Stress produced by contaminated sediments with nickel in a pond with rainbow trout Oncorhynchus mykiss (Pisces: Salmonidae)

Previous studies indicated the presence of high concentrations of nickel in the "El truchón" ponds (in México), so it was necessary to find a possible correlation between sediment physicochemical properties of this water body and the damage that could be produced on the rainbow trout which culture is done in a rustic reservoir. The study was initiated with the determination of the physicochemical properties of the trout farm sediments, which are; granular composition, total nitrogen percentage, organic matter content, pH, redox potential, cationic interchange capacity, and Ni concentration. LC50 of Ni at 96 h was determined on Onchorynchus mykiss in the system water-sediment from the trout farm at equilibrium time. In the same system the sublethal toxicity of Ni was evaluated by the determination of metallotioneine (MT) levels, o-demethylase activity (OD) and protein concentration. The results showed a significant increment in the three-biochemical parameters. A linear relation was observed between Ni concentration and MT, OD and protein concentration, so these damage biomarkers are recommended in order to evaluate Ni toxicity. Probably these effects were due to the physicochemical characteristics of the sediments, which may give a high capacity to store metal in it. According to the obtained results it was suggested not to use rustic ponds in the fish culture, and use concrete ponds to avoid the accumulation of toxic compounds or make periodic sediments remotion.     

Mitochondrial glycerol-3-phosphate acyltransferase-deficient mice have reduced weight and liver triacylglycerol content and altered glycerolipid fatty acid composition

Microsomal and mitochondrial isoforms of glycerol-3-phosphate acyltransferase (GPAT; E.C. 2.3.1.15) catalyze the committed step in glycerolipid synthesis. The mitochondrial isoform, mtGPAT, was believed to control the positioning of saturated fatty acids at the sn-1 position of phospholipids, and nutritional, hormonal, and overexpression studies suggested that mtGPAT activity is important for the synthesis of triacylglycerol. To determine whether these purported functions were true, we constructed mice deficient in mtGPAT. mtGPAT(-/-) mice weighed less than controls and had reduced gonadal fat pad weights and lower hepatic triacylglycerol content, plasma triacylglycerol, and very low density lipoprotein triacylglycerol secretion. As predicted, in mtGPAT(-/-) liver, the palmitate content was lower in triacylglycerol, phosphatidylcholine, and phosphatidylethanolamine. Positional analysis revealed that mtGPAT(-/-) liver phosphatidylethanolamine and phosphatidylcholine had about 21% less palmitate in the sn-1 position and 36 and 40%, respectively, more arachidonate in the sn-2 position. These data confirm the important role of mtGPAT in the synthesis of triacylglycerol, in the fatty acid content of triacylglycerol and cholesterol esters, and in the positioning of specific fatty acids, particularly palmitate and arachidonate, in phospholipids. The increase in arachidonate may be functionally significant in terms of eicosanoid production.     

Retroviral transfer of the p16INK4a cDNA inhibits C6 glioma formation in Wistar rats

BACKGROUND: The p16INK4A gene product halts cell proliferation by preventing phosphorylation of the Rb protein. The p16INK4a gene is often deleted in human glioblastoma multiforme, contributing to unchecked Rb phosphorylation and rapid cell division. We show here that transduction of the human p16INK4a cDNA using the pCL retroviral system is an efficient means of stopping the proliferation of the rat-derrived glioma cell line, C6, both in tissue culture and in an animal model. C6 cells were transduced with pCL retrovirus encoding the p16INK4a, p53, or Rb genes. These cells were analyzed by a colony formation assay. Expression of p16INK4a was confirmed by immunohistochemistry and Western blot analysis. The altered morphology of the p16-expressing cells was further characterized by the senescence-associated beta-galactosidase assay. C6 cells infected ex vivo were implanted by stereotaxic injection in order to assess tumor formation. RESULTS: The p16INK4a gene arrested C6 cells more efficiently than either p53 or Rb. Continued studies with the p16INK4a gene revealed that a large portion of infected cells expressed the p16INK4a protein and the morphology of these cells was altered. The enlarged, flat, and bi-polar shape indicated a senescence-like state, confirmed by the senescence-associated beta-galactosidase assay. The animal model revealed that cells infected with the pCLp16 virus did not form tumors. CONCLUSION: Our results show that retrovirus mediated transfer of p16INK4a halts glioma formation in a rat model. These results corroborate the idea that retrovirus-mediated transfer of the p16INK4a gene may be an effective means to arrest human glioma and glioblastoma.     

Onchocerca volvulus keratitis (river blindness) is exacerbated in BALB/c IL-4 gene knockout mice

To determine the outcome of Onchocerca volvulus keratitis in IL-4(-/-) BALB/c mice, animals were immunized subcutaneously and injected into the corneal stroma with soluble O. volvulus antigens. IL-4(-/-) BALB/c mice had a deviated cellular response, with decreased serum IgE and IgG1 and elevated IgG2a compared with control BALB/c mice. In marked contrast to control BALB/c, C57BL/6, and IL-4(-/-) C57BL/6 mice, IL-4(-/-) BALB/c mice developed severe corneal opacification and neovascularization that was associated with a pronounced neutrophil infiltrate to the corneal stroma. STAT-6(-/-) BALB/c mice had the same phenotype as IL-4(-/-) BALB/c mice, and complement depletion had no effect on the severity of O. volvulus keratitis in these mice. These findings indicate that on a BALB/c background, IL-4 has a critical role in regulating neutrophil recruitment to the cornea and development of O. volvulus keratitis.     

Targeted disruption of the Akap4 gene causes defects in sperm flagellum and motility

A-kinase anchoring proteins (AKAPs) tether cyclic AMP-dependent protein kinases and thereby localize phosphorylation of target proteins and initiation of signal-transduction processes triggered by cyclic AMP. AKAPs can also be scaffolds for kinases and phosphatases and form macromolecular complexes with other proteins involved in signal transduction. Akap4 is transcribed only in the postmeiotic phase of spermatogenesis and encodes the most abundant protein in the fibrous sheath, a novel cytoskeletal structure present in the principal piece of the sperm flagellum. Previous studies indicated that cyclic AMP-dependent signaling processes are important in the regulation of sperm motility, and gene targeting was used here to test the hypothesis that AKAP4 is a scaffold for protein complexes involved in regulating flagellar function. Sperm numbers were not reduced in male mice lacking AKAP4, but sperm failed to show progressive motility and male mice were infertile. The fibrous sheath anlagen formed, but the definitive fibrous sheath did not develop, the flagellum was shortened, and proteins usually associated with the fibrous sheath were absent or substantially reduced in amount. However, the other cytoskeletal components of the flagellum were present and appeared fully developed. We conclude that AKAP4 is a scaffold protein required for the organization and integrity of the fibrous sheath and that effective sperm motility is lost in the absence of AKAP4 because signal transduction and glycolytic enzymes fail to become associated with the fibrous sheath.     

First evidence for phototropin-related blue-light receptors in prokaryotes

A prokaryotic protein, YtvA from Bacillus subtilis, was found to possess a light, oxygen, voltage (LOV) domain sharing high homology with the photoactive, flavin mononucleotide (FMN)-binding LOV domains of phototropins (phot), blue-light photoreceptors for phototropism in higher plants. Computer-based three-dimensional modeling suggests that YtvA-LOV binds FMN in a similar pocket as phot-LOVs. Recombinant YtvA indeed exhibits the same spectroscopical features and blue-light-induced photochemistry as phot-LOVs, with the reversible formation of a blue-shifted photoproduct, assigned to an FMN-cysteine thiol adduct (Thio383). By means of laser-flash photolysis and time-resolved optoacoustic experiments, we measured the quantum yield of formation for Thio383, Phi(Thio) = 0.49, and the enthalpy change, DeltaH(Thio) = 135 kJ/mol, with respect to the parent state. The formation of Thio383 is accompanied by a considerable volume contraction, DeltaV(Thio) = -13.5 ml/mol. Similar to phot-LOVs, Thio383 is formed from the decay of a red-shifted transient species, T650, within 2 micros. In both YtvA and free FMN, this transient has an enthalpy content of approximately 200 kJ/mol, and its formation is accompanied by a small contraction, DeltaV(T) approximately -1.5 ml/mol, supporting the assignment of T650 to the FMN triplet state, as suggested by spectroscopical evidences. These are the first studies indicating that phototropin-related, blue-light receptors may exist also in prokaryotes, besides constituting a steadily growing family in plants.     

Seasonal and daily changes in the capacity for nonshivering thermogenesis in the golden hamsters housed under semi-natural conditions

Nonshivering thermogenesis (NST) is a main source of heat for many small mammals. It undergoes seasonal changes, being the highest in winter and the lowest in summer. Such acclimatization can ensure winter survival for species living in moderate or cold climates. Nevertheless, not only seasonal, but also daily changes in the capacity for NST seem to be of great importance. In this study, the effects of season and time of day on the temperature of brown adipose tissue (T(BAT)), preferred ambient temperature (PT(a)) and activity after noradrenaline (NA) injections in golden hamsters (Mesocricetus auratus) housed under semi-natural conditions were investigated. Animals were kept in outdoor enclosures and experienced natural changes in both, photoperiod and ambient temperature (T(a)). NA-induced hyperthermia was the largest during autumn (mean increase in T(BAT) by 0.74+/-0.04 degrees C), while during summer increase in T(BAT) was similar to that recorded in control (saline-injected) animals (0.16+/-0.05 degrees C and 0.24+/-0.04 degrees C, respectively). In spring hyperthermia was intermediary (0.57+/-0.05 degrees C). Daily variations in the response to NA depended on the season. In summer, the largest increase in T(BAT) (0.45+/-0.1 degrees C) was recorded during the first part of the day, while in autumn-in the middle of the day and night (1.1+/-0.1 degrees C and 0.9+/-0.1 degrees C, respectively). In spring, all NA injections induced large increase in T(BAT) except for the injection in the middle of the night. The largest decrease in PT(a) after NA administration was recorded in autumn (mean decrease by 1.5+/-0.3 degrees C). Both, seasonal and daily changes in the capacity for NST reflect different demands for heat dependently on the time of the year and time of the day. It can be concluded that although long history of breeding in captivity, golden hamsters preserved ability to survive in natural environment.     

Seasonal changes in the thermoregulation of laboratory golden hamsters during acclimation to seminatural outdoor conditions

Proper adjustments of the thermoregulatory mechanisms ensure survival in the natural environment. In the present study, we tested the hypothesis that laboratory golden hamsters (Mesocricetus auratus) housed under seminatural outdoor conditions are able to acclimatize to daily and seasonal changes in the environment despite their long history of breeding in captivity. The animals experienced natural changes in the photoperiod and ambient temperature characteristic for central Poland. During experiments in the thermal gradient system, the daily rhythms of body temperature (measured as the temperature of brown adipose tissue, TBAT), preferred ambient temperature (PTa) and activity were measured in summer, autumn and spring. We found that mean TBAT was highest in autumn and least in summer, reflecting seasonal changes in the capacity for nonshivering thermogenesis (NST). In summer, TBAT followed the robust daily rhythm with the amplitude of 1.1+/-0.1 degrees C. This amplitude was depressed in autumn (0.2+/-0.1 degrees C) and partially restored in spring (0.4+/-0.1 degrees C). Seasonal changes in the daily amplitude of TBAT recorded during both transitional periods, i.e., in autumn and spring, seem to be associated with hamsters' hibernation. In autumn, mean daily PTa was lower than in summer and spring, indicating the lowering of a set point for core body temperature (Tb) regulation. Locomotor activity was much higher in spring than in summer and autumn, and it always predominated at night. We conclude that laboratory golden hamsters housed under seminatural conditions express daily and seasonal changes in the thermoregulatory mechanisms that, despite long history of breeding in captivity, enable proper acclimatization to seasonally changing environment and ensure successful hibernation and winter survival.     

Metabolic reconstruction of sulfur assimilation in the extremophile <Emphasis Type="Italic">Acidithiobacillus ferrooxidans</Emphasis> based on genome analysis

Background  

Acidithiobacillus ferrooxidansis a gamma-proteobacterium that lives at pH2 and obtains energy by the oxidation of sulfur and iron. It is used in the biomining industry for the recovery of metals and is one of the causative agents of acid mine drainage. Effective tools for the study of its genetics and physiology are not in widespread use and, despite considerable effort, an understanding of its unusual physiology remains at a rudimentary level. Nearly complete genome sequences ofA. ferrooxidansare available from two public sources and we have exploited this information to reconstruct aspects of its sulfur metabolism.