Improved Accuracy and Precision in Diagnosing Nonobstructive Azoospermia (NOA) With Spermatogenic Cell Type Specific Gene Expression Profiling

Non-obstructive azoospermia (NOA) is a form of male infertility affecting 1% of men overall and constituting 10%-15% of male infertility cases. Most cases of NOA are idiopathic. Current NOA diagnostic methods rely on testis biopsies which are assumed to represent the entire testis, thus introducing bias. Histological identification of spermatogenic cell types requires extensive training, is tedious, time-consuming, and can be ambiguous. Recently, we defined a panel of 33 genes uniquely expressed by 11 different cell types within the human spermatogenic lineage based on results from single-cell RNA-seq. Here, I sought to verify that measuring mRNA abundance of these spermatogenic cell type specific genes would accurately diagnose which cells are present in biopsies. For this purpose, I confirmed the medical diagnosis of testis specimens from patients with normal spermatogenesis and those with NOA using marker labeling and compared results with quantitative RT-PCR (qRT-PCR) detection of mRNA abundance for the 33 gene panel. Cell type specific mRNA expression identified the same misdiagnoses as marker labeling (3:46 samples). Cell type specific gene expression also distinguished cases of Sertoli cell only (SCO) from maturational arrest and normal testes, and also identify the point of arrest in the spermatogenic lineage. Surprisingly, one definitive SCO case exhibited expression of all spermatogenic marker mRNAs, demonstrating the possibility of false-positive results. Overall, our results indicate that detection of spermatogenic cell type specific gene expression provides a less-tedious and time-consuming, and more comprehensive, accurate and precise approach to diagnosing NOA.