No abstract

In vitro maturation (IVM) and vitrification have been widely used to prepare oocytes before fertilization; however, potential effects of these procedures, such as expression profile changes, are poorly understood. In this study, mouse oocytes were divided into four groups and subjected to combinations of in vitro maturation and/or vitrification treatments. RNA-seq and in silico pathway analysis were used to identify differentially expressed genes (DEGs) that may be involved in oocyte viability after in vitro maturation and/or vitrification. Our results showed that 1) 69 genes were differentially expressed after IVM, 66 of which were up-regulated. Atp5e and Atp5o were enriched in the most significant gene ontology term “mitochondrial membrane part”; thus, these genes may be promising candidate biomarkers for oocyte viability after IVM. 2) The influence of vitrification on the transcriptome of oocytes was negligible, as no DEGs were found between vitrified and fresh oocytes. 3) The MII stage is more suitable for oocyte vitrification with respect to the transcriptome. This study provides a valuable new theoretical basis to further improve the efficiency of in vitro maturation and/or oocyte vitrification.

Cryopreservation damages permeability of sperm mitochondrial membranes, with formation of a mitochondrial permeability transition pore (mPTP). Mitochondria are both a primary synthesis site and principle target for melatonin, which can directly inhibit mPTP formation. The objective was to determine effects of melatonin on mPTP opening of frozen‐thawed ram sperm and elucidate underlying pathways by antagonist and agonists of melatonin receptors (MTs), and antagonists of PI3K and GSK 3β treatments; furthermore, plasma membrane integrity, mitochondrial membrane potential (ΔΨm), mitochondrial cytochrome c (Cyt c) release and fertilization were analysed to assess the effect of mPTP status mediated by melatonin on quality of frozen‐thawed sperm. Fresh ram semen was diluted in glucose‐egg yolk buffer with 0 or 10–7 M melatonin (frozen and frozen + melatonin groups, respectively) and slow‐frozen. In frozen‐thawed sperm, melatonin added at initiation of 4°C equilibration was most effective for inhibiting mPTP opening, decreasing peptidyl‐prolyl‐cis/trans isomerase activity of cyclophilin D and increasing plasma membrane integrity, ΔΨm, mitochondrial Cyt c concentration and fertilizing ability (p < .05). In a mechanistic study, the melatonin receptor (MT)1 antagonist eliminated inhibition of melatonin on mPTP opening, whereas MT1 agonist had opposite effects (p < .05). Neither MT2 antagonist nor agonist had significant effect, but PI3K and/or GSK 3β antagonist decreased inhibition of MT1 agonist on mPTP opening (p < .05). In conclusion, melatonin improved sperm cryopreservation, perhaps by acting on MT1 via the PI3K‐Akt‐GSK 3β pathway to inhibit mPTP opening.

Summary Sertoli cells are the major component of the spermatogonial stem cell (SSC) niche; however, regulatory mechanisms in Sertoli cells that dictate SSC fate decisions remain largely unknown. Here we revealed features of the N 6 -methyladenosine (m 6 A) mRNA modification in Sertoli cells and demonstrated the functions of WTAP, the key subunit of the m 6 A methyltransferase complex in spermatogenesis. m 6 A-sequencing analysis identified 21,909 m 6 A sites from 15,365 putative m 6 A-enriched transcripts within 6,122 genes, including many Sertoli cell-specific genes. Conditional deletion of Wtap in Sertoli cells resulted in sterility and the progressive loss of the SSC population. RNA sequencing and ribosome nascent-chain complex-bound mRNA sequencing analyses suggested that alternative splicing events of transcripts encoding SSC niche factors were sharply altered and translation of these transcripts were severely dysregulated by Wtap deletion. Collectively, this study uncovers a novel regulatory mechanism of the SSC niche and provide insights into molecular interactions between stem cells and their cognate niches in mammals.

No abstract

Background Spermatogenesis is a cellular differentiation process that includes three major events: mitosis of spermatogonia, meiosis of spermatocytes and spermiogenesis. Steady-state spermatogenesis relies on functions of spermatogonial stem cells (SSCs). Establishing and maintaining a foundational SSC pool is essential for continued spermatogenesis in mammals. Currently, our knowledge about SSC and spermatogenesis is severely limited in domestic animals. Results In the present study, we examined transcriptomes of testes from domestic yaks at four different stages (3, 5, 8 and 24 months of age) and attempted to identify genes that are associated with key developmental events of spermatogenesis. Histological analyses showed that the most advanced germ cells within seminiferous tubules of testes from 3, 5, 8 and 24 months old yaks were gonocytes, spermatogonia, spermatocytes and elongated spermatids, respectively. RNA-sequencing (RNA-seq) analyses revealed that 11904, 4381 and 2459 genes were differentially expressed during the gonocyte to spermatogonia transition, the mitosis to meiosis transition and the meiosis to post-meiosis transition. Further analyses identified a list of candidate genes than may regulate these important cellular processes. CXCR4 , a previously identified SSC niche factor in mouse, was one of the up-regulated genes in the 5 months old yak testis. Results of immunohistochemical staining confirmed that CXCR4 was exclusively expressed in gonocytes and a subpopulation of spermatogonia in the yak testis. Conclusions Together, these findings demonstrated histological changes of postnatal testis development in the domestic yak. During development of spermatogonial lineage, meiotic and haploid germ cells are supported by dynamic transcriptional regulation of gene expression. Our transcriptomic analyses provided a list of candidate genes that potentially play crucial roles in directing the establishment of SSC and spermatogenesis in yak. Electronic supplementary material The online version of this article (10.1186/s40104-019-0360-7) contains supplementary material, which is available to authorized users.

No abstract

No abstract