Project description:In order to establish an obese mouse model, female mice were continuously fed with a high-fat diet (HFD) or a normal diet (control) for 16 weeks beginning at three weeks of age. In this paper, these mice are termed ‘HFD mice’ and ‘control mice’, respectively. Accordingly, we call their oocytes ‘HFD oocytes’ and ‘control oocytes’. Substantial evidence indicates that the effects of maternal obesity on embryo/offspring development can be attributed to factors within the oocyte (9). To identify such potential effectors, we performed a comparative proteomic analysis of ovulated MII oocytes from control and HFD mice.
Project description:The cumulus cells (CCs) that envelope antral and ovulated oocytes have been regarded as putative source of non-invasive markers of the oocyte developmental competence and a number of studies have observed a correlation between CCs gene expression, embryo quality and final pregnancy outcome. With the aim of identifying marker transcripts, we isolated CCs from antral mouse oocytes of known developmental incompetence (NSN-CCs) or competence (SN-CCs) and compared their transcriptomes.
Project description:The cumulus cells (CCs) that envelope antral and ovulated oocytes have been regarded as putative source of non-invasive markers of the oocyte developmental competence and a number of studies have observed a correlation between CCs gene expression, embryo quality and final pregnancy outcome. With the aim of identifying marker transcripts, we isolated CCs from antral mouse oocytes of known developmental incompetence (NSN-CCs) or competence (SN-CCs) and compared their transcriptomes. Total RNA was isolated from cumulus cells derived from oocytes with a SN or NSN chromatin organization.
Project description:Chromosome aneuploidy increases in oocytes with maternal age, and is considered the leading cause for the increased incidence of infertility, miscarriage, and birth defects. Using mRNA-Sequencing of oocytes from 12 month old mouse versus 3 month young mouse, we identified a spindle assembly checkpoint gene, BubR1, whose expression was significantly decreased. We employed a mRNA microinjection based approach to increase BubR1 expression in aging oocytes. We find that increased expression of BubR1 protects against aneuploidy and chromosome misalignment in aging oocytes. After in vitro fertilization, the embryos derived from BubR1 increased expression aging oocytes exhibited chromosome stability as robust as those of the young ones. Furthermore, following embryo transfer, these embryos showed greatly improved developmental competency, with comparable levels of full-term development to those of the young ones. These results indicate that the decline in oocyte quality may be reversible and could lead to treatments that prolong female fertility. Examination of the effect of maternal aging on the mRNA expression in the mature oocytes of the female mice. Naturally ovulated mature oocytes (MII stage) were collected from 6 young (3 month) and 6 aging (12 month) female mice (3 oocytes per mice, 18 oocytes for each group).
Project description:The success of human reproduction relies on high quality oocytes. Oocyte quality is manifested by the competence to complete meiosis, to be fertilized, and to support embryonic development. This meiotic and developmental competence is gradually established during the course of oocyte and follicle development, and is determined in large part by the autonomous gene expression program intrinsic to the oocyte. In order to explore the regulatory role of LSM14B in oocyte, we analyzed the effect of Lsm14b KO on gene expression in GV-stage fully-grown oocytes (FGOs) in mice by comparing the corresponding transcriptomes via RNA-Seq Analysis.
Project description:The goal of this study is to identify the differentially expressed genes in Zp3-Cre mediated Mtor oocyte-specific knockout (CKO) Metaphase II (MII) oocytes by comparing the transcriptomes of Wild-Type (WT) and CKO Mouse MII oocytes via RNA-Seq Analysis. Methods: mRNA profiles of 3 sets of super-ovulated MII oocytes collected from 24-day old WT- and Mtor CKO female mice were generated by deep sequencing using an Illumina Hiseq2500 platform with 41bp single read. All RNA-seq reads passed filter were trimmed to remove low-quality bases and adaptor sequences. Reads then were aligned to the mm10 reference genome using tophat2 (v2.0.8b). Reads aligned to genes are counted by cufflinks (v2.1.1). The FPKMs are normalized usingcuffnorm. Differentially expressed genes are calculated using cuffdiff.
Project description:To investigate the effect of mitochondrial fission factor (MFF) on oocyte competence and female fertility using an oocyte-specific Mff knockout mouse model. Oocyte-specific Mff deficiency hindered oocyte maturation and early embryonic development via regulation of mitochondrial dynamics, resulting in female subfertility. What is known already: Emerging studies have demonstrated that mitochondrial dynamics play a vital role in oocyte maturation and early embryo development. MFF is one of the crucial proteins regulating mitochondrial dynamics. Our previous studies have indicated that Mff is necessary for oocyte competence and fertility using a global Mff knockout mouse model. However, further investigation is needed to determine whether the deletion of Mff in oocytes could affect oocyte competence and early embryonic development by regulating mitochondrial dynamics.The Mfffl/fl; Gdf9-Cre mice consistently exhibited reduced fertility with a significant decrease in litter size, number of litters per female, and number of pups per female compared to the Mfffl/fl mice. The numbers of GV oocytes, ovulated oocytes and 2-cell embryos were significantly lower in the Mfffl/fl; Gdf9-Cre mice. The blastocyst (32.32% vs. 68.44%, p<0.05) embryo development rate per 2-cell embryos was also significantly lower. In vitro maturation revealed that of Mfffl/fl; Gdf9-Cre GV oocytes had significantly lower rates of GV break-down and polar body extrusion . The ratios of aberrant spindle and misaligned chromosomesin ovulated oocytes from Mfffl/fl; Gdf9-Cre mice were significantly increased. In terms of mitochondrial dynamics, the mitochondrial size and the ratio of abnormal mitochondrial distribution (unevenly aggregated) in Mfffl/fl; Gdf9-Cre oocytes significantly elevated. This study found that the Mff played a crucial role in oocyte maturation and early embryonic development. These results shed light on the important role played by MFF in ensuring proper mitochondrial dynamics in oocytes and underscore the significance of this protein in female fertility. This study provides insights into potential therapeutic targets for infertility and related diseases in humans.
Project description:After the library was qualified, the library was pooled according to the effective concentration and the demand of target offline data, and was sequenced by Illumina platform. Since PABPN1L can bind the poly(A) tail, we propose that it participates in the process of post-transcriptional regulation and degradation of maternal mRNA. We investigated the mRNA changes in GV oocytes, MII oocytes, and fertilized eggs using RNA-seq. GV oocytes, MII oocytes and zygotes were collected and mixed from three mice of each genotypes separately.
Project description:Once ovulated, the oocyte has to be fertilized in a short time window, or it will undergo post-ovulation aging (POA), whose underlying mechanisms are still not elucidated. Here, we optimized single-cell proteomics methods and performed single-cell proteomic and phosphoproteomic analysis of fresh, POA, melatonin-treated and MG132-treated POA oocytes.