Multiple ageing effects on testicular/epididymal germ cells lead to decreased male fertility in mice.

In mammals, females undergo reproductive cessation with age, whereas male fertility gradually declines but persists almost throughout life. However, the detailed effects of ageing on germ cells during and after spermatogenesis, in the testis and epididymis, respectively, remain unclear. Here we comprehensively examined the in vivo male fertility and the overall organization of the testis and epididymis with age, focusing on spermatogenesis, and sperm function and fertility, in mice.

In vitro spermatogenesis in isolated seminiferous tubules of immature mice.

Mouse spermatogenesis, from spermatogonial stem cell proliferation to sperm formation, can be reproduced in vitro by culturing testis tissue masses of neonatal mice. However, it remains to be determined whether this method is also applicable when testis tissues are further divided into tiny fragments, such as segments of the seminiferous tubule (ST), a minimal anatomical unit for spermatogenesis. In this study, we investigated this issue using the testis of an Acrosin-GFP/Histone H3.

SPATA33 localizes calcineurin to the mitochondria and regulates sperm motility in mice.

Calcineurin is a calcium-dependent phosphatase that plays roles in a variety of biological processes including immune responses. In spermatozoa, there is a testis-enriched calcineurin composed of PPP3CC and PPP3R2 (sperm calcineurin) that is essential for sperm motility and male fertility. Because sperm calcineurin has been proposed as a target for reversible male contraceptives, identifying proteins that interact with sperm calcineurin widens the choice for developing specific inhibitors.

Rubicon prevents autophagic degradation of GATA4 to promote Sertoli cell function.

Autophagy degrades unnecessary proteins or damaged organelles to maintain cellular function. Therefore, autophagy has a preventive role against various diseases including hepatic disorders, neurodegenerative diseases, and cancer. Although autophagy in germ cells or Sertoli cells is known to be required for spermatogenesis and male fertility, it remains poorly understood how autophagy participates in spermatogenesis.

Sertoli cell replacement in explanted mouse testis tissue supporting host spermatogenesis†.

Spermatogenesis takes place in the seminiferous tubules, starting from the spermatogonial stem cell and maturing into sperm through multiple stages of cell differentiation. Sertoli cells, the main somatic cell constituting the seminiferous tubule, are in close contact with every germ cell and play pivotal roles in the progression of spermatogenesis. In this study, we developed an in vitro Sertoli cell replacement method by combining an organ culture technique and a toxin receptor-mediated cell knockout system.

Rat in vitro spermatogenesis promoted by chemical supplementations and oxygen-tension control.

In vitro spermatogenesis (IVS) using air-liquid interphase organ culture method is possible with mouse testis tissues. The same method, however, has been hardly applicable to animals other than mice, only producing no or limited progression of spermatogenesis. In the present study, we challenged IVS of rats with modifications of culture medium, by supplementing chemical substances, including hormones, antioxidants, and lysophospholipids.

Antioxidant vitamins and lysophospholipids are critical for inducing mouse spermatogenesis under organ culture conditions.

In vitro mouse spermatogenesis using a classical organ culture method became possible by supplementing basal culture medium with only the product of bovine serum albumin purified by chromatography (AlbuMAX), which indicated that AlbuMAX contained every chemical factor necessary for mouse spermatogenesis. However, since the identity of these factors was unclear, improvements in culture media and our understanding of the nutritional and signal substances required for spermatogenesis were hindered. In the present study, chemically defined media (CDM) without AlbuMAX was used to evaluate each supplementary factor and their combinations for the induction of spermatogenesis.

PITHD1 is a proteasome-interacting protein essential for male fertilization.

The proteasome is a protein-degrading molecular complex that is necessary for protein homeostasis and various biological functions, including cell cycle regulation, signal transduction, and immune response. Proteasome activity is finely regulated by a variety of proteasome-interacting molecules. PITHD1 is a recently described molecule that has a domain putatively capable of interacting with the proteasome.

CRISPR/Cas9-mediated genome editing reveals 30 testis-enriched genes dispensable for male fertility in mice†.

More than 1000 genes are predicted to be predominantly expressed in mouse testis, yet many of them remain unstudied in terms of their roles in spermatogenesis and sperm function and their essentiality in male reproduction. Since individually indispensable factors can provide important implications for the diagnosis of genetically related idiopathic male infertility and may serve as candidate targets for the development of nonhormonal male contraceptives, our laboratories continuously analyze the functions of testis-enriched genes in vivo by generating knockout mouse lines using the CRISPR/Cas9 system. The dispensability of genes in male reproduction is easily determined by examining the fecundity of knockout males.

An azoospermic factor gene, Ddx3y and its paralog, Ddx3x are dispensable in germ cells for male fertility.

About 10% of male infertile patients show abnormalities in spermatogenesis. The microdeletion of azoospermia factor a (AZFa) region of the Y chromosome is thought to be a cause of spermatogenic failure. However, candidate gene responsible for the spermatogenic failure in AZFa deleted patients has not been elucidated yet.

Seminal vesicle secretory protein 7, PATE4, is not required for sperm function but for copulatory plug formation to ensure fecundity†.

Seminal vesicle secretions (SVSs), together with spermatozoa, are ejaculated into the female reproductive tract. SVS7, also known as PATE4, is one of the major SVS proteins found in the seminal vesicle, copulatory plug, and uterine fluid after copulation. Here, we generated Pate4 knockout (-/-) mice and examined the detailed function of PATE4 on male fecundity.

Chimeric analysis with newly established EGFP/DsRed2-tagged ES cells identify HYDIN as essential for spermiogenesis in mice.

The CRISPR/Cas9 system can efficiently introduce biallelic mutations in ES cells (ESCs), and its application with fluorescently-tagged ESCs enables phenotype analysis in chimeric mice. We have utilized ESCs that express EGFP in the cytosol and acrosome [EGR-G101 129S2 × (CAG/Acr-EGFP) B6] in previous studies; however, the EGFP signal in the sperm cytosol is weak and the signal in the acrosome is lost after the acrosome reaction, precluding analysis between wild type and ESC derived spermatozoa. In this study, we established an ESC line from RBGS (Red Body Green Sperm) transgenic mice [B6D2-Tg (CAG/Su9-DsRed2, Acr3-EGFP) RBGS002Osb] whose spermatozoa exhibit green fluorescence in the acrosome and red fluorescence in the mitochondria within the flagellar midpiece that is retained after the acrosome reaction.

FGFRL1 deficiency reduces motility and tumorigenic potential of cells derived from oesophageal squamous cell carcinomas.

Oesophageal squamous cell carcinoma (ESCC) is an aggressive cancer that resulted in ~400,000 mortalities worldwide in 2012. It was reported previously that fibroblast growth factor receptor-like 1 (FGFRL1) is highly expressed in ESCC patients with lymph node metastasis and poor prognosis accordingly. FGFRL1 is an FGFR that lacks tyrosine kinase activity, whereas the activity is critical for other FGFRs to activate intracellular signalling.