Structural organization of the beta-globin locus of B-haplotype sheep

Goats and some sheep synthesize a juvenile hemoglobin, Hb C (alpha 2 beta C2), at birth and produce this hemoglobin exclusively during severe anemia. Sheep that synthesize this juvenile hemoglobin are of the A haplotype. Other sheep, belonging to a separate group, the B haplotype, do not synthesize hemoglobin C and during anemia continue to produce their adult hemoglobin. To understand the basis for this difference we have determined the structural organization of the beta- globin locus of B-type sheep by constructing and isolating overlapping genomic clones. These clones have allowed us to establish the linkage map 5' epsilon I-epsilon II-psi beta I-beta B-epsilon III-epsilon IV- psi beta II-beta F3' in this haplotype. Thus, B sheep lack four genes, including the BC gene, and have only eight genes, compared with the 12 found in the goat globin locus. The goat beta-globin locus is as follows: 5' epsilon I-epsilon II-psi beta X-beta C-epsilon III-epsilon IV-psi beta Z-beta A-epsilon V-epsilon VI-psi beta Y-beta F3'. Southern blot analysis of A-type sheep reveals that these animals have a beta- globin locus similar to that of goat, i.e., 12 globin genes. Thus, the beta-globin locus of B-haplotype sheep resembles that of cows and may have retained the duplicated locus of the ancestor of cows and sheep. Alternatively, the B-sheep locus arrangement may be the result of a deletion of a four-gene set from the triplicated locus.      

Intracellular pH Regulation during NO(3) Assimilation in Shoot and Roots of Ricinus communis

Ricinus communis L. was used to test the Dijkshoorn-Ben Zioni hypothesis that NO₃⁻ uptake by roots is regulated by NO₃⁻ assimilation in the shoot. The fate of the electronegative charge arising from total assimilated NO₃⁻ (and SO₄²⁻) was followed in its distribution between organic anion accumulation and HCO₃⁻ excretion into the nutrient solution. In plants adequately supplied with NO₃⁻, HCO₃⁻ excretion accounted for about 47% of the anion charge, reflecting an excess nutrient anion over cation uptake. In vivo nitrate reductase assays revealed that the roots represented the site of about 44% of the total NO₃⁻ reduction in the plants. To trace vascular transport of ionic and nitrogenous constituents within the plant, the composition of both xylem and phloem saps was thoroughly investigated. Detailed dry tissue and sap analyses revealed that only between 19 and 24% of the HCO₃⁻ excretion could be accounted for from oxidative decarboxylation of shoot-borne organic anions produced in the NO₃⁻ reduction process. The results obtained in this investigation may be interpreted as providing direct evidence for a minor importance of phloem transport of cation-organate for the regulation of intracellular pH and electroneutrality, thus practically eliminating the necessity for the Dijkshoorn-Ben Zioni recycling process.     

Selective inhibition of prostaglandin biosynthesis by gold salts and phenylbutazone

Gold salts and phenylbutazone selectively inhibit the synthesis of PGF_2α and PGE₂ respectively. Lowered production of one prostaglandin species is accompanied by an increased production of the other. Selective inhibition by these drugs was observed in the presence of adrenaline, reduced glutathione and copper sulphate under conditions when most anti-inflammatory compounds inhibited PGE₂ and PGF_2α syntheses equally. It is postulated that selective inhibitors may have a different mode of action and beneficial effects may be related to the endogenous ratio of PGE to PGF required for normal function.     

Prenatal Vitamin D Supplementation and Child Respiratory Health: A Randomised Controlled Trial

Background

Observational studies suggest high prenatal vitamin D intake may be associated with reduced childhood wheezing. We examined the effect of prenatal vitamin D on childhood wheezing in an interventional study.

Methods

We randomised 180 pregnant women at 27 weeks gestation to either no vitamin D, 800 IU ergocalciferol daily until delivery or single oral bolus of 200,000 IU cholecalciferol, in an ethnically stratified, randomised controlled trial. Supplementation improved but did not optimise vitamin D status. Researchers blind to allocation assessed offspring at 3 years. Primary outcome was any history of wheeze assessed by validated questionnaire. Secondary outcomes included atopy, respiratory infection, impulse oscillometry and exhaled nitric oxide. Primary analyses used logistic and linear regression.

Results

We evaluated 158 of 180 (88%) offspring at age 3 years for the primary outcome. Atopy was assessed by skin test for 95 children (53%), serum IgE for 86 (48%), exhaled nitric oxide for 62 (34%) and impulse oscillometry of acceptable quality for 51 (28%). We found no difference between supplemented and control groups in risk of wheeze [no vitamin D: 14/50 (28%); any vitamin D: 26/108 (24%) (risk ratio 0.86; 95% confidence interval 0.49, 1.50; P = 0.69)]. There was no significant difference in atopy, eczema risk, lung function or exhaled nitric oxide between supplemented groups and controls.

Conclusion

Prenatal vitamin D supplementation in late pregnancy that had a modest effect on cord blood vitamin D level, was not associated with decreased wheezing in offspring at age three years.

Trial Registration

Controlled-Trials.com ISRCTN68645785     

Two silicone nontoxic fouling release coatings: Hydrosilation cured PDMS and CaCO3 filled,ethoxysiloxane cured RTV11

Two silicone coatings have been evaluated for barnacle adhesion. One coating is an unfilled hydrosilation cured polydimethylsiloxane (PDMS) network, while the other is a room temperature vulcanized (RTV), filled, ethoxysiloxane cured PDMS elastomer, RTV11^?. The adhesion strength of one species of barnacle, Balanus eburneus, to the hydrosilation coatings is in the range of 0.37–0.60 kg cm^‐2 while the corresponding range for RTV11 is 0.64–0.90 kg cm^‐2. The easier release of B. eburneus from the hydrosilation cured network compared to RTV11 is discussed in relationship to differences in bulk and surface properties. Preliminary results suggest bulk modulus may be the most important parameter in determining barnacle adhesion strength. In light or mechanical property analysis, a re‐evaluation of surface properties and chemical stability is presented.     

LC-MS/MS Confirms That COX-1 Drives Vascular Prostacyclin Whilst Gene Expression Pattern Reveals Non-Vascular Sites of COX-2 Expression

There are two schools of thought regarding the cyclooxygenase (COX) isoform active in the vasculature. Using urinary prostacyclin markers some groups have proposed that vascular COX-2 drives prostacyclin release. In contrast, we and others have found that COX-1, not COX-2, is responsible for vascular prostacyclin production. Our experiments have relied on immunoassays to detect the prostacyclin breakdown product, 6-keto-PGF_1α and antibodies to detect COX-2 protein. Whilst these are standard approaches, used by many laboratories, antibody-based techniques are inherently indirect and have been criticized as limiting the conclusions that can be drawn. To address this question, we measured production of prostanoids, including 6-keto-PGF_1α, by isolated vessels and in the circulation in vivo using liquid chromatography tandem mass spectrometry and found values essentially identical to those obtained by immunoassay. In addition, we determined expression from the Cox2 gene using a knockin reporter mouse in which luciferase activity reflects Cox2 gene expression. Using this we confirm the aorta to be essentially devoid of Cox2 driven expression. In contrast, thymus, renal medulla, and regions of the brain and gut expressed substantial levels of luciferase activity, which correlated well with COX-2-dependent prostanoid production. These data are consistent with the conclusion that COX-1 drives vascular prostacyclin release and puts the sparse expression of Cox2 in the vasculature in the context of the rest of the body. In doing so, we have identified the thymus, gut, brain and other tissues as target organs for consideration in developing a new understanding of how COX-2 protects the cardiovascular system.     

Thrombosis is reduced by inhibition of COX-1, but unaffected by inhibition of COX-2, in an acute model of platelet activation in the mouse

Background

Clinical use of selective inhibitors of cyclooxygenase (COX)-2 appears associated with increased risk of thrombotic events. This is often hypothesised to reflect reduction in anti-thrombotic prostanoids, notably PGI₂, formed by COX-2 present within endothelial cells. However, whether COX-2 is actually expressed to any significant extent within endothelial cells is controversial. Here we have tested the effects of acute inhibition of COX on platelet reactivity using a functional in vivo approach in mice.

Methodology/Principal Findings

A non-lethal model of platelet-driven thromboembolism in the mouse was used to assess the effects of aspirin (7 days orally as control) diclofenac (1 mg.kg⁻¹, i.v.) and parecoxib (0.5 mg.kg⁻¹, i.v.) on thrombus formation induced by collagen or the thromboxane (TX) A₂-mimetic, U46619. The COX inhibitory profiles of the drugs were confirmed in mouse tissues ex vivo. Collagen and U46619 caused in vivo thrombus formation with the former, but not latter, sensitive to oral dosing with aspirin. Diclofenac inhibited COX-1 and COX-2 ex vivo and reduced thrombus formation in response to collagen, but not U46619. Parecoxib inhibited only COX-2 and had no effect upon thrombus formation caused by either agonist.

Conclusions/Significance

Inhibition of COX-1 by diclofenac or aspirin reduced thrombus formation induced by collagen, which is partly dependent upon platelet-derived TXA₂, but not that induced by U46619, which is independent of platelet TXA₂. These results are consistent with the model demonstrating the effects of COX-1 inhibition in platelets, but provide no support for the hypothesis that acute inhibition of COX-2 in the circulation increases thrombosis.     

A mathematical model of the cell cycle of a hybridoma cell line

A one-dimensional age-based population balance model of the cell cycle is proposed for a mouse-mouse hybridoma cell line (mm321) producing immunoglobulin G antibody to paraquat. It includes the four conventional cell cycle phases, however, G1 is divided into two parts (G1a and G1b). Two additional phases have been added, a non-cycling state G1', and a pre-death phase D. The duration of these additional phases is determined by cumulative glutamine content and ammonia concentration, respectively. It is assumed that glutamine is only consumed during G1 and antibody is only produced during G1b and S, the kinetics are assumed to be zero-order. Glucose is consumed throughout the cell cycle at a rate that is dependent upon its prevalent concentration. Ammonia and lactate are produced in direct proportion to glutamine and glucose consumption, respectively. Parameters in the model have been determined from experimental data or from fitting the model to post-synchronisation data. The model thus fitted has been used to successfully predict this cell lines behaviour in conventional batch culture at different initial glutamine concentrations, and in chemostat culture at steady-state and in response to a glutamine pulse. The model predicts viable cell, glutamine, glucose and lactate kinetics well, but there are some discrepancies in the prediction for ammonia and antibody. Overall, the results obtained support the assumptions made in the model relating to the regulation of cell cycle progression. It is concluded that this approach has the potential to be exploited with other cell lines and used in a model-based control scheme.     

Spatial and Temporal Analysis of Contact Rates in Female White-Tailed Deer

Abstract: White-tailed deer (Odocoileus virginianus) are important game mammals and potential reservoirs of diseases of domestic livestock; thus, diseases of deer are of great concern to wildlife managers. Contact, either direct or indirect, is necessary for disease transmission, but we know little about the ecological contexts that promote intrasexual contact among deer. Using pair-wise direct contacts estimated from Global Positioning System collar locations and joint utilization distributions (JUDs), we assessed habitats in which contacts occur to test whether direct contact rates among female white-tailed deer in different social groups differs among land-cover types. We also tested whether contact rates differed among seasons, lunar phases, and times of day. We obtained locations from 27 female deer for periods of 0.5–17 months during 2002–2006. We designated any simultaneous pair of locations for 2 deer <25 m apart as a direct contact. For each season, we used compositional analysis to compare land-cover types where 2 deer had contact to available land-cover weighted by their JUD. We used mixed-model logistic regression to test for effects of season, lunar phase, and time of day on contact rates. Contact rates during the gestation season were greater than expected from random use in forest and grassland cover, whereas contact rates during the fawning period were greater in agricultural fields than in other land-cover types. Contact rates were greatest during the rut and lowest in summer. Diel patterns of contact rates varied with season, and contact rates were elevated during full moon compared to other lunar periods. Both spatial and temporal analyses suggest that contact between female deer in different social groups occurs mainly during feeding, which highlights the potential impact of food distribution and habitat on contact rates among deer. By using methods to associate contacts and land-cover, we have created beneficial tools for more elaborate and detailed studies of disease transmission. Our methods can offer information necessary to develop spatially realistic models of disease transmission in deer.