formatted with nine previously reported probes and nine STSs developed from the sequence of probes and end fragments of YACs. YAC end fragments were obtained by subcloning, Alu-vector PCR, or primer-ligation PCR methods. Eighteen of the YACs were recovered from a library specific for Xq24-q28; two that fill a gap were obtained from a second library made from total human DNA. One region, containing probes pX78c and 2A1.1, was unstable in YACs, but it was possible to generate a self-consistent map of DNA over the entire contig. Overlaps were confirmed by Southern blot analyses of YAC DNAs, and pulsed-field gel electrophoresis confirmed the extent of the contig and identified at least four CpG islands in the region.