Isolation and characterization of immortalized porcine aortic endothelial cell lines

Primary porcine endothelial cells have a limited life span in culture. After four to five passages, they tend to de-differentiate and eventually reach senescence. The aim of this work was to establish immortalized porcine aortic endothelial cell lines (AOCs) to facilitate in vitro studies of different pathological process involving the endothelium. Primary porcine aortic endothelial cells (PAECs) were transfected with a plasmid containing the SV40 genome and selected on the basis of morphological and phenotypical features. Flow cytometry analysis demonstrated uptake of acetylated low density lipoproteins (Ac-LDL) and constitutive expression of SLA class I, CD29, CD31, CD41/61, CD80/86, CD46, SWC3, and LAMP-1 antigens by all analyzed lines and showed little differences to primary cells. The functional similarity between primary and immortalized endothelial cells was demonstrated in a cytotoxicity assay using a human natural killer cell line (NKL) as effector. The AOCs cell lines should be valuable tools for in vitro study of the human immune response against pig endothelial cells. In addition, they would be very useful to gain insight in the pathogenesis of some viral haemorrhagic diseases of pig such as African swine fever (ASF) or classical swine fever (CSF).     

Weed growth interference from temperate cereals: the effect of a hydroxamic-acids-exuding rye (Secale cereale L.) cultivar

Under field conditions, a plot sown with Forrajero-Baer, a rye (Secale cereale L.) cultivar exuding hydroxamic acids through its roots, resulted in 83% less weed biomass than wheat (Triticum aestivum L.) and 76% less than forage oats (Avena strigosa L.). Forage oats reduced broadleaf weed biomass by 40% compared to wheat but failed to suppress wild oats (Avena fatua L.). Rye interference reduced wild oat biomass by 84% and 86% compared to wheat and forage oats, respectively. Bioassays carried out with 2-O-ß-glucopyranosyl-4-hydroxy-l,4-benzoxazin-3-one (DIBOA-glc), the main hydroxamic acid found in rye, its aglucone 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA), and its breakdown product benzoxazolin-2-one (BOA) showed inhibition of root and coleoptile growth of A. fatua seedlings at concentrations above 0–25 mM l^?1. These results indicate that hydroxamic acids from Forrajero-Baer rye probably play a significant role in the weed-suppressing ability of this cultivar. Interférence entre la croissance des adventices et les cereales de zones tempérées: l'effet d'un cultivar de seigle (Secale cereale L.) secrétant des acides hydroxamiques Sous conditions de plein champ, une parcelle semée avec Forrajero-Baer, un cultivar de seigle (Secale cereale L.) secrétant des acides hydroxamiques à travers ses racines, a aboutit à une biomasse en adventices de 83% de moins qu'un blé (Triticum aestivum L.) et 76% de moins que de l'avoine fourragère (Avena strigosa L.). Les avoines fourragères réduisent la biomasse des adventices dicotylédones de 40% en comparaison du blé, mais n'éliminent pas les folles avoines (Avena fatua L.). L'inter-férence du seigle a réduit la biomasse de folle avoine par 84% et 86% en comparaison du blé et de l'avoine fourrageé respectivement. Des bioessais conduits avec du 2-O-ß-glucopyrano-glucopyranosyl- 4 -hydroxy- 1,4- benzoxazin -3-one (DIBOA-glc), le principal acide hydroxamique trouvé dans le seigle, son aglycone 2,4-dihydroxy-1-4 benzoxazin 3-one (DIBOA) et son produit de dégradation benzaxazolin-2-one (BOA) ont mis en évidence une inhibition de la croissance des racines et du coléoptile des plantules d'A. fatuaà des concentrations supérieures à 0,25 mM 1^?1. Ces résultats montrent que les acides hydroxamiques du seigle Forrajero-Baer jouent un rôle significatif dans l'aptitude à combattre les adventices de ce cultivar. Beeinträchtigung des Unkrautwachstums durch Getreide: die Wirkung einer Hydroxamsäuren ausscheidenden Sorte des Roggens (Secale cereale L.) In einem Feldversuch war unter der Roggen-Sorge ‘Forrajero-Baer’ (Secale cereale L.), die öber die Wurzeln Hydroxamsäuren ausscheidet, die Unkrautbiomasse 83% geringer als unter Weizen (Triticum aestivum L.) und 76% geringer als unter Sand-Hafer (Avena strigosa L.). Der Sand-Hafer reduzierte die Biomasse dikotyler Unkrautarten im Vergleich zum Weizen um 40%, konnte Flug-Hafer (Avena fatua L.) aber nicht unterdrücken. Der Roggen reduzierte die Biomasse von Flug-Hafer im Vergleich zu Weizen und Sand-Hafer um 84 bzw. 86%. In Biotests hemmten 2-O-β-Gluco-pyranosyl - 4 - hydroxy -1,4- benzoxazin - 3 - on (DOBIA-glc), die hauptsächlich in Roggen gefundene Hydroxamsäure, sein Aglukon 2,4-Hydroxy-l,4-benzoxazin-3-on (DIBOA) und sein Abbauprodukt Benzoxazolin-2-on (BOA) in Konzentrationen über 0,25 mm1^?1 das Wachstum der Wurzeln und der Koleoptile von Avena-fatua-Keimpflanzen. Diese Ergebnisse zeigen, daß Hydroxamsäuren aus der Roggen-Sorte ‘Forrajero-Baer’ offensichtlich eine wichtige Rolle bei der unkrautunterdrückenden Wirkung dieser Sorte spielen.     

The development of forage production and utilization gradients around livestock watering points

Large herbivores can impose spatial patterns on otherwise homogeneous vegetation, but how these patterns change through time is poorly understood. Domestic livestock pastures are model systems for studying how foraging behavior influences the development of coupled grazing and vegetation patterns. We sampled forage production and utilization by cattle along distance-from-water gradients to provide a snapshot of grazing and vegetation patterns, and then evaluated the ability of simulation models to qualitatively reproduce these patterns. In the field, forage production increased with distance from water, as expected, but utilization peaked at intermediate distances from water in two of three study areas. Likewise, simulations based on a variety of foraging strategies produced gradients in forage production and, after forage availability near water declined sufficiently, peaks in utilization at intermediate distances. Distance-from-water gradients thus represent cumulative but not necessarily present day gradients in grazing intensity. The model with a foraging strategy based on time minimization produced slightly more realistic patterns in forage abundance than a model based on energy maximization, although results were sensitive to the value of the threshold for rejecting sites of low forage biomass. However, all models produced implausible thresholds in grazing and forage distribution, suggesting that factors besides resource distribution influence herbivore distributions. Moreover, different foraging rules produced similar vegetation gradients, especially on point water source landscapes, illustrating the difficulty of inferring foraging processes from vegetation patterns.     

Ultrasonographic measurement of abomasal volume, location, and emptying rate in calves

OBJECTIVE: To develop and validate an ultrasonographic method for measuring abomasal volume, location, and emptying rate in suckling calves. ANIMALS: 9 male Holstein calves < 40 days of age. PROCEDURE: Before and after calves were fed different volumes of milk replacer or 2 L of oral electrolyte solutions, ultrasonographic measurements of abomasal dimensions (width, length, and height) were obtained by applying a 3.5-MHz sector probe to the ventral aspect of the abdomen in the transverse and sagittal planes. Abomasal volume was calculated from the ultrasonographic measurements by modeling the abomasum as an ellipsoid and by use of a power exponential equation to calculate the half-time of abomasal emptying (t1/2). RESULTS: Preprandial abomasal volume was 20 to 137 mL. All 3 abomasal dimensions increased during feeding and after suckling, and the abomasum was symmetrically located about the midline of the ventral aspect of the abdomen. Strong linear relationships were identified between ultrasonographic and suckled volumes, between ultrasonographic and scintigraphic heights, and between ultrasonographic and scintigraphic lengths. Ultrasonographic t1/2 was linearly related to scintigraphic t1/2; the latter is regarded as the gold standard measure of gastric emptying rate. CONCLUSIONS AND CLINICAL RELEVANCE: Ultrasonographic evaluation of the abomasum appears to provide a practical, rapid, noninvasive, and accurate method for determination of abomasal volume, location, and emptying rate in suckling calves.     

Azotemia and mortality among Babesia microti-like infected dogs

Babesia microti-like piroplasms are a recently recognized cause of illness in dogs in northwest Spain. Our objective was to describe the clinical characteristics and investigate the risk factors for azotemia and death among 58 B microti-like infected dogs. Twenty-one of the 58 (36%) dogs were azotemic at the time that the infection was diagnosed. The case fatality rate during the following week was 22%. Dogs with azotemia at the time of diagnosis were 10 times (95% CI, 3.26-28.8) more likely to die during the following week. Azotemia was the main cause of death for B microti-like infected dogs (attributable fraction = 90%). Severe anemia was present in 45 of the 58 (78%) dogs. Azotemic dogs also presented with hyperphosphatemia, hypoalbuminemia, hypercholesterolemia, proteinuria, and high urine protein: creatinine ratios, suggesting a glomerular component to the disease. Age was the only factor significantly associated with the risk of azotemia (P = .042): on average, a 4-year age increase doubled the risk of an infected dog being azotemic. The only factor significantly associated with mortality was azotemia (P = .001). We concluded that B microti-like infection is associated with a high risk of azotemia and mortality.     

Investigation of pulmonary deposition of a nebulized radiopharmaceutical agent in awake cats

OBJECTIVE: To determine whether conscious, unsedated cats will inhale a nebulized material administered via a facemask and whether this material will reach the lower airways. ANIMALS: 20 healthy adult cats. PROCEDURE: Technetium Tc 99m-diaminetriaminopentaacetic acid (99mTc-DTPA) was nebulized into a spacer and administered to the cats via a closely fitting facemask. By use of a gamma camera, images were then immediately obtained to determine the distribution of 99mTc-DTPA within the lower airways. RESULTS: Images obtained by use of the gamma camera revealed that all 20 cats had inhaled 99mTc-DTPA from the facemask. In each cat, deposition of the radiopharmaceutical agent was evident throughout the lung fields. CONCLUSIONS AND CLINICAL RELEVANCE: Awake cats that were not used to the application of a facemask did inhale substances from such a device. Aerosolization of medications may be a feasible route of administration for cats with lower airway disease.     

In vitro effect of classical swine fever virus on a porcine aortic endothelial cell line

The effect of classical swine fever (CSF) virus on some phenotypic and functional features of an established porcine aortic endothelial cell (AOC) line was investigated. AOC cells show most of the characteristics of primary endothelial cells, avoiding the alterations and senescence that these cells undergo after a few passages in culture. AOC cells were susceptible to CSF virus infection to a high degree, reaching 90% of CSF virus positive cells after 24 h of infection; however as with other porcine susceptible cells, no cytopathic effect could be observed. In these conditions none of the surface molecules studied, including SLA-II MHC antigens, adhesion or co-stimulatory molecules, were altered by virus infection after 24 or 48 h. Functionally CSF virus infection induced a decrease in the pro-coagulant activity of the AOC cells, determined by the increase in the clot formation time shown by the lysates of these cells. This contrasts with the increase observed in the expression of mRNA corresponding to IL-1 alpha and IL-6, two proinflammatory and pro-coagulant cytokines, in CSF virus-infected AOC cells.     

Population genomics of early events in the ecological differentiation of bacteria

Genetic exchange is common among bacteria, but its effect on population diversity during ecological differentiation remains controversial. A fundamental question is whether advantageous mutations lead to selection of clonal genomes or, as in sexual eukaryotes, sweep through populations on their own. Here, we show that in two recently diverged populations of ocean bacteria, ecological differentiation has occurred akin to a sexual mechanism: A few genome regions have swept through subpopulations in a habitat-specific manner, accompanied by gradual separation of gene pools as evidenced by increased habitat specificity of the most recent recombinations. These findings reconcile previous, seemingly contradictory empirical observations of the genetic structure of bacterial populations and point to a more unified process of differentiation in bacteria and sexual eukaryotes than previously thought.     

Adenosine deaminase activity in serum, erythrocytes and lymphocytes of rats infected with Leptospira icterohaemorrhagiae

Leptospirosis is a systemic disease of humans and domestic animals, mainly dogs, cattle and swine. The course of human leptospirosis varies from mild to severe fatal forms and the most severe form of human leptospirosis is principally caused by Leptospira interrogans serovar icterohaemorrhagiae (L. icterohaemorrhagiae). The enzyme adenosine deaminase (ADA) plays an important role in the production and differentiation of blood cells. The aim of this study was to evaluate the activity of ADA in serum, erythrocytes and lymphocytes of rats infected with L. icterohaemorrhagiae, as compared with non-infected rats. Twenty-four adult rats, divided into two uniform groups (A and B) were used for the enzymatic assays. The animals in Group B were inoculated intraperitoneally with 2×10(8) leptospires/rat, and the rodents in Group A (control) were not-inoculated. Blood collection was performed on days 5 and 15 post-infection (PI) and the blood used to assess the ADA activity. The infection by L.icterohaemorrhagiae altered erythrocyte count, hemoglobin concentration and hematocrit, causing a decrease in all these parameters on day 15 PI. Lymphocytes decreased significantly on day 15 PI, and ADA activity in serum was inhibited in infected rats on days 5 and 15 PI and its activity in erythrocytes were increased on day 5 PI. On day 5 PI, we found an increase in ADA activity in erythrocytes of infected rats. No correlation was observed between hematocrit and erythrocyte ADA activity on days 5 and 15 PI. The ADA activity was inhibited in rats infected on day 15 PI. A positive correlation (r(2)=60) was also observed between the number of lymphocytes and ADA activity in lymphocytes on day 15 PI (P<0.05). In conclusion, our results showed that the ADA activity is altered in serum, lymphocytes and erythrocytes in experimental infection by L.icterohaemorrhagiae in rats, concomitantly with hematological parameters.     

Iridoid, phenylethanoid and flavonoid glycosides from Sideritis trojana

From the MeOH extract of Sideritis trojana, a new iridoid glycoside, 10-O-(E)-feruloylmelittoside (1) was obtained in addition to four known iridoid glycosides [melittoside (2), 10-O-(E)-p-coumaroylmelittoside (3), stachysosides E (4) and G (5)]. Moreover, five phenylethanoid glycosides [verbascoside (6), isoacteoside (7), lamalboside (8), leonoside A (9), isolavandulifolioside (10), three flavone glycosides (isoscutellarein 7-O-[6'-O-acetyl-β-allopyranosyl-(1→2)]-β-glucopyranoside (11), 4'-O-methyisoscutellarein 7-O-[6'-O-acetyl-β-allopyranosyl-(1→2)]-β-glucopyranoside (12), 3'-hydroxy-4'-O-methyisoscutellarein 7-O-[6'-O-acetyl-β-allopyranosyl-(1→2)]-β-glucopyranoside (13) and a benzylalcohol derivative (di-O-methylcrenatin) were obtained and identified. The structures were elucidated on the basis of NMR and HRMS data. All compounds were tested for their antioxidant activity by in vitro TEAC assay and some of them exhibited moderate activity (0.97-1.44 mM) when compared with the reference compound (quercetin 1.86 mM). Glycosides 6-13, the most active compounds in the TEAC assay, were also tested by flow cytometry to evaluate their ability to affect the levels of reactive oxygen species (ROS) in human prostate cancer cells (PC3).