Anti-Müllerian hormone (AMH) and progesterone (P4) levels in diagnosing canine ovarian remnant syndrome | VETgirl Veterinary Continuing Education Podcasts

In this VETgirl online veterinary continuing education podcast, we review the use of combining serum anti-Müllerian hormone (AMH) and progesterone (P4) levels when diagnosing canine ovarian remnant syndrome (ORS). ORS is a condition in which ovarian tissue is left behind after a female dog has been spayed. Unfortunately, when this happens, the tissue can continue to release hormones that manifest clinically as signs of estrus. As many of our small animal clinicians have learned, suspecting ORS can be easy at times, but diagnosing ORS can prove quite challenging! The diagnostic options for ORS, including vaginal cytology, estrogen levels, various stimulation and other hormonal tests, luteinizing hormone (LH), and imaging, can be overwhelming and possess individual limitations.

So, Place et al wanted to evaluate this in a study entitled Evaluation of combined assessments of serum anti-Müllerian hormone and progesterone concentrations for the diagnosis of ovarian remnant syndrome in dogs. In this study, the investigators evaluated the efficacy of combined AMH and P4 levels when diagnosing ORS. If you are at all like this VETgirl, theriogenology class seems like a LONG time ago! If you reach way back, you may remember that AMH is secreted by Sertoli cells in the testes. However, the lack of AMH in female fetuses is what allows those ole’ Mullerian ducts to develop into various female reproductive parts, such as the uterus, portions of the vagina, and the oviducts (Josso). An important note for our discussion today is that the granulosa cells of the ovaries will then begin to secrete AMH after development is complete (Weenen). Since no other part of the body secretes AMH in a female, finding positive serum levels of AMH in a “spayed” female dog suggests the presence of functional ovarian tissue in the body. Unfortunately a negative AMH test result cannot rule out ORS, however. For instance, if the ovarian remnant primarily consists of luteal tissue, AMH may not be present in sufficient amounts to be detectable. Therefore in this study, the authors measure both AMH and P4 in spayed bitches with suspected ORS, hypothesizing that the combined testing would be superior in detection of ORS than either test alone.

602 serum samples were submitted over two years to a diagnostic laboratory for AMH testing in dogs with suspected ORS, all of which were included in the study. Residual serum from these samples was also used for P4 testing. Both tests were performed on commercially available and previously validated assays, and each assay had the option of giving positive, negative, or inconclusive results. Veterinarians who elected to participate in the present study also submitted any tissues removed surgically to the anatomic pathology laboratory for histopathology. The investigators then assessed agreement between the testing methods. It is important to note that the study was not designed to evaluate the sensitivity and specificity of these diagnostics. Of the 602 cases, there were 74 cases that did have surgical exploration performed, but only 2 of the surgical cases had negative results on both AMH and P4 testing. Such a low number made calculation of the tests’ specificity impossible, though the study was able to determine the percentage of correct identifications per test by using cases with histologically confirmed ORS.

Of the 74 dogs that had surgical exploration, 2 did not have tissue submitted, and 19 were excluded for other reasons. This leaves 53 dogs that had an AMH test, a P4 test, and surgical exploration for ORS with surgical tissue submitted. Of these 53 dogs, there were 48 with ORS confirmed via histopathology! So how did the AMH and P4 tests fare in these pups? Well first of all, there were NO dogs with histologically confirmed ORS that had negative results on both tests. That’s great! Fifty-two percent of dogs with ORS had positive results on both tests, and 35% had a positive result on one test. This means there were 35% of cases in which performing only AMH or P4 could have shown a false negative result! There were 6 dogs that had an inconclusive result on one or both tests, but again, none that were true negatives on both. Again, specificities of the tests could not be determined since only 2 dogs with negative test results had surgical exploration to rule out ORS. There were also 5 dogs that had tissue removed that was not histologically confirmed as ovarian. However all of these dogs had a positive AMH, and 3 had a positive P4. For the 48 dogs with confirmed ORS, the kappa coefficient was 0.16, but it was 0.41 if looking at the total 602 samples. Just as a quick reminder of what this means – the kappa coefficient is a statistical measure to determine the degree of agreement between these tests. In a nutshell, a value of 1 indicates complete agreement, whereas a value of 0 indicates a level of agreement comparable to what would occur by chance (McHugh). The authors explain that the reason there is better agreement of AMH and P4 from the total population compared to only dogs with confirmed ORS is because the latter does not account for all of the agreement between negative test results.

So what can we take away from this VETgirl podcast? Well, if you have a “spayed” patient that begins showing signs of heat, remember that ORS can absolutely happen, even with the best of surgeons! If you suspect ORS, combining AMH and P4 testing can be effective in making the diagnosis, and the combined approach lowers the risk of a false negative result. Recall that the production of such hormones has cyclic fluctuation, so there is risk of false negative results in testing either of these alone. While false positives can occur for P4 from exogenous sources, false positives for AMH is considered rare. Furthermore, make sure that if you take an ORS suspect to surgical explore, it is essential to submit any tissue retrieved for histopathology!

AMH: Anti-Müllerian hormone
LH: Luteinizing hormone
P4: Progesterone

1. Place N, Cheraskin J-L, Hansen B. Evaluation of combined assessments of serum anti-Müllerian hormone and progesterone concentrations for the diagnosis of ovarian remnant syndrome in dogs. J Am Vet Med Assoc 2019;254(9):1067-1072.
2. Josso N, Lamarre I, Picard JY, et al. Anti-Müllerian hormone in early human development. Early Hum Dev 1993;33:91–99.
3. Weenen C, Laven JS, Von Bergh AR, et al. Anti-Müllerian hormone expression pattern in the human ovary: potential implications for initial and cyclic follicle recruitment. Mol Hum Reprod 2004;10:77–83.
4.McHugh M. Interrater reliability: the kappa statistic. Biochem Med 2012;22(3):276–282.

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