Diet modulates proteinuria in heterozygous female dogs with X-linked hereditary nephropathy

Young adult heterozygous (carrier) female dogs with X-linked hereditary nephropathy (XLHN) have glomerular proteinuria but are otherwise healthy. Because data regarding dietary influences on the magnitude of proteinuria in dogs with spontaneous glomerular disease are not available, 12 such dogs were studied in a double crossover experiment intended to determine effects of altering dietary protein intake for up to 6 weeks. Dogs were blocked by urine protein : creatinine ratio (UPC) and randomly assigned to receive 2 diets: high protein (34.6% dry matter [DM], HP) or low protein (14.1% DM, LP) fed in HP-LP-HP or LP-HP-LP sequence. Food intake was measured daily, body weight (BW) was measured twice weekly, and UPC, plasma creatinine, blood urea nitrogen, phosphorus, albumin, and protein concentrations were measured at 2-week intervals. Nutrient digestibility was measured during the third treatment period. Diet had a significant effect (P < .0001) on all measured variables except plasma phosphorus (P > .5), but unintended differences in digestibility of protein and energy (P < or = .01) prevented assignment of the diet effect exclusively to protein. Proteinuria was greater (UPC 4.7 +/- 2.2 versus 1.8 +/- 1.1, P < .0001) when the HP diet was fed, but the LP diet did not maintain starting BW or plasma albumin concentration within the normal reference range. Diet greatly affects the magnitude of proteinuria in XLHN carrier females. Dietary protein restriction can reduce proteinuria in dogs with glomerular disease, but BW and blood protein concentrations may not be maintained if the restriction is too severe.     

Integration and modelling of pharmacokinetic and pharmacodynamic data to optimize dosage regimens in veterinary medicine

In veterinary drug development procedures, pharmacokinetic (PK) and pharmacodynamic (PD) data have generally been established in separate, parallel studies to assist in the design of dosage schedules for subsequent evaluation in clinical trials. This review introduces the concept of PK/PD modelling, an approach in which PK and PD data are generated in the same study, and used to derive numerical values for PD parameters based on drug plasma concentrations. The PD parameters define the efficacy, potency and slope (sensitivity) of the concentration-effect relationship. It is proposed that the parameters derived from PK/PD modelling may be used as an alternative and preferred approach to dose titration studies for selecting rational dosage regimens (both dose and dosing interval) for further evaluation in clinical trials. In PK/PD modelling, the explicative variable for effect is the plasma concentration profile. The PK/PD approach provides several advantages over dose-titration studies, including determination of a projected dosage regimen by investigation of a single dose, in contrast to dose-ranging studies which by definition require testing of multiple dosage. Implementation of PK/PD modelling in the veterinary drug development process is currently constrained by the limited number of veterinary studies performed to date, and the consequently limited understanding of PK/PD concepts and their absence from regulatory authority guidelines. Nevertheless, PK/PD modelling has major potential for rational dosage regimen determination, as it considers and quantifies the two main sources of interspecies variability (PK and PD). It is therefore applicable to interspecies extrapolation and to multiple species drug development. As well as the currently limited appreciation of PK/PD principles in the veterinary scientific community, a further constraint in implementing PK/PD modelling is the need to validate PK/PD approaches and thereby gain confidence in its value by pharmaceutical companies and regulatory authorities.     

Cutaneous onchocerciasis in the horse: five cases in southwestern british columbia

Five horses were presented because of a dermatitis of the forehead. Unlike previous reports, ventral midline dermatitis was not the major problem, and was present in only two of five cases. All five horses responded to levamisole therapy at a daily dosage of 5.5 g for one week. Owners were cautioned that repeat therapy may be necessary.     

Platelet activating factor as a mediator of equine cell locomotion

Equine polymorphonuclear (PMN) and mononuclear (MN) leucocytes were separated on Percoll gradients and used to study the chemoattractant properties of the polar ether-linked phospholipid, platelet activating factor (PAF). Six concentrations of PAF ranging from 1 ng/ml to 100 g/ml were studied in each of two in vitro assay systems, the agarose microdroplet and a microfilter technique. Very significant (p<0.01) increases in the movement of both PMN and MN cells were obtained with most concentrations of PAF. In two instances there was no apparent concentration-response relationship, although the action of PAF was approximately bell-shaped in two others. The possible significance of these findings for equine inflammatory conditions is discussed.     

Agonist-induced adherence of equine neutrophils to fibronectin- and serum-coated plastic is CD18 dependent.

Adherence to vascular endothelium and extracellular matrix proteins is a pre-requisite for neutrophil accumulation at sites of inflammation. In this study, equine neutrophil adherence to fibronectin and autologous serum-coated plastic in response to PAF, hrIL-8, hrC5a and PMA has been measured. In addition, the mechanisms involved have been investigated using monoclonal antibodies (MoAbs) against the beta2 integrin CD18. PAF and hrC5a caused similar, concentration dependent, increases in adherence to fibronectin- and serum-coated plastic (maximum responses 19 +/- 4% and 19 +/- 3% for PAF and 15 +/- 4% and 16 +/- 2% for hrC5a on fibronectin- and serum-coated plastic, respectively). Adherence in response to PMA, although not reaching a maximum over the time course studied, was of a similar magnitude on the two surfaces (41 +/- 1% and 38 +/- 2% with 10(-7) M PMA on fibronectin- and serum-coated plastic, respectively). In contrast, the maximum adherence caused by hrIL-8 was significantly lower on fibronectin- than on serum-coated plastic (9 +/- 3% vs. 17 +/- 2%; 10(8) x M hrIL-8). Pre-incubation with MoAbs against CD18 (H20A and 6.5E) caused concentration related inhibition of stimulus-induced adherence to both fibronectin- and serum-coated plastic. Equine neutrophil adherence in response to PAF, hrIL-8, hrC5a and PMA therefore appears to be mediated by a CD18 dependent mechanism.     

Detection of Colletotrichum coccodes from soil and potato tubers by conventional and quantitative real-time PCR

Colletotrichum coccodes is the causal agent of the potato blemish disease black dot. Two PCR primer sets were designed to sequences of the ribosomal internal transcribed spacer (ITS1 and ITS2) regions for use in a nested PCR. The genus-specific outer primers (Cc1F1/Cc2R1) were designed to regions common to Colletotrichum spp., and the species-specific nested primers (Cc1NF1/Cc2NR1) were designed to sequences unique to C . coccodes . The primer sets amplified single products of 447 bp (Cc1F1/Cc2R1) and 349 bp (Cc1NF1/Cc2NR1) with DNA extracted from 33 European and North American isolates of C. coccodes. The specificity of primers Cc1NF1/Cc2NR1 was confirmed by the absence of amplified product with DNA of other species representing the six phylogenetic groups of the genus Colletotrichum and 46 other eukaryotic and prokaryotic plant pathogenic species. A rapid procedure for the direct extraction of DNA from soil and potato tubers was used to verify the PCR assay for detecting C. coccodes in environmental samples. The limit of sensitivity of PCR for the specific detection of C. coccodes when inoculum was added to soils was 3·0 spores per g, or the equivalent of 0·06 microsclerotia per g soil, the lowest level of inoculum tested. Colletotrichum coccodes was also detected by PCR in naturally infested soil and from both potato peel and peel extract from infected and apparently healthy tubers. Specific primers and a TaqMan fluorogenic probe were designed to perform quantitative real-time (TaqMan) PCR to obtain the same levels of sensitivity for detection of C. coccodes in soil and tubers during a first-round PCR as with conventional nested PCR and gel electrophoresis. This rapid and quantitative PCR diagnostic assay allows an accurate estimation of tuber and soil contamination by C. coccodes .     

Study of the influence of alcoholic fermentation and distillation on the oxygen-18/oxygen-16 isotope ratio of ethanol

A laboratory procedure for the analysis of the oxygen-18/oxygen-16 isotope ratios of ethanol derived from sugars and fruit juices by pyrolysis-isotope ratio mass spectrometry (IRMS) has been applied to the study of isotopic fractionation induced by the isotope effects of fermentation and distillation. For both processes, an experimental model has been established to describe and explain the observed fractionation phenomena. It is shown that reproducible results can be obtained when appropriate analytical conditions are used. Moreover, the ability of ethanol to act as a reliable indicator of the (18)O/(16)O ratio of sugars in orange juice (and therefore to be used as an internal reference for detecting water addition) is demonstrated both in theory and in practice.     

Effects-based Assays in the Earthworm Aporrectodea caliginosa: Their Utilisation for Evaluation of Contaminated Sites before and after Remediation (8 pp)

Goal, Scope and Background   In a preliminary ecological risk assessment, potential adverse effects of contaminants are often evaluated by measuring chemical residues and comparing these with regulatory guidelines. However limitations with this approach with regards to establishing actual effects have resulted in the increasing usage of sublethal effects-based assays, including biomarkers, to evaluate the hazard posed by contaminants in the environment. In this study a number of effects-based endpoints in the earthworm Aporrectodea caliginosa were evaluated to determine their comparative sensitivity for assessment of adverse effects of soil contaminated with petroleum hydrocarbons. Methods   Adult and juvenile earthworms were exposed for 4 weeks to sublethal concentrations of soil collected before and after remediation of a petroleum-contaminated site. A suite of endpoints were measured in these earthworms, including mortality, fecundity, growth, and juvenile maturation, and two less traditional endpoints, the biomarker, the neutral red retention assay (NRRA) and an avoidance behaviour test. Results and Discussion   Cocoon viability in this species is not a reliable parameter to measure, due to low viability in controls and a high coefficient of variation. Growth in adult earthworms was a more sensitive parameter than cocoon production. Maturation and growth of juveniles have been proposed as more sensitive endpoints than adult cocoon production and growth respectively. This was not apparent in the growth parameters, but maturation of juveniles did appear to be more sensitive than cocoon production by adults. The NRRA was a more sensitive parameter than cocoon production, and the NRRA and growth were both affected at the lowest concentration tested. The NRRA response appeared to be more sensitive than growth, but NRRT was only evaluated in one soil only, while the other parameters were assessed in two soils. However, the NRRA has previously been found to be more sensitive than growth after exposure to a number of contaminants. The avoidance behaviour assay exhibited similar sensitivity to growth and fecundity and could therefore be useful as a simple pre-screening test. Conclusion   The chronic endpoints, growth, cocoon production, and juvenile maturation parameters, were all sensitive endpoints for detecting exposure to the petroleum-hydrocarbon-contaminated soil. The NRRA was the most sensitive of the endpoints assessed and could be used as an early-warning indicator to predict adverse impacts. Avoidance behaviour could be used as a simple pre-screening test to evaluate contaminated soils prior to more extensive and invasive testing. Recommendations and Perspective   Measuring chemical concentrations in environmental samples is not always useful, as the toxicological impacts of exposure to these concentrations are not always discernible. However, the use of effects-based endpoints, either in situ or in the laboratory using laboratory-reared earthworms, can account for the bioavailability of chemicals in the soil, and can therefore provide information on the toxicological impacts of exposure. The assays tested in this research were sensitive indicators of exposure, and therefore can be used to determine potential ecological risks at contaminated sites and to monitor the progress of remediation at these sites.