Single-cell mass cytometry of differential immune and drug responses across a human hematopoietic continuum

Flow cytometry is an essential tool for dissecting the functional complexity of hematopoiesis. We used single-cell "mass cytometry" to examine healthy human bone marrow, measuring 34 parameters simultaneously in single cells (binding of 31 antibodies, viability, DNA content, and relative cell size). The signaling behavior of cell subsets spanning a defined hematopoietic hierarchy was monitored with 18 simultaneous markers of functional signaling states perturbed by a set of ex vivo stimuli and inhibitors. The data set allowed for an algorithmically driven assembly of related cell types defined by surface antigen expression, providing a superimposable map of cell signaling responses in combination with drug inhibition. Visualized in this manner, the analysis revealed previously unappreciated instances of both precise signaling responses that were bounded within conventionally defined cell subsets and more continuous phosphorylation responses that crossed cell population boundaries in unexpected manners yet tracked closely with cellular phenotype. Collectively, such single-cell analyses provide system-wide views of immune signaling in healthy human hematopoiesis, against which drug action and disease can be compared for mechanistic studies and pharmacologic intervention.     

Spatial variation of tracer distribution in a structured clay field soil

Within the sensitive soils of the River Oder Basin (E Brandenburg, Germany), chloride‐tracer transport was studied with respect to soil‐surface conditions of the well structured clayey topsoil (disturbed vs. undisturbed) and irrigation mode (flooding vs. sprinkling). The spatial variation of chloride and dye distribution was sampled in a regular grid within different soil depths. Different methods were used for the analysis of spatial heterogeneity: a heterogeneity index HI derived from fitting parameters of the cumulative distribution function, semivariogram analyses to identify the spatial representativity of observations and to classify the spatial variation, and Spearman's rank correlations to examine the spatial similarity of tracer distribution across different soil depths. Soil aggregation was obvious throughout the soil profile, and macropores and fractures were preferred flow paths for the tracer. Flood irrigation resulted in more “uniform” distribution than sprinkling did. However, preferential flow was identified for all treatments, where, once established below the surface layer, flow paths led to heterogeneity indices manifesting nonuniform flow and reduced lateral mixing between macropores and soil matrix. Within the flooded plot, spatial structure of chloride concentration was moderate unlike the strongly structured variation within the sprinkled plots. For purposes to generalize and to assess regional risk of the water and solute transport within the topsoil of the River Oder Basin, spatial autocorrelation ranges of about 15 cm should be considered and included into concepts of soil protection and land‐use management, soil‐sampling strategies, or modeling approaches.     

Constituents in evening primrose oil with radical scavenging,cyclooxygenase, and neutrophil elastase inhibitory activities

Cold-pressed, non-raffinated evening primrose oil was found to contain lipophilic radical scavengers. A highly enriched fraction of these compounds could be obtained from the oil by extraction with aqueous ethanol and subsequent liquid-liquid partitioning with petroleum. LC-DAD-MS analysis revealed that the fraction contained three aromatic compounds with identical UV and ESI-MS spectra. The compounds were isolated by RP-HPLC and their structures established by chemical and spectroscopic means as 3-O-trans-caffeoyl derivatives of betulinic, morolic, and oleanolic acid. The morolic acid derivative was a new compound. The three esters exhibited pronounced radical scavenging activity against the stable 2,2-diphenyl-1-picrylhydrazyl radical and were potent inhibitors of neutrophil elastase and cyclooxygenase-1 and -2 in vitro. Commercial samples of evening primrose oils contained only traces of these lipophilic antioxidants.     

Managing lead exposure and toxicity in cow-calf herds to minimize the potential for food residues.

Lead poisoning is commonly diagnosed in cattle. In this study, 3 groups of cattle from different herds accidentally exposed to discarded lead batteries on pasture were intensively studied to determine the extent and severity of exposure. The losses from acute death due to lead toxicity were substantial in all the 3 study groups at 12%, 17%, and 4%. Blood samples were taken from all cattle around the time of the first diagnosis and then later in 2 of the 3 herds to monitor the change in lead concentrations over time. Asymptomatic lead toxicosis was observed in these herds. In these 3 groups, between 4% and 12% of asymptomatic cattle had blood lead concentrations consistent with acute lead poisoning (> 0.35 ppm), and between 7% and 40% of these asymptomatic animals were in the high-normal range (0.1-0.35 ppm). Because of the consistently high number of asymptomatic cattle with elevated lead levels, all cattle potentially exposed to a lead source should be tested before sale or slaughter to minimize the entry of lead into the food chain. The blood lead concentrations, which were monitored for months after the initial diagnosis, decreased slowly after the cattle were removed from the lead source. The prolonged retention of lead may be due to continued release and absorption of lead from metal particles in the reticulum or rumen. The mean reduction in the lead level was 0.046 ppm (95% CI, 0.017-0.075 ppm) every 30 days for these 2 herds. Using a single-component exponential model, the half-life of lead in the animals retested from Herds 1 and 2 was highly variable. The median half-life was 63 days (interquartile range, 34-107 days). One out of 8 pregnant heifers with high blood levels had a stillborn calf. There were no abortions or calf mortalities in this group. Blood samples were'collected from the calves around the time of birth. The concentrations of lead in the blood of the calves exposed in utero were low (0.010-0.095 ppm).     

Effects of breed and intake level on growth and feed efficiency in ram lambs

Effects of breed and intake level on growth and feed efficiency in ram lambs were tested with 52 Rambouillet, 46 Dorset and 53 Finnish Landrace (Finn) rams. Rams were individually fed at 100, 85 or 70% of ad libitum or at maintenance. The experimental diet contained 79% digestible dry matter (DM), 3.28% N and 2.95 Mcal metabolizable energy/kg DM. Rams were serially slaughtered every 35 d for 175 d. The efficiency of deposition of body weight, protein, energy and trimmed cuts was determined by breed and intake level over constant time, weight, fat and maturity intervals. Ranking of the breeds for feed efficiency depended upon both the interval and the criterion of evaluation. Efficiency of protein and trimmed cut gain was positively related to mature size in the time and weight intervals, but all differences in efficiency were small in the fat or maturity intervals. Gross energetic efficiency did not differ greatly among breeds in any interval. Dorsets appeared to have a somewhat greater propensity to fatten than the other breeds and to be less able to retain N at a fixed N intake. In the constant time interval, maintenance requirements were estimated to be 115 kcal/kg.75, and the estimated partial efficiencies of energy deposition in fat and protein were .49 and .26, respectively.     

Restricted energy intake and elevated calcium and phosphorus intake for gilts during growth. IV. Characterization of metacarpal, metatarsal, femur, humerus and turbinate bones of sows during three parities

The effect of early nutrition on subsequent bone development was studied using gilts that had previously been fed ad libitum or 75% ad libitum intake and 100 or 150% National Research Council-recommended daily Ca and P from weaning to 100 kg. During the three-parity reproductive study, sows were fed a 14% protein diet. Metacarpals and metatarsals were taken from sows culled due to lameness or failure to breed and from sows after completing three parities. Femur and humerus articular cartilage and turbinates were described at necropsy. Metacarpals and metatarsals were heavier and tended to have thicker walls when sows were previously fed ad libitum or fed 150% Ca and P. Energy intake produced the greater response. Metacarpal breaking strength was greatest for sows previously fed ad libitum. Metatarsals were not affected by energy intake. Stiffness, Young's modulus of elasticity (YME) and flexural modulus for metacarpals and metatarsals were not affected by energy intake. Previously fed Ca and P intakes did not affect any of the mechanical bone characteristics. Metacarpals were heavier, had a greater breaking strength, were more elastic and exhibited slightly less resistance to bending than the metatarsals. The ether extract, ash, Ca and P content and the Ca:P ratio of metacarpals and metatarsals were not affected by previously fed energy or Ca and P intakes. The ether extract content tended to decrease and the ash, Ca and P content tended to increase with age. Articular cartilage and turbinate scores were not influenced by previously fed energy or Ca and P intakes. The YME and flexural modulus were the only bone characteristics that were even poorly correlated (average r = .25) with soundness scores, suggesting that high values result in poorer soundness scores. Energy and Ca and P intakes during growth and development had only minimal effects on bone characteristics and no apparent effect on structural soundness and longevity of sows kept for three parities.     

Cauda equina compression syndrome (CECS): retrospective study of surgical treatment with partial dorsal laminectomy in 86 dogs with lumbosacral stenosis

We report our results of partial lumbosacral laminectomy for treatment of canine Cauda equina Compression Syndrome due to a lumbosacral stenosis. Opposite to conventional techniques of dorsal laminectomy, only widening of the Spatium interarcuale is performed. This is achieved by exstirpation of the Lig. flavum and partial dorsal laminectomy of the first sacral segment. The Proc. spinosi and integrity of facet joints are fully maintained by this technique. In 96.5% of 86 dogs treated with this method relief of dorsal pressure and permanent rapid regression of clinical symptoms was achieved. In two cases recurrence of clinical symptoms was observed during follow up and one case showed no improvement at all. In conclusion partial dorsal laminectomy is a minimal invasive technique for treatment of Cauda equina compression syndrome expressed by pain reaction accompanied by minor neurological deficits caused by lumbosacral stenosis. Maintained spinal stability allows short reconvalescence and the unrestricted use of dogs immediately post operation.     

Rationale and perspectives on the success of vaccination against bovine herpesvirus-1

Several characteristics of BHV-1 have contributed to the successful development of both conventional and marker vaccines. BHV-1 is a stable virus, which grows to high titers in vitro, has a limited host range and causes acute viremic infections. Furthermore, the protective antigens, as well as the antigens that are suitable as marker, are present in the predominant virus isolates and induce significant and long-lasting immune responses, both in na?ve and in previously vaccinated animals. In many parts of the world including North-America control of BHV-1 is achieved by vaccination with conventional attenuated or inactivated vaccines. With parts of Europe being BHV-1 free, the ability to differentiate infected from vaccinated animals has become critical as a trading tool. Live and killed gE-deleted marker vaccines are now widely used in Europe, in combination with gE-based diagnostic tests to monitor cattle. However, several issues remain to be resolved. BHV-1 causes latency, which creates a need for stringent management practices in case eradication is to be achieved. Since intramuscular delivery with a syringe and needle leads to considerable tissue damage, needle-free delivery methods should be adopted for beef cattle. Furthermore, conventional inactivated and attenuated vaccines are less efficacious in neonates, so alternative vaccine types such as CpG adjuvanted protein vaccines or DNA vaccines are required for effective vaccination of this age group.