Cryptorchidism In Horses Research Articles

Diagnostic of cryptorchidism in horses.

Cryptorchidism in horses is the most common congenital malformation, caused by a failure in the descent of testicles from the abdominal cavity into the scrotum. As a result, there is a partial or complete loss of reproductive potential although the production and secretion of steroid sex hormones and anti-Mueller hormone, as well as behavioral characteristics of the stallion, are maintained. The exact location of the undescended testis is established through a thorough clinical examination of the interior (of the cryptorchid horse), supported by an ultrasound examination of the inguinal region, and the transrectal ultrasound visualization of the bladder area and the inner ring of the inguinal canal. The results of several genetic studies suggest a genetic basis for cryptorchidism. In particularly difficult cases of suspected cryptorchidism in horses (hemi-castrated animals without a known clinical history), it is necessary to determine androgenic hormone levels and the anti-Mueller hormone level and to perform a testosterone production stimulation test with hCG induction (Cox test). The exact location of the retained testicle needs to be known before surgical removal so that optimal decisions can be made regarding the technique of the procedure and selection of appropriate anaesthesia.

The G-Protein-Coupled Membrane Estrogen Receptor Is Present in Horse Cryptorchid Testes and Mediates Downstream Pathways.

Cryptorchidism in horses is a commonly occurring malformation. The molecular basis of this pathology is not fully known. In addition, the origins of high intratesticular estrogen levels in horses remain obscure. In order to investigate the role of the G-protein-coupled membrane estrogen receptor (GPER) and establish histological and biochemical cryptorchid testis status, healthy and cryptorchid horse testes were subjected to scanning electron microscopy analysis, histochemical staining for total protein (with naphthol blue black; NBB), acid content (with toluidine blue O; TBO), and polysaccharide content (with periodic acid–Schiff; PAS). The expression of GPER was analyzed by immunohistochemistry and Western blot. GPER-mediated intracellular cAMP and calcium (Ca2+) signaling were measured immunoenzymatically or colorimetrically. Our data revealed changes in the distribution of polysaccharide content but not the protein and acid content in the cryptorchid testis. Polysaccharides seemed to be partially translocated from the interstitial compartment to the seminiferous tubule compartment. Moreover, the markedly decreased expression of GPER and GPER downstream molecules, cAMP and Ca2+, suggests their potential role in testis pathology. Increased estrogen levels in cryptorchid conditions may be linked to disturbed GPER signaling. We postulate that GPER is a prominent key player in testis development and function and may be used as a new biomarker of horse testis in health and disease.

Changes in circulating concentrations of testosterone and estrone sulfate after human chorionic gonadotropin administration and subsequent to castration of 2-year-old stallions

Reproductive steroids testosterone (T) and estrone sulfate (E1S) are used as diagnostic markers for cryptorchidism in horses. The human chorionic gonadotropin (hCG) stimulation test is used as a diagnostic aid because administration of this hormone results in greater incremental differences in circulating steroid concentrations. Thoughts regarding optimal sampling times following hCG administration, however, are inconsistent. Additionally, determination of half-life of these steroids is important in postsurgical samples to confirm complete removal of testicular tissue. Objectives of this study, therefore, were to determine optimal sampling periods for peak T and E1S after hCG administration and half-life of these steroids after castration. Eight pony stallions were randomly assigned to control or treatment groups (5000 IU hCG). Blood samples were collected following hCG administration. Subsequently, stallions were castrated and blood samples were collected post-castration. The T concentrations were greatest at 72 h after hCG and were greater (P < 0.02) in samples from hCG-treated than control animals: 9,903.4 ± 384 and 784.0 ± 192 pg/mL, respectively (Mean ± SEM). The T concentrations were also greater at 1, 12, 24, 48 and 96 h. The E1S concentrations did not change after administration of hCG. The T response to hCG administration was biphasic with a maximal response between 48–96 h after administration. Half-lives of T and E1S were 1.1 and 0.7 h, respectively, and concentration of T and E1S was similar to that of geldings at 24 h post-castration, which, therefore, should be considered an optimal time to ensure complete castration has occurred.

Serum concentrations and testicular expressions of insulin-like peptide 3 and Anti-Müllerian hormone in normal and cryptorchid male horses

Insulin-like peptide 3 (INSL3) is an important hormone for testicular descent during embryonic development and a factor for assessing functional status of Leydig cells of testes, but there is limited number of equine studies. Anti-Müllerian hormone (AMH) is a useful diagnostic marker for cryptorchidism in horses. This study aimed to compare serum concentrations and testicular expression intensity of INSL3 and AMH in intact and cryptorchid male horses. Serum INSL3 concentrations in intact (n = 9; mean ± SEM, 19.9 ± 5.9 ng/mL) and noncastrated unilateral cryptorchid (UC) male horses (n = 16; mean ± SEM, 16.8 ± 4.1 ng/mL) were higher compared with hemicastrated unilateral cryptorchid (HCUC) male horses (n = 9; mean ± SEM, 3.8 ± 0.7 ng/mL) (P < 0.05). And serum INSL3 in bilateral cryptorchid (BC) male horses (n = 4; 1.9 ± 0.4; mean ± SEM, ng/mL) were lower compared with intact male horses (P < 0.05). Serum AMH concentrations in BC male horses (n = 3; mean ± SEM, 30.6 ± 4.8 ng/mL) were higher compared with intact male horses (n = 5; mean ± SEM, 12.2 ± 3.9 ng/mL) (P < 0.05). Immunostaining of scrotal and cryptorchid testis showed that Sertoli cells were positive for AMH, and Leydig cells were positive for INSL3. Staining intensity of AMH was higher in cryptorchid testis than in scrotal testis (P < 0.05). Furthermore, AMH expression intensity was higher in abdominal testis than in inguinal testis (P < 0.05). Immunostaining intensity of INSL3 in the testis was positively correlated with serum INSL3 (r, 0.7; P < 0.01), seminiferous tubule area (r, 0.727; P < 0.01), and Johnsen score for spermatogenesis (r, 0.604; P < 0.05), whereas immunostaining intensity of AMH in the testis was negatively correlated with seminiferous tubule area (r, −0.814; P < 0.01) and Johnsen score for spermatogenesis (r, −0.807; P < 0.01). Our findings suggested that AMH is a good biomarker for diagnosing cryptorchidism in male horses, in addition to INSL3 values to assess the testis of intact and cryptorchid male horses.

Comparative Transcriptomics Analysis of Testicular miRNA from Cryptorchid and Normal Horses.

Simple SummaryThe testis is an important organ for mammals, and testicular microRNA expression is associated with male fertility to a certain extent. Cryptorchidism is the failure of one or both testes to descend into the scrotal sac. It is a common congenital malformation in horses. The major clinical consequence of this abnormality is impaired fertility. The expression of testicular microRNAs is influenced by many factors, including high temperature and disease, in cryptorchid horses. Here, we investigated the microRNA expression levels of normal and retained testes. Their expression patterns showed significant differences. In addition, we obtained comprehensive expression data for equine testicular microRNA, which is fundamental information for further analysis.In the biological process of testicular spermatogenesis, the expression and interaction of many genes are regulated by microRNAs (miRNAs). However, comparisons of miRNA expression between descended testes (DTs) and undescended testes (UDTs) are rarely done in horses. In this study, we selected two UDTs (CKY2b and GU4b) from Chakouyi (CKY) and Guanzhong (GU) horses and eight DTs (GU1–3, CKY1, CKY3, CKY2a, GU4a, and GU5). Three groups were compared to evaluate expression patterns of testicular miRNA in stallion testes. Group 1 compared normal CKY horses and GU horses (CKY1 and CKY3 vs. GU1–3). Group 2 (CKY2a and GU4a (DTs) vs. CKY2b and GU4b (UDTs)) and group 3 (GU1–3, CKY1, CKY3 (DTs) vs. CKY2b and GU4b (UDTs)) compared the expression levels in unilateral retained testes to normal testes. The results show that 42 miRNAs (7 upregulated and 35 downregulated) had significantly different expression levels in both comparisons. The expression levels of eca-miR-545, eca-miR-9084, eca-miR-449a, eca-miR-9024, eca-miR-9121, eca-miR-8908e, eca-miR-136, eca-miR-329b, eca-miR-370, and eca-miR-181b were further confirmed by quantitative real-time PCR assay. The target genes of differentially expressed miRNAs in three comparisons were predicted, and the functions were annotated. The putative target genes of the 42 co-differentially expressed miRNAs were annotated to 15 functional terms, including metal ion binding, GTPase activator activity, zinc ion binding, intracellular, cytoplasm, and cancer pathways, and osteoclast differentiation. Our data indicate that the differentially expressed miRNAs in undescended testis suggests a potential role in male fertility and a relationship with cryptorchidism in horses. The discovery of miRNAs in stallion testes might contribute to a new direction in the search for biomarkers of stallion fertility.

Diagnostische benadering van cryptorchidie bij de hengst

De diagnose van cryptorchidie bij het paard is vaak een uitdaging aangezien niet altijd definitief uitsluitsel kan gegeven worden op basis van de anamnese, het klinische onderzoek en het rectaal en echografisch onderzoek. Verschillende endocrinologische diagnostische testen zoals de bepaling van het testosteron-, androstenedione-, oestrogenen-, urinaire steroïden- en het antimülleriaans hormoongehalte, die de aanwezigheid van testiculair weefsel aantonen, zijn beschreven. In dit artikel wordt getracht om de voor- en nadelen van deze testen te vergelijken, zodat practici een idee krijgen welke testen in de praktijk gebruikt kunnen worden.

Anti-Müllerian hormone as an indicator of hemi-castrated unilateral cryptorchid horses.

ABSTRACTAnti-Müllerian hormone (AMH), a glycoprotein secreted from the fetal testis, is responsible for regression of the Müllerian duct in the male fetus. The aim of this study was to evaluate the usefulness of serum AMH as a biomarker for diagnosis of cryptorchidism in horses. Serum AMH concentrations were measured in intact stallions, hemi-castrated unilateral cryptorchid stallions, and geldings. In addition, expression of AMH was characterized in cryptorchid testes by immunohistochemistry. Serum AMH was detected in intact stallions (n=11, 13.3 ± 1.8 ng/ml) and in hemi-castrated cryptorchid stallions (n=8, 17.6 ± 3.0 ng/ml), but not in geldings (n=6, all data were below the limit of detection). Immunolabeling for AMH was detected in Sertoli cells of undescended testes from cryptorchid horses as well as those of normal testes. Our findings indicate that the cryptorchid testis after hemi-castration secretes AMH and that serum AMH concentrations may be a useful biomarker for diagnosis of equine cryptorchidism.

Surgical Treatment of 4 Horses for Cryptorchidism Caused by Failure of Regression of the Cranial Suspensory Ligament of the Testis

To report surgical management of 4 horses with cryptorchidism caused by failure of regression of the cranial suspensory ligament (CSL). Retrospective case series. Cryptorchid horses (n = 4). Horses with unilateral or bilateral cryptorchidism caused by failure of regression of the CSL were treated by removing the retained testes through a standing laparoscopic approach (2), flank laparotomy (1), or paramedian celiotomy (1). After identification of the retained testis attached to the caudal pole of the kidney by the CSL, the vascular pedicle and ductus deferens were ligated and removed. Two horses had bilateral cryptorchidism and 2 horses had unilateral cryptorchidism. Standing laparoscopic cryptorchidectomy was performed successfully in 2 horses and in 2, the retained testes were removed using a flank or paramedian celiotomy. All testes were located in the dorsal aspect of the abdomen just caudal to the kidney and had a well-developed CSL. All horses recovered successfully from surgery. Failure of regression of the CSL is an uncommon cause of cryptorchidism in horses; however, affected horses can be treated using surgical approaches that facilitate exploration of the dorsocaudal aspect of the abdomen.

Identification of cryptorchidism in horses by analysing urine samples with gas chromatography/mass spectrometry

Currently there are two common radioimmunoassay-based methods for the detection of equine cryptorchidism; one measures testosterone concentrations in peripheral blood samples taken before and after an intravenous injection of human chorionic gonadotrophin (hCG) and the other measures plasma estrone sulfate. However, each of these invasive methods has its own shortfalls and neither gives unequivocal results. In this article a highly reliable gas chromatography/mass spectrometry (GC/MS) method is described based on the analysis of urine samples for the identification of cryptorchidism in horses, some as young as 2 years old.

Clinical Diagnosis of the Cryptorchid Stallion

Cryptorchidism, or failure of testicular descent, can be a challenging clinical diagnosis, particularly in horses with an unknown castration history. Etiology of cryptorchidism is multifactorial and includes a genetic component. Diagnosis is centered on a detailed physical examination. Transrectal ultrasonography and endocrinologic assays including baseline testosterone, estrone sulfate, and human chorionic gonadotrophin stimulation are diagnostic aids. Treatment of cryptorchidism in horses is surgical removal of testicular tissue. A review of testicular descent, proposed etiologic mechanisms of cryptorchidism, as well as diagnosis and treatment options are presented.

Cryptorchidism in horses

Equine Veterinary EducationVolume 11, Issue 2 p. 77-86 Cryptorchidism in horses P. O. E. Mueller, P. O. E. Mueller The Department of Large Animal Medicine, College of Veterinary Medicine, The University of Georgia, Athens, Georgia 30602, USA.Search for more papers by this authorA. H. Parks, A. H. Parks The Department of Large Animal Medicine, College of Veterinary Medicine, The University of Georgia, Athens, Georgia 30602, USA.Search for more papers by this author P. O. E. Mueller, P. O. E. Mueller The Department of Large Animal Medicine, College of Veterinary Medicine, The University of Georgia, Athens, Georgia 30602, USA.Search for more papers by this authorA. H. Parks, A. H. Parks The Department of Large Animal Medicine, College of Veterinary Medicine, The University of Georgia, Athens, Georgia 30602, USA.Search for more papers by this author First published: 26 April 2010 https://doi.org/10.1111/j.2042-3292.1999.tb00926.xCitations: 9AboutPDF ToolsExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume11, Issue2April 1999Pages 77-86 RelatedInformation

Faecal steroid analysis for non-invasive monitoring of reproductive status in farm, wild and zoo animals

Abstract Non-invasive faecal oestrogen and progesterone metabolite evaluations are well established approaches for monitoring reproductive function in a variety of mammalian species. The route of excretion of steroid hormone metabolites varies considerably among species, and also between steroids within the same species. Steroid concentrations in faeces exhibit a similar pattern to those in plasma, but have a lag time, which depending upon the species, can be from 12 h to more than 2 days. Faecal steroid metabolites in mammals are mainly unconjugated compounds. Faecal oestrogens consist predominantly of oestrone and/or oestradiol-17α or -17β. Therefore, specific oestrogen antibodies or antibodies against total oestrogens can be used for their determination. Progesterone is metabolised to several 5α- or 5β-reduced pregnanediones and hydroxylated pregnanes prior to its faecal excretion. Therefore, relevant antibodies for their determination show considerable cross-reactivities with several pregnane metabolites, whereas specific progesterone antibodies are less suitable. Faecal oestrogen evaluations have been used as reliable indicators of pregnancy in several ungulate and some primate species. They have also been used to determine the preovulatory period in carnivores, corpus luteum activity in New World primates, and to diagnose cryptorchidism in horses. Faecal progesterone metabolite analysis has been successfully used for monitoring corpus luteum function and pregnancy, abortion, seasonality and treatment therapies in an ever expanding list of species.

Measuring fecal estrogens for the diagnosis of cryptorchidism in horses

Abstract The measurement of androgens and estrogens in blood has been widely utilized to differentiate geldings from horses with testicular tissue. Estrogens are excreted in part via feces and can be quantified by immunoassay. Therefore a diagnosis of cryptorchidism based on the determination of fecal estrogens was established. The concentration of unconjugated estrogens was measured in fecal samples of normal stallions (n=63), cryptorchid horses (n=34) and geldings (n=125). In addition the concentration of conjugated estrogens and testosterone in blood samples was measured. Mature stallions had fecal estrogen concentrations in the range of 59 to 398 nmol/kg (median: 130) whereas that of geldings ranged from 1 to 38 nmol/kg (median: 12). Stallions with one undescended testis (n=18) had concentrations of 60 to 322 nmol/kg (median: 113), which were indistinguishable from those of normal stallions. Fecal estrogen concentrations of bilateral (n=9) and hemicastrated (n=7) cryptorchid horses ranged from 34 to 222 nmol/kg (median: 64) and differed (p

Diagnostic ultrasonography for evaluation of cryptorchidism in horses

Summary The location and size of 11 retained testes were accurately determined ultrasonographically. There was 100% correlation between the location of the testis determined by ultrasound vs that determined by surgery. Testicular size determined presurgically in all cases closely approximated the actual size obtained by gross measurement of the excised testis. The cryptorchid testicular echotexture was less dense than that of the normal descended testicles, but was easily identified. Ultrasonographic evaluations were completed by use of an ultrasound base unit with attached 5-MHz transrectal transducer. Rectal scans were started at the pelvic brim and continued in a to-and-fro pattern between the midline and the lateral abdominal wall. When the testis was located, the image was froze to allow measurement. All testicular locations were ascertained ultrasonographically either by rectal or external inguinal scans.

Comparison of hormonal methods for diagnosis of cryptorchidism in horses

Thirty-eight horses, including unilateral cryptorchid (n=20), “phantom” (n=16), and intact stallions (n=2), were used to evaluate hormonal methods of diagnosis for cryptorchidism. Blood sera were obtained for determination of resting concentrations of testosterone and estrone sulphate. The horses were then treated with either human Chorionic Gonadotropin or Gonadotropin Releasing hormone or were subjected to stimulation by a marein estrus. Blood samples were collected at 24 hours before and immediately before treatment, 5 times during an hour after treatment, at 3-hour intervals until 12 hours post-treatment, and then 24 hours post-treatment to determine changes in testosterone and estrone sulphate concentrations. Exploratory inguinal surgery was performed to confirm the presence and location of testicular tissue. Resting testosterone concentration was the most reliable method for differentiating geldings from animals with testicular tissue. The geldings had a mean testosterone value of 0.12 ng/ml, while animals with testicular tissue had values ranging from means of 0.72 to 0.98 ng/ml (p <0.05). Five percent of the animals were incorrectly diagnosed using resting testosterone concentrations as the test, when animals of all ages were compared. Resting estrone sulphate concentration, however, was useful for differentiating geldings from animals with testicular tissue, especially in animals ≥ 3 years of age. When using the resting estrone sulphate test, 9% of the animals were incorrectly diagnosed when includingt animals of all ages, but when only including animals ≥ 3 years of age, the percent incorrectly diagnosed dropped down to 5%. Human Chorionic Gonadotropin administration consistently stimulated increases in hormone concentrations in intact stallions, and unilateral cryptorchids, however, the response of bilateral cryptorchid and hemicastrate stallions in terms of increases in hormone concentrations was minimal.