Haplotypes of the bovine IgG2 heavy gamma chain in tick-resistant and tick-susceptible breeds of cattle

Bovines present contrasting, heritable phenotypes of infestations with the cattle tick, Rhipicephalus (Boophilus) microplus. Tick salivary glands produce IgG-binding proteins (IGBPs) as a mechanism for escaping from host antibodies that these ectoparasites ingest during blood meals. Allotypes that occur in the constant region of IgG may differ in their capacity to bind with tick IGBPs; this may be reflected by the distribution of distinct allotypes according to phenotypes of tick infestations. In order to test this hypothesis, we investigated the frequency of haplotypes of bovine IgG2 among tick-resistant and tick-susceptible breeds of bovines. Sequencing of the gene coding for the heavy chain of IgG2 from 114 tick-resistant (Bos taurus indicus, Nelore breed) and tick-susceptible (B. t. taurus, Holstein breed) bovines revealed SNPs that generated 13 different haplotypes, of which 11 were novel and 5 were exclusive of Holstein and 3 of Nelore breeds. Alignment and modeling of coded haplotypes for hinge regions of the bovine IgG2 showed that they differ in the distribution of polar and hydrophobic amino acids and in shape according to the distribution of these amino acids. We also found that there was an association between genotypes of the constant region of the IgG2 heavy chain with phenotypes of tick infestations. These findings open the possibility of investigating if certain IgG allotypes hinder the function of tick IGBPs. If so, they may be markers for breeding for resistance against tick infestations.     

Biochemical characterization of recombinant guaA-encoded guanosine monophosphate synthetase (EC 6.3.5.2) from Mycobacterium tuberculosis H37Rv strain

Administration of the current tuberculosis (TB) vaccine to newborns is not a reliable route for preventing TB in adults. The conversion of XMP to GMP is catalyzed by guaA-encoded GMP synthetase (GMPS), and deletions in the Shiguella flexneri guaBA operon led to an attenuated auxotrophic strain. Here we present the cloning, expression, and purification of recombinant guaA-encoded GMPS from Mycobacterium tuberculosis (MtGMPS). Mass spectrometry data, oligomeric state determination, steady-state kinetics, isothermal titration calorimetry (ITC), and multiple sequence alignment are also presented. The homodimeric MtGMPS catalyzes the conversion of XMP, MgATP, and glutamine into GMP, ADP, PP(i), and glutamate. XMP, NH(4)(+), and Mg(2+) displayed positive homotropic cooperativity, whereas ATP and glutamine displayed hyperbolic saturation curves. The activity of ATP pyrophosphatase domain is independent of glutamine amidotransferase domain, whereas the latter cannot catalyze hydrolysis of glutamine to NH(3) and glutamate in the absence of substrates. ITC data suggest random order of binding of substrates, and PP(i) is the last product released. Sequence comparison analysis showed conservation of both Cys-His-Glu catalytic triad of N-terminal Class I amidotransferase and of amino acid residues of the P-loop of the N-type ATP pyrophosphatase family.     

Morphologic evaluation and expression of matrix metalloproteinases-2 and 9 and nitric oxide during experimental periodontal disease in rat

The immunopathologic and inflammatory mechanisms involved in periodontal disease (PD) include the participation of host resident, inflammatory cells and chemical mediators. Metalloproteinases (MMPs) and nitric oxide (NO) play essential role in extracellular matrix turnover of periodontal tissue destruction. In this study, by means of RT-PCR through semi-quantitative densitometric scanning methods, the expression of MMPs -2 and -9 and inducible NO synthase (iNOS) was temporally and spatially investigated during the destructive mechanisms of experimentally induced PD in rats. Samples from different periods were microscopically analyzed and compared with the contralateral side (control). Our results showed significant expression of MMP-9 and iNOS in tissues affected by PD, as compared with controls, three days after PD induction, simultaneously with the beginning of alveolar bone loss. At 7 days post induction, only the MMP-9 mRNA presented a significantly higher expression, as compared with the respective controls. Thus, in the rat ligature-induced PD, MMP-9 and iNOS might importantly participate in the early stages of the disease, including inflammatory cell migration, tissue destruction and alveolar bone resorption. Also, we may suggest that the exuberant presence of PMNs may be related to the important expression of iNOS and MMP-9 found at 3 days post induction.     

Cardiotonic steroids: potential endogenous sodium pump ligands with diverse function

The highly conserved cardiotonic steroid (CS) binding site present on the ubiquitous membrane sodium pump, sodium, potassium-ATPase, appears to have been conserved by no force other than its capacity to bind CS: a family that includes plant-derived cardiac glycosides and putative endogenous vertebrate counterparts. Binding of ligand is inhibited by increased extracellular potassium. This implies functional coordination because inhibition of the sodium pump would be counterproductive when extracellular potassium is elevated. The interesting biology of the CS binding site continues to stimulate investigations into the identity of endogenous ligands, their role as pump regulators at the cellular level, and as mediators of body fluid balance and blood pressure regulation. In addition to inhibition of sodium and potassium transport, there is considerable recent evidence suggesting that the sodium pump may act as a cell signaling receptor activated by CS binding and responding by coordination of intracellular signaling pathways that can be dependent on and also independent of the reduction in transmembrane ion flux resulting directly from pump inhibition. This signaling may influence cell survival, growth, and differentiation. Recent insight into the biology of pump regulation by CS is reviewed.     

Preparation of a beef-extract as a laboratory reference material for the determination of heterocyclic amines

The present paper describes the preparation of a suitable laboratory reference material (LRM) to validate analytical methods for the determination of heterocyclic amines (HAs) in foods. Three different lots of reference material were prepared using a beef extract which was contaminated with a well-known quantity of amines at different levels ranging from 10 to 75 ng/g. These materials were then lyophilised under determined conditions and, after grinding and sieving, homogenised and, finally, bottled and labelled. Homogeneity and stability studies were performed and no statistical differences were observed in the analysis of variances for within- and between-bottle results, thus demonstrating the homogeneity of the material. Stability at different storage temperatures (-18, +4, +25 and +40 degrees C) and times (1, 2, 3 and 6 months) was also tested. Therefore, the material can be considered homogeneous and stable and can be proposed for use in inter-comparison exercises for the determination of HAs.     

Modulation of auxin signalling through DIAGETROPICA and ENTIRE differentially affects tomato plant growth via changes in photosynthetic and mitochondrial metabolism

Auxin modulates a range of plant developmental processes including embryogenesis, organogenesis, and shoot and root development. Recent studies have shown that plant hormones also strongly influence metabolic networks, which results in altered growth phenotypes. Modulating auxin signalling pathways may therefore provide an opportunity to alter crop performance. Here, we performed a detailed physiological and metabolic characterization of tomato (Solanum lycopersicum) mutants with either increased (entire) or reduced (diageotropica—dgt) auxin signalling to investigate the consequences of altered auxin signalling on photosynthesis, water use, and primary metabolism. We show that reduced auxin sensitivity in dgt led to anatomical and physiological modifications, including altered stomatal distribution along the leaf blade and reduced stomatal conductance, resulting in clear reductions in both photosynthesis and water loss in detached leaves. By contrast, plants with higher auxin sensitivity (entire) increased the photosynthetic capacity, as deduced by higher V_cmax and J_max coupled with reduced stomatal limitation. Remarkably, our results demonstrate that auxin‐sensitive mutants (dgt) are characterized by impairments in the usage of starch that led to lower growth, most likely associated with decreased respiration. Collectively, our findings suggest that mutations in different components of the auxin signalling pathway specifically modulate photosynthetic and respiratory processes.     

Inhibitory effect of combinations of digoxin and endogenous cardiotonic steroids on Na+/K+-ATPase activity in human kidney membrane preparation

AimsCardiac glycosides have been extensively used in the treatment of congestive heart failure for more than 200 years. Recently, cardenolides and bufadienolides were isolated from mammalian tissue and are considered as a new class of steroidal hormones. The aim of the present work was to characterize the interaction between the most clinical used cardiac glycoside digoxin and the cardiac glycosides known to exist endogenously, i.e., ouabain, marinobufagin and telocinobufagin, on human kidney Na⁺/K⁺-ATPase.Main methodsInhibition of Na⁺/K⁺-ATPase activity from crude membrane preparations of human kidney was performed using increasing concentrations of the drugs alone or mixtures of ouabain:digoxin, telocinobufagin:digoxin and marinobufagin:digoxin in a fixed ratio 1:4, 2:3 and 3:2, respectively. The colorimetric method of Fiske and Subbarow was used to measure the inorganic phosphate released.Key findingsAnalyses of inhibition curves showed that the experimental curves for all combinations were superimposed on the theoretical additive curves indicating that an additive effect occurs among distinct cardenolides and bufadienolides combinations on the human α1β1 Na⁺/K⁺-ATPase protomer.SignificanceConsidering the extensive use of digoxin in the treatment of heart failure and the recent findings that endogenous cardiac glycosides may have altered levels in many diseases, including heart failure, the demonstration of additive effect between cardiac glycosides can help in the understanding of recent clinical observations, including that lower than usual doses of cardiac glycosides are necessary for decreasing mortality in these patients.     

Maternal-Fetal Distribution of Calcium,Iron, Copper,and Zinc in Pregnant Teenagers and Adults

Adolescence is marked by intensive growth and development. When pregnancy occurs during this period of the mother’s growth, there is an increase in her nutritional needs. The aim of this study was to determine the levels of calcium, iron, copper, and zinc in maternal plasma, the placenta, and in the cord plasma of pregnant teenagers and adults. A total of 80 sets of maternal plasma, placentas, and cord plasma (40 from teenagers and 40 from adults) were analyzed using synchrotron radiation total reflection X-ray fluorescence. The levels of calcium, copper, and zinc in the maternal and cord plasma from teenagers were not significantly different than those from adults. Iron levels in the teenagers maternal and cord plasma were higher than in the adults. All of the minerals analyzed were present at higher levels in the placentas from adults than those from teenagers. However, the low quantities of placental calcium, iron, copper, and zinc in the teenagers do not compromise the levels of these minerals in the cord plasma. Future research regarding the placental transport of these minerals is recommended to investigate the efficiency of mechanisms of transfer of these minerals in pregnant teenagers.     

Naturally occurring radionuclides in food and drinking water from a thorium-rich area

This paper focuses on a survey of uranium and thorium decay chain radionuclides in food and drinking water from the thorium-rich (monazite-bearing) region of Buena, which is located in the state of Rio de Janeiro, Brazil. The radionuclide concentration values in the food and drinking water from Buena reached values higher than 100-fold the international reference values. The daily intake of radionuclides by the local population is similar to that of another high background radiation area in Brazil, but the intake is higher than that of residents from a normal background radiation area. Approximately 58?% of the food consumed by Buena inhabitants is produced locally. Based on that figure, locally produced food and the dilution of total radionuclides in the diet of residents caused by food importation are both highly relevant to a population’s intake of radionuclides. The concentration values for ²¹⁰Pb and the radium isotopes in drinking water from Buena are among the highest values to be reported in the literature. ²²⁸Ra is the most important radionuclide ingested with both food and water among the inhabitants of Buena.