Evaluating genetic causes of azoospermia: What can we learn from a complex cellular structure and single-cell transcriptomics of the human testis?
Azoospermia is a condition defined as the absence of spermatozoa in the ejaculate, but the testicular phenotype of men with azoospermia may be very variable, ranging from full spermatogenesis, through arrested maturation of germ cells at different stages, to completely degenerated tissue with ghost tubules. Hence, information regarding the cell-type-specific expression patterns is needed to prioritise potential pathogenic variants that contribute to the pathogenesis of azoospermia. Thanks to technological advances within next-generation sequencing, it is now possible to obtain detailed cell-type-specific expression patterns in the testis by single-cell RNA sequencing. However, to interpret single-cell RNA sequencing data properly, substantial knowledge of the highly sophisticated data processing and visualisation methods is needed. Here we review the complex cellular structure of the human testis in different types of azoospermia and outline how known genetic alterations affect the pathology of the testis. We combined the currently available single-cell RNA sequencing datasets originating from the human testis into one dataset covering 62,751 testicular cells, each with a median of 2637 transcripts quantified. We show what effects the most common data-processing steps have, and how different visualisation methods can be used. Furthermore, we calculated expression patterns in pseudotime, and show how splicing rates can be used to determine the velocity of differentiation during spermatogenesis. With the combined dataset we show expression patterns and network analysis of genes known to be involved in the pathogenesis of azoospermia. Finally, we provide the combined dataset as an interactive online resource where expression of genes and different visualisation methods can be explored (https://testis.cells.ucsc.edu/).
Assisted reproductive techniques
Congenital absence of the vas deferens
Differentially expressed gene
GEnetics of Male INfertility Initiative
Genome-wide association studies
Intracytoplasmic sperm injection
International Male infertility Genomics Consortium
Mutual nearest neighbour
Principal component analysis
Single-cell RNA sequencing
Testicular sperm extraction
T-distributed stochastic nearest neighbour
Uniform manifold approximation
Unique molecular identifier
World Health Organisation
The authors thank the editor, Prof. Csilla Krausz and the anonymous referees for their constructive comments and suggestions.
The Danish Council for Independent Research | Natural Sciences (Grant Number 6108-00385A), and the Novo Nordisk Foundation (Grant Number NNF17OC0031004) to MHS.
Compliance with ethical standards
Conflict of interest
On behalf of all authors, the corresponding author states that there is no conflict of interest.
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