






































Type of the Paper (Article


 

  

 

 
Cluj Vet J 2024, 29, 3 http://clujveterinaryjournal.ro 

Article 

Analysis of gonadocorticoids profile in canine patients  
diagnosed with ovarian cysts 
Zoltán-Miklós GÁL 1, Ana Hîruța 2,*, Alexandru-Raul POP 1, †), Alexandra IRIMIE 3, Ioan Ștefan GROZA 1 

1 Faculty of Veterinary Medicine, Reproduction Department, University of Agricultural Sciences and Veteri-
nary Medicine, Cluj-Napoca, 400372, Cluj, Romania 

2 Faculty of Veterinary Medicine, Pathology Department, University of Agricultural Sciences and Veterinary 
Medicine, Cluj-Napoca, 400372, Cluj, Romania 

3 Faculty of Veterinary Medicine, Anatomy Department, University of Agricultural Sciences and Veterinary 
Medicine, Cluj-Napoca, 400372, Cluj, Romania. 

* Correspondence: ana.hiruta@usamvcluj.ro 
†) These authors contributed equally to this work. 
 

Abstract: Veterinarians can obtain a more comprehensive understanding of the progression of ovarian cystic pathology in dogs by 
assessing hormonal levels. This study aimed to examine the gonadocorticoid profile of dogs diagnosed with ovarian cysts and com-
pare these levels with those of healthy female dogs. The study included 96 female dogs split into two groups: a control group of 
healthy females and a study group of females with ovarian cysts larger than 4.5–5 mm in diameter and in diestrus or anestrus phase, 
determined by progesterone measurements. The control group had no ovarian pathologies and matched the estrus cycle phase of 
the study group. Ovarian cysts were diagnosed using the Esaote MyLab X5 ultrasound machine, while sex hormone levels (testos-
terone, progesterone, and estradiol) were measured with the Biomerieux MiniVidas analyzer. The analyzer utilizes the Enzyme 
Linked Fluorescent Assay (ELFA) technique for hormonal assessments. The study group (ovarian cysts) had significantly higher 
estradiol levels (21.83 pg/ml) compared to the control group (10.70 pg/ml). However, there was no significant difference in proges-
terone levels between the two groups. The study group females showed significantly higher estradiol levels in both the anestrus 
phase (p = 0.0105) and the diestrus phase (p = 0.0105). There was also a significant difference in estradiol levels between control group 
females in diestrus and anestrus, with significantly higher levels observed in the anestrus phase (p = 0.0105). In summary, female 
dogs with ovarian cysts had higher estradiol levels in both estrus and anestrus phases compared to the control group. The authors 
recommend assessing hormonal profiles in dogs with ovarian cysts for better treatment planning. 

Keywords: canine ovarian cysts; estradiol; progesterone; gonadocorticoids  
 

1. Introduction 
Female canines are a monoestrous, non-seasonal, polytocous species, with an aver-

age interestrus period of 6 months, divided into 4 phases. The 4 phases of the sexual cycle 
for this species are: proestrus, with a duration of approximately 7-10 days, followed by 
estrus lasting 5-10 days, diestrus lasting 20-90 days, and anestrus, with a variable duration 
of 15-150 days [1,2]. 

In canine species, ovarian cysts are clinically significant as they are a major source of 
hyperestrogenism in bitches, potentially leading to prolonged estrus [3,4]. These follicular 
cysts are endocrinologically active, secreting estradiol and progesterone. Persistent follic-
ular cysts are believed to induce hyperestrogenism in bitches with estrus extending up to 
three months [2,3]. 

The ovarian cysts are classified into the following types: follicular cysts, luteal cysts, 
cystic corpus luteum, cystic rete ovarii and cysts of the subsurface epithelium [5,6]. 

Functional ovarian cysts manifest as single or multiple fluid-filled structures of var-
ying sizes, which can be unilateral or bilateral in bitches aged 6-8 years. The pathogenesis 
is not well understood, but possible causes include insufficient LH surge, changes in gon-
adotropin receptors within the follicles, and growth factors [6]. Inadequate LH peaks to 

Received: 05.08.2024 

Accepted: 10.08.204 

Published: 12.09.2024 

DOI: 10.52331/vsa8e911 

 

 

 

Copyright: © 2024 by the authors. 

Submitted for possible open access 

publication under the terms and 

conditions of the Creative Commons 

Attribution (CC BY) license 

(http://creativecommons.org/licenses

/by/4.0/). 

https://doi.org/10.52331/vsa8e911


Cluj Vet J 2024, 29, 3 3 of 29 
 

trigger ovulation of dominant follicles or insufficient receptor responsiveness can lead to ovarian cyst for-
mation. Some researchers suggest that the inhibition of aromatase, an enzyme critical for the conversion of 
androgens to estrogens, may be an intraovarian disruption linked to the pathophysiology of polycystic ovary 
syndrome [7]. 

Clinically, follicular cysts must be differentiated from other ovarian cystic pathologies, including rete 
ovarii hyperplasia, superficial epithelial cysts, and ovarian neoplasms. Although the life expectancy of dif-
ferent dog breeds varies, there is an increased incidence of ovarian cysts in dogs over 6-8 years of age [6]. 

The primary endocrine lesion causing anovulation is the central failure to produce timely GnRH pulses 
and an LH surge, which are essential for final follicular growth and ovulation, or peripheral molecular ho-
meostatic imbalances within the follicles that impair the ovary's response to the preovulatory LH surge. The 
former hypothesis explains the development of multiple follicular cysts, while the latter is more applicable 
to the occurrence of solitary follicular cysts [5]. 

As large cysts lose their structure and the arrangement of cellular layers changes under the pressure 
exerted by the cystic fluid, identifying the origin of the cyst becomes challenging. Follicular and luteal cysts 
are the two main types of functional ovarian cysts. Additionally, non-follicular cysts often arise from surface 
epithelium, underlying cells, or the mesonephric tubules of the ovary [8]. 

In humans, the etiology of polycystic ovarian syndrome (PCOS) has a strong genetic component. Genes 
such as CAPN10, the Cytochrome P450 family, the insulin gene, AR, FTO, and FSHR have been implicated 
[9]. Similarly, in dairy cows, variations in the bovine PCOS-related DENND1A gene have also been identi-
fied [10]. 

The prolonged presence of steroid hormones in the blood, which underlie the formation of ovarian 
cysts, not only predisposes the organism to uterine pathologies but also affects the ovulation rate, leading to 
a reduction in the size of the follicles. Histopathologically, the predominant uterine changes include cystic 
endometrial hyperplasia, periglandular fibrosis, lymphoplasmacytic endometritis, and adenomyosis, with 
incidences of 19.7%, 14.5%, 4.0%, and respectively 2.6% [11]. 

By evaluating the hormonal levels, veterinarians can gain a deeper understanding of how this pathol-
ogy progresses in dogs. The aim of this research was to analyze the gonadocorticoids profile of canine pa-
tients diagnosed with ovarian cysts and compare the values with healthy canine females. 

2. Materials and Methods 

2.1. Study design 
The study included a total number of 96 canine females. The dogs were segregated into two following 

two groups in accordance with the results of the clinical and paraclinical evaluation: control group for the 
females which were clinically healthy and study group for the females which were diagnosed with ovarian 
cysts. The study group inclusion criteria required the presence of ovarian cysts with sizes bigger than 4.5–5 
mm in diameter, and the subjects needed to be in either the diestrus or anestrus phase. The sexual cycle 
phase was determined through routine progesterone measurement. For the control group, the inclusion cri-
teria were the absence of pathological ovarian structures and being in the same estrus cycle phase (diestrus 
or anestrus) as the study group. 

The diagnosis of females with ovarian cysts, as well as the collection of samples for endocrine profiling 
of these patients, was conducted at the Faculty of Veterinary Medicine in Cluj-Napoca within the Depart-
ment of Reproduction, Obstetrics, and Reproductive Pathology (small animal clinic), as well as at the spe-
cialized veterinary practice Quantas Repro Vet in Cluj-Napoca. 

The clinical history of patients suspected of having ovarian cysts primarily included infertility (repeated 
mating without successful conception), irregular heat cycles (prolonged estrus with shortened anestrus pe-
riods), pregnancy checks, routine gynecological examinations, essential vaginal hemorrhages, as well as 
other reproductive tract pathologies such as cystic endometrial hyperplasia, pyometra, mammary gland tu-
mors, or vaginal tumors. 

The imaging diagnosis of ovarian cystic formations was performed using a stationary Esaote MyLab X5 
ultrasound machine with a microconvex probe. Subsequently, the cystic structures were measured using the 
ultrasound machine’s software. In some cases, the diagnosis was confirmed through macroscopic examina-
tion of the ovaries, for the patients following ovariohysterectomy for therapeutic purposes or routine ovari-
ohysterectomy (at the owner ´s request). It is noteworthy that in these females, clinical signs were absent, 
and preoperative clinical examination revealed that the patients were clinically healthy. 



Cluj Vet J 2024, 29, 3 4 of 29 
 

2.2 Sample Collection 
Blood samples were collected from all patients for routine blood work. The blood samples were centri-

fuged at 5000 rpm for 10 minutes and the serum was stored at −18° C until the evaluations were done. The 
Biomerieux MiniVidas analyzer was used to measure sex hormones such as testosterone, progesterone, and 
estradiol. This automatic analyzer uses the Enzyme Linked Fluorescent Assay (ELFA) technique for hormo-
nal assays. 

All owners provided informed written consent, granting permission for sample collection and its use 
in research, as well as for the anonymous publication of the results. 

2.3. Statistical analysis 
The obtained data were examined for normality using the Shapiro-Wilk test. For the normal distribution 

results comparison, the independent samples t-test was applied. In case the normality was rejected for the 
examined data, the Mann-Whitney test was applied. Values of p < 0.05 were considered as statistically sig-
nificant. The statistical analysis was performed using MedCalc® Statistical Software version 22.032 (Med-
Calc Software Ltd, Ostend, Belgium). 

3. Results 
From the total of 96 canine females, 48 were included in the control group (healthy females) and 48 in 

the study group (ovarian cysts). Within the control group, 26 females were in the diestrus phase and 22 were 
in the anestrus phase. In the study group, 23 females were in the diestrus phase and 25 were in the anestrus 
phase. 

The values resulting from the statistical analysis are presented in Table 1. The results for the testosterone 
levels were not included since all the females had values < 0.05 ng/ml, except for one female from the study 
group which had 0.09 ng/ml. The females from the study group (ovarian cysts) had significantly higher 
estradiol values (p < 0.0001) with an arithmetic mean of 21.83 pg/ml (95% CI: 14.65–25.37), compared with 
the control group, which had a median of 10.70 pg/ml (95% CI: 9.86–12.30). For the progesterone values 
comparison between the control group (median: 7 ng/ml) and the study group (median: 2.96 ng/ml), there 
wasn’t a statistically significant difference (p = 0.6735), hence no additional comparisons involving proges-
terone were done. Estradiol values in diestrus and anestrus were further compared between and within the 
study and control groups. 

Table 1. Statistical analysis results for the estradiol and progesterone levels in the control and study groups 

Hormones Estral 
phase 

n Mean ± SD Median Min. Max. Confidence Interval 
for Mean 95% 

Estradiol 
study 
group 

Diestrus 23 23.75 ± 13.41 23.17 9 55.86 17.95–29.55 
Anestrus 25 24.79 ± 20.10 21.19 9 95.40 16.49–33.09 
Overall 48 24.29 ± 17.05 21.83 9 95.40 19.34–29.25 

Estradiol 
control 
group 

Diestrus 26 10.51 ± 2.01 9.63 8.21 15.5 9.69–11.32 
Anestrus 22 13.24 ± 3.50 12.95 8.47 21.08 11.69–14.79 
Overall 48 11.76 ± 3.08 10.7 8.21 21.08 10.86–12.66 

Progesteron 
study 
group 

Diestrus 23 36.75 ± 23.69 38.26 4.07 80.00 26.50–47.00 
Anestrus 25 1.38 ± 0.92 0.95 0.25 2.98 1.00–1.77 
Overall 48 39.6 ± 22.78 2.96 0.25 80.00 11.32–25.34 

Progesteron 
control 
group 

Diestrus 26 37.57 ± 19.76 37.19 3.08 80.00 29.59–45.56 
Anestrus 22 1.22 ± 0.77 1.05 0.21 2.57 0.88–1.56 
Overall 48 20.91 ± 23.30 7.00 0.21 80.00 14.15–27.68 

Legend: SD - standard deviation; n – number of individuals. The estradiol values are expressed pg/ml. Proges-
teron levels are expressed as ng/ml. 
 



Cluj Vet J 2024, 29, 3 5 of 29 
 

For the comparison between the estradiol values in the anestrum phase between the control group and 
the study group, it was noticed that there was a statistically significant difference. The females from the 
study group had a significantly higher (p = 0.0105) estradiol value in the anestrum phase with a median of 
21.19 pg/ml (95% CI: 13.44–26.09), compared with the control group which had a median level of 12.95 pg/ml 
(95% CI: 10.46–15.72). 

For the comparison between the estradiol values in the diestrum phase between the control group and 
the study group, it was noticed that there was a statistically significant difference. The females from the 
study group had a significantly higher (p = 0.0105) estradiol value in the diestrum phase with a median of 
23.17 pg/ml (95% CI: 11.75–28.77), compared with the control group which had a median level of 9.63 pg/ml 
(95% CI: 9.02–10.92). 

For the comparison of estradiol values between the control group females in diestrus and anestrus, it 
was noticed that there was a statistically significant difference. The females in the anestrus phase had a sig-
nificantly higher (p = 0.0105) estradiol value with a median of 12.95 pg/ml (95% CI: 10.46–15.72), compared 
with the female in the diestrus phase which had a median level of 9.63 pg/ml (95% CI: 9.02–10.92). 

For the comparison of estradiol values between the study group females (ovarian cysts) in diestrus and 
anestrus, it was noticed that there wasn’t a statistically significant difference (p = 0.8607). The females in the 
anestrus phase had a median value for estradiol of 21.19 pg/ml (95% CI: 13.44–26.09), compared with the 
females in the diestrus phase which had a median level of 23.17 pg/ml (95% CI: 11.74–28.77). 

4. Discussion 
Female dogs exhibiting estrus behavior with a proestrus or estrus period extending beyond 30 days 

should undergo ovarian imaging using ultrasonography to determine the presence of follicular cysts. Follic-
ular cysts are typically large (8-12 mm in diameter), with thin walls and containing an anechoic fluid. These 
cysts must be carefully differentiated from normal follicles, cavitary corpora lutea, and parovarian cysts. The 
consequences of untreated follicular cysts are unclear; however, they may lead in most cases to pyometra 
and neoplasms such as mammary gland tumors and vaginal tumors, in addition to that in some cases, bone 
marrow suppression may occur due to persistently elevated estrogen levels. Consequently, in most cases, 
attempts are made to induce luteinization using HCG (500 IU per bitch) administered three times. In cases 
unresponsive to treatment, progesterone administration was attempted to be used to achieve cyst regression 
(though progesterone increases the risk of pyometra in a uterus influenced by estrogen) or an ovariectomy 
may be performed [12]. 

In examining the prevalence of cystic structures among 400 specimens, one study found a diverse dis-
tribution of cyst types. Follicular cysts were identified in 41 specimens (10.3%), corpus luteum cysts were 
present in 9 specimens, representing 2.3%, more than 360 specimens, or over 90%, had rete ovarii cysts and 
cysts of subsurface epithelial structures were found in 20 specimens, comprising 5.0% of the total. [13] 

The endocrine potential of each ovarian cyst can be determined by analyzing the concentrations of es-
tradiol-17ß and progesterone in the cystic fluid. In a study, the levels of estradiol-17ß in cystic fluid ranged 
from 2.0 to 568,000.0 pg/ml (median 545.0 pg/ml), while progesterone concentrations ranged from 0.1 to 
20,138.0 ng/ml (mean 31.0 ng/ml). Additionally, hormone levels vary among different types of cysts present 
on the same ovary [14]. 

A study involving a retrospective examination of ovaries removed during ovariohysterectomy per-
formed for the treatment of pyometra over a 12-month period revealed that 20.7% (17/82) of bitches had 
follicular cysts. The presence of ovarian cysts showed no significant association with cystic endometrial hy-
perplasia in Fisher's exact test (P = .78). Approximately 35.3% of bitches with ovarian cysts also had pyom-
etra, while 58.8% had pyometra without cystic endometrial changes. Only 5.9% of females developed ovar-
ian cysts without uterine pathology [15]. Although hyperestrogenism is less common in bitches with follic-
ular cysts, the development of the cystic endometrial hyperplasia-pyometra complex is frequent, leading to 
polyuria and polydipsia, mucoid or purulent vaginal discharge, vomiting, abdominal distension, abdominal 
stress, or pain [16]. Clinical signs associated with hyperestrogenism syndrome include pancytopenia, ane-
mia, granulocytopenia, thrombocytopenia, or hemorrhagic vulvar discharge. In chronic cases, females may 
develop typical bilateral symmetrical alopecia, lichenification, hyperkeratosis, and bone marrow suppres-
sion [17]. 



Cluj Vet J 2024, 29, 3 6 of 29 
 

In females, testosterone is primarily produced by the ovaries and the adrenal glands. However, as this 
study suggests, the ovaries do not produce significant amounts of testosterone in females with ovarian cysts 
or in healthy females.  

The pathogenesis, diagnosis, and treatment of cystic ovarian diseases in bitches remain unclear. There-
fore, it is crucial to establish an early diagnosis and implement appropriate treatment strategies promptly to 
prevent disease progression [6]. 

5. Conclusions 
In conclusion, estradiol levels were significantly elevated in female dogs with ovarian cysts during 

both the diestrus and anestrus phases compared to the control group. In female dogs without ovarian cysts, 
estradiol levels were notably higher during the anestrus phase than in the diestrus phase, likely due to in-
creased ovarian responsiveness to gonadotrophins in late anestrus. 

The ovaries do not typically produce significant amounts of testosterone in females with ovarian cysts 
or in healthy females. 

The presence of ovarian cysts does not alter significantly serum progesterone levels during either the 
diestrus or anestrus phases. Progesterone mainly regulates the estrous cycle in this species and is a reliable 
marker for identifying the estrous period in female dogs, even when cystic formations are present. This is 
corroborated by the observation that active corpora lutea can coexist with cysts in canines, particularly dur-
ing the diestrus phase. The authors recommend hormonal profile determination in canine patients with 
ovarian cysts, for a better understanding regarding the evolution of this pathology and for a more accurate 
treatment plan. This proactive approach not only aims to improve the health and well-being of the affected 
dogs but also enhances the overall outcomes of treatment strategies. 

 
Author Contributions: Conceptualization, Z.-M.G. and I.Ș.G.; methodology, A.-R.P., A.H. and A.I.; statistical analysis, 
Z.-M.G.; resources, A.-R.P.; data curation, A.H.; writing—original draft preparation, Z.-M.G., A.H. and A.I.; writing—
review and editing, A.I.; supervision, I.Ș.G. All authors have read and agreed to the published version of the manuscript. 

Funding: This research was founded by the Discipline of Animal Reproduction at the Faculty of Veterinary Medicine 
Cluj-Napoca.  

Acknowledgments: The authors extend their gratitude to the Discipline of Animal Reproduction at the Faculty of Vet-
erinary Medicine Cluj-Napoca, and the private practice reproduction referral clinic Quantas Repro Vet SRL in Cluj-
Napoca for their help in collecting the samples. 

Conflicts of Interest: The authors declare no conflict of interest. 

References 

1.  Concannon, P.W. Reproductive Cycles of the Domestic Bitch. Animal Reproduction Science 2011, 124, 200–210, 

doi:10.1016/j.anireprosci.2010.08.028. 

2.  Groza, I.Ş.; Bogdan, L.M.; Cătană, R. Ginecologie, andrologie şi obstetrică veterinară: compendiu; Editura Academiei Române: 

Bucureşti, 2006; ISBN 978-973-27-1445-4. 

3.  Johnston, S.D.; Kustritz, M.V.R.; Olson, P.N.S. Canine and Feline Theriogenology; W.B. Saunders: Philadelphia London New York, 

2001; ISBN 978-0-7216-5607-6. 

4.  Arlt, S.; Spankowsky, S.; Heuwieser, W. Follicular Cysts and Prolonged Oestrus in a Female Dog after Administration of a 

Deslorelin Implant. New Zealand Veterinary Journal 2011, 59, 87–91, doi:10.1080/00480169.2011.552858. 

5.  Knauf, Y.; Köhler, K.; Knauf, S.; Wehrend, A. Histological Classification of Canine Ovarian Cyst Types with Reference to 

Medical History. J Vet Sci 2018, 19, 725, doi:10.4142/jvs.2018.19.6.725. 

6.  Sasidharan, J.K.; Patra, M.K.; Singh, L.K.; Saxena, A.C.; De., U.K.; Singh, V.; Mathesh, K.; Kumar, H.; Krishnaswamy, N. 

Ovarian Cysts in the Bitch: An Update. Topics in Companion Animal Medicine 2021, 43, 100511, doi:10.1016/j.tcam.2021.100511. 

7.  Corbin, C.J.; Trant, J.M.; Walters, K.W.; Conley, A.J. Changes in Testosterone Metabolism Associated with the Evolution of 

Placental and Gonadal Isozymes of Porcine Aromatase Cytochrome P4501. Endocrinology 1999, 140, 5202–5210, 

doi:10.1210/endo.140.11.7140. 



Cluj Vet J 2024, 29, 3 7 of 29 
 

8.  Payan-Carreira, R.; Pires, M.A. Ovarian Cysts in Dogs´ Practice. In Advances in Medicine and Biology; Nova Science Publishers 

Inc, 2016; Vol. 94, pp. 65–88. 

9.  Ajmal, N.; Khan, S.Z.; Shaikh, R. Polycystic Ovary Syndrome (PCOS) and Genetic Predisposition: A Review Article. European 

Journal of Obstetrics & Gynecology and Reproductive Biology: X 2019, 3, 100060, doi:10.1016/j.eurox.2019.100060. 

10.  Zheng, J.; Deng, T.; Jiang, E.; Li, J.; Wijayanti, D.; Wang, Y.; Ding, X.; Lan, X. Genetic Variations of Bovine PCOS-Related 

DENND1A Gene Identified in GWAS Significantly Affect Female Reproductive Traits. Gene 2021, 802, 145867, 

doi:10.1016/j.gene.2021.145867. 

11.  Maya-Pulgarin, D.; Gonzalez-Dominguez, M.S.; Aranzazu-Taborda, D.; Mendoza, N.; Maldonado-Estrada, J.G. 

Histopathologic Findings in Uteri and Ovaries Collected from Clinically Healthy Dogs at Elective Ovariohysterectomy: A 

Cross-Sectional Study. J Vet Sci 2017, 18, 407, doi:10.4142/jvs.2017.18.3.407. 

12.  BSAVA Manual of Canine and Feline Reproduction and Neonatology; England, G.C.W., Heimendahl, A. von, British Small Animal 

Veterinary Association, Eds.; 2nd ed.; British Small Animal Veterinary Association: Quedgeley, Gloucester [England], 2010; 

ISBN 978-1-905319-19-0. 

13.  Arlt, S.; Haimerl, P. Cystic Ovaries and Ovarian Neoplasia in the Female Dog – a Systematic Review. Reprod Domestic Animals 

2016, 51, 3–11, doi:10.1111/rda.12781. 

14.  Knauf, Y.; Bostedt, H.; Failing, K.; Knauf, S.; Wehrend, A. Gross Pathology and Endocrinology of Ovarian Cysts in Bitches. 

Reprod Domestic Animals 2014, 49, 463–468, doi:10.1111/rda.12311. 

15.  De Bosschere, H.; Ducatelle, R.; Vermeirsch, H.; Van Den Broeck, W.; Coryn, M. Cystic Endometrial Hyperplasia- Pyometra 

Complex in the Bitch: Should the Two Entities Be Disconnected? Theriogenology 2001, 55, 1509–1519, doi:10.1016/S0093-

691X(01)00498-8. 

16.  Ortega-Pacheco, A.; Gutiérrez-Blanco, E.; Jiménez-Coello, M. Common Lesions in the Female Reproductive Tract of Dogs and 

Cats. Veterinary Clinics of North America: Small Animal Practice 2012, 42, 547–559, doi:10.1016/j.cvsm.2012.01.011. 

17.  Schlafer, D.H.; Miller, R.B. Pathology of the Ovary (Nondevelopmental Lesions). In Jubb, Kennedy, and Palmer’s Pathology of 

Domestic Animals; Elsevier Saunders: Philadelphia, 2007; Vol. 3, pp. 431–563. 
 
 
  


