ScRNA-seq and scATAC-seq reveal that sertoli cell mediate spermatogenesis disorders through stage-specific communications in non-obstructive azoospermia

Reviewer #1 (Public Review):

Summary:

The manuscript is dedicated heavily to cell type mapping and identification of sub-type markers in the human testis but does not present enough results from cross-investigation between NOA cases versus control. Their findings are mostly based on transcriptome and the authors do not make enough use of the scATAC-seq data in their analyses as they put forward in the title. Overall, the authors should do more to include the differential profile of NOA cases at the molecular level - specific gene expression, chromatin accessibility, TF binding, pathway, and signaling that are perturbed in NOA patients that may be associated with azoospermia.

Strengths:

(1) The establishment of single-cell data (both RNA and ATAC) from the human testicular tissues is noteworthy.

(2) The manuscript includes extensive mapping of sub-cell populations with some claimed as novel, and reports marker gene expression.

(3) The authors present inter-cellular cross-talks in human testicular tissues that may be important in adequate sperm cell differentiation.

Weaknesses:

(1) A low sample size (2 OA and 3 NOA cases). There are no control samples from healthy individuals.

(2) Their argument about interactions between germ and Sertoli cells is not based on statistical testing.

(3) Rationale/logic of the study. This study, in its present form, seems to be more about the role of sub-Sertoli population interactions in sperm cell development and does not provide enough insights about NOA.

(4) The authors do not make full use of the scATAC-seq data.

Reviewer #2 (Public Review):

Summary:

Shimin Wang et al. investigated the role of Sertoli cells in mediating spermatogenesis disorders in non-obstructive azoospermia (NOA) through stage-specific communications. The authors utilized scRNA-seq and scATAC-seq to analyze the molecular and epigenetic profiles of germ cells and Sertoli cells at different stages of spermatogenesis.

Strengths:

By understanding the gene expression patterns and chromatin accessibility changes in Sertoli cells, the authors sought to uncover key regulatory mechanisms underlying male infertility and identify potential targets for therapeutic interventions. They emphasized that the absence of the SC3 subtype would be a major factor contributing to NOA.

Weaknesses:

Although the authors used cutting-edge techniques to support their arguments, it is difficult to find conceptual and scientific advances compared to Zeng S et al.'s paper (Zeng S, Chen L, Liu X, Tang H, Wu H, and Liu C (2023) Single-cell multi-omics analysis reveals dysfunctional Wnt signaling of spermatogonia in non-obstructive azoospermia. Front. Endocrinol. 14:1138386.). Overall, the authors need to improve their manuscript to demonstrate the novelty of their findings in a more logical way.

Reviewer #3 (Public Review):

Summary:

This study profiled the single-cell transcriptome of human spermatogenesis and provided many potential molecular markers for developing testicular puncture-specific marker kits for NOA patients.

Strengths:

Perform single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) on testicular tissues from two OA patients and three NOA patients.

Weaknesses:

Most results are analytical and lack specific experiments to support these analytical results and hypotheses.