Spermatozoa recovered from testicular biopsies can be used through intracytoplasmic sperm injection (ICSI) to achieve a pregnancy. To assess the likelihood of successful testicular sperm extraction (TESE) in men suffering from severe oligo- or azoospermia, bilateral biopsy specimens were obtained. Following semi-thin sectioning, the morphology of testicular samples was graded according to a modified Johnsen score. TESE was performed in parallel to this histological examination. The number of isolated spermatozoa was assessed in a semiquantitative way. From 103 patients investigated, 64 (62.1%) showed azoospermia in a preceding semen analysis and 29 (28.2%) patients had sperm concentrations between 0.1 and 1 x 10(6)/ml. In 10 patients who had higher sperm counts, most spermatozoa were non-motile. Spermatozoa could be detected after TESE in the testicular tissue of 49 (77%) azoospermic men. When follicle stimulating hormone (FSH) concentration was normal, most patients had detectable spermatozoa after TESE. Nearly one-third of patients with mildly elevated FSH had no spermatozoa. Thirty-nine percent of patients in whom FSH was elevated to more than twice normal and 50% of patients with grossly elevated FSH had no detectable spermatozoa. In all, 82.8% of men with sperm concentrations between 0.1 and 1x10(6)/ml in their ejaculate showed spermatozoa in the tissue sample after TESE. Our data demonstrate that, contrary to previous recommendations, infertile men with azoospermia and high FSH values should be reconsidered for testicular biopsy, provided that tissue samples can be cryopreserved for later TESE/ICSI treatment.

Non-obstructive azoospermia (NOA) is the most severe form of male infertility and typically incurable. Defining the genetic basis of NOA has proven challenging, and the most advanced classification of NOA subforms is not based on genetics, but simple description of testis histology. In this study, we exome-sequenced over 1000 clinically diagnosed NOA cases and identified a plausible recessive Mendelian cause in 20%. We find further support for 21 genes in a 2-stage burden test with 2072 cases and 11,587 fertile controls. The disrupted genes are primarily on the autosomes, enriched for undescribed human “knockouts”, and, for the most part, have yet to be linked to a Mendelian trait. Integration with single-cell RNA sequencing data shows that azoospermia genes can be grouped into molecular subforms with synchronized expression patterns, and analogs of these subforms exist in mice. This analysis framework identifies groups of genes with known roles in spermatogenesis but also reveals unrecognized subforms, such as a set of genes expressed across mitotic divisions of differentiating spermatogonia. Our findings highlight NOA as an understudied Mendelian disorder and provide a conceptual structure for organizing the complex genetics of male infertility, which may provide a rational basis for disease classification.

A key step in studying the biology of spermatogonia is to determine their global gene expression profile. However, disassociation of these cells from the testis may alter their profile to a considerable degree. To characterize the molecular phenotype of human spermatogonia, including spermatogonial stem cells (SSCs), within their cognate microenvironment, a rare subtype of human defective spermatogenesis was exploited in which spermatogonia were the only germ cell type. The global expression profile of these samples was assessed on the Affymetrix microarray platform and compared with tissues showing homogeneous Sertoli-cell-only appearance; selected genes were validated by quantitative real-time PCR and immunohistochemistry on disparate sample sets. Highly significant differences in gene expression levels correlated with the appearance of spermatogonia, including 239 best candidates of human spermatogonially expressed genes. Specifically, fibroblast growth factor receptor 3 (FGFR3), desmoglein 2 (DSG2), E3 ubiquitin ligase c-CBL (casitas B-cell lymphoma), cancer/testis antigen NY-ESO-1 (CTAG1A/B), undifferentiated embryonic cell transcription factor 1 (UTF1) and synaptosomal-associated protein, 91 kDa homolog (SNAP91) were shown to represent specific biomarkers of human spermatogonia. These biomarkers, specifically the surface markers FGFR3 and DSG2, may facilitate the isolation and enrichment of human stem and/or progenitor spermatogonia and thus lay a foundation for studies of long-term maintenance of human SSCs/progenitor cells, spermatogonial self-renewal, clonal expansion and differentiation.

Shah R, et al. World J Mens Health. 2023 Jan;41(1):164-197. https://doi.org/10.5534/wjmh.220048