Temperature and salinity effects on the respiratory metabolism of the first zoeal stage of Macrobrachium holthuisi Genofre & Lobão (decapoda: Palaemonidae)

Oxygen consumption rates of stage I Macrobrachium holthuisi Genofre & Lobão zoeae were measured in 24 different temperature and salinity combinations using Cartesian diver microrespirometers. Metabolic rates varied little with salinity at 15°C while at 20°C a marked elevation occurred in 0 and 35‰ At 25°C, a slight elevation occurred in 0‰; rates remained constant, however, in the other salinities. At 30°C, respiratory rates were similar to those recorded at 25°C except for decreases at 0 and 28‰ salinity. Q₁₀ values in the different salinities were usually highest between 15 and 20°C. Statistical analyses showed that while both temperature, salinity and their interaction significantly influenced larval respiratory rates, temperature had the more pronouced effect. Larval metabolism is salinity independent over the salinity range encountered in the larval biotope (7–21‰) at temperatures of 15–30°C.     

Midgut proteinases in three divergent species of Coleoptera

Cysteine proteinases have been found in some families of Coleoptera and, based on this, these enzymes were supposed to be characteristic of Coleoptera. To test this hypothesis, we studied midgut homogenates of three phylogenetically distant Coleoptera species: Tenebrio molitor (Tenebrionidae) larvae, Pyrearinus termitilluminans (Elateridae) larvae, and Pheropsophus aequinoctialis (Carabidae) adults. T. molitor display two cysteine proteinases (pHo 6.8) resolved in Superose (FPLC) with Mr 31,000 and 51,000. These enzymes are inhibited by E-64 and pHMB, are activated by EDTA + cysteine and hydrolyze benzoyl-DL-arginine-β-naphthylamide. T. molitor enzymes differ from a cysteine proteinase (Mr 64,000 using Superose) present in the wheat meal ingested by the insect. The cysteine proteinases predominate in the anterior two thirds of T. molitor midgut, probably because they are unstable in the higher luminal pH observed in the posterior third of the midgut. P. termitilluminans and P. aequinoctialis do not display cysteine proteinases, although they have trypsins (Mr 15,000, 25,000 and 41,000 for P. termitilluminans; Mr 26,000, 33,000 and 52,000 for P. aequinoctialis) and chymotrypsins (Mr 38,000 and 25,000 for P. aequinoctialis and Mr 15,000 for P. termitilluminans). Our results, together with literature data, suggest that cysteine proteinases occur in the Cucujiformia ancestor, which corresponds to the ancestor of most Coleoptera which ingest seeds rich in serine proteinase inhibitors.     

Influence of fetal bovine serum and hormones on primary vs. long-term cultures of human breast cancers

Summary We investigated the influence of fetal bovine serum, complemented or otherwise with estradiol or insulin or both, on the proliferation of mammary cancer cells from long-term and primary cultures. The long-term culture corresponded to mouse MXT and MCF-7 cell lines whereas the primary culture corresponded to primitive breast cancers squashed onto histologic slides and maintained in cultures for between 12 and 48 h. Cell proliferation was evaluated by means of digital cell image analysis of Feulgen-stained nuclei. Our results show that the addition of estradiol and insulin slightly but nevertheless significantly increases the proportion of cells still living at Hour 48 of culture. Fetal bovine serum, necessary for the growth of MXT and MCF-7 mammary cells, was highly cytotoxic with respect to the primary cultures of the 20 breast cancers under study. We are now conducting new experiments using chemically defined media to study the influence of new antineoplastic compounds on primary cultures of breast cancers.     

Apoptosis vs. necrosis: glutathione-mediated cell death during rewarming of rat hepatocytes

Hypothermia induces injury in its own right, but the mechanisms involved in the cell damage are still unclear. The aim of this study was to test the effects that glutathione (GSH) depletion induces on cell death in isolated rat hepatocytes, kept at 4 degrees C for 20 h, by modulating intracellular GSH concentration with diethylmaleate and buthionine sulfoximine (DEM and BSO). Untreated hepatocytes showed Annexin V stained cells (AnxV(+)), scarce propidium iodide stained cells (PI(+)) and presented a low level of lactate dehydrogenase (LDH) leakage after 20 h at 4 degrees C and rewarming at 37 degrees C. When DEM and BSO were added before cold storage, we observed a few AnXV(+) cells and an increase in PI(+) cells associated with LDH release in the incubation medium. Conversely, the addition of DEM and BSO only during rewarming caused a marked increase in cell death by apoptosis. Production of reactive oxygen species (ROS) and thiobarbituric acid species (TBARS), associated with a decrease in GSH concentrations, was higher when DEM and BSO were added before cold storage. Cells treated with DEM and BSO before cold storage showed lower ATP energy stores than hepatocytes treated with DEM and BSO only during rewarming. Pretreatment of hepatocytes with deferoxamine protected against apoptotic and necrotic morphology in conditions of GSH depletion. These results suggest that pretreatment of hepatocytes with DEM and BSO before cold storage induces necrosis, while the treatment of hepatocytes only during rewarming increases apoptosis. In both conditions, iron represents a crucial mediator of cell death.     

Adding denture cleanser to microwave disinfection regimen to reduce the irradiation time and the exposure of dentures to high temperatures

Background: The microwave energy is an efficient disinfection method; however, it can generate high temperatures that can result in distortion of the dentures. objectives: To evaluate whether the addition of an enzymatic cleanser to microwave disinfection regimen would disinfect dentures with shorter irradiation time. Materials and methods: Seven resin discs colonized with Candida albicans biofilm were placed on the palatal surface of sterile dentures to be randomly assigned to the following treatments: immersion in distilled water for 3 min with 0 (DW), 1 (DW + M1), 2 (DW + M2), or 3 min (DW + M3) of microwave irradiation; or immersion in denture cleanser for 3 min with 0 (DC), 1 (DC + M1), 2 (DC + M2) or 3 min (DC + M3) of irradiation. After the treatments, the viable cells were counted by a blinded examiner. The temperature was measured immediately after irradiation. The data were analyzed by ANOVA and Tukey post hoc tests (α = 0.05). Results: No viable cells were found after DC + M2, DC + M3, and DW + M3 treatments, of which DC + M2 achieved the lowest temperature. No significant difference was found between the effectiveness of DW, DW + M1 and DC treatments (p > 0.05). Conclusion: Within the limits of this study, the association of a denture cleanser and microwave energy is efficient to disinfect dentures in lower irradiation time and temperature.     

Proteome analysis of the plasma protein layer adsorbed to a rough titanium surface

In this study a label-free proteomic approach was used to investigate the composition of the layer of protein adsorbed to rough titanium (Ti) after exposure to human blood plasma. The influence of the protein layer on the surface free energy (SFE) of the Ti was evaluated by contact angle measurements. Ti discs were incubated with blood plasma for 180?min at 37?°C, and the proteins recovered were subjected to liquid chromatography coupled to tandem mass spectrometry analysis. A total of 129 different peptides were identified and assigned to 25 distinct plasma proteins. The most abundant proteins were fibronectin, serum albumin, apolipoprotein A-I, and fibrinogen, comprising 74.54% of the total spectral counts. Moreover, the protein layer increased the SFE of the Ti (p?<?0.05). The layer adsorbed to the rough Ti surface was composed mainly of proteins related to cell adhesion, molecule transportation, and coagulation processes, creating a polar and hydrophilic interface for subsequent interactions with host cells.     

Forest structure and carbon dynamics in Amazonian tropical rain forests

Living trees constitute one of the major stocks of carbon in tropical forests. A better understanding of variations in the dynamics and structure of tropical forests is necessary for predicting the potential for these ecosystems to lose or store carbon, and for understanding how they recover from disturbance. Amazonian tropical forests occur over a vast area that encompasses differences in topography, climate, and geologic substrate. We observed large differences in forest structure, biomass, and tree growth rates in permanent plots situated in the eastern (near Santarém, Pará), central (near Manaus, Amazonas) and southwestern (near Rio Branco, Acre) Amazon, which differed in dry season length, as well as other factors. Forests at the two sites experiencing longer dry seasons, near Rio Branco and Santarém, had lower stem frequencies (460 and 466 ha^–1 respectively), less biodiversity (Shannon–Wiener diversity index), and smaller aboveground C stocks (140.6 and 122.1 Mg C ha^–1) than the Manaus site (626 trees ha^–1, 180.1 Mg C ha^–1), which had less seasonal variation in rainfall. The forests experiencing longer dry seasons also stored a greater proportion of the total biomass in trees with >50 cm diameter (41–45 vs 30% in Manaus). Rates of annual addition of C to living trees calculated from monthly dendrometer band measurements were 1.9 (Manaus), 2.8 (Santarém), and 2.6 (Rio Branco) Mg C ha^–1 year^–1. At all sites, trees in the 10–30 cm diameter class accounted for the highest proportion of annual growth (38, 55 and 56% in Manaus, Rio Branco and Santarém, respectively). Growth showed marked seasonality, with largest stem diameter increment in the wet season and smallest in the dry season, though this may be confounded by seasonal variation in wood water content. Year-to-year variations in C allocated to stem growth ranged from nearly zero in Rio Branco, to 0.8 Mg C ha^–1 year^–1 in Manaus (40% of annual mean) and 0.9 Mg C ha^–1 year^–1 (33% of annual mean) in Santarém, though this variability showed no significant relation with precipitation among years. Initial estimates of the C balance of live wood including recruitment and mortality as well as growth suggests that live wood biomass is at near steady-state in Manaus, but accumulating at about 1.5 Mg C ha^–1 at the other two sites. The causes of C imbalance in living wood pools in Santarém and Rio Branco sites are unknown, but may be related to previous disturbance at these sites. Based on size distribution and growth rate differences in the three sites, we predict that trees in the Manaus forest have greater mean age (~240 years) than those of the other two forests (~140 years).