Long-term Hyperandrogenemia and/or Western-style Diet in Rhesus Macaque Females Impairs Preimplantation Embryogenesis.

Hyperandrogenemia and obesity are common in women with polycystic ovary syndrome, but it is currently unclear how each alone or in combination contribute to reproductive dysfunction and female infertility. To distinguish the individual and combined effects of hyperandrogenemia and an obesogenic diet on ovarian function, prepubertal female rhesus macaques received a standard control (C) diet, testosterone (T) implants, an obesogenic Western-style diet (WSD), or both (T + WSD). After 5 to 6 years of treatment, the females underwent metabolic assessments and controlled ovarian stimulations. Follicular fluid (FF) was collected for steroid and cytokine analysis and the oocytes fertilized in vitro. Although the T + WSD females exhibited higher insulin resistance compared to the controls, there were no significant differences in metabolic parameters between treatments. Significantly higher concentrations of CXCL-10 were detected in the FF from the T group, but no significant differences in intrafollicular steroid levels were observed. Immunostaining of cleavage-stage embryos revealed multiple nuclear abnormalities in the T, WSD, and T + WSD groups. Single-cell DNA sequencing showed that while C embryos contained primarily euploid blastomeres, most cells in the other treatment groups were aneuploid. Despite yielding a higher number of mature oocytes, T + WSD treatment resulted in significantly reduced blastocyst formation rates compared to the T group. RNA sequencing analysis of individual blastocysts showed differential expression of genes involved in critical implantation processes between the C group and other treatments. Collectively, we show that long-term WSD consumption reduces the capacity of fertilized oocytes to develop into blastocysts and that the addition of T further impacts gene expression and embryogenesis.

Metabolomics analysis of follicular fluid coupled with oocyte aspiration reveals importance of glucocorticoids in primate periovulatory follicle competency.

Gonadotropin administration during infertility treatment stimulates the growth and development of multiple ovarian follicles, yielding heterogeneous oocytes with variable capacity for fertilization, cleavage, and blastocyst formation. To determine how the intrafollicular environment affects oocyte competency, 74 individual rhesus macaque follicles were aspirated and the corresponding oocytes classified as failed to cleave, cleaved but arrested prior to blastulation, or those that formed blastocysts following in vitro fertilization. Metabolomics analysis of the follicular fluid (FF) identified 60 unique metabolites that were significantly different between embryo classifications, of which a notable increase in the intrafollicular ratio of cortisol to cortisone was observed in the blastocyst group. Immunolocalization of the glucocorticoid receptor (GR, NR3C1) revealed translocation from the cytoplasm to nucleus with oocyte maturation in vitro and, correlation to intrafollicular expression of the 11-hydroxy steroid dehydrogenases that interconvert these glucocorticoids was detected upon an ovulatory stimulus in vivo. While NR3C1 knockdown in oocytes had no effect on their maturation or fertilization, expansion of the associated cumulus granulosa cells was inhibited. Our findings indicate an important role for NR3C1 in the regulation of follicular processes via paracrine signaling. Further studies are required to define the means through which the FF cortisol:cortisone ratio determines oocyte competency.

Short-term Western-style diet negatively impacts reproductive outcomes in primates.

A maternal Western-style diet (WSD) is associated with poor reproductive outcomes, but whether this is from the diet itself or underlying metabolic dysfunction is unknown. Here, we performed a longitudinal study using regularly cycling female rhesus macaques (n = 10) that underwent 2 consecutive in vitro fertilization (IVF) cycles, one while consuming a low-fat diet and another 6-8 months after consuming a high-fat WSD. Metabolic data were collected from the females prior to each IVF cycle. Follicular fluid (FF) and oocytes were assessed for cytokine/steroid levels and IVF potential, respectively. Although transition to a WSD led to weight gain and increased body fat, no difference in insulin levels was observed. A significant decrease in IL-1RA concentration and the ratio of cortisol/cortisone was detected in FF after WSD intake. Despite an increased probability of isolating mature oocytes, a 44% reduction in blastocyst number was observed with WSD consumption, and time-lapse imaging revealed delayed mitotic timing and multipolar divisions. RNA sequencing of blastocysts demonstrated dysregulation of genes involved in RNA binding, protein channel activity, mitochondrial function and pluripotency versus cell differentiation after WSD consumption. Thus, short-term WSD consumption promotes a proinflammatory intrafollicular microenvironment that is associated with impaired preimplantation development in the absence of large-scale metabolic changes.

Influenza virus infection causes global RNAPII termination defects.

Viral infection perturbs host cells and can be used to uncover regulatory mechanisms controlling cellular responses and susceptibility to infections. Using cell biological, biochemical, and genetic tools, we reveal that influenza A virus (IAV) infection induces global transcriptional defects at the 3' ends of active host genes and RNA polymerase II (RNAPII) run-through into extragenic regions. Deregulated RNAPII leads to expression of aberrant RNAs (3' extensions and host-gene fusions) that ultimately cause global transcriptional downregulation of physiological transcripts, an effect influencing antiviral response and virulence. This phenomenon occurs with multiple strains of IAV, is dependent on influenza NS1 protein, and can be modulated by SUMOylation of an intrinsically disordered region (IDR) of NS1 expressed by the 1918 pandemic IAV strain. Our data identify a strategy used by IAV to suppress host gene expression and indicate that polymorphisms in IDRs of viral proteins can affect the outcome of an infection.