Index   |   Contributors   |   Search

HORMONE ASSESSMENT

It is not necessary to perform hormone measurements routinely. Follicle-stimulating hormone (FSH) levels in patients with azoospermia, normal testicular volume, and normal virilization may help distinguish genital tract obstruction from a spermatogenic disorder. The most useful FSH value for the upper limit in reproductively normal young men is ~8 IU/L.^{89, 90} However, some men with primary seminiferous tubule failure have normal FSH levels. Normal FSH is common with germ cell arrest at the primary spermatocyte stage. Rarely, high FSH levels are seen with normal spermatogenesis.⁹¹ Measurement of FSH, luteinizing hormone, and testosterone is useful in men with reduced testicular volume and signs of androgen deficiency, to distinguish primary from secondary hypogonadism. Inhibin B measurement may provide additional information about the state of spermatogenesis.⁹² but is rarely used in routine practice.

Prolactin should be measured in men with galactorrhea or androgen deficiency and loss of libido.⁹³ Other hormone investigations are occasionally required, such as thyroid function tests with hyperprolactinemia, 17-hydroxyprogesterone measurements with congenital adrenal hyperplasia, estradiol with liver disease or tumors, iron studies to exclude hemochromatosis, human chorionic gonadotropin with tumors and estrogen excess, and pituitary function tests for panhypopituitarism.⁴³

CHROMOSOME AND GENETIC STUDIES

Chromosomal anomalies are 8-10X more common in infertile than fertile populations, and in many cases there are no other phenotypic changes. The prevalence is inversely correlated to the sperm density, being 14% of azoospermia.^{94, 95} Accordingly a routine assessment is recommended in men with unexplained infertility and sperm densities < 5-10 million/ml A karyotypes is performed in men with clinical evidence of primary testicular failure and small testes to confirm a clinical diagnosis of Klinefelter syndrome. Usually the karyotype is 47,XXY, but there may be higher numbers of X chromosomes or a sex-reversal 46,XX karyotype.^96-98 Although most men with Klinefelter syndrome produce no sperm in the semen, some are oligospermic and very rarely fertile.⁹⁶ Also, sperm for ICSI may be obtained by testicular biopsy in about 50% of patients.^{97, 98} Defective spermatogenesis may occur with 47,XYY, but the clinical picture is much less uniform than it is for Klinefelter syndrome. The extra Y chromosome is deleted early in gametogenesis because the sperm, embryos, and children generally have normal karyotypes. However, an increased rate of sex chromosomal and autosomal aneuploidy has been noted in studies of sperm from XXY and XYY men.^{98, 99} Some Y chromosome abnormalities, such as an isochromosome of two short arms, are associated with absences of spermatogenesis.

Infertile men have much higher rate of aneuploidies compared to fertile men and most of them have no other phenotypic features. An increased frequency of autosomal abnormalities is found with defective spermatogenesis, particularly balanced autosomal translocations (reciprocal and Robertsonian), which may be transmitted in unbalanced form to their offspring.¹⁰⁰ As part of their infertility investigation, it is imperative to screen severely oligospermic men as the result may affect treatment outcome.

Cystic fibrosis gene studies are important for evaluation of patients with congenital absence of the vas and their partners.¹⁰¹ If the woman has a cystic fibrosis gene mutation, preimplantation genetic diagnosis of their embryos can be offered. Microdeletions in the long arm of the Y chromosome (AZF regions) have been found in 3% to 15% of men with severe primary spermatogenic disorders.^{18, 19, 27, 36, 37} Sons of men with these microdeletions have the same microdeletions.¹⁰²

Another gene involved in spermatogenesis (histones replacement) and different from the AZF regions Yq deletions is CDY.¹⁰³ TSPY gene is located on the short arm of Y chromosome and may regulate the timing of spermatogenesis by signaling spermatogonia to enter meiosis.²⁹

The sex hormone-binding globulin (SHBG) gene has been studied for possible role in spermatogenesis.¹⁰⁴ Other genes that have been investigated for a possible involvement in fertility are DAZL (sperm count) MTHFR and the estrogen receptors genes (ESR1 and ESR2). Polymorphisms of the promoter region of the estrogen receptor gene have been shown to be related to sperm production. Men with higher numbers of TA repeats have lower sperm counts.²⁹

Androgen receptor defects have also been found in some men with unexplained primary spermatogenic disorders. Mutations in the gene impairing androgen receptor activity produce androgen insensitivity, which has a variable phenotypic expression from testicular feminization to otherwise normal males with gynecomastia or hypospermatogenesis and oligospermia.³³ Increases in the number of CAG repeats in exon 1 over approximately 40 are associated with Kennedy disease (progressive spinobulbar atrophy) and men with this condition may be infertile.

The field of epigenetic errors has also been studied for its possible contribution to male infertility.¹⁰⁵ (Table 4)

Other specific genetic tests and family studies may be indicated on clinical grounds (see Table 4).

At present, it is reasonable to screen all infertile men with otherwise unexplained primary spermatogenic defects with average sperm concentrations less than 5-15 million/mL by karyotype and Yq microdeletion testing.¹⁰⁶ All patients should be counseled about the possibility of transmitting known and unknown genetic defects.

Androgen receptor defects have also been found in some men with unexplained primary spermatogenic disorders. Mutations in the gene impairing androgen receptor activity produce androgen insensitivity, which has a variable phenotypic expression from testicular feminization to otherwise normal males with gynecomastia or hypospermatogenesis and oligospermia.³³ Increases in the number of CAG repeats in exon 1 over approximately 40 are associated with Kennedy disease (progressive spinobulbar atrophy) and men with this condition may be infertile. The field of epigenetic errors has also been studied for its possible contribution to male infertility.¹⁰⁵ (Table 4) Other specific genetic tests and family studies may be indicated on clinical grounds (see Table 4). At present, it is reasonable to screen all infertile men with otherwise unexplained primary spermatogenic defects with average sperm concentrations less than 5-15 million/mL by karyotype and Yq microdeletion testing.¹⁰⁶ All patients should be counseled about the possibility of transmitting known and unknown genetic defects.

TESTICULAR BIOPSY

Testicular biopsies are necessary to assess spermatogenesis in men with presumed genital tract obstruction. A significant proportion of men with azoospermia, normal testicular size, and normal FSH are found to have severe spermatogenic disorders.¹⁴ Some severe spermatogenic defects may be incomplete, and because ICSI can be performed if sperm can be obtained from the testes, diagnostic testicular biopsies should be offered to men with severe primary spermatogenic tubule disorders with persistent azoospermia. If any elongated spermatids can be found, it should be possible to perform ICSI. However, if no elongated spermatids are seen in the diagnostic biopsies it still may be possible to find spermatids by more extensive sampling of testicular tissue with open biopsies (see later). It is most important that tissue for histology is removed from the testes with minimal damage and placed in a suitable fixative, such as Bouin’s or Steive’s solution. Standard formalin fixatives destroy the cytoarchitecture.

Testis biopsies may be performed under local or general anesthesia. Needle biopsy many obtain only isolated cells but these may be sufficient for diagnosis based on cytology or for flow cytometry. The technique shown in Fig. 4 usually provides sufficient material for a histologic diagnosis of the state of the seminiferous epithelium despite some deformation artifacts.¹⁰⁷ It is also useful for obtaining testicular sperm for ICSI.¹⁰⁸ Complications are rare and include minor bleeding in the skin and testis, and rarely hematoma or reactions to the local anesthetic. Failure to obtain tissue occurs particularly with fibrosed or small (<5mL) testes In the presence of azoospermia, an open testicular biopsy might not just be a therapeutic approach in the way of sperm retrieval, but also the only way of excluding early testicular germ cell neoplasia or even an overt testicular cancer.

Infertile men are more likely to develop testicular cancer compared to men with normal fertility.¹⁰⁹ Testicular intra-tubular germ cell neoplasia of the unclassified type (ITGCNU) also called carcinoma in situ (see below) is the precursor of testicular germ cell tumors in which the neoplastic cells are confined within the seminiferous tubules. This makes it a non-invasive stage of the disease. It can be found in testicular tissue adjacent to germ cell tumours in more than 90 percent of adult cases.^{110, 111}

The incidence of ITGCNU in men undergoing fertility evaluation ranges between 0.4 - 1.1 percent.^{112, 113} As ITGCNU is asymptomatic, patients remain undiagnosed until an overt tumour can be identified usually by palpation. A sample from a sperm retrieval open biopsy should be sent to histopathology routinely.

For clinical purposes, testicular histology is classified as follows: normal or hypospermatogenesis (all the cellular elements of spermatogenesis are present but in reduced numbers), germ cell arrest (the earlier cellular elements of spermatogenesis are present but at a certain stage, the process stops, most often at the primary spermatocytes), Sertoli-cell-only syndrome or germ cell aplasia (the tubules contain Sertoli cells but no germ cells), hyalinization (the cellular elements have disappeared, leaving only thickened seminiferous tubule walls as in Klinefelter syndrome), and immature testis (no gonadotropin stimulation, prepubertal appearance).¹¹⁴ Examples are shown in Figure 5. Other classifications such as partial or incomplete maturation arrest and partial germ cell aplasia cause confusion in the literature and should not be used.¹¹⁵

OTHER INVESTIGATIONS

Ultrasonography is useful to check for tumors in the testes, particularly when the testes are difficult to palpate because of a tense hydrocele.¹¹⁶ Other abnormalities may also be found (Figs 6-8). It can also be used to measure testicular size and confirm the presence and nature of cysts or other abnormalities in the scrotum. Some argue scrotal ultrasound should be performed routinely in infertile men to measure testicular volume, assessing the texture and exclude impalpable malignant tumors in the testes.¹¹⁷ However, some clinical guidelines do not support this.¹¹⁸ Doppler blood flow assessment is valuable in assessing a painful swollen testis for torsion or inflammation and for evaluating varicoceles. Other tests of a varicocele, including thermography, technetium scans, and venography may be performed but, as pointed out later, the value of treating varicoceles to improve fertility is uncertain. Rectal ultrasound may demonstrate cysts in the prostate, enlarged seminal vesicles, or dilated ejaculatory ducts associated with distal genital tract obstructions (Figure 8).¹¹⁹ Clinical suspicion of the presence of a pituitary tumor should be followed up by appropriate radiology. Abdominal imaging is necessary to check the position of impalpable testes.

FIGURE 8 Ultrasound of undescended testes. A - In neck of scrotum at external inguinal ring with small tumour; B - In inguinal canal; Incidental prostatic cyst in a healthy man: C - transverse, D - sagittal; Ejaculatory duct obstruction: E - prostate transverse - dilated ejaculatory duct; F - transverse, dilated seminal vesicle; G - sagittal, dilated duct and seminal vesicle