Nonsyndromic Cryptorchidism Research Articles

The Inhibin-B Level at Orchidopexy and Follow-up of 280 Boys With Non-syndromic Unilateral Cryptorchid Testes

PurposeAn impaired germ cell number per tubular cross-section (G/T) at orchidopexy indicates a high risk of infertility. A recent study in boys with bilateral cryptorchidism showed a very high predictive value for a low serum inhibin-B level, indicating bilateral impaired G/T. Several other studies have shown a fairly strong correlation between inhibin-B and G/T. We aimed to evaluate if inhibin-B levels at orchidopexy improved at follow-up in boys with unilateral cryptorchidism. MethodsWe included 280 boys with unilateral non-syndromic cryptorchidism at the median age of 1 year (4 months-9 years) who underwent orchidopexy. They were evaluated for serum FSH, LH and inhibin-B levels at surgery and at follow-up (median 16 months later), including multiple of the median (MoM) estimations of inhibin-B due to the age dependency of normal levels. ResultsThe inhibin-B MoM score improved significantly at follow-up. At orchidopexy, 59 (21%) boys had inhibin-B levels below the normal 2.5-percentile indicating impaired G/T bilaterally. At follow-up, 36% of the boys still had low inhibin-B. At orchidopexy, 221 (79%) boys had inhibin-B levels above normal 2.5-percentile and only 5% had low inhibin-B levels at follow-up. The risk of low inhibin-B levels at follow-up was significantly different between the two groups (p < 0.0001). At follow-up, totally, 32 (11%) boys had low inhibin-B levels, hereof only 3 patients with increased FSH. ConclusionsOrchidopexy benefits the fertility potential. About 10% of boys with unilateral non-syndromic cryptorchidism may have a bilateral testicular disease reducing their fertility potential. Insufficient gonadotropin stimulation may possibly be the cause.

The safety of neoadjuvant hormonal treatment in infants with cryptorchidism

The standard treatment for boys with non-syndromic cryptorchidism is an early orchidopexy. It is unclear if surgical intervention alone is enough for future fertility. Recent studies show benefit of neoadjuvant or adjuvant hormonal treatment with gonadorelin (GnRH) for spermatogonia maturation based on testicular biopsy. The aim of this prospective study was to assess the safety of this treatment in infants with undescended testis at the recommended timing of early gonadorelin administration and timing of orchidopexy. Unilateral cryptorchid full term boys were initially examined (including hormonal, physical and ultrasound examination) at the age of 2.5-3.5 months. At 6 months of age, cryptorchidism was confirmed. Those with non-syndromic cryptorchidism and palpable or sonographically detected testis were randomly assigned into two groups: with and without intranasal gonadorelin treatment. Inclusion criteria were met by 36 boys (21 in GNRH and 15 in the control groups). The following orchidopexy was performed before 12 months of age with repeated examination at time of surgery. Penile size and testicular volume (using ultrasound) and basal serum levels of LH, FSH, testosterone, Inhibin B and AMH were recorded at age of 3.0 (mean) months and 11.0 (mean) months (date of surgery). The stimulation hormonal levels were checked during GnRH administration. Between minipuberty (mean 3 months) and time of orchidopexy (mean 11 months of age) the penile size increased significantly and similarly in both groups. There was no significant difference in the change of the volume of descended testis between the groups nor of the volume of undescended testis. In addition, we did not find any significant difference in the change (drop) of hormonal levels of LH, FSH, Testosterone, Inhibin B and AMH (Table 1a) CONCLUSION: The neoadjuvant gonadorelin stimulation in infants with unilateral undescended testis has not shown any specific effect on the development of penile size, testicular volume and hormonal levels at time of orchidopexy in comparison with boys without stimulation, and in the mid-term, this treatment can be considered safe. Further follow-up is necessary to evaluate the long-term effect of this early treatment.

Re: Fertility Potential Is Compromised in 20% to 25% of Boys with Nonsyndromic Cryptorchidism Despite Orchidopexy Within the First Year of Life

Re: Fertility Potential Is Compromised in 20% to 25% of Boys with Nonsyndromic Cryptorchidism Despite Orchidopexy Within the First Year of Life

The Majority of Boys Having Orchidopexy for Congenital Nonsyndromic Cryptorchidism during Minipuberty Exhibited Normal Reproductive Hormonal Profiles

The Majority of Boys Having Orchidopexy for Congenital Nonsyndromic Cryptorchidism during Minipuberty Exhibited Normal Reproductive Hormonal Profiles

The Majority of Boys Having Orchidopexy for Congenital Nonsyndromic Cryptorchidism during Minipuberty Exhibited Normal Reproductive Hormonal Profiles

The activation of the hypothalamic-pituitary-gonadal axis that occurs in male minipuberty during the first 5 months of life is important for early germ cell development. Orchidopexy during minipuberty may improve fertility potential as the germinative epithelium may benefit from the naturally occurring gonadotropin stimulation. We hypothesize that most boys with congenital nonsyndromic cryptorchidism display normal reproductive hormonal profiles and histological findings during minipuberty. We included boys with congenital nonsyndromic cryptorchidism who underwent orchidopexy at less than 160 days of age, having no potential for spontaneous resolution clinically. At surgery, testicular biopsies and reproductive hormones were collected and compared with normal reference values. We measured the germ cells (G/T) and type A dark spermatogonia number per tubule. Thirty-five boys aged 37 to 159 (median age: 124) days at orchidopexy were included, five were bilateral. G/T was below the normal lower range in 26% (9/35) of the cases. In six of these cases, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were above 97.5 percentile, whereas one case had FSH below 2.5 percentile. Totally, 97% (33/34, one FSH was missing) exhibited a normal LH/FSH ratio. LH was more often above 97.5 percentile than FSH: 34% (12/35) versus 3% (1/34, p < 0.001). Inhibin B was below 2.5 percentile in 17% (6/35) of cases who all proved FSH above normal mean and four had LH above normal mean. Generally, reproductive hormonal profiles of the cryptorchid boys exhibited normal minipubertal pattern. Thus, 26% of the boys had reduced germ cell number, and transient hypogonadotropic hypogonadism could be suspected in few cases.

Analysis of the CAG tract length in the Androgen Receptor gene in Mexican patients with nonsyndromic cryptorchidism

Abstract Objectives Cryptorchidism is the most common genitourinary birth defect in live newborn males and is considered as an important risk factor for testicular germ cell tumors and infertility. The Androgen Receptor gene is important in this pathology due to its participation, mainly, in the inguinoscrotal phase of testicular descent. We determine the length of the CAG tract in the Androgen Receptor (AR) gene in Mexican patients with nonsyndromic cryptorchidism. Methods One hundred and 15 males were included; of these, 62 had nonsyndromic cryptorchidism and 53 were healthy volunteers. DNA was extracted from a peripheral blood samples, subsequently, the CAG tract in exon 1 of AR gene was amplified by PCR and sequenced. Results Mexican patients with nonsyndromic cryptorchidism presented 25.03 ± 2.58 repeats of CAG tract in the AR gene compared to 22.72 ± 3.17 repeats of CAG tract in Mexican healthy individuals (p≤0.0001; t value of 4.3). Furthermore, the deletion of codon 57 that corresponds to the deletion of a leucine residue at position 57 (Del L57) in the AR gene was found for the first time in a nonsyndromic cryptorchidism patient. This molecular alteration has been related previously to testicular germ cell tumor (TGCT). Conclusions The CAG tract in the AR gene is longer in patients with nonsyndromic cryptorchidism than in healthy individuals, supporting the association between this polymorphism of the AR gene and nonsyndromic cryptorchidism in the Mexican population.

The impact of early and successful orchidopexy on hormonal follow-up for 208 boys with bilateral non-syndromic cryptorchidism.

Inhibin-B is produced by Sertoli cells and decreased values might be associated with impaired fertility potential. The aim of the study was to evaluate the impact of bilateral orchidopexy on serum inhibin-B and follicle-stimulating hormone (FSH). A cohort study including 208 bilateral cryptorchid boys (median age: 1.7year) was evaluated with serum inhibin-B and FSH in relation to histological parameters. Based on the fertility potential, the boys were divided into three subgroups. At follow-up (median age: 2.7years) the boys were evaluated with FSH and in case of inhibin-B using multiple of the median (MoM). Inhibin-B MoM improved significantly at follow-up. In 32 boys with high FSH at orchidopexy 63% normalized FSH and 59% increased MoM inhibin-B, but 31% had impaired inhibin-B at follow-up. In 105 boys with transient hypogonadotropic hypogonadism, 52% increased inhibin-B MoM but 31% had impaired inhibin-B at follow-up. In 71 boys with normal FSH, inhibin-B, and G/T, 54% increased inhibin-B MoM and 15% had impaired inhibin-B at follow-up. The effect of the surgery was best in patients younger than 1year. Orchidopexy, especially before 1year of age, improves the fertility potential in bilateral cryptorchidism. At follow-up, 26% (54/208) had a risk of infertility based on inhibin-B.

Parental Acceptance Rate of Testicular Tissue Cryopreservation in Danish Boys with Cryptorchidism

Despite orchidopexy within the first year of life, 20-25% of boys with nonsyndromic cryptorchidism may risk infertility according to histological and hormonal data obtained during surgery. The aim of this study was to evaluate the acceptance rate of testicular tissue cryopreservation among parents of prepubertal boys with cryptorchidism. Fourteen boys with cryptorchidism and high infertility risk were offered cryopreservation as an additional procedure after orchidopexy based on abnormal histopathological findings at primary surgery, whereas 27 boys with bilateral cryptorchidism were offered cryopreservation at the initial orchidopexy. A total of 90% of parents (37/41, 13/14, and 24/27) gave consent to perform cryopreservation, despite being well-informed that the procedural efficacy is largely unproven and may only be needed in about 20% of cases. The number of germ cells per tubule cross-section was 0.03-1.70 (median 0.37) and 22 boys (54%, 22/41) had a value below the lower range. Twelve boys (29%, 12/41) had no type A dark spermatogonia in their biopsy. Cryopreservation of testicular tissue is the first step to introduce spermatogonial stem cell-based therapy into clinical male infertility treatment. At the time of orchidopexy, a testicular biopsy can be collected to ascertain the infertility risk, and it may be an option for boys with bilateral cryptorchidism to have spermatogonial stem cells frozen as a fertility reserve.

Fertility Potential is Compromised in 20% to 25% of Boys with Nonsyndromic Cryptorchidism despite Orchiopexy within the First Year of Life

One of the concerns surrounding cryptorchidism is the risk of impaired fertility. Current guidelines recommend orchiopexy at age 6 to 12 months to optimize fertility outcome. We evaluated the fertility potential of boys with nonsyndromic cryptorchidism who underwent orchiopexy within the recommended age range to clarify the need for eventual supplemental treatment modalities. We retrospectively evaluated mini-puberty hormones (follicle-stimulating hormone, luteinizing hormone and inhibin B) and testicular biopsies from boys with cryptorchidism who underwent orchiopexy within the first year of life between 2010 and 2019. We histologically analyzed germ cell number and type A dark spermatogonia number per seminiferous tubule cross-section in relation to normal values. Of the 333 boys with nonsyndromic cryptorchidism 83 (25%, 21% with bilateral cryptorchidism) had a reduced number of germ cells. A total of 70 boys (21%) had low serum inhibin B, of whom 32 (46%) had a decreased number of germ cells and 23 (33%) had a decreased number of type A dark spermatogonia (p <0.01). Overall, 75 boys (23%) had no type A dark spermatogonia present. Despite early and successful orchiopexy, 20% to 25% of boys with cryptorchidism may be at risk for infertility based on hormonal and histological data. Blood test and testicular biopsy are mandatory to identify boys at high risk for infertility, in whom additional treatment modalities and followup may be needed.

Novel combined insulin-like 3 variations of a single nucleotide in cryptorchidism.

Background Insulin-like 3 hormone (INSL3) is involved in the process of testicular descent, and has been thoroughly studied in cryptorchidism. However, INSL3 allelic variations found in the human genome were heterozygous and only a few of them were found exclusively in patients with cryptorchidism. Under this perspective, we aimed to study the presence of INSL3 allelic variations in a cohort of patients with cryptorchidism and to estimate their potential consequences. Methods Blood samples were collected from 46 male patients with non-syndromic cryptorchidism and from 43 age-matched controls. DNA extraction and polymerase chain reaction (PCR) were performed for exons 1 and 2 of the INSL3 gene in all subjects. Sequencing analysis was carried out on the PCR products. All data were grouped according to testicular location. Results Seven variations of a single nucleotide (SNVs) were identified both in patients with cryptorchidism and in controls: rs2286663 (c.27G > A), rs1047233 (c.126A > G) and rs6523 (c.178A > G) at exon 1, rs74531687 (c.191-30C > T) at the intron, rs121912556 (c.305G > A) at exon 2 and rs17750642 (c.*101C > A) and rs1003887 (c.*263G > A) at the untranslated region (UTR). The allelic variants rs74531687 and rs121912556 were found for the first time in the Greek population. The novel homozygotic combination of the three allelic variants rs1047233-rs6523-rs1003887 seemed to present a stronger correlation with more severe forms of cryptorchidism. Conclusions The combination of specific INSL3 SNVs rather than the existence of each one of them alone may offer a new insight into the involvement of allelic variants in phenotypic variability and severity.

Postnatal germ cell development during first 18 months of life in testes from boys with non-syndromic cryptorchidism and complete or partial androgen insensitivity syndrome

IntroductionNeonatal testicular germ cells/gonocytes, transform into stem cells for spermatogenesis during ‘minipuberty’, driving change in timing of surgery. This study examined gonocyte transformation in cryptorchid testes in children ≤18 months of age with unilateral, bilateral undescended testes (UDT), complete or partial androgen insensitivity syndrome (CAIS, PAIS) [3,4]. Material and methodsTesticular biopsies were taken from patients with unilateral or bilateral UDT, PAIS or CAIS, aged 10 days–18 months. These testicular sections underwent immunohistochemistry with antibodies (Oct4, Ki67, C-Kit, Sox9) followed by confocal imaging, cell counting and statistical analysis. ResultsBoth Sertoli cells/tubule and germ cells (GC)/tubule decreased with age, and % empty tubules (no GC) increased with age but with no significant differences between patient groups. Oct4+ germ cells/tubule decreased with age. There are some GCs and Sertoli cells proliferating during the first year and most proliferating Oct4+ germ cells (Oct4+/Ki67+) were located off tubular basement membrane. ConclusionOur study showed that Oct4 expression gradually decreased after minipuberty and transformation into spermatogonia. Germ cells and Sertoli cells undergo mitosis during the first 12 months although not abundantly. We propose that Oct4+ gonocyte transformation into spermatogonia via proliferation and migration to the basement membrane may be delayed in UDT.

Perspective: A Neuro-Hormonal Systems Approach to Understanding the Complexity of Cryptorchidism Susceptibility.

Nonsyndromic cryptorchidism is a common multifactorial, condition with long-term risks of subfertility and testicular cancer. Revealing the causes of cryptorchidism will likely improve prediction and prevention of adverse outcomes. Herein we provide our current perspective of cryptorchidism complexity in a synthesis of cumulative clinical and translational data generated by ourselves and others. From our recent comparison of genome-wide association study (GWAS) data of cryptorchidism with or without testicular germ cell tumor, we identified RBFOX family genes as candidate susceptibility loci. Notably, RBFOX proteins regulate production of calcitonin gene-related peptide (CGRP), a sensory neuropeptide linked to testicular descent in animal models. We also re-analyzed existing fetal testis transcriptome data from a rat model of inherited cryptorchidism (the LE/orl strain) for enrichment of Leydig cell progenitor genes. The majority are coordinately downregulated, consistent with known reduced testicular testosterone levels in the LE/orl fetus, and similarly suppressed in the gubernaculum. Using qRT-PCR, we found dysregulation of dorsal root ganglia (DRG) sensory transcripts ipsilateral to undescended testes. These data suggest that LE/orl cryptorchidism is associated with altered signaling in possibly related cell types in the testis and gubernaculum as well as DRG. Complementary rat and human studies thus lead us to propose a multi-level, integrated neuro-hormonal model of testicular descent. Variants in genes encoding RBFOX family proteins and/or their transcriptional targets combined with environmental exposures may disrupt this complex pathway to enhance cryptorchidism susceptibility. We believe that a systems approach is necessary to provide further insight into the causes and consequences of cryptorchidism.

Postnatal Germ Cell Development in the Cryptorchid Testis: The Key to Explain Why Early Surgery Decreases the Risk of Malignancy.

Cryptorchidism is a risk factor for testicular malignancy and surgical treatment lowers this risk. This study aimed to investigate the germ cell behavior in prepubertal cryptorchid testes using immunohistochemical markers for germ cell malignancy to understand how early orchiopexy may possibly prevent cancer developing. Histology sections from 1,521 consecutive testicular biopsies from 1,134 boys aged 1 month to 16.5 years operated for cryptorchidism were incubated with antibodies including antiplacental-like alkaline phosphatase (PLAP), anti-Oct3/4, anti-C-kit, and anti-D2-40. Oct3/4 and D2-40-positive germ cells are found throughout the first 2 years of life, with declining frequency thereafter. After 2 years, they should have disappeared and may indicate neoplasia. PLAP-positive cells were seen in 57 to 82% and C-kit-positive cells in 5 to 21% of cryptorchid testes between 4 and 13 years. Not until puberty did PLAP and C-kit-positive undifferentiated spermatogonial stem cells vanish. Only 0.3% of the present material had obvious prepubertal intratubular germ cell neoplasia (ITGCN) and they all had syndromic cryptorchidism. An additional three boys (0.3%) older than 2 years had weak Oct3/4 expression in undescended testes, but all cases were D2-40 negative. Prepubertal ITGCN was rare and mostly seen in syndromic cryptorchidism. In nonsyndromic cryptorchidism PLAP-positive undifferentiated spermatogonial stem cells persisted in a significant proportion of nontreated undescended testes and they will be especially sensitive to long-lasting abnormally high temperature that may be the single most important cause facilitating the accumulation of mutations during cell replication and the development of ITGCN to be prevented by orchiopexy.

Single nucleotide polymorphisms associated with nonsyndromic cryptorchidism in Mexican patients.

Cryptorchidism is a frequent genitourinary malformation considered as an important risk factor for infertility and testicular malignancy. The aetiology of cryptorchidism is multifactorial in which certain SNPs, capable of inhibiting the development of the gubernaculum, are implicated. We analysed 16 SNPs by allelic discrimination and automated sequencing in 85 patients and 99 healthy people, with the objective to identify the association between these variants and isolated cryptorchidism. In two different patients with unilateral cryptorchidism, we found the variants rs121912556 and p.R105R of INSL3 gene in a heterozygous form associated with cryptorchidism, so we could considered them as risk factors for cryptorchidism. On the other hand, SNPs rs10421916 of INSL3 gene, as well as the variants rs1555633 and rs7325513 in the RXFP2 gene, and rs3779456 variant of the HOXA10 gene were statistically significant, when the patients and controls were compared and could be considered as protective factors since are predominantly present in controls. The genotype-phenotype correlation did not show statistical significance. With these results, we could conclude that these polymorphisms can be considered as important variants in our population and would contribute in the future knowledge of the aetiology and physiopathology of cryptorchidism.

Genomic copy number variation association study in Caucasian patients with nonsyndromic cryptorchidism.

BackgroundCopy number variation (CNV) is a potential contributing factor to many genetic diseases. Here we investigated the potential association of CNV with nonsyndromic cryptorchidism, the most common male congenital genitourinary defect, in a Caucasian population.MethodsGenome wide genotyping were performed in 559 cases and 1772 controls (Group 1) using Illumina HumanHap550 v1, HumanHap550 v3 or Human610-Quad platforms and in 353 cases and 1149 controls (Group 2) using the Illumina Human OmniExpress 12v1 or Human OmniExpress 12v1-1. Signal intensity data including log R ratio (LRR) and B allele frequency (BAF) for each single nucleotide polymorphism (SNP) were used for CNV detection using PennCNV software. After sample quality control, gene- and CNV-based association tests were performed using cleaned data from Group 1 (493 cases and 1586 controls) and Group 2 (307 cases and 1102 controls) using ParseCNV software. Meta-analysis was performed using gene-based test results as input to identify significant genes, and CNVs in or around significant genes were identified in CNV-based association test results. Called CNVs passing quality control and signal intensity visualization examination were considered for validation using TaqMan CNV assays and QuantStudio® 3D Digital PCR System.ResultsThe meta-analysis identified 373 genome wide significant (p < 5X10−4) genes/loci including 49 genes/loci with deletions and 324 with duplications. Among them, 17 genes with deletion and 1 gene with duplication were identified in CNV-based association results in both Group 1 and Group 2. Only 2 genes (NUCB2 and UPF2) containing deletions passed CNV quality control in both groups and signal intensity visualization examination, but laboratory validation failed to verify these deletions.ConclusionsOur data do not support that structural variation is a major cause of nonsyndromic cryptorchidism.Electronic supplementary materialThe online version of this article (doi:10.1186/s12894-016-0180-4) contains supplementary material, which is available to authorized users.

Positive Oct -3/4 and D2-40 Immunohistochemical Expression in Germ Cells and Suspected Histology Pattern of Intratubular Germ Cell Neoplasia in Boys with Cryptorchidism Vanish after the Age of 2 Years.

Introduction Intratubular germ cell neoplasia (ITGCN) is a precursor to testicular germ cell cancer. Adult germ cell cancer immunohistochemical markers may fail to detect ITGCN in prepubertal boys with congenital cryptorchidism, because positive immunohistochemistry is commonly seen in boys younger than the age of 2 years, where most orchiopexies are performed. The aim of the study was to evaluate the diagnostic challenge to differentiate between a histological pattern of ITGCN and a histological pattern with some atypical germ cells and all positive cancer immunohistochemical markers, but no increased risk of malignancy. Materials and Methods Histology sections from 373 testicular biopsies from 289 boys aged 1 month to 2 years operated for cryptorchidism were incubated with primary antibodies including anti-placental-like-alkaline phosphatase, antiOct-3/4, anti-C-kit, anti-D2-40, and in case of repeat biopsy with anti-stem cell factor (SCF) receptor. Results The prevalence of Oct-3/4 and D2-40-positive staining of germ cells in testicular biopsies were in age groups less than 6 months, 100% and 50%; 6-12 months, 60% and 17%; and 1-2 years, 12% and 4%. A 1 year, 1-month-old boy with Prader-Willi syndrome treated with growth hormone had ITGCN in both cryptorchid testes. In another three bilateral nonsyndromic cases, 8 months, 8 months and 1-year-old, a histological pattern in accordance with ITGCN was found. These three boys had a repeat biopsy from both testes performed at the age of 3 years, 4 months, 3.5 years, and 3 years, 10months, respectively. In all cases, the Oct-3/4 and D2-40 positive germ cells turned negative and the histological pattern normalized completely. The primary biopsies had SCF negative germ cells. Conclusion This study is valuable in identifying the age-related change in Oct-3/4 or D2-40 immunopositive germ cells in seminiferous tubules. An ITGCN-like histological pattern in nonsyndromic cryptorchidism will vanish after the age of 3 years. Even when immunohistochemistry is applied, prepubertal ITGCN is so rarely demonstrated in cryptorchid testes, that it is not plausible that ITGCN generally originates during fetal development in cryptorchidism.

The diagnostic impact of testicular biopsies for intratubular germ cell neoplasia in cryptorchid boys and the subsequent risk of testicular cancer in men with prepubertal surgery for syndromic or non-syndromic cryptorchidism

IntroductionCryptorchidism is a risk factor for testicular cancer in adult life. It remains unclear how prepubertal surgery for cryptorchidism impacts later development of adult testicular cancer. The aim of study was to investigate tools to identify the cryptorchid boys who later develop testicular cancer. MethodsThe study cohort consisted of 1403 men operated prepubertally/pubertally for undescended testis between 1971 and 2003. At surgery testicular biopsies were taken from the cryptorchid testes. The boys were followed for occurrence of testicular cancer. The testicular cancer risk was compared to the risk in the Danish Population. Testicular biopsies from the boys who developed testicular cancer during follow-up underwent histological examination with specific diagnostic immunohistochemical markers for germ cell neoplasia. ResultsThe cohort was followed for 33,627 person years at risk. We identified 16 cases with testicular cancer in adulthood. The standardized incidence ratio was 2.66 (95% CI: 1.52–4.32). At time of primary surgery in prepubertal/pubertal age Intratubular Germ Cell Neoplasia (ITGCN) was diagnosed in 5 cases and the boys were unilaterally orchiectomized. At follow-up new immunohistochemical staining indicated ITGCN in two of the 16 cancer cases at reevaluation of the original biopsies from time of prepubertal/pubertal surgery. One had syndromic cryptorchid and developed seminoma, and another showed nonsyndromic cryptorchidism and developed embryonic teratocarcinoma. Totally, ITGCN was diagnosed in 0.5% (7/1403) of prepubertal cryptorchid boys, whereof 57% (4/7) in syndromic-cryptorchidism. DiscussionITGCN is predominantly observed prepubertally in boys with syndromic-cryptorchidism. In nonsyndromic cryptorchidism testicular cancer develops postpubertally, generally not based on dormant germ cells of ITGCN caused by an early fetal maldevelopment. Levels of EvidenceLEVEL I.

Polygenic inheritance of cryptorchidism susceptibility in the LE/orl rat.

Susceptibility to inherited cryptorchidism in the LE/orl rat may be associated with genetic loci that influence developmental patterning of the gubernaculum by the fetal testis. Cryptorchidism in the LE/orl rat is associated with a unique combination of homozygous minor alleles at multiple loci, and the encoded proteins are co-localized with androgen receptor (AR) and Leydig cells in fetal gubernaculum and testis, respectively. Prior studies have shown aberrant perinatal gubernacular migration, muscle patterning defects and reduced fetal testicular testosterone in the LE/orl strain. In addition, altered expression of androgen-responsive, cytoskeletal and muscle-related transcripts in the LE/orl fetal gubernaculum suggest a role for defective AR signaling in cryptorchidism susceptibility. The long-term LE/orl colony and short-term colonies of outbred Crl:LE and Crl:SD, and inbred WKY/Ncrl rats were maintained for studies. Animals were intercrossed (LE/orl X WKY/Ncrl), and obligate heterozygotes were reciprocally backcrossed to LE/orl rats to generate 54 F2 males used for genotyping and/or linkage analysis. At least five fetuses per gestational time point from two or more litters were used for quantitative real-time RT-PCR (qRT-PCR) and freshly harvested embryonic (E) day 17 gubernaculum was used to generate conditionally immortalized cell lines. We completed genotyping and gene expression analyses using genome-wide microsatellite markers and single nucleotide polymorphism (SNP) arrays, PCR amplification, direct sequencing, restriction enzyme digest with fragment analysis, whole genome sequencing (WGS), and qRT-PCR. Linkage analysis was performed in Haploview with multiple testing correction, and qRT-PCR data were analyzed using ANOVA after log transformation. Imaging was performed using custom and commercial antibodies directed at candidate proteins in gubernaculum and testis tissues, and gubernaculum cell lines. LE/orl rats showed reduced fertility and fecundity, and higher risk of perinatal death as compared with Crl:LE rats, but there were no differences in breeding outcomes between normal and unilaterally cryptorchid males. Linkage analysis identified multiple peaks, and with selective breeding of outbred Crl:LE and Crl:SD strains for alleles within two of the most significant (P < 0.003) peaks on chromosomes 6 and 16, we were able to generate a non-LE/orl cryptorchid rat. Associated loci contain potentially functional minor alleles (0.25-0.36 in tested rat strains) including an exonic deletion in Syne2, a large intronic insertion in Ncoa4 (an AR coactivator) and potentially deleterious variants in Solh/Capn15, Ankrd28, and Hsd17b2. Existing WGS data indicate that homozygosity for these combined alleles does not occur in any other sequenced rat strain. We observed a modifying effect of the Syne2(del) allele on expression of other candidate genes, particularly Ncoa4, and for muscle and hormone-responsive transcripts. The selected candidate genes/proteins are highly expressed, androgen-responsive and/or co-localized with developing muscle and AR in fetal gubernaculum, and co-localized with Leydig cells in fetal testis. The present study identified multiple cryptorchidism-associated linkage peaks in the LE/orl rat, containing potentially causal alleles. These are strong candidate susceptibility loci, but further studies are needed to demonstrate functional relevance to the phenotype. Association data from both human and rat models of spontaneous, nonsyndromic cryptorchidism support a polygenic etiology of the disease. Both the present study and a human genome-wide association study suggest that common variants with weak effects contribute to susceptibility, and may exist in genes encoding proteins that participate in AR signaling in the developing gubernaculum. These findings have potential implications for the gene-environment interaction in the etiology of cryptorchidism. Sequences were deposited in the Rat Genome Database (RGD, http://rgd.mcw.edu/). This work was supported by: R01HD060769 from the Eunice Kennedy Shriver National Institute for Child Health and Human Development (NICHD), 2P20GM103446 and P20GM103464 from the National Institute of General Medical Sciences (NIGMS), and Nemours Biomedical Research. The authors have no competing interests to declare.

Pathway analysis supports association of nonsyndromic cryptorchidism with genetic loci linked to cytoskeleton-dependent functions.

What are the genetic loci that increase susceptibility to nonsyndromic cryptorchidism, or undescended testis? A genome-wide association study (GWAS) suggests that susceptibility to cryptorchidism is heterogeneous, with a subset of suggestive signals linked to cytoskeleton-dependent functions and syndromic forms of the disease. Population studies suggest moderate genetic risk of cryptorchidism and possible maternal and environmental contributions to risk. Previous candidate gene analyses have failed to identify a major associated locus, although variants in insulin-like 3 (INSL3), relaxin/insulin-like family peptide receptor 2 (RXFP2) and other hormonal pathway genes may increase risk in a small percentage of patients. This is a case-control GWAS of 844 boys with nonsyndromic cryptorchidism and 2718 control subjects without syndromes or genital anomalies, all of European ancestry. All boys with cryptorchidism were diagnosed and treated by a pediatric specialist. In the discovery phase, DNA was extracted from tissue or blood samples and genotyping performed using the Illumina HumanHap550 and Human610-Quad (Group 1) or OmniExpress (Group 2) platform. We imputed genotypes genome-wide, and combined single marker association results in meta-analyses for all cases and for secondary subphenotype analyses based on testis position, laterality and age, and defined genome-wide significance as P = 7 × 10(-9) to correct for multiple testing. Selected markers were genotyped in an independent replication group of European cases (n = 298) and controls (n = 324). We used several bioinformatics tools to analyze top (P < 10(-5)) and suggestive (P < 10(-3)) signals for significant enrichment of signaling pathways, cellular functions and custom gene lists after multiple testing correction. In the full analysis, we identified 20 top loci, none reaching genome-wide significance, but one passing this threshold in a subphenotype analysis of proximal testis position (rs55867206, near SH3PXD2B, odds ratio = 2.2 (95% confidence interval 1.7, 2.9), P = 2 × 10(-9)). An additional 127 top loci emerged in at least one secondary analysis, particularly of more severe phenotypes. Cytoskeleton-dependent molecular and cellular functions were prevalent in pathway analysis of suggestive signals, and may implicate loci encoding cytoskeletal proteins that participate in androgen receptor signaling. Genes linked to human syndromic cryptorchidism, including hypogonadotropic hypogonadism, and to hormone-responsive and/or differentially expressed genes in normal and cryptorchid rat gubernaculum, were also significantly overrepresented. No tested marker showed significant replication in an independent population. The results suggest heterogeneous, multilocus and potentially multifactorial susceptibility to nonsyndromic cryptorchidism. The present study failed to identify genome-wide significant markers associated with cryptorchidism that could be replicated in an independent population, so further studies are required to define true positive signals among suggestive loci. As the only GWAS to date of nonsyndromic cryptorchidism, these data will provide a basis for future efforts to understand genetic susceptibility to this common reproductive anomaly and the potential for additive risk from environmental exposures. This work was supported by R01HD060769 (the Eunice Kennedy Shriver National Institute for Child Health and Human Development (NICHD)), P20RR20173 (the National Center for Research Resources (NCRR), currently P20GM103464 from the National Institute of General Medical Sciences (NIGMS)), an Institute Development Fund to the Center for Applied Genomics at The Children's Hospital of Philadelphia, and Nemours Biomedical Research. The authors have no competing interests to declare.

This Month in Pediatric Urology

No AccessJournal of UrologyThis Month in Pediatric Urology1 May 2015This Month in Pediatric Urology Laurence S. Baskin Laurence S. BaskinLaurence S. Baskin More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.008AboutFull TextPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail "This Month in Pediatric Urology." The Journal of Urology, 193(5), pp. 1457–1458 References 1 : Phenotype specific association of the TGFBR3 locus with nonsyndromic cryptorchidism. J Urol2015; 193. 1637. Google Scholar 2 : Congenital penile pathology is associated with abnormal development of the dartos muscle: a prospective study of primary penile surgery at a tertiary referral center. J Urol2015; 193. 1620. Google Scholar 3 : Genetic polymorphisms in ESR1 and ESR2 genes, and risk of hypospadias in a multiethnic study population. J Urol2015; 193. 1625. Google Scholar 4 : Serum inhibin B values in boys with unilateral vanished testis or unilateral cryptorchidism. J Urol2015; 193. 1632. Google Scholar © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 5May 2015Page: 1457-1458 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Laurence S. Baskin More articles by this author Expand All Advertisement PDF downloadLoading ...