Hyperadrenocorticism (HAC) is a common endocrine

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1 J Vet Intern Med 2002;16: The Use of 17-Hydroxyprogesterone in the Diagnosis of Canine Hyperadrenocorticism J.M.E. Ristic, I.K. Ramsey, F.M. Heath, H.J. Evans, and M.E. Herrtage A number of dogs are seen with clinical signs consistent with hyperadrenocorticism (HAC), supporting CBC and biochemical findings, but the disease cannot be confirmed with either the ACTH stimulation test or the low-dose dexamethasone suppression test (LDDST). Therefore, another screening test is required to aid diagnosis in these atypical cases of HAC. The aim of this study was to investigate whether measuring 17-hydroxyprogesterone (OHP) concentrations could be used in this role. Plasma cortisol and OHP concentrations were measured in dogs with clinical signs suggestive of HAC before and after administration of exogenous ACTH. In dogs with HAC, plasma OHP showed an exaggerated response to ACTH stimulation. This was seen in both typical cases of HAC with a positive cortisol response to ACTH administration and in atypical cases with negative screening test results. The test can be performed on plasma already taken for a conventional ACTH stimulation test. Post-ACTH OHP concentrations decreased after treatment with mitotane or adrenalectomy. These results suggest that OHP measurements can be used as an aid to diagnose and manage canine HAC. Key words: ACTH stimulation test; Cushing s syndrome; Dog. Hyperadrenocorticism (HAC) is a common endocrine disease of dogs. 1,2 Most dogs present with classic clinical signs including polyuria and polydipsia, polyphagia, hepatomegaly, a pendulous abdomen, and dermatologic changes including skin thinning, loss of elasticity, and bilaterally symmetric alopecia. 3 Affected dogs have CBC and biochemical abnormalities that support the diagnosis, and specific endocrine tests, including the ACTH stimulation test and the low-dose dexamethasone suppression test (LDDST), usually confirm the diagnosis. However, some dogs have negative results with these tests, despite other suggestive features of the disease. Published work suggests that the ACTH stimulation test will detect 83% of dogs with HAC and the LDDST about 92% of dogs with HAC. 1 The ACTH stimulation test has been reported to detect 88% of patients with pituitary-dependent hyperadrenocorticism (PDH) and about 57% of patients with adrenal-dependent hyperadrenocorticism (ADH). 1 The LDDST confirms 96% of PDH patients and up to 100% of ADH patients. 1 Another report found 3 false-positive responses and 1 false-negative response on an ACTH stimulation test and also 3 falsepositive responses and 1 false-negative response on an LDDST. 4 There is a dilemma regarding confirmation of the diagnosis of HAC in dogs that are negative on both of these tests. In a series of 64 dogs with HAC reported by Feldman, 1 no dog was negative on both screening tests, but such cases do appear to exist. From the Queen s Veterinary School Hospital, Department of Clinical Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge, UK (Ristic, Heath, Herrtage); the Glasgow Veterinary School, University of Glasgow, Bearsden Road, Glasgow, Scotland (Ramsey); and Cambridge Specialist Laboratories Services, Sawston, Cambridge, UK (Evans). Part of the work has been presented at the British Small Animal Veterinary Association Congress 2000, the European Society for Veterinary Internal Medicine Congress 2000, and the American College of Veterinary Internal Medicine Forum Reprint requests: Jelena Ristic, BVetMed, Wey Referrals, Chertsey Road, Woking, Surrey, UK GU21 5BP; jelnbri@ aol.com. Submitted July 24, 2001; Revised October 16, 2001; Accepted January 24, Copyright 2002 by the American College of Veterinary Internal Medicine /02/ /$3.00/0 In the past, we have examined 4 dogs that had classic clinical signs of HAC CBC and biochemical findings that were consistent with the disease and yet were negative on both ACTH stimulation and LDDST. All 4 dogs were treated with mitotane, a their clinical signs resolved, and they survived for up to 2 years on maintenance therapy. This response to therapy was seen as evidence that they had an atypical form of HAC. These dogs provided the initial impetus for the authors to investigate the role of cortisol precursors in HAC. Traditionally, HAC is thought of as a disease in which excess cortisol is produced, either because of a pituitary tumor producing excess ACTH, which leads to overproduction of cortisol, or because of direct oversecretion of cortisol from an adrenal tumor. It is possible that some dogs have a derangement of the steroid production pathway and that some of the precursors may be abnormally increased, even though the cortisol response to ACTH or dexamethasone administration is normal. In humans, adrenal adenomas with a relative blockade of the 11 hydroxylase enzyme have been reported to produce deoxycortisol, 17- hydroxyprogesterone (OHP), and progesterone in response to ACTH in vitro in the absence of a cortisol response. 5 In certain breeds of dogs, an enzyme deficiency is thought to lead to reduced aldosterone or cortisol production, which in turn stimulates ACTH production and consequential overproduction of sex hormones. This, in turn, leads to a congenital adrenal hyperplasia-like syndrome with alopecia. 6,7 OHP is a precursor in the steroid production pathway (Fig 1). Normal ranges for OHP, both before and after administration of synthetic ACTH, have been established for dogs in a variety of breeds. 8,9 The aim of this study was to investigate whether this steroid precursor could be useful in the diagnosis of atypical HAC. Materials and Methods This study included dogs with a diagnosis of HAC seen at the Queen s Veterinary School Hospital, University of Cambridge, between 1998 and In all dogs, a full history and physical examination, CBC, biochemistry, and cortisol and OHP measurements in response to ACTH stimulation were performed. In most dogs, an LDDST also was performed. Further discriminatory tests including

2 434 Ristic et al Fig 1. Steroid production pathway (modified from Cerundolo et al 6 ). endogenous ACTH measurement, abdominal radiography, and ultrasonography were used to differentiate ADH and PDH. Screening tests were performed by the same method in all dogs as detailed below. All cortisol and OHP assays were performed at Cambridge Specialist Laboratories. ACTH Stimulation Test Dogs were fasted overnight, and a heparinized blood sample was taken. Synthetic ACTH in the form of tetracosactrin acetate b was injected IV at a dose of 250 g for dogs 5 kg and 125 g for dogs 5 kg. A 2nd heparinized blood sample was taken 30 minutes later. Cortisol was assayed on all samples by radioimmunoassay c ; the lower limit for the assay was 0.72 g/dl (20 nmol/l), and the interassay variation was 3 7%. The results were considered positive for HAC if the cortisol concentration post-acth administration was 21.7 g/dl (600 nmol/l). The LDDST The LDDST was performed on a day different from the ACTH stimulation test. Dogs were fasted overnight, and a resting heparinized blood sample was taken. Dexamethasone d was injected IV at a dosage of 0.01 mg/kg, and heparinized blood samples were taken at 3 and 8 hours postinjection. Cortisol was assayed on all samples by radioimmunoassay. c The LDDST was considered positive if the cortisol concentration in the 8-hour sample was 1.45 g/dl (40 nmol/l). OHP Assays OHP was assayed on the heparinized plasma samples from the ACTH stimulation tests by solid-phase 121 I radioimmunoassay. e The lower limit of sensitivity for this assay was 0.33 ng/ml (1.0 nmol/l), and the interassay variation was 3 11%. Results of the OHP tests were considered positive if the post-acth OHP concentration was 1.32 ng/ml (4 nmol/l). 8 Dogs were allocated to 2 groups according to their cortisol response to ACTH stimulation. 1 This group consisted of 11 dogs that had a positive cortisol response to the ACTH stimulation test.

3 Hydroxyprogesterone in Hyperadrenocorticism 435 Fig 2. 2 Low-dose dexamethasone suppression tests (LDDSTs). Table 1. Clinical Sign Polyuria/Polydipsia Polyphagia Hepatomegaly Pendulous abdomen Skin and coat changes Seizures Diabetes mellitus 2 2 consisted of 13 dogs, including 3 seen at Glasgow Veterinary School, that had a negative cortisol response to the ACTH stimulation test. These dogs were further subdivided on the basis of their cortisol response to the LDDST (Fig 2). 2a. This group consisted of 6 dogs that had a negative ACTH stimulation test and a positive LDDST. 2b. This group consisted of 4 dogs that had a negative ACTH stimulation test and a negative LDDST. 2c. This group consisted of 3 dogs that had a negative ACTH stimulation test and low plasma cortisol concentrations throughout an LDDST. Controls The OHP both pre- and post-acth administration was compared to concentrations for normal dogs as reported by Curtis et al. 8 The laboratory technique and personnel were the same. The dogs were all healthy pet dogs without evidence of systemic disease. They were aged from 3 to 9 years and were a mixture of breeds. Values from both minimum and maximum photoperiod were used to eliminate any errors due to seasonal variation in hormone concentrations. All dogs had a normal cortisol response to ACTH stimulation. Treatment Confirmed cases of HAC in all groups were treated with either mitotane a or trilostane f or by surgical adrenalectomy. Statistical Methods Statistical Package for the Social Sciences (SPSS) 6.1 for the Power Macintosh was used for statistical analysis. 2 was treated as 1 group because the numbers in each subgroup were small. The Kruskal- Wallis one-way analysis of variance was used to compare data from Clinical signs of the dogs in each group. 1 8/11 7/11 9/11 10/11 10/11 1/11 1/11 2a 5/6 4/6 6/6 6/6 6/6 2b 2/4 4/4 3/4 3/4 4/4 2c 2/3 normal dogs, group 1, and group 2 simultaneously. The Mann-Whitney U-test was used to compare groups 1 and 2 with each other and to compare each individual group with the controls, both pre- and post- ACTH stimulation. When OHP was undetectable, an arbitrary value of ng/ml (0.5 nmol/l) was used. In all comparisons, the statistic was corrected for ties. P.05 was considered significant. Results 1 Eleven dogs had typical HAC, including a positive result on the ACTH stimulation test. All of these dogs displayed 3 or more major clinical signs suggestive of HAC (Table 1). Nine dogs had at least some changes typical of HAC on CBC: 2 had thrombocytosis, 1 had neutrophilia, 7 had lymphopenia, 8 had eosinopenia, and 1 had monocytosis. Biochemical abnormalities also were suggestive of HAC: 2 dogs had low blood urea nitrogen (BUN), 8 had fasting hyperglycemia, 9 had high alkaline phosphatase and alanine aminotransferase activities, and 6 had hypercholesterolemia. Plasma OHP concentrations before ACTH administration in these dogs are shown in Table 2; only 2 dogs had high basal OHP concentrations. The prestimulation concentrations were significantly different (P.009) from those reported for normal dogs 8 (Fig 3). All 11 dogs had high OHP concentrations after ACTH stimulation (Table 2). The dog with a concentration of ng/ml (38 nmol/l) had concurrent diabetes mellitus. Without this dog, the highest post-acth OHP concentration was 5.54 ng/ml (16.8 nmol/l). The post-acth results in group 1 were much higher than in normal dogs 8 (Fig 3), and this finding was highly statistically significant (P.001). Seven of these dogs subsequently were confirmed to have PDH, and 2 were found to have adrenal tumors on the basis of a combination of endogenous plasma ACTH measurement and ultrasonography. 10 The 2 remaining dogs did not undergo discriminatory tests. Nine of the dogs in group 1 were treated medically. At the endpoint of induction therapy or after 7 days (whichever was sooner), an ACTH stimulation test was performed. Cortisol was assayed in all dogs, and OHP was assayed in 8. In 1 dog diagnosed with ADH that was treated with mitotane, there was a marked reduction in both post-acth cortisol from 47.2 g/dl (1,303 nmol/l) to g/dl

4 436 Ristic et al Table 2. OHP concentrations in response to ACTH stimulation. Pre-ACTH OHP (ng/ml) Post-ACTH OHP (ng/ml) Pre-ACTH OHP (nmol/l) Post-ACTH OHP (nmol/l) Controls 1 2a 2b 2c ( 0.33) (0.46) (0.46) (1.1) ( 0.33) (0.83) (4.22) (1.9) (3.7) (1.6) ( 1) (1.4) (1.4) 1 6 (3.3) ( 1) (2.5) (12.8) (5.85) (11.2) (4.9) OHP, 17-hydroxyprogesterone. (335 nmol/l) and post-acth OHP from 3.76 ng/ml (11.4 nmol/l) to 1.19 ng/ml (3.6 nmol/l). Another dog with ADH showed little response to mitotane. Five dogs with PDH treated with mitotane showed a reduction in both cortisol and OHP concentrations post-acth stimulation. Post- ACTH cortisol for the 5 dogs decreased from a median of 29.9 g/dl (824 nmol/l) to 0.72 g/dl (20 nmol/l) postmitotane, and the median post-acth OHP decreased from 4.8 ng/ml (14.6 nmol/l) to 0.33 ng/ml (1.0 nmol/ L) after therapy. One dog that was treated with trilostane and showed clinical improvement demonstrated a reduction in the post-acth cortisol concentrations from g/ dl (1,066 nmol/l) to 0.72 g/dl (20 nmol/l) but an increase in the post-acth OHP from 4.22 ng/ml (12.8 nmol/l) to ng/ml (33 nmol/l) after 2 months of treatment. 2 Fig 3. Plasma 17-hydroxyprogesterone (OHP) before and after ACTH administration in control and diseased dogs. Thirteen dogs were seen with classic clinical signs of HAC CBC and biochemical findings that were consistent with the disease but with a negative result on ACTH stimulation testing and varied responses to LDDST (Fig 2). Dogs in all 3 subgroups displayed a combination of at least 3 of the classic clinical signs, as in group 1 (Table 1). Biochemistry, and in some cases CBC, was suggestive of HAC in all of these dogs: CBC disclosed neutrophilia in 2 dogs (1 each in group 2a,c), lymphopenia in 9 dogs (4 in group 2a, 3 in 2b, and 1 in 2c), and eosinopenia in 6 dogs (4 in group 2a and 1 each in 2b and 2c). One dog in group 2a had thrombocytosis. Two dogs in group 2a did not show any evidence of a steroid response on CBC. Biochemistry disclosed high alkaline phosphatase activity in all dogs and high alanine aminotransferase activity in 7 dogs (4 in group 2a, 1 in 2b, and 2 in 2c). BUN was low in 1 dog from each group, and hyperglycemia was seen in 9 dogs (5 in group 2a and 2 in group 2b,c). Cholesterol was high in the 7 dogs in which it was measured. The results of the OHP concentrations pre- and post- ACTH stimulation are shown in Table 2 for groups 2a c. All 3 groups demonstrated an exaggerated response to ACTH administration. The response of the individual groups is illustrated in Figure 3, but statistical analysis was not possible because of the small numbers of dogs, and group 2 was analyzed as a whole. The basal plasma OHP concentrations were significantly different (P.01) from those previously reported in normal dogs 8 (Fig 3). After ACTH administration, all 13 dogs had exaggerated increases in plasma OHP concentrations, and these were markedly different from normal dogs (Fig 3). This difference was statistically significant (P.001). When groups 1 and 2 were compared to each other, there was no significant difference between the pre-acth OHP concentrations, but there was a significant difference in the results post-acth stimulation (P.02). With a combination of endogenous plasma ACTH concentrations and adrenal ultrasonography, 10 it was shown that all dogs in group 2a had PDH and that all dogs in group 2c had ADH. 2b contained 3 dogs with PDH and 1 with ADH. Ten dogs in group 2 were treated, and an additional dog was returned to the referring veterinary surgeon for treatment. Plasma cortisol and OHP concentrations were measured at the endpoint of mitotane induction therapy and 2 months after beginning treatment in some dogs. In group 2a, 3 mitotane-treated PDH dogs showed a reduction in both cortisol and OHP concentrations post-acth administration. Cortisol concentrations decreased from a median of 12.8 g/dl (354 nmol/l) to 0.72 g/dl (20 nmol/l), and OHP decreased from 3.0 ng/ml (9.2 nmol/l) to 0.33 ng/ml (1.0 nmol/l). Another dog showed clinical improvement with mitotane. In group 2b, 1 trilostane-treated dog

5 Hydroxyprogesterone in Hyperadrenocorticism 437 with PDH showed a reduction in cortisol concentrations post-acth (from g/dl [493 nmol/l] to 0.72 g/ dl [20 nmol/l]) but an increase in plasma OHP concentrations from 4.39 ng/ml (13.3 nmol/l) to 11.8 ng/ml (36 nmol/l) after 2 months of treatment, which is similar to the dog treated with trilostane in group 1. The other 2 dogs with PDH showed clinical improvement on mitotane. One dog in group 2b had an adrenal tumor that was removed surgically and showed a reduction in both plasma cortisol and OHP concentrations post-acth stimulation. Two dogs with ADH from group 2c had an initial response to mitotane therapy, eventually had postmortem examinations, and on histopathology, adrenal tumors were confirmed. Discussion Cortisol, sex hormones, and aldosterone all are produced in the canine adrenal gland (Fig 1). The syndrome of HAC is traditionally thought to be due to overproduction of cortisol either from an adrenal tumor or as a result of overproduction of ACTH from a pituitary tumor. However, there are a number of animals in which a definitive diagnosis of HAC cannot be made by currently recommended tests. There are some reports in the literature of adrenocortical tumors secreting hormones other than cortisol. For example, there has been a single case report of a dog with an adrenocortical tumor secreting deoxycorticosterone. 11 Two dogs with HAC due to adrenocortical neoplasia have previously been reported to have an exaggerated stimulation of plasma OHP concentrations in response to ACTH stimulation. 12 Abnormal production of sex hormones has been reported in Pomeranians and Chows due to a congenital adrenal hyperplasia-like syndrome mediated by a deficiency of 1 enzyme (usually 21 hydroxylase) in the steroid production pathway. 7 Until recently, there have been no reports of plasma OHP concentrations being used to diagnose PDH. A series of 11 dogs with an exaggerated cortisol response to ACTH, including 5 PDH patients, recently has been published, and 6 dogs in that study had an exaggerated OHP response to ACTH administration. 13 All of the dogs in our study presented with classic signs of HAC. 1 had positive conventional test results and also high OHP concentrations after ACTH stimulation, and group 2 dogs were negative on 1 or more conventional tests but had high OHP concentrations after ACTH stimulation. 2b dogs were negative on both ACTH stimulation and LDDST, and group 2c had equivocal LDDST results, but both groups had high OHP concentrations after ACTH stimulation and therefore had an atypical form of HAC. Dogs from all groups showed resolution of clinical signs with standard therapeutic regimens for HAC. Adrenal ultrasonography, endogenous plasma ACTH concentrations, and postmortem examination confirmed a mixture of dogs with PDH and ADH. All of the dogs in this study displayed at least 3 of the classic clinical signs of HAC. CBC was supportive in all but 4 dogs, with evidence of neutrophilia, lymphopenia, or eosinopenia. 3 Biochemistry showed a high plasma alkaline phosphatase activity in all dogs, with the exception of 2 dogs in group 1, and this finding is consistent with previous reports, suggesting that 90% of dogs with HAC show this biochemical abnormality. 3 Other abnormal biochemical results were present in varying combinations. Therefore, clinically, there was a high index of suspicion for HAC. An ACTH stimulation test was used as the 1st screening test because it is easy to perform, requires only 2 blood samples, and provides useful information for monitoring therapy. 1 In group 1, dogs already had a diagnosis of HAC from their conventional ACTH stimulation tests. The plasma OHP measurements were made to confirm that this hormone would also increase in classic HAC, as might be predicted. Results of all of the dogs with typical HAC were supportive of this hypothesis, and the findings were similar in both ADH and PDH patients. The good clinical response and the fall in plasma OHP concentrations after mitotane treatment (in 5 PDH patients and 1 ADH patient) are evidence that the measurement of plasma OHP concentrations can be useful as an aid to the diagnosis and management of HAC. 2a consisted of 6 dogs with normal cortisol responses to ACTH stimulation, but these dogs had exaggerated OHP responses with the same plasma samples as well as positive LDDST results for HAC. Four dogs responded well clinically to mitotane therapy, and 3 had a documented decrease in their OHP concentrations, further confirming that OHP may be useful in both the diagnosis and the monitoring of HAC. In group 2b, the 4 dogs with negative ACTH stimulation and LDDST showed marked increases in OHP in response to exogenous ACTH administration, which suggests an abnormality in the cortisol production pathway. Their good clinical response to treatment (1 with adrenalectomy, 2 on mitotane, and 1 on trilostane) is supportive of the diagnosis of an atypical form of HAC (ie, with HAC causing secretion of excess OHP rather than cortisol). 2c consisted of 3 dogs that had a negative ACTH stimulation test result and low plasma cortisol concentrations throughout their LDDST (Fig 2). Cortisol concentrations were either below or close to 1.45 g/dl (40 nmol/ L) for the duration of the test. Two of these dogs had a cortisol concentration 1.45 g/dl (40 nmol/l) at 8 hours, and 1 had a concentration of 1.63 g/dl (45 nmol/l). Hence, it was difficult to make a convincing diagnosis of HAC in these dogs. The plasma OHP response to ACTH was exaggerated, endogenous ACTH was undetectable, abdominal ultrasound confirmed the presence of an adrenal mass, and, in 2 dogs, postmortem confirmation of an adrenal tumor was obtained. Therefore, there was no doubt regarding the diagnosis of ADH in these dogs, and plasma OHP was useful in confirming the diagnosis in these dogs with low plasma cortisol concentrations throughout the LDDST, as suggested by Norman et al. 12 Hence, these dogs also had an atypical form of HAC with excess OHP secretion rather than cortisol secretion. Two dogs were treated with trilostane, 1 in group 1 and 1 in group 2b, and both showed a decrease in cortisol and an increase in OHP concentrations post-acth administration. Trilostane is reported to be a steroid synthesis inhibitor, acting at the level of 3 hydroxysteroid dehydrogenase, and would be expected to cause a decrease in OHP

6 438 Ristic et al concentrations (Fig 1). The opposite was true in this study, and OHP does not appear to be a useful marker for monitoring response to this drug. It is possible there is crossreactivity between 17-hydroxypregnenolone and OHP in the radioimmunoassay, and further work is necessary to investigate this possibility. In group 1, the cortisol response to ACTH was more marked than in group 2, and the OHP response to ACTH mirrored this finding. There was no significant difference in cortisol or OHP between the 2 groups pre-acth, but post-acth, there was a significant difference in both hormones. Perhaps this finding is due to the linear production pathway of cortisol. Nonetheless, there was a significant difference in OHP post-acth in both groups 1 and 2 compared to the controls. At this stage, it is not clear whether OHP contributes to the clinical signs in affected dogs, especially the atypical patients in group 2b,c, or whether another hormone in the pathway is involved. Progesterone can cause clinical signs similar to those associated with HAC in cats. 14 Progestins act as glucocorticoid agonists and suppress the hypothalamic-pituitary-adrenocortical axis in the dog. 15 There are reports of humans with incidental adrenal tumors who demonstrate exaggerated OHP responses to ACTH, and yet these patients have no clinical signs of HAC, 16,17 suggesting that another hormone may be responsible for the clinical signs. However, other reports in the human literature suggest that some patients with incidental adrenal tumors have various hormonal abnormalities, including an exaggerated OHP response to ACTH, a low dihydroepiandrostenedione response to ACTH, and a high morning cortisol concentration. Although they do not have classic signs of HAC, they do have abnormalities such as obesity, hypertension, type II diabetes mellitus, and abnormal serum lipid concentrations, which resolve after adrenalectomy. 18 Also, the activity of these precursors may be different in humans. Furthermore, the response of cortisol to ACTH administration is affected by stress, and this may also be the case with OHP, because it is part of a linear production pathway. Hence, OHP response to administration of exogenous ACTH may be a useful method for confirming HAC in atypical cases that have features suggestive of HAC but normal ACTH and LDDST results. OHP also appears to be overstimulated in typical HAC diagnosed by the usual methods. The test can be performed on plasma samples already taken for a conventional ACTH stimulation test; this is not invasive for the animal or time-consuming for the clinician and owner. Also, the cost is approximately the same as for the cortisol assay, making this a very practical test. Results must be interpreted in conjunction with clinical findings and other supportive laboratory results. From these preliminary data, the OHP response to ACTH also appears useful for monitoring dogs receiving mitotane treatment or after surgical adrenalectomy. Footnotes a Lysodren, Bristol-Myers Squibb, Hounslow, UK b Synacthen, Alliance Pharmaceuticals Limited, Chippenham, UK c Cambridge Specialist Laboratories Services, Unit C, Peek House, Dales Manor Business Park, Sawston, Cambridge, UK CB2 4TJ d Dexadreson, Intervet UK Limited, Milton Keynes, UK e Coat-A-Count 17 -OH Progesterone, Diagnostic Products Corporation, Caernarfon, UK f Modrenal, Wanskerne Limited, St Austell, UK Acknowledgments We would like to thank the referring veterinary surgeons and all the staff and students at the Cambridge and Glasgow Universities who helped to care for these dogs, particularly the veterinary surgeons who had primary responsibility for some of the dogs included. Also, we thank Cathy Curtis for allowing us to use her control data and Brian Catchpole for his constructive comments on the manuscript. References 1. Feldman EC. Comparison of ACTH response and dexamethasone suppression as screening tests in canine hyperadrenocorticism. J Am Vet Med Assoc 1983;182: Feldman EC, Nelson RW, Feldman MS. Use of low- and highdose dexamethasone tests for distinguishing pituitary-dependent from adrenal tumor hyperadrenocorticism in dogs. J Am Vet Med Assoc 1996;209: Herrtage ME. Canine hyperadrenocorticism. In: Torrance AG, Mooney CT, eds. Manual of Small Animal Endocrinology, 2nd ed. Gloucester, UK: British Small Animal Veterinary Association; 1998: Van Liew CH, Greco DS, Salman MD. Comparison of results of adrenocorticotrophic hormone stimulation and low-dose dexamethasone suppression tests with necropsy findings in dogs: 81 cases ( ). J Am Vet Med Assoc 1997;211: Lamberts SWJ, Zuiderwijk J, Uitterlinden P, et al. Characterisation of adrenal autonomy in Cushing s syndrome: A comparison between in vivo and in vitro responsiveness of the adrenal gland. J Clin Endocrinol Metab 1989;70: Cerundolo R, Lloyd DH, McNeil PE, Evans H. An analysis of factors underlying hypotrichosis and alopecia in Irish Water Spaniels in the United Kingdom. Vet Dermatol 2000;11: Schmeitzel LP, Lothrop CD, Rosenkrantz WS. Congenital adrenal hyperplasia-like syndrome. In: Bonagura JD, Kirk RW, eds. Kirk s Current Veterinary Therapy XII. Philadelphia, PA: WB Saunders; 1995: Curtis CF, Evans H, Lloyd DH. Investigation of the reproductive and growth hormone status of dogs affected by idiopathic recurrent flank alopecia. J Small Anim Pract 1996;37: Schmeitzel LP, Lothrop CD. Hormonal abnormalities in Pomeranians with normal coat and in Pomeranians with growth hormoneresponsive dermatosis. J Am Vet Med Assoc 1990;197: Gould SM, Baines EA, Mannion PA, et al. Use of endogenous ACTH concentration and adrenal ultrasonography to distinguish the cause of canine hyperadrenocorticism. J Small Anim Pract 2001;42: Reine NJ, Hohenhaus AE, Peterson ME, Patnaik AK. Deoxycortisone secreting adrenocortical carcinoma in a dog. 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