Expression and localization of NRF2/Keap1 signalling pathway genes in mouse preimplantation embryos exposed to free fatty acids.

Obese women experience greater incidence of infertility, with reproductive tracts exposing preimplantation embryos to elevated free fatty acids (FFA) such as palmitic acid (PA) and oleic acid (OA). PA treatment impairs mouse preimplantation development in vitro, while OA co-treatment rescues blastocyst development of PA treated embryos. In the present study, we investigated the effects of PA and OA treatment on NRF2/Keap1 localization, and relative antioxidant enzyme (Glutathione peroxidase; Gpx1, Catalase; Cat, Superoxide dismutase; Sod1 and γ-Glutamylcysteine ligase catalytic unit; Gclc) mRNA levels, during in vitro mouse preimplantation embryo development. Female mice were superovulated, mated, and embryos cultured in the presence of bovine Serum albumin (BSA) control or PA, or OA, alone (each at 100 µM) or PA + OA combined (each at 100 µM) treatment. NRF2 displayed nuclear localization at all developmental stages, whereas Keap1 primarily displayed cytoplasmic localization throughout control mouse preimplantation development in vitro. Relative transcript levels of Nrf2, Keap1, and downstream antioxidants significantly increased throughout control mouse preimplantation development in vitro. PA treatment significantly decreased blastocyst development and the levels of nuclear NRF2, while OA and PA + OA treatments did not. PA and OA treatments did not impact relative mRNA levels of Nrf2, Keap1, Gpx1, Cat, Sod1 or Gclc. Our outcomes demonstrate that cultured mouse embryos display nuclear NRF2, but that PA treatment reduces nuclear NRF2 and thus likely impacts NRF2/KEAP1 stress response mechanisms. Further studies should investigate whether free fatty acid effects on NRF2/KEAP1 contribute to the reduced fertility displayed by obese patients.

Effects of palmitic acid on localization of embryo cell fate and blastocyst formation gene products.

As obese and overweight patients commonly display hyperlipidemia and are increasingly accessing fertility clinics for their conception needs, our studies are directed at understanding the effects of hyperlipidemia on early pregnancy. We have focused on investigating palmitic acid (PA) and oleic acid (OA) treatment alone and in combination from the mouse two-cell stage embryos as a model for understanding their effects on the mammalian preimplantation embryo. We recently reported that PA exerts a negative effect on mouse two-cell progression to the blastocyst stage, whereas OA co-treatment reverses that negative effect. In the present study, we hypothesized that PA treatment of mouse embryos would disrupt proper localization of cell fate determining and blastocyst formation gene products and that co-treatment with OA would reverse these effects. Our results demonstrate that PA treatment significantly (P < 0.05) reduces blastocyst development and cell number but did not prevent nuclear localization of YAP in outer cells. PA treatment significantly reduced the number of OCT4+ and CDX2+ nuclei. PA-treated embryos had lower expression of blastocyst formation proteins (E-cadherin, ZO-1 and Na/K-ATPase alpha1 subunit). Importantly, co-treatment of embryos with OA reversed PA-induced effects on blastocyst development and increased inner cell mass (ICM) and trophectoderm (TE) cell numbers and expression of blastocyst formation proteins. Our findings demonstrate that PA treatment does not impede cell fate gene localization but does disrupt proper blastocyst formation gene localization during mouse preimplantation development. OA treatment is protective and reverses PA's detrimental effects. The results advance our understanding of the impact of FFA exposure on mammalian preimplantation development.

Oleic Acid Counters Impaired Blastocyst Development Induced by Palmitic Acid During Mouse Preimplantation Development: Understanding Obesity-Related Declines in Fertility.

Obesity is associated with altered fatty acid profiles, reduced fertility, and assisted reproductive technology (ART) success. The effects of palmitic acid (PA), oleic acid (OA), and their combination on mouse preimplantation development, endoplasmic reticulum (ER) stress pathway gene expression, lipid droplet formation, and mitochondrial reactive oxygen species (ROS) were characterized. Two-cell stage mouse embryos collected from superovulated and mated CD1 females were placed into culture with KSOMaa medium, or PA alone or in combination with OA for 46 h. PA significantly reduced blastocyst development in a concentration-dependent manner, which was prevented by co-treatment with OA. PA and OA levels in mouse reproductive tracts were assessed by liquid chromatography coupled to mass spectrometry (LC-MS). LC-MS indicated higher concentrations of PA in the mouse oviduct than the uterus. Transcript analysis revealed that PA alone groups had increased ER stress pathway (ATF3, CHOP, and XBP1 splicing) mRNAs, which was alleviated by OA co-treatment. OA co-treatment significantly increased lipid droplet accumulation and significantly decreased mitochondrial ROS from PA treatment alone. PA treatment for only 24 h significantly reduced its impact on blastocyst development from the 2-cell stage. Thus, PA affects ER stress pathway gene expression, lipid droplet accumulation, and mitochondrial ROS in treated preimplantation embryos. These mechanisms may serve to offset free fatty acid exposure effects on preimplantation development, but their protective ability may be overwhelmed by elevated PA.

Venous Gas Embolism during Hysteroscopic Endometrial Ablation: Report of 5 Cases and Review of the Literature.

STUDY OBJECTIVE: To highlight the circumstances, presentation, and treatment of venous gas embolism (VGE) and provide guidance and propose potential changes in surgical practice and perioperative monitoring to minimize the adverse consequences and sequalae of this potentially serious complication. DESIGN: A case series. SETTING: A university-affiliated teaching hospital. PATIENTS: Five women developed VGE during hysteroscopic endometrial ablation. INTERVENTIONS: From 1990 through 2014, the principle author (G.A.V.) performed 5249 primary and 458 repeat hysteroscopic endometrial ablations under general anesthesia using a monopolar 26F (9-mm) resectoscope connected to a peristaltic pump-driven active inflow and outflow irrigation and distension system (1.5% glycine) and an 8-mm monopolar loop electrode at a 120-W continuous (cut) and/or a 3- to 5-mm rollerball interrupted (coagulation) waveform or a combination of them. MEASUREMENTS AND MAIN RESULTS: Among 5707 procedures, we encountered 5 (0.09%, 1/1140) incidents of VGE during primary ablations. All patients exhibited the same symptoms of ventilatory and hemodynamic decompensation, beginning with a reduction in end-tidal carbon dioxide and arterial oxygen desaturation. All patients recovered after immediate cessation of the surgery and resuscitation including ventilatory support with 100% O2 and intravenous fluids. CONCLUSIONS: Although entrainment of some air/gas bubbles is common during hysteroscopy, life-threatening/fatal VGE is rare (1/1140 cases). Situational awareness and strict adherence to certain principles including understanding the conditions, prerequisites, and pathophysiology of VGE; attention to surgical principles and operative technique; close communication with the anesthesiologist; and early therapeutic intervention are of paramount importance to avoid this rare but potentially serious complication.