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  • Author or Editor: Jorg M. Steiner x
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Abstract

Objective—To describe the kinetics of demethylation of 13C-aminopyrine in healthy dogs for use in determining the most appropriate time for collection of blood samples for a 13C-aminopyrine demethylation blood test for evaluation of hepatic function.

Animals—9 healthy dogs.

Procedures—A 2-mL baseline blood sample was collected into an evacuated heparinized tube, and 13Caminopyrine was administered to each dog (2 mg/kg, IV). Additional 2-mL blood samples were collected 15, 30, 45, 60, 75, 90, 105, 120, 135, 150, 180, 240, 300, and 360 minutes after 13C-aminopyrine administration. The CO2 was extracted from blood samples by addition of a strong acid, and the percentage dose of 13CO2 (PCD) in the extracted gas was determined by fractional mass spectrometry.

Results—No dogs had gross evidence of adverse effects, and all had an increase in PCD after IV administration of 13C-aminopyrine. The PCD had the least variability among 5 variables used to evaluate hepatic demethylating capacity. Peak PCD was detected at 30 minutes in 1 dog, 45 minutes in 5 dogs, 60 minutes in 2 dogs, and 75 minutes in 1 dog. The mean PCD for the 9 dogs peaked at 45 minutes after 13C-aminopyrine administration.

Conclusions and Clinical Relevance—PCD appears to be the preferable variable for evaluation of hepatic demethylating capacity. Intravenous administration of 13C-aminopyrine leads to a consistent increase in PCD. Mean PCD peaked 45 minutes after administration, suggesting that blood sample collection 45 minutes after 13C-aminopyrine administration may be appropriate for use in estimating hepatic demethylating capacity. (Am J Vet Res 2004;65:159–162)

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in American Journal of Veterinary Research

Abstract

OBJECTIVE To develop and validate a sandwich ELISA for the measurement of α1-proteinase inhibitor (α1-PI) concentrations in serum and fecal samples obtained from common marmosets (Callithrix jacchus).

SAMPLE Leftover serum (n = 42) and fecal (23) samples submitted for diagnostic testing; paired serum and fecal samples obtained from 30 common marmosets at 2 research colonies.

PROCEDURES A sandwich ELISA was developed and analytically validated by determining the lower limit of detection, linearity, accuracy, precision, and reproducibility. Reference intervals for α1-PI concentrations in serum and feces of common marmosets were calculated.

RESULTS The standard curve was generated for concentrations between 1 and 100 ng/mL. Mean ± SD observed-to-expected ratio for serial dilutions of serum and fecal samples was 117.1 ± 5.6% (range, 112.2% to 123.0%) and 106.1 ± 19.7% (range, 82.6% to 130.2%), respectively. Mean observed-to-expected ratio for spiking recovery of serum and fecal samples was 102.9 ± 12.1% (range, 86.8% to 115.8%) and 97.9 ± 19.0% (range, 83.0% to 125.1%), respectively. Reference interval for serum concentrations of α1-PI was 1,254 to 1,813 μg/mL, for 3-day mean fecal concentrations was 11.5 to 42.2 μg/g of feces, and for 3-day maximum fecal concentrations was 13.2 to 51.2 μg/g of feces.

CONCLUSIONS AND CLINICAL RELEVANCE The ELISA was linear, accurate, precise, and reproducible for quantification of α1-PI concentrations in serum and feces of common marmosets. However, the ELISA had limited linearity and accuracy for spiking recovery of fecal samples.

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in American Journal of Veterinary Research

Abstract

Objective—To purify and partially characterize feline pepsinogen (fPG) from the gastric mucosa and compare fPG with PGs of other species.

Sample Population—Stomachs of 6 cats.

Procedure—A crude protein extract was prepared from the gastric mucosa of feline stomachs. Feline PG A was purified by ammonium sulfate precipitation, weak-anion-exchange chromatography, size-exclusion chromatography, and strong-anion exchange chromatography. Partial characterization consisted of estimation of molecular weights (MWs) and isoelectric points, N-terminal amino acid sequencing, and investigation of susceptibility to pepstatin inhibition.

Results—Several fPG A-group isoforms were identified. The MWs of the isoforms ranged from 37,000 to 44,820. Isoelectric points were all < pH 3.0. The proteolytic activity of the activated PGs was inhibited completely by pepstatin in a range of equimolar to 10- fold molar excess. The specific absorbance of fPG A was 1.29. The N-terminal amino acid sequence of the first 25 residues of the predominant fPG A7 had 75%, 72%, 64%, and 56% homology with PG A of dogs, rabbits, cattle, and humans, respectively. Sequences of 4 other fPG A-group isoforms were similar to fPG A7. All isoforms were immunologically cross-reactive with sheep anti-fPG A7 antiserum.

Conclusions and Clinical Relevance—PG A is the only identified type of PG in cats and, similar to pg in other species, comprises multiple isoforms. The availability of fPG A may be used to facilitate the development of an immunoassay to quantify serum fPG A as a potential marker for gastric disorders in cats. (Am J Vet Res 2004;65:1195–1199)

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in American Journal of Veterinary Research

Abstract

Objective—To evaluate the specificity of a canine pancreas-specific lipase (cPSL) assay for diagnosing pancreatitis in dogs without clinical or histologic evidence of the disease.

Animals—20 dogs from another study with macroscopic evidence of pancreatitis and 44 dogs surrendered for euthanasia or expected to die.

Procedures—Prior to death, physical examination of each dog was performed and blood samples were collected for serum biochemical, serum cPSL, and hematologic analyses. After death, the pancreas was removed, sectioned in 1- to 2-cm slices, and evaluated by a pathologist. Dogs were classified by whether they had clinical or macroscopic pancreatitis. Each pancreatic section was histologically examined, and mean cumulative scores (MCSs) were assigned for 8 histologic characteristics. For each characteristic, comparisons were made between dogs with and without pancreatitis to establish histologic criteria for lack of evidence of pancreatitis.

Results—For all histologic characteristics except lymphocytic infiltration, the median MCS differed significantly between dogs with and without pancreatitis. Dogs were categorized as having no histologic evidence of pancreatitis when the MCSs for neutrophilic infiltration, pancreatic necrosis, peripancreatic fat necrosis, and edema were 0.0. On the basis of these criteria, 40 dogs were classified as having no evidence of pancreatitis. The cPSL concentration was within reference limits in 38 of these 40 dogs and was less than the cutoff value for diagnosing pancreatitis (400 μg/L) in 39 of the 40 dogs, resulting in a specificity of 97.5% (95% confidence interval, 86.8% to 99.9%).

Conclusions and Clinical Relevance—The cutoff cPSL value used in this study may be useful for diagnosing pancreatitis in dogs with a lack of histologic lesions consistent with pancreatitis and for which pancreatitis is not considered a major differential diagnosis.

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in American Journal of Veterinary Research

Abstract

Objective—To determine the prevalence of hypocobalaminemia or methylmalonic acidemia (or both) in dogs with chronic gastrointestinal disease.

Sample—Serum samples from 56 dogs with chronic gastrointestinal disease and 43 control dogs.

Procedures—Serum cobalamin and methylmalonic acid (MMA) concentrations were measured in all samples and compared between groups. A correlation between serum cobalamin and MMA concentrations and the canine chronic enteropathy clinical activity index was evaluated via the Spearman rank correlation.

Results—20 of 56 (36%) dogs with gastrointestinal disease had hypocobalaminemia. Serum cobalamin concentrations were significantly lower in dogs with gastrointestinal disease than in control dogs. Five of 56 (9%) dogs with chronic gastrointestinal disease and 5 of 20 (25%) hypocobalaminemic dogs had increased MMA concentrations. There was a significant negative correlation (Spearman r = −0.450) between serum cobalamin and MMA concentrations in dogs with gastrointestinal disease. No correlation was found between the canine chronic enteropathy clinical activity index and serum cobalamin or MMA concentrations.

Conclusions and Clinical Relevance—These data indicated the prevalence of hypocobalaminemia in dogs with chronic gastrointestinal disease was 20 of 56 (36%). Five of 20 (25%) hypocobalaminemic dogs had increased serum MMA concentrations, which indicated that although hypocobalaminemia was common in these dogs, it did not always appear to be associated with a deficiency of cobalamin on a cellular level. Hypocobalaminemia is a risk factor for negative outcome in dogs with chronic gastrointestinal disease and should be considered in every patient with corresponding clinical signs.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To provide values for gastrointestinal permeability and absorptive function tests (GIPFTs) with chromium 51 (51Cr)-labeled EDTA, lactulose, rhamnose, d-xylose, 3-O-methyl-d-glucose, and sucrose in Beagles and to evaluate potential correlations between markers.

Animals—19 healthy adult male Beagles.

Procedures—A test solution containing 3.7 MBq of 51Cr-labeled EDTA, 2 g of lactulose, 2 g of rhamnose, 2 g of d-xylose, 1 g of 3-O-methyl-d-glucose, and 8 g of sucrose was administered intragastrically to each dog. Urinary recovery of each probe was determined 6 hours after administration.

Results—Mean ± SD (range) percentage urinary recovery was 6.3 ± 1.6% (4.3% to 9.7%) for 51Cr-labeled EDTA, 3.3 ± 1.1% (1.7% to 5.3%) for lactulose, 25.5 ± 5.0% (16.7% to 36.9%) for rhamnose, and 58.8% ± 11.0% (40.1% to 87.8%) for 3-O-methyl-d-glucose. Mean (range) recovery ratio was 0.25 ± 0.06 (0.17 to 0.37) for 51Cr-labeled EDTA to rhamnose, 0.13 ± 0.04 (0.08 to 0.23) for lactulose to rhamnose, and 0.73 ± 0.09 (0.60 to 0.90) for d-xylose to 3-O-methyl-d-glucose. Median (range) percentage urinary recovery was 40.3% (31.6% to 62.7%) for d-xylose and 0% (0% to 0.8%) for sucrose.

Conclusions and Clinical Relevance—Reference values in healthy adult male Beagles for 6 of the most commonly used GIPFT markers were determined. The correlation between results for 51Cr-labeled EDTA and lactulose was not as prominent as that reported for humans and cats; thus, investigators should be cautious in the use and interpretation of GIPFTs performed with sugar probes in dogs with suspected intestinal dysbiosis.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine the optimal sample handling and processing conditions for the carbon 13 (13C)-labeled aminopyrine demethylation blood test (ADBT; phase 1) and determine the reference range for test results (phase 2) in apparently healthy dogs.

Animals—44 apparently healthy dogs (phase 1, 19 dogs; phase 2, 44 dogs).

Procedures—In phase 1, a blood sample from each dog was collected before and 45 minutes after (day 0) IV administration of 13C-labeled aminopyrine (2 mg/kg); aliquots were immediately transferred into tubes containing sodium heparin and hydrochloric acid (samples A and B), sodium heparin alone (samples C, D, and E), or sodium fluoride (sample F). Hydrochloric acid was added to samples C through F at days 7, 14, 21, and 21, respectively. The baseline and 45-minute samples' absolute 13C:12C ratios were determined via fractional mass spectrometry on day 0 (control sample A) or 21 (samples B through F) and used to calculate the percentage dose of 13C recovered in CO2 extracted from samples (PCD). In phase 2, blood samples from each dog were collected into tubes containing sodium fluoride and processed within 3 weeks.

Results—Compared with the control sample value, PCDs for samples C through E differed significantly, whereas PCD in sample F did not. The 13C-ADBT–derived PCD reference range (central 95th percentile) for apparently healthy dogs was 0.08% to 0.2%.

Conclusions and Clinical Relevance—Glycolytic CO2 production in canine blood samples collected during 13C-ADBTs was sufficiently inhibited by sodium fluoride to allow delayed sample analysis and avoid transportation of hydrochloric acid–treated samples.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To indirectly assess the pancreatic response in healthy dogs that were fed diets of different fat compositions with or without supplemental pancreatic enzymes and medium-chain triglycerides (MCTs).

Animals—10 healthy adult dogs.

Procedures—Dogs were fed 4 diets once in random order at 1-week intervals; food was withheld from the dogs for ≥ 12 hours prior to the feeding of each diet. Diets A and B contained 16% and 5% crude fat, respectively; diet C was composed of diet A with pancreatic enzymes; and diet D was composed of diet B with pancreatic enzymes and MCTs. Serum canine trypsin–like immunoreactivity (cTLI) and canine pancreatic lipase immunoreactivity (cPLI) concentrations were measured before (0 hours) and at 1 to 2 and 6 hours after feeding. Serum gastrin concentration was measured at 0 hours and at 5 to 10 minutes and 1 to 2 hours after feeding. A gastrin assay validation study was performed to confirm accuracy of test results in dogs. Data were analyzed by use of a repeated-measures general ANOVA.

Results—Serum cTLI, cPLI, or gastrin concentrations in the dogs did not differ among the different diets fed, among dogs, or over time. When multiple comparisons were analyzed, diet D caused the least amount of measurable pancreatic response, although this difference was not significant.

Conclusions and Clinical Relevance—Results did not indicate a significant effect of dietary fat content or addition of supplemental MCT oil or pancreatic enzymes in diets on serum cTLI, cPLI, or gastrin concentrations in healthy dogs.

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in American Journal of Veterinary Research

Abstract

Objective—To evaluate stability of canine pancreatic lipase immunoreactivity (cPLI) in serum samples and to determine the effect of long-term administration of prednisone on serum cPLI concentrations.

Sample Population—8 canine serum samples for the stability evaluation and serum samples obtained from 6 healthy young adult heterozygous (carrier) dogs with X-linked hereditary nephritis for determining the effect of prednisone administration.

Procedures—To evaluate stability of serum cPLI concentration, an aliquot of each serum sample was stored at each of 4 temperatures between −80° and 24°C; samples were analyzed on days 0, 3, 7, 14, and 21. To determine the effect of long-term prednisone administration, pretreatment serum samples were obtained (days 0 and 14) and prednisone was administered (2.2 mg/kg, q 24 h, PO) on days 15 through 42, with serum samples obtained on days 28 and 42. Additional serum samples were obtained on days 56 and 70.

Results—Mean serum cPLI concentrations did not change significantly from day 0 to day 21 regardless of storage temperature. Serum cPLI concentrations in dogs after prednisone administration were within the reference range for all dogs at all time points, and results of repeated-measures ANOVA revealed that serum cPLI concentrations did not change significantly over time.

Conclusions and Clinical Relevance—Serum cPLI concentrations measured in canine serum samples stored at room temperature, in a refrigerator, or in a freezer at −20° or −80°C were stable for at least 21 days. Also, long-term prednisone administration to dogs did not significantly affect serum cPLI concentrations.

Full access
in American Journal of Veterinary Research

Abstract

Objective—To develop and validate an ELISA for measurement of serum canine pepsinogen A (cPG A) as a diagnostic marker of gastric disorders in dogs and to measure serum cPG A in healthy dogs after food deprivation and after feeding.

Sample Population—Sera from 72 healthy dogs.

Procedure—A sandwich ELISA was developed and validated. The reference range for serum concentrations of cPG A was determined in 64 healthy dogs. Postprandial changes in serum concentrations of cPG A were evaluated in 8 healthy dogs.

Results—Assay sensitivity was 18 µg/L, and the maximum detectable concentration was 1,080 µg/L. The observed-to-expected ratio (O:E) for 3 serial dilutions of 3 serum samples ranged from 69.3 to 104.1%. The O:E for 3 serum samples spiked with 8 concentrations of cPG A ranged from 58.8 to 120.4%. Coefficients of variation for intra- and interassay variability of 3 serum samples ranged from 7.6 to 11.9% and from 10.1 to 13.1%, respectively. Mean ± SD serum concentration of cPG A in healthy dogs was 63.8 ± 31.0 µg/L and the reference range was 18 to 129 µg/L. Significant increases in serum concentrations of cPG A were observed between 1 and 7 hours after feeding.

Conclusions and Clinical Relevance—The ELISA for measuring cPG A was sufficiently sensitive, linear, accurate, precise, and reproducible for clinical use. Serum concentrations of cPG A increase substantially after feeding, which should be taken into account when conducting clinical studies. (Am J Vet Res 2003;64:1146–1150)

Full access
in American Journal of Veterinary Research