Alteration of the embryonic microenvironment and sex-specific responses of the preimplantation embryo related to a maternal high-fat diet in the rabbit model.

The maternal metabolic environment can be detrimental to the health of the offspring. In a previous work, we showed that maternal high-fat (HH) feeding in rabbit induced sex-dependent metabolic adaptation in the fetus and led to metabolic syndrome in adult offspring. As early development representing a critical window of susceptibility, in the present work we aimed to explore the effects of the HH diet on the oocyte, preimplantation embryo and its microenvironment. In oocytes from females on HH diet, transcriptomic analysis revealed a weak modification in the content of transcripts mainly involved in meiosis and translational control. The effect of maternal HH diet on the embryonic microenvironment was investigated by identifying the metabolite composition of uterine and embryonic fluids collected in vivo by biomicroscopy. Metabolomic analysis revealed differences in the HH uterine fluid surrounding the embryo, with increased pyruvate concentration. Within the blastocoelic fluid, metabolomic profiles showed decreased glucose and alanine concentrations. In addition, the blastocyst transcriptome showed under-expression of genes and pathways involved in lipid, glucose and amino acid transport and metabolism, most pronounced in female embryos. This work demonstrates that the maternal HH diet disrupts the in vivo composition of the embryonic microenvironment, where the presence of nutrients is increased. In contrast to this nutrient-rich environment, the embryo presents a decrease in nutrient sensing and metabolism suggesting a potential protective process. In addition, this work identifies a very early sex-specific response to the maternal HH diet, from the blastocyst stage.

Prenatal air pollution exposure to diesel exhaust induces cardiometabolic disorders in adulthood in a sex-specific manner.

BACKGROUND: Results from observational and experimental studies indicate that exposure to air pollutants during gestation reduces birth weight, whereas little is known on potential cardiometabolic consequences for the offspring at adulthood. OBJECTIVES: Our aim was to evaluate the long-term effects of gestational exposure to diesel engine exhaust (DE) on adult offspring phenotype in a rabbit model. METHODS: The protocol was designed to mimic human exposure in large European cities. Females rabbits were exposed to diluted (1 mg/m3) DE (exposed, n = 9) or clean air (controls, n = 7), from 3 days after mating, 2 h/d and 5 d/wk in a nose-only inhalation system throughout gestation (gestation days 3-27). After birth and weaning, 72 offspring (47 exposed and 25 controls) were raised until adulthood (7.5 months) to evaluate their cardio-metabolic status, including the monitoring of body weight and food intake, fasting biochemistry, body composition (iDXA), cardiovascular parameters and glucose tolerance. After a metabolic challenge (high fat diet in males and gestation in females), animals were euthanized for postmortem phenotyping. RESULTS: Sex-specific responses to maternal exposure were observed in adult offspring. Age-related increases in blood pressure (p = 0.058), glycaemia (p = 0.029), and perirenal fat mass (p = 0.026) as well as reductions in HDL-cholesterol (p = 0.025) and fat-to-body weight ratio (p = 0.011) were observed in exposed males, suggesting a metabolic syndrome. Almost only trends were observed in exposed females with higher triglycerides and decreased bone density compared to control females. Metabolic challenges triggered or amplified some biological responses, especially in females. CONCLUSIONS: In utero exposure to air pollution predisposed rabbit offspring to cardiometabolic disorders in a sex-specific manner.

Metabolomic differences in blastocoel and uterine fluids collected in vivo by ultrasound biomicroscopy on rabbit embryos†.

The success of embryo development and implantation depends in part on the environment in which the embryo evolves. However, the composition of the uterine fluid surrounding the embryo in the peri-implantation period remains poorly studied. In this work, we aimed to develop a new strategy to visualize, collect, and analyze both blastocoelic liquid and juxta-embryonic uterine fluid from in vivo peri-implantation rabbit embryos. Using high-resolution ultrasound biomicroscopy, embryos were observed as fluid-filled anechoic vesicles, some of which were surrounded by a thin layer of uterine fluid. Ultrasound-guided puncture and aspiration of both the blastocoelic fluid contained in the embryo and the uterine fluid in the vicinity of the embryo were performed. Using nuclear magnetic resonance spectroscopy, altogether 24 metabolites were identified and quantified, of which 21 were detected in both fluids with a higher concentration in the uterus compared to the blastocoel. In contrast, pyruvate was detected at a higher concentration in blastocoelic compared to uterine fluid. Two acidic amino acids, glutamate and aspartate, were not detected in uterine fluid in contrast to blastocoelic fluid, suggesting a local regulation of uterine fluid composition. To our knowledge, this is the first report of simultaneous analysis of blastocoelic and uterine fluids collected in vivo at the time of implantation in mammals, shedding new insight for understanding the relationship between the embryo and its local environment at this critical period of development.

Maternal exposure to diluted diesel engine exhaust alters placental function and induces intergenerational effects in rabbits.

BACKGROUND: Airborne pollution is a rising concern in urban areas. Epidemiological studies in humans and animal experiments using rodent models indicate that gestational exposure to airborne pollution, in particular diesel engine exhaust (DE), reduces birth weight, but effects depend on exposure duration, gestational window and nanoparticle (NP) concentration. Our aim was to evaluate the effects of gestational exposure to diluted DE on feto-placental development in a rabbit model. Pregnant females were exposed to diluted (1 mg/m(3)), filtered DE (NP diameter ≈ 69 nm) or clean air (controls) for 2 h/day, 5 days/week by nose-only exposure (total exposure: 20 days in a 31-day gestation). RESULTS: DE exposure induced early signs of growth retardation at mid gestation with decreased head length (p = 0.04) and umbilical pulse (p = 0.018). Near term, fetal head length (p = 0.029) and plasma insulin and IGF1 concentrations (p = 0.05 and p = 0.019) were reduced. Placental function was also affected, with reduced placental efficiency (fetal/placental weight) (p = 0.049), decreased placental blood flow (p = 0.009) and fetal vessel volume (p = 0.002). Non-aggregated and "fingerprint" NP were observed at various locations, in maternal blood space, in trophoblastic cells and in the fetal blood, demonstrating transplacental transfer. Adult female offspring were bred with control males. Although fetoplacental biometry was not affected near term, second generation fetal metabolism was modified by grand-dam exposure with decreased plasma cholesterol (p = 0.008) and increased triglyceride concentrations (p = 0.015). CONCLUSIONS: Repeated daily gestational exposure to DE at levels close to urban pollution can affect feto-placental development in the first and second generation.

In Vivo Evaluation of Cervical Stiffness Evolution during Induced Ripening Using Shear Wave Elastography, Histology and 2 Photon Excitation Microscopy: Insight from an Animal Model.

Prematurity affects 11% of the births and is the main cause of infant mortality. On the opposite case, the failure of induction of parturition in the case of delayed spontaneous birth is associated with fetal suffering. Both conditions are associated with precocious and/or delayed cervical ripening. Quantitative and objective information about the temporal evolution of the cervical ripening may provide a complementary method to identify cases at risk of preterm delivery and to assess the likelihood of successful induction of labour. In this study, the cervical stiffness was measured in vivo in pregnant sheep by using Shear Wave Elastography (SWE). This technique assesses the stiffness of tissue through the measurement of shear waves speed (SWS). In the present study, 9 pregnant ewes were used. Cervical ripening was induced at 127 days of pregnancy (term: 145 days) by dexamethasone injection in 5 animals, while 4 animals were used as control. Elastographic images of the cervix were obtained by two independent operators every 4 hours during 24 hours after injection to monitor the cervical maturation induced by the dexamethasone. Based on the measurements of SWS during vaginal ultrasound examination, the stiffness in the second ring of the cervix was quantified over a circular region of interest of 5 mm diameter. SWS was found to decrease significantly in the first 4-8 hours after dexamethasone compared to controls, which was associated with cervical ripening induced by dexamethasone (from 1.779 m/s ± 0.548 m/s, p < 0.0005, to 1.291 m/s ± 0.516 m/s, p < 0.000). Consequently a drop in the cervical elasticity was quantified too (from 9.5 kPa ± 0.9 kPa, p < 0.0005, to 5.0 kPa ± 0.8 kPa, p < 0.000). Moreover, SWE measurements were highly reproducible between both operators at all times. Cervical ripening induced by dexamethasone was confirmed by the significant increase in maternal plasma Prostaglandin E2 (PGE2), as evidenced by the assay of its metabolite PGEM. Histological analyses and two-photon excitation microscopy, combining both Second Harmonic Generation (SHG) and Two-photon Fluorescence microscopy (2PF) contrasts, were used to investigate, at the microscopic scale, the structure of cervical tissue. Results show that both collagen and 2PF-active fibrillar structures could be closely related to the mechanical properties of cervical tissue that are perceptible in elastography. In conclusion, SWE may be a valuable method to objectively quantify the cervical stiffness and as a complementary diagnostic tool for preterm birth and for labour induction success.

Sex and breed-dependent organ development and metabolic responses in foetuses from lean and obese/leptin resistant swine.

The present study aimed to determine the effects of breed and sex on growth patterns and metabolic features of advanced-pregnancy foetuses exposed to the same environmental conditions. Thus, at Day 62 of pregnancy, swine foetuses from an obese breed with leptin resistance (Iberian breed) were compared to lean crossbred foetuses (25% Large White ×25% Landrace ×50% Pietrain). There were differential developmental patterns in foetuses with leptin resistance, mainly a higher relative weight of the brain resembling "brain-sparing effect". Prioritization of brain growth may be protective for the adequate growth and postnatal survival of the Iberian individuals, an ancient breed reared in extensive semi-feral conditions for centuries. There were also clear sex-related differences in foetal development and metabolism in the Iberian breed. Female Iberian foetuses were similar in size and weight to male littermates but had a significantly higher relative liver to body weight ratio resembling "liver-sparing effect" and a trend for a higher relative intestine to body ratio. Moreover, the availability of triglycerides, cholesterol and IL-6 in female Iberian foetuses was similar to that of lean crossbred foetuses. Overall, these features may favour a better postnatal survival and development of females, the sex more critical for the species survival. These findings set the basis for future translational studies aimed at increasing the knowledge on the interaction between genetic and environmental factors in the early programming of the adult phenotype.