Stefan Schellenberg

Urinary catecholamine and metanephrine to creatinine ratios in dogs with hyperadrenocorticism or pheochromocytoma, and in healthy dogs.

BACKGROUND Urinary catecholamines and metanephrines are used for the diagnosis of pheochromocytoma (PHEO) in dogs. Hyperadrenocorticism (HAC) is an important differential diagnosis for PHEO. OBJECTIVES To measure urinary catecholamines and metanephrines in dogs with HAC. ANIMALS Fourteen dogs with HAC, 7 dogs with PHEO, and 10 healthy dogs. METHODS Prospective clinical trial. Urine was collected during initial work-up in the hospital; in dogs with HAC an additional sample was taken at home 1 week after discharge. Parameters were measured using high-pressure liquid chromatography and expressed as ratios to urinary creatinine concentration. RESULTS Dogs with HAC had significantly higher urinary epinephrine, norepinephrine and normetanephrine to creatinine ratios than healthy dogs. Urinary epinephrine, norepinephrine, and metanephrine to creatinine ratios did not differ between dogs with HAC and dogs with PHEO, whereas the urinary normetanephrine to creatinine ratio was significantly higher (P= .011) in dogs with PHEO (414, 157.0-925.0, median, range versus (117.5, 53.0-323.0). Using a cut-off ratio of 4 times the highest normetanephrine to creatinine ratio measured in controls, there was no overlap between dogs with HAC and dogs with PHEO. The variables determined in urine samples collected at home did not differ from those collected in the hospital. CONCLUSION AND CLINICAL IMPORTANCE Dogs with HAC might have increased concentrations of urinary catecholamines and normetanephrine. A high concentration of urinary normetanephrine (4 times normal), is highly suggestive of PHEO.

Endocrine hypertension in small animals.

Hypertension is classified as idiopathic or secondary. In animals with idiopathic hypertension, persistently elevated blood pressure is not caused by an identifiable underlying or predisposing disease. Until recently, more than 95% of cases of hypertension in humans were diagnosed as idiopathic. New studies have shown, however, a much higher prevalence of secondary causes, such as primary hyperaldosteronism. In dogs and cats, secondary hypertension is the most prevalent form and is subclassified into renal and endocrine hypertension. This review focuses on the most common causes of endocrine hypertension in dogs and cats.

Microbiologic evaluation of gallbladder bile of healthy dogs and dogs with iatrogenic hypercortisolism: a pilot study.

BACKGROUND In people, hypercortisolism (HC) has been associated with acalculous cholecystitis and biliary dyskinesia, which may potentiate ascending biliary infections. In dogs, an association between HC and gallbladder disease recently has been documented, although the role of bacteria remains controversial. Furthermore, there is no information on the gallbladder bile microbial flora in healthy dogs. OBJECTIVES To investigate the microbial flora in gallbladder bile in healthy dogs, the relationship between iatrogenic hyperadrenocorticism and bactibilia and possible changes in biliary microbial flora after cortisol withdrawal in dogs. ANIMALS Six control dogs and 6 dogs treated with hydrocortisone. METHODS Gallbladder bile obtained by percutaneous ultrasound-guided cholecystocentesis was cultured aerobically and anaerobically and examined cytologically before (d0), during (d28, d56, d84), and after (d28p, d56p, d84p) administration of hydrocortisone (8 mg/kg PO q12h). RESULTS In the control group, 2/42 bile cultures yielded bacterial growth (Enterococcus sp.; Escherichia coli on d0) and 1/42 bile smears had cytological evidence of bacteria (d28). In the HC group, 2/42 bile cultures yielded bacterial growth (Enterococcus sp. on d28; Bacillus sp. on d28p) and 3/42 bile smears had cytological evidence of bacteria (d84, d84, d28p). All dogs remained healthy throughout the study period (168d). CONCLUSIONS AND CLINICAL IMPORTANCE Based on the results of conventional bacterial culture techniques, gallbladder bile of healthy dogs periodically may harbor bacteria, which do not appear to be clinically relevant. A 3-month period of iatrogenic HC was not associated with bactibilia. A higher prevalence of bactibilia may be detected with micromolecular techniques.

Salivary cortisol concentrations in healthy dogs and dogs with hypercortisolism.

BACKGROUND Measurement of salivary cortisol is a useful diagnostic test for hypercortisolism (HC) in humans. OBJECTIVES To determine whether measurement of salivary cortisol concentration is a practical alternative to plasma cortisol to diagnose HC, to validate the use of salivary cortisol, and to examine the effect of time of day and sampling location on salivary cortisol. ANIMALS Thirty healthy dogs and 6 dogs with HC. METHODS Prospective, observational clinical trial including healthy volunteer dogs and dogs newly diagnosed with HC. Salivary and plasma cortisol concentrations were measured with an immunoassay analyzer. Intra- and interassay variability, linearity, and correlation between salivary and plasma cortisol concentrations were determined. RESULTS The required 300 microL of saliva could not be obtained in 88/326 samples from healthy dogs and in 15/30 samples from dogs with HC. The intra-assay variability for measurement of salivary cortisol was 5-17.7%, the interassay variability 8.5 and 17.3%, and the observed to expected ratio 89-125%. The correlation (r) between salivary and plasma cortisol was 0.98. The time of day and location of collection did not affect salivary cortisol concentrations. Dogs with HC had significantly higher salivary cortisol values than healthy dogs (10.2 +/- 7.3 nmol/L versus 1.54 +/- 0.97 nmol/L; P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE The ROCHE Elecsys immunoassay analyzer correctly measured salivary cortisol in dogs. However, a broad clinical application of the method seems limited, because of the large sample volume required.

Remission of diabetes mellitus in cats cannot be predicted by the arginine stimulation test

BACKGROUND Cats with diabetes mellitus frequently achieve clinical remission, suggesting residual β-cell function. Responsiveness of β-cells to arginine persists the longest during diabetes progression, making the intravenous arginine stimulation test (IVAST) a useful tool to assess residual insulin and glucagon secretion. HYPOTHESIS Diabetic cats with and without remission will have different arginine-induced insulin or glucagon response. ANIMALS Seventeen cats with diabetes, 7 healthy cats. METHODS Blood samples collected on admission and during subsequent IVAST. Glucose, insulin, and glucagon were measured. Response to IVAST was assessed by calculating the insulin and glucagon area under the curve (AUC) and the AUC glucagon-to-insulin ratio. Diabetic cats were treated with insulin and were followed for 18 weeks. Remission was defined as normoglycemia and disappearance of clinical signs of diabetes for ≥4 weeks, without requiring insulin. RESULTS Seven diabetic cats (41%) achieved remission. On admission, blood glucose concentration was significantly lower in cats with remission (median, 389 mg/dL; range, 342-536 mg/dL) than in those without remission (median, 506 mg/dL; range, 266-738 mg/dL). After IVAST, diabetic cats with remission had higher AUC glucagon-to-insulin ratios (median, 61; range, 34-852) than did cats without remission (median, 26; range, 20-498); glucose, insulin, and glucagon AUCs were not different. Diabetic cats had lower insulin AUC than did healthy cats but comparable glucagon AUC. CONCLUSIONS AND CLINICAL IMPORTANCE Diabetic cats with and without remission have similar arginine-stimulated insulin secretion on admission. Although cats with remission had lower blood glucose concentrations and higher AUC glucagon-to-insulin ratios, large overlap between groups prevents use of these parameters in clinical practice.

Evaluation of four methods used to measure plasma insulin-like growth factor 1 concentrations in healthy cats and cats with diabetes mellitus or other diseases

OBJECTIVE To evaluate 4 methods used to measure plasma insulin-like growth factor (IGF) 1 concentrations in healthy cats and cats with diabetes mellitus or other diseases. ANIMALS 39 healthy cats, 7 cats with diabetes mellitus, and 33 cats with other diseases. PROCEDURES 4 assays preceded by different sample preparation methods were evaluated, including acid chromatography followed by radioimmunoassay (AC-RIA), acid-ethanol extraction followed by immunoradiometry assay (AEE-IRMA), acidification followed by immunochemiluminescence assay (A-ICMA), and IGF-2 excess followed by RIA (IE-RIA). Validation of the methods included determination of precision, accuracy, and recovery. The concentration of IGF-1 was measured with all methods, and results were compared among cat groups. RESULTS The intra-assay coefficient of variation was < 10% for AC-RIA, A-ICMA, and AEE-IRMA and 14% to 22% for IE-RIA. The linearity of dilution was close to 1 for each method. Recovery rates ranged from 69% to 119%. Five healthy cats had IGF-1 concentrations > 1,000 ng/mL with the AEE-IRMA, but < 1,000 ng/mL with the other methods. Compared with healthy cats, hyperthyroid cats had significantly higher concentrations of IGF-1 with the A-ICMA method, but lower concentrations with the IE-RIA method. Cats with lymphoma had lower IGF-1 concentrations than did healthy cats regardless of the method used. CONCLUSIONS AND CLINICAL RELEVANCE Differences in the methodologies of assays for IGF-1 may explain, at least in part, the conflicting results previously reported in diabetic cats. Disorders such as hyperthyroidism and lymphoma affected IGF-1 concentrations, making interpretation of results more difficult if these conditions are present in cats with diabetes mellitus.

Effect of long-term adaptation on indirect measurements of systolic blood pressure in conscious untrained beagles

To evaluate the effect of an adaptation period on systemic blood pressure readings, systolic blood pressure was measured in 12 young adult untrained beagles over several weeks by means of a Doppler flow detector and oscillometric devices. The pressure decreased gradually and significantly, and levelled out after 14 days. The median (range) of values obtained by Doppler were 166 (149 to 200) mmHg initially, 145 (119 to 176) mmHg on day 9, 138 (118 to 165) mmHg on day 10, 127 (111 to 139) mmHg on day 35, 124 (115 to 143) mmHg on day 94 and 127 (114 to 142) mmHg on day 161. All the later measurements were significantly lower than the initial measurement. Male dogs had higher blood pressures than females on each occasion. The blood pressure readings obtained with one of the oscillometric devices and the Doppler device were comparable and correlated significantly.

Effects of iatrogenic hypercortisolism on gallbladder sludge formation and biochemical bile constituents in dogs

An association between gallbladder mucoceles and hypercortisolism (HC) was recently described in dogs. Because the formation of a mucocele from clear bile without the transitional formation of microprecipitates appears unlikely, the aim of this study was to investigate the effects of iatrogenic HC on sludge formation and changes in the biochemical composition of bile. Bile samples from 6 dogs obtained by percutaneous ultrasound-guided cholecystocentesis before (day 0), during (days 28, 56, and 84), and after (days 28p, 56p, and 84p) oral administration of hydrocortisone (8 mg/kg every 12 h) were analysed for calcium, cholesterol and bilirubin concentrations and pH. In addition the gallbladder was examined ultrasonographically for sludge. Six dogs receiving a placebo served as controls. Although gallbladder sludge was observed in all treated dogs at day 56, it was also noted in 50% of control dogs, and no significant differences were seen between groups at any sampling time. Bilirubin and cholesterol concentrations decreased significantly and reversibly during treatment, and calcium concentration showed a similar trend. Bile pH was consistently slightly alkaline during iatrogenic HC, whereas it was slightly acidic in control animals. A 3-month period of iatrogenic HC does not lead to ultrasonographically detectable gallbladder sludge or to an increase in bile constituents that are commonly implicated in sludge formation in humans.

Effect of twice-daily oral administration of hydrocortisone on the bile acids composition of gallbladder bile in dogs.

OBJECTIVE To investigate the effects of twice-daily oral administration of hydrocortisone on the bile acids composition of gallbladder bile in dogs. ANIMALS 6 placebo-treated control dogs and 6 hydrocortisone-treated dogs. PROCEDURES Dogs received hydrocortisone (median dose, 8.5 mg/kg) or a gelatin capsule (control group) orally every 12 hours for 84 days. Gallbladder bile samples were obtained via percutaneous ultrasound-guided cholecystocentesis from each dog before (day 0 [baseline]), during (days 28, 56, and 84), and after (days 28p, 56p, and 84p) treatment for differentiated quantification of unconjugated bile acids and taurine-conjugated and glycine-conjugated bile acids via high-performance liquid chromatography-tandem mass spectrometry. RESULTS Treatment with hydrocortisone for 84 days resulted in significant and reversible increases in the concentrations of unconjugated bile acids (ie, cholic, chenodeoxycholic, and deoxycholic acids) and a significant and reversible decrease in the concentration of total taurine-conjugated bile acids, compared with baseline or control group values. Treatment with hydrocortisone had no effect on bile concentrations of glycine-conjugated bile acids. CONCLUSIONS AND CLINICAL RELEVANCE In dogs, hydrocortisone administration caused reversible shifts toward higher concentrations of the more hydrophobic unconjugated bile acids (chenodeoxycholic acid and deoxycholic acid) and toward lower concentrations of the amphipathic taurine-conjugated bile acids in gallbladder bile. These data suggest that similar bile acids changes could cause major alterations in gallbladder structure or function over time in hypercortisolemic dogs.

Validation of an enzyme-linked immunosorbent assay (ELISA) for the measurement of canine S100A12.

BACKGROUND Canine S100 calcium-binding protein A12 (cS100A12) shows promise as biomarker of inflammation in dogs. A previously developed cS100A12-radioimmunoassay (RIA) requires radioactive tracers and is not sensitive enough for fecal cS100A12 concentrations in 79% of tested healthy dogs. An ELISA assay may be more sensitive than RIA and does not require radioactive tracers. OBJECTIVE The purpose of the study was to establish a sandwich ELISA for serum and fecal cS100A12, and to establish reference intervals (RI) for normal healthy canine serum and feces. METHODS Polyclonal rabbit anti-cS100A12 antibodies were generated and tested by Western blotting and immunohistochemistry. A sandwich ELISA was developed and validated, including accuracy and precision, and agreement with cS100A12-RIA. The RI, stability, and biologic variation in fecal cS100A12, and the effect of corticosteroids on serum cS100A12 were evaluated. RESULTS Lower detection limits were 5 μg/L (serum) and 1 ng/g (fecal), respectively. Intra- and inter-assay coefficients of variation were ≤ 4.4% and ≤ 10.9%, respectively. Observed-to-expected ratios for linearity and spiking recovery were 98.2 ± 9.8% (mean ± SD) and 93.0 ± 6.1%, respectively. There was a significant bias between the ELISA and the RIA. The RI was 49-320 μg/L for serum and 2-484 ng/g for fecal cS100A12. Fecal cS100A12 was stable for 7 days at 23, 4, -20, and -80°C; biologic variation was negligible but variation within one fecal sample was significant. Corticosteroid treatment had no clinically significant effect on serum cS100A12 concentrations. CONCLUSIONS The cS100A12-ELISA is a precise and accurate assay for serum and fecal cS100A12 in dogs.