Project description:MicroRNAs (miRNAs) are involved in nearly every biological process examined to date. Mounting evidence show that some spermatozoa specific miRNAs play important roles in the regulation of spermatogenesis and germ cells development, but little is known of the exact identity and function of miRNA in sperm cells or their potential involvement in spermatogenesis and germ cells development. Here, we investigated the spermatozoa miRNA profiles using illumina deep sequencing combined with bioinformatic analysis using zebrafish as a model system. Deep sequencing of small RNAs yielded 12 million raw reads from zebrafish spermatozoa. Analysis showed that the noncoding RNA of the spermatozoa included tRNA, rRNA, snRNA, snoRNA and miRNA. By mapping to the zebrafish genome, we identified 400 novel and 204 conserved miRNAs which could be grouped into 104 families, including zebrafish specific families, such as mir-731, mir-724, mir-725, mir-729 and mir-2185. We report the first characterization of the miRNAs profiling in zebrafish spermatozoa. The obtained spermatozoa miRNAs profiling will serve as valuable resources to systematically study spermatogenesis in fish and vertebrate.
Project description:We reported RNA profiles of mice spermatozoa, a total of 35,288,825 reads matching 33,039 transcripts, including 27,310 coding transcripts, were obtained.
Project description:MicroRNAs (miRNAs) are involved in nearly every biological process examined to date. Mounting evidence show that some spermatozoa specific miRNAs play important roles in the regulation of spermatogenesis and germ cells development, but little is known of the exact identity and function of miRNA in sperm cells or their potential involvement in spermatogenesis and germ cells development. Here, we investigated the spermatozoa miRNA profiles using illumina deep sequencing combined with bioinformatic analysis using zebrafish as a model system. Deep sequencing of small RNAs yielded 12 million raw reads from zebrafish spermatozoa. Analysis showed that the noncoding RNA of the spermatozoa included tRNA, rRNA, snRNA, snoRNA and miRNA. By mapping to the zebrafish genome, we identified 400 novel and 204 conserved miRNAs which could be grouped into 104 families, including zebrafish specific families, such as mir-731, mir-724, mir-725, mir-729 and mir-2185. We report the first characterization of the miRNAs profiling in zebrafish spermatozoa. The obtained spermatozoa miRNAs profiling will serve as valuable resources to systematically study spermatogenesis in fish and vertebrate. Examination of small RNA populations in zebrafish spermatozoa
Project description:We reported RNA profiles of mice spermatozoa, a total of 35,288,825 reads matching 33,039 transcripts, including 27,310 coding transcripts, were obtained. RNA profiles of the spermatozoa of 9-10 weeks adult mice were sequenced by RNA-seq,using Illumina GAIIx.
Project description:Proteins play an important role in many reproductive functions such as sperm maturation and physiology, sperm transit in the female genital tract or sperm-oocyte interaction. However, little information concerning reproductive features is available in the case of aquatic animals. The present study aims to characterize the proteome of both spermatozoa and seminal plasma of bottlenose dolphins (Tursiops truncatus) as a model organism for cetaceans. Ejaculate samples were obtained voluntarily from two trained dolphins housed in an aquarium. Spermatozoa and seminal plasma were analyzed by means of proteomic analyses using a LC-MS/MS and a list with the gene symbols corresponding to each protein was submitted to DAVID database. A total of 423 and 307 proteins were identified in spermatozoa and seminal plasma, respectively, and the samples shared 111 proteins. Furthermore, 70 proteins were identified as involved in reproductive processes, 39 in spermatozoa, and 31 in seminal plasma. The top-5 most abundant proteins were also identified in these samples: AKAP3, ODF2, TUBB, GST3, ROPN1 for spermatozoa and CST11, LTF, ALB, HSP90B1, PIGR for seminal plasma. In conclusion, this proteomic study provides the first characterization of proteomic expression patterns in cetacean sperm and seminal plasma, opening a window to future research on the discovery of biomarkers, analysis of conservation capacity or additional applications in the field of assisted reproduction technologies.
Project description:Our objective was to determine whether oxidative damage of rhesus macaque sperm induced by reactive oxygen species (ROS) in vitro would affect embryo development following intracytoplasmic sperm injection (ICSI) of metaphase II (MII) oocytes. Fresh rhesus macaque spermatozoa were treated with ROS as follows: 1 mM xanthine and 0.1 U/ml xanthine oxidase (XXO) at 37°C and 5% CO? in air for 2.25 h. Sperm were then assessed for motility, viability, and lipid peroxidation. Motile ROS-treated and control sperm were used for ICSI of MII oocytes. Embryo culture was evaluated for 3 days for development to the eight-cell stage. Embryos were fixed and stained for signs of cytoplasmic and nuclear abnormalities. Gene expression was analyzed by RNA-Seq in two-cell embryos from control and treated groups. Exposure of sperm to XXO resulted in increased lipid peroxidation and decreased sperm motility. ICSI of MII oocytes with motile sperm induced similar rates of fertilization and cleavage between treatments. Development to four- and eight-cell stage was significantly lower for embryos generated with ROS-treated sperm than for controls. All embryos produced from ROS-treated sperm demonstrated permanent embryonic arrest and varying degrees of degeneration and nuclear fragmentation, changes that are suggestive of prolonged senescence or apoptotic cell death. RNA-Seq analysis of two-cell embryos showed changes in transcript abundance resulting from sperm treatment with ROS. Differentially expressed genes were enriched for processes associated with cytoskeletal organization, cell adhesion, and protein phosphorylation. ROS-induced damage to sperm adversely affects embryo development by contributing to mitotic arrest after ICSI of MII rhesus oocytes. Changes in transcript abundance in embryos destined for mitotic arrest is evident at the two-cell stage of development.
Project description:The objective of the present study is to investigate if females have the ability to recognise X or Y chromosome bearing spermatozoa and present a different response to different spermatozoa. In order to investigate if females have the ability to recognise X or Y chromosome bearing spermatozoa and present a different response to different spermatozoa. Samples of semen were sorted for chromosomal sex by M-oM-,M-^Bow cytometric-sorting. Using laparoscopic insemination one oviduct was inseminated with X spermatozoa and the contralateral oviduct was inseminated with Y spermatozoa.This allowed us to obtain samples from the oviduct in contact with X sperm and Y sperm in the the contrlateral oviduct from the same sow. Oviduct pig samples were collected for RNA isolation and hybridization on Affymetrix microarrays. Four biological replicates were performed (n= 4 sows) and a total of 8 arrays were used for microarrays study (4 arrays for oviduct samples in contact with X spermatozoa and 4 arrays for oviduct samples in contact with Y spermatozoa.
Project description:Small noncoding RNAs (sncRNA) are becoming recognized for their participation in a diverse range of cellular functions. In this study, their global characterization in human spermatozoa from donors with proven fertility was undertaken. Reads were analyzed in two classes, those mapping to unique locations and those that could be aligned to up to 10 genomic locations. All libraries showed comparable distribution of reads between intergenic, intronic and exonic genomic regions. Analysis of the sequences revealed the presence of multiple small RNA classes. The miRNAs retained in spermatozoa were found to be associated with promoter regions, suggestive of a role at the transcriptional level. Piwi-interacting sncRNAs as well as repeat-associated small RNAs were identified for the first time in mature spermatozoa from a mammalian species. Human spermatozoa retain a multifaceted population of small non-coding RNAs. Their presence is consistent with the view that they may function to stabilize the genome as part of the confrontation and consolidation of the genomes at fertilization. Examination of sperm samples from 3 individuals, sequenced individually