Seminiferous tubule molecular imaging for evaluation of male fertility: Seeing is believing

Yao, Chencheng; Zhao, Liangyu; Tian, Ruhui; Zhu, Zijue; Xue, Yi; Chen, Huixing; Gong, Yuehua; Liu, Nachuan; Yang, Chao; He, Zuping; Li, Zheng

doi:10.1016/j.tice.2019.04.003

Cited by 4 publications

(4 citation statements)

References 36 publications

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“…DAZL and VASA both increased advancement during the meiosis. However, ectopic expression of DAZL generated the most haploid cells, with no indication of synergy with ectopic expression of VASA 47 . However, researchers found that both DAZL and VASA overexpression increased the proportion of ACROSIN-positive cells, suggesting that both proteins may be involved in the growth of this late male germ cell marker.…”

Section: Discussionmentioning

confidence: 94%

VASA protein and gene expression analysis of human non-obstructive azoospermia and normal by immunohistochemistry, immunocytochemistry, and bioinformatics analysis

Amirian

Azizi

Karoii

et al. 2022

Sci Rep

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VASA, also known as DDX4, is a member of the DEAD-box proteins and an RNA binding protein with an ATP-dependent RNA helicase. The VASA gene expression, which is required for human germ cell development, may lead to infertility. Immunocytochemistry and immunohistochemistry were used to examine the expression of VASA protein in the human testis sections of azoospermic patients, in-vitro and in-silico models. Some studies of fertile humans showed VASA expression in the basal and adluminal compartments of seminiferous tubules. Our Immunocytochemistry and immunohistochemistry in infertile humans showed expression of VASA in the luminal compartments of the seminiferous tubule. The immunohistochemical analysis of three human cases with different levels of non-obstructive azoospermia revealed a higher expression of VASA-positive cells. For this purpose, Enrichr and Shiny Gene Ontology databases were used for pathway enrichment analysis and gene ontology. STRING and Cytoscape online evaluation were applied to predict proteins' functional and molecular interactions and performed to recognize the master genes, respectively. According to the obtained results, the main molecular functions of the up-regulated and downregulated genes include the meiotic cell cycle, RNA binding, and differentiation. STRING and Cytoscape analyses presented seven genes, i.e., DDX5, TNP2, DDX3Y, TDRD6, SOHL2, DDX31, and SYCP3, as the hub genes involved in infertility with VASA co-function and protein–protein interaction. Our findings suggest that VASA and its interacting hub proteins could help determine the pathophysiology of germ cell abnormalities and infertility.

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Section: Discussionmentioning

confidence: 94%

VASA protein and gene expression analysis of human non-obstructive azoospermia and normal by immunohistochemistry, immunocytochemistry, and bioinformatics analysis

Amirian

Azizi

Karoii

et al. 2022

Sci Rep

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“…Intact seminiferous tubes were dissected and isolated from the testis of adult (15 weeks old) homozygous Nutm1-T2A-Luc2TdTomato reporter mice following published protocols 12 . Segments of live seminiferous tubes were placed in a 50ul drop of M2 medium (Cytospring) on a 3.5cm glass-bottom culture dish (MatTek Inc.).…”

Section: Methodsmentioning

confidence: 99%

“…Intact seminiferous tubes were dissected and isolated from the testis of adult (15 weeks old) homozygous Nutm1-T2A-Luc2TdTomato reporter mice following published protocols 12 .…”

Section: Whole-mount Live Imaging Of Seminiferous Tubesmentioning

confidence: 99%

The generation of a Nutm1 knock-in reporter mouse line for imaging post-meiotic spermatogenesis

Hakun

Rossant

2022

Preprint

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Spermiogenesis, the post-meiotic stage of sperm development, is critical for normal male fertility. Many genetic defects and environmental assaults that affect spermiogenesis have been shown to be associated with male infertility. In addition, this later stage of spermatogenesis has been proposed to be an ideal target for male contraceptive development. The mouse is a widely used model for studying the mechanisms of spermatogenesis and spermiogenesis. However, due to the complexity and the asynchronous nature of spermatogenesis in adult testis, it is challenging to study molecular processes restricted to this specific developmental stage. It is also challenging to monitor the spermiogenesic activity in live mice, which is critical for screening for fertility-modulating interventions such as contraceptives. Here we reported the development of a Nutm1-T2A- luciferase 2(Luc2)-tandem Tomato(TdTomato) knock-in reporter mouse model that specifically labels post-meiotic spermatids. Homozygous reporter mice are healthy and fully fertile, demonstrating no interference with the normal functions of the Nutm1 gene by the reporter. We demonstrated the visualization of post-meiotic spermatids by fluorescent imaging of the TdTomato reporter in both live and fixed testis tissues. We also demonstrated bioluminescence imaging of Nutm1 expressing cells in live mice. The Nutm1-T2A-Luc2TdTomato reporter mouse can serve as a valuable tool for studying spermiogenesis.

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“…However, in some SCO phenotypes of unknown etiology, the rare presence of spermatogonia in a small percentage of tubules are detected [9,10,[16][17][18]. Additionally, recent studies using next generation sequencing [19], whole mount staining of testicular biopsy [20], and 3D-agar/ methylcellulose-based culture system [21] have unequivocally demonstrated the presence of spermatogonia and SSCs in SCO testes. Nonetheless, OA and NOA can now be distinguished by FSH levels and/or testicular size based on detailed retrospective analysis of 153 azoospermic men between 1995 and 2000, including patient history, physical examination findings, endocrine profiles, testicular histology and sperm retrieval rates [22].…”

Section: Introductionmentioning

confidence: 99%

Defects of microtubule cytoskeletal organization in NOA human testes

Yun

Sang

et al. 2022

Reprod Biol Endocrinol

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The importance of actin and microtubule (MT) cytoskeletons in testis function in rodents is known to some extent, but its role in the etiology of azoospermia in humans remains unexplored. Here, we examined if MT cytoskeleton was defective in NOA (non-obstructive azoospermia) testes versus normal human testes based on histopathological, immunofluorescence (IF), and scRNA-Seq transcriptome profiling. Testis biopsy samples from n = 6 normal men versus n = 3 Sertoli cell only (SCO) and n = 3 MA (meiotic arrest) of NOA patients were used for histopathological analysis. IF analysis was also used to examine MT organization across the seminiferous epithelium, investigating the likely involvement of microtubule-associated proteins (MAPs). scRNA-Seq transcriptome profiling datasets from testes of 3 SCO patients versus 3 normal men in public domain in Gene Expression Omnibus (GEO) Sample (GSM) with identifiers were analyzed to examine relevant genes that regulate MT dynamics. NOA testes of MA and SCO patients displayed notable defects in MT organization across the epithelium with extensive truncation, mis-alignments and appeared as collapsed structures near the base of the tubules. These changes are in contrast to MTs in testes of normal men. scRNA-Seq analyses revealed considerable loss of spermatogenesis capacity in SCO testes of NOA patients versus normal men. An array of genes that support MT dynamics displayed considerable changes in expression and in spatial distribution. In summary, defects in MT cytoskeleton were noted in testes of NOA (SCO) patients, possibly mediated by defective spatial expression and/or distribution of MAPs. These changes, in turn, may impede spermatogenesis in SCO testes of NOA patients.

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Seminiferous tubule molecular imaging for evaluation of male fertility: Seeing is believing

Cited by 4 publications

References 36 publications

VASA protein and gene expression analysis of human non-obstructive azoospermia and normal by immunohistochemistry, immunocytochemistry, and bioinformatics analysis

VASA protein and gene expression analysis of human non-obstructive azoospermia and normal by immunohistochemistry, immunocytochemistry, and bioinformatics analysis

The generation of a Nutm1 knock-in reporter mouse line for imaging post-meiotic spermatogenesis

Defects of microtubule cytoskeletal organization in NOA human testes

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