Choosing the Right Diagnostic Tests for Hyperadrenocorticism
World Small Animal Veterinary Association World Congress Proceedings, 2003
Carmel T. Mooney, MVB, MPhil, PhD, DECVIM-CA, MRCVS
Department of Small Animal Clinical Studies, Faculty of Veterinary Medicine, University College Dublin, Belfield
Dublin, Ireland

There are numerous diagnostic tests recommended for hyperadrenocorticism. Unfortunately there is no true gold standard. There is a wide variation in veterinary input, invasiveness and time and financial costs between the tests and each differs significantly with regard to diagnostic sensitivity, specificity and efficiency. Some of the tests are capable of differentiating the site of the lesion (pituitary versus adrenal) while others may be useful in monitoring the response to therapy. Knowledge of the characteristics and an understanding of the advantages and disadvantages of each test allows for selection of the most appropriate one in the individual patient under investigation.

Basal circulating cortisol concentration

Basal circulating cortisol measurement is of little diagnostic value alone because of the wide overlap in results from healthy dogs and those with hypo-and hyperadrenocorticism and non-adrenal illness.

Adrenocorticotropic hormone stimulation test

The adrenocorticotropic hormone (ACTH) stimulation test is generally used to screen for the presence of hyperadrenocorticism. It distinguishes between iatrogenic and spontaneous hyperadrenocorticism and is useful in the monitoring of adrenocorticolytic therapy.

It is a simple and quick test easily performed in the practice environment. Both natural and synthetic (tetracosactrin, cosyntropin) ACTH produce similar cortisol responses and can be used interchangeably.(6,27) The standard protocol for synthetic ACTH is measurement of circulating cortisol concentration before and 1-hour after a single intramuscular or intravenous injection of 250 µg/dog.(10) While a lower dose of tetracosactrin (1.0 µg/kg) can be used, care must be taken to time the poststimulation cortisol sample precisely so as not to miss peak cortisol concentration.(15) Tetracosactrin can be reconstituted and stored frozen at -20°C in plastic syringes for six months with no adverse effects on bioactivity.(7)

Dogs with hyperadrenocorticism theoretically have an exaggerated response to ACTH. The absolute post-ACTH cortisol concentration is most frequently used to assess the response during an ACTH stimulation test. Although values above the reference range are frequently cited as abnormal, most clinicians use a value that significantly exceeds this range, often between 600 and 650 nmol/l. Approximately 80 % of dogs with pituitary dependent hyperadrenocorticism have an exaggerated cortisol response to ACTH while only approximately 60 % of dogs with adrenal tumours (AT) have such results.(3,13,18,20) A diagnosis of hyperadrenocorticism cannot be excluded based on a reference range test result. However, an advantage of the ACTH stimulation test is that it is specific (approximately 0.9).(13,25) Occasionally, dogs with AT have a subnormal cortisol response to exogenous ACTH (17,20)

Low-dose dexamethasone suppression test

In healthy dogs, glucocorticoids exert negative feedback inhibition on ACTH secretion. A low-dose of dexamethasone (0.01-0.015 mg/kg) administered intravenously to healthy dogs, causes inhibition of ACTH secretion and reduced plasma cortisol concentrations within 2 to 3 hours lasting up to 8 hours. In hyperadrenocorticism, the 8 hour cortisol value is not sufficiently suppressed and remains above 30 to 40 nmol/l. The low dose dexamethasone suppression test is an extremely sensitive diagnostic test. In approximately 95 % of dogs with PDH and 100 % of dogs with AT, cortisol suppression is inadequate.(3,5,25) Unfortunately, false-positive test results may occur in dogs with non-adrenal disease and the specificity of the test has been reported to be as low as 0.44.(13) In general, the more severe the non-adrenal illness, the more likely that cortisol suppression will not occur.

An additional value in performing a low dose dexamethasone suppression test is its ability to distinguish between PDH and AT in up to 60 % of cases when circulating cortisol concentrations are measured at 3 or 4 and at 8 hours. Criteria that indicate a diagnosis of PDH include a decrease of circulating cortisol concentration to less than a laboratory reference value at 3 or 4 hours, less than 50 % of the baseline value at 3 or 4 hours, or less than 50 % of the baseline value at 8 hours.(5,16) However, PDH cannot be ruled out if suppression does not occur. Only rarely, do AT exhibit cortisol suppression.(17)

Urinary Cortisol:Creatinine Ratio

The determination of the cortisol (corticoid):creatinine ratio (UCCR )in urine samples taken in the morning can be used in the investigation of hyperadrenocorticism in dogs. It is the least stressful of all the diagnostic tests as owners can obtain samples in the home environment. It is also extremely sensitive with a (4,12,21,23,24) reported range of 0.75 to However, it lacks specificity with values as low as 0.21 especially if the animal is stressed or concurrent moderate to severe non-adrenal illness is present.(13,23,26) Overall, its high negative predictive value suggests that hyperadrenocorticism is unlikely if the UCCR is within the reference range but that further investigation for hyperadrenocorticism is warranted if it is elevated.

Extremely elevated UCCR values occur almost exclusively in PDH. Suppression to greater than 50 % of baseline following oral administration of three doses of dexamethasone (0.1 mg/kg) is similarly consistent with PDH. Overall, it is considered an inappropriate test for accurate monitoring of adrenocorticolytic therapy.(1,9,19)

Measurement of 17a-Hydroxyprogesterone

Measurement of 17a-hydroxyprogesterone (17-OHP) is considered useful as a marker for alopecia associated with a sex hormone imbalance in the dog. Its measurement has also proven useful in the investigation of dogs with AT that have subnormal cortisol responses to exogenous ACTH.(17)

The 17-α-hydroxylase enzyme that catalyses the conversion of pregnenolone to 17-OHP in the adrenal cortex is ACTH responsive and as such concentrations are also likely to increase in dogs with hyperadrenocorticism. 17-OHP is further metabolised by 21-hydroxylase and 11-β-hydroxylase to produce 11deoxycortisol and cortisol, respectively.

Recent research has suggested that measurement of 17-OHP during the ACTH stimulation test is extremely useful particularly in dogs with hyperadrenocorticism that do not exhibit classical results using traditional tests.(22) However, because of the overlap in test results between healthy and sick dogs and those with hyperadrenocorticism, it cannot be advocated as a routine screening test for hyperadrenocorticism.(2)

Plasma adrenocorticotropic hormone concentration

Measurement of circulating ACTH concentration is a specific test to discriminate between PDH and AT but has no role to play in the diagnosis of the condition. Dogs with PDH release large amounts of ACTH but dogs with AT have reduced ACTH output. In dogs, inappropriately elevated plasma ACTH concentrations are consistent with PDH while low values are consistent with AT.(3,8,20) The measurement of plasma ACTH concentration is significantly influenced by sample handling procedures.(11) Aprotinin has a profound preservative effect upon canine plasma ACTH and it may be possible to submit unfrozen plasma samples to which aprotinin has been added.(14)

High-dose dexamethasone suppression test

A higher dose of dexamethasone (0.1 mg/kg) can suppress cortisol concentrations in dogs with PDH but not in dogs with AT.(5) Suppression of plasma cortisol concentrations below the reference range, or below 50 % of the baseline concentration at 3, 4 or 8 hours is consistent with PDH. Approximately 20 to 30 % of dogs with PDH do not exhibit cortisol suppression in response to a high dose of dexamethasone, so failure to adequately suppress cannot be interpreted as confirmation of ADH thus necessitating the use of another discriminatory test. Administration of high doses of dexamethasone to dogs with pre-existing hyperadrenocorticism could potentially increase the risk of gastrointestinal haemorrhage. Because of this and because the test neither confirms AT nor significantly increases the number of dogs diagnosed with PDH, it has largely fallen out of favour.

References

1.  Angles JM, Feldman EC, Nelson RW & Feldman MS (1997) Use of urine cortisol:creatinine ratio versus adrenocorticotropic hormone stimulation testing for monitoring mitotane treatment of pituitary-dependent hyperadrenocorticism in dogs. Journal of the American Veterinary Medical Association 211: 10021004.

2.  Chapman, P.S., Mooney, C.T., Ede, J., Evans, H., O'Connor, J., Pfeiffer, D.U. and Neiger, R. Evaluation of basal and post-ACTH serum 17hydroxyprogesterone concentrations for the diagnosis of canine hyperadrenocorticism. Veterinary Record (Accepted for publication).

3.  Feldman EC (1983) Comparison of ACTH response and dexamethasone suppression as screening tests in canine hyperadrenocorticism. Journal of the American Veterinary Medical Association 182: 506-510.

4.  Feldman EC & Mack RE (1992) Urine cortisol:creatinine ratio as a screening test for hyperadrenocorticism in dogs. Journal of the American Veterinary Medical Association 200: 16371641.

5.  Feldman EC, Nelson RW & Feldman MS (1996) Use of low-and high-dose dexamethasone tests for distinguishing pituitary-dependent from adrenal tumour hyperadrenocorticism in dogs. Journal of the American Veterinary Medical Association 209: 772775.

6.  Feldman EC, Stabenfeldt GH, Farver TB & Addiego LA (1982) Comparison of aqueous porcine ACTH with synthetic ACTH in adrenal stimulation tests of the female dog. American Journal of Veterinary Research 43: 522-524.

7.  Frank LA & Oliver JW (1998) Comparison of serum cortisol concentrations in clinically normal dogs after administration of freshly reconstituted versus reconstituted and frozen cosyntropin. Journal of the American Veterinary Medical Association 212: 15691571.

8.  Gould SM, Baines EA, Mannion PA, Evans H & Herrtage ME (2001) Use of endogenous ACTH concentration and adrenal ultrasonography to distinguish the cause of canine hyperadrenocorticism. Journal of Small Animal Practice 42: 113-121.

9.  Guptill L, Scott-Moncrieff JC, Bottoms G, Glickman L, Johnson M, Glickman N, Nelson R & Bertoy E (1997) use of the urine cortisol:creatinine ratio to monitor treatment response in dogs with pituitarydependent hyperadrenocorticism. Journal of the American Veterinary Medical Association 210: 11581161.

10. Hansen BL, Kemppainen RJ & MacDonald JM (1994) Synthetic ACTH (cosyntropin) stimulation tests in normal dogs: comparison of intravenous and intramuscular administration. Journal of the AmericanAnimalHospital Association 30: 38-41.

11. Hegstad RL, Johnston SD & Pasternak DM (1990) Effects of sample handling on adrenocorticotropin concentration measured in canine plasma, using a commercially available radioimmunoassay kit. American Journal of Veterinary Research 51: 19411947.

12. Jensen AL, Iversen L, Hoier R & Petersen TK (1997) Evaluation of the urinary cortisol: creatinine ratio in the diagnosis of hyperadrenocorticism in dogs. Journal of Small animal Practice 38: 99-102.

13. Kaplan AJ, Peterson ME & Kemppainen RJ (1995) Effects of disease on the results of the diagnostic tests for use in detecting hyperadrenocorticism in dogs. Journal of the American Veterinary Medical Association 207: 445-451.

14. Kemppainen RJ, Clark TP & Peterson ME (1994) Preservative effect of aprotinin on canine plasma immunoreactive adrenocorticotropin concentrations. Domestic Animal Endocrinology 11: 355-362.

15. Kerl ME, Peterson ME, Wallace MS, Melian C & Kemppainen RJ (1999) Evaluation of a low-dose synthetic adrenocorticotropic hormone stimulation testing clinically normal dogs and dogs with naturally developing hyperdrenocorticism. Journal of the American Veterinary Medical Association 180: 542-544.

16. Mack RE & Feldman EC (1990) Comparison of two low-dose dexamethasone suppression protocols as screening and discrimination tests in dogs with hyperadrenocorticism. Journal of the American Veterinary Medical Association 197: 1603-1606.

17. Norman EJ, Thompson H & Mooney CT (1999) Dynamic adrenal function testing in eight dogs with hyperadrenocorticism associated with adrenocortical neoplasia. Veterinary Record 144: 551-554.

18. Peterson ME, Gilbertson SR & Drucker WD (1982) Plasma cortisol response to exogenous ACTH in 22 dogs with hyperadrenocorticism caused by adrenocortical neoplasia. Journal of the American Veterinary Medical Association 180: 542-544.

19. Randolph JF, Toomey J, Center SA, Scarlett JM, Reimers T, Graham P & Nachreiner RF (1998) Use of the urine cortisol-to-creatinine ratio for monitoring dogs with pituitary-dependent hyperadrenocorticism during induction treatment with mitotane (o,p'-DDD). American Journal of Veterinary Research 59, 258-261.

20. Reusch CE & Feldman EC (1991) Canine hyperadrenocorticism due to adrenocortical neoplasia. Journal of Veterinary Internal Medicine 5: 3-10.

21. Rijnberk A, Van Wees A & Mol JA (1988) Assessment of two tests for the diagnosis of canine hyperadrenocorticism. Veterinary Record 122: 178180.

22. Ristic JME, Ramsey IK, Heath FM, Evans HJ & Herrtage ME (2002) The use of 17-hydroxyprogesterone in the diagnosis of canine hyperadrenocorticism. Journal of Veterinary Internal Medicine 16: 433-439.

23. Smiley LE & Peterson ME (1993) Evaluation of a urine cortisol: creatinine ratio as a screening test for hyperadrenocorticism in dogs. Journal of Veterinary Internal Medicine 7: 163-168.

24. Stolp R, Rijnberk A, Meijer JC & Croughs RJM (1983) Urinary corticoids in the diagnosis of canine hyperadrenocorticism. Research in Veterinary Science 34: 141-144.

25. Van Liew CH, Greco DS & Mowafak DS (1997) Comparison of results of adrenocorticotropic hormone stimulation and low-dose dexamethasone suppression tests with necropsy findings in dogs: 81 cases. Journal of the American Veterinary Medical Association 211: 322-325.

26. Van Vonderen IK, Kooistra HS & Rijnberk A (1998) Influence of veterinary care on the urinary corticoid:creatinine ratio in dogs. Journal of Veterinary Internal Medicine 12: 431-435.

27. Watson ADJ, Church DB, Emslie DR & Foster SF (1998) Plasma cortisol response to three corticotrophic preparations in normal dogs. Australian Veterinary Journal 76: 255-257.

Speaker Information
(click the speaker's name to view other papers and abstracts submitted by this speaker)

Carmel T. Mooney, MVB, MPhil, PhD, DECVIM-CA, MRCVS
Department of Small Animal Clinical Studies
Faculty of Veterinary Medicine
University College Dublin, Belfield
Dublin, Ireland


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