Non-invasive evaluation of placental blood flow: lessons from animal models.

In human obstetrics, placental vascularisation impairment is frequent as well as linked to severe pathological events (preeclampsia and intrauterine growth restriction), and there is a need for reliable methods allowing non-invasive evaluation of placental blood flow. Uteroplacental vascularisation is complex, and animal models are essential for the technical development and safety assessment of these imaging tools for human clinical use; however, these techniques can also be applied in the veterinary context. This paper reviews how ultrasound-based imaging methods such as 2D and 3D Doppler can provide valuable insight for the exploration of placental blood flow both in humans and animals and how new approaches such as the use of ultrasound contrast agents or ultrafast Doppler may allow to discriminate between maternal (non-pulsatile) and foetal (pulsatile) blood flow in the placenta. Finally, functional magnetic resonance imaging could also be used to evaluate placental blood flow, as indicated by studies in animal models, but its safety in human pregnancy still requires to be confirmed.

Uterine wound healing after caesarean section: A systematic review.

The rate of caesarean section (CS) is increasing worldwide. Defects in uterine healing have a major gynaecological and obstetric impact (uterine rupture, caesarean scar defect, caesarean scar pregnancy, placenta accreta spectrum). The complex process of cellular uterine healing after surgery, and specifically after CS, remains poorly understood in contrast to skin wound healing. This literature review on uterine wound healing was mainly based on histological observations, particularly after CS. The primary objective of the review was to examine the effects of CS on uterine tissue at the cellular level, based on histological observations. The secondary objectives were to describe the biomechanical characteristics and the therapies used to improve scar tissue after CS. This review was performed using PRISMA criteria, and PubMed was the data source. The study included all clinical and animal model studies with CS and histological analysis of the uterine scar area (macroscopic, microscopic, immunohistochemical and biomechanical). Twenty studies were included: 10 human and 10 animal models. In total, 533 female humans and 511 female animals were included. Review articles, meeting abstracts, case series, case reports, and abstracts without access to full-text were excluded. The search was limited to studies published in English. No correlation was found between cutaneous and uterine healing. The histology of uterine scars is characterized by disorganized smooth muscle, fibrosis with collagen fibres and fewer endometrial glands. As for skin healing, the initial inflammation phase and mediation of some growth factors (particularly connective tissue growth factor, vascular endothelial growth factor, platelet-derived growth factor, tumour necrosis factor α and tumour necrosis factor ß) seem to be essential. This initial phase has an impact on the subsequent phases of proliferation and maturation. Collagen appears to play a key role in the initial granulation tissue to replace the loss of substance. Subsequent maturation of the scar tissue is essential, with a decrease in collagen and smooth muscle restoration. Unlike skin, the glandular structure of uterine tissue could be responsible for the relatively high incidence of healing defects. Uterine scar defects after CS are characterized by an atrophic disorganized endometrium with atypia and a fibroblastic highly collagenic stromal reaction. Concerning immunohistochemistry, one study found a decrease in tumour necrosis factor ß in uterine scar defects. No correlation was found between biomechanical characteristics (particularly uterine strength) and the presence of a collagenous scar after CS. Based on the findings of this review, an illustration of current understanding about uterine healing is provided. There is currently no validated prevention of caesarean scar defects. Various treatments to improve uterine healing after CS have been tested, and appeared to have good efficacy in animal studies: alpha lipoic acid, growth factors, collagen scaffolds and mesenchymal stem cells. Further prospective studies are needed.

PIPAC nebulizer: How to test the new devices in the market, expert recommendations.

Pressurized intraperitoneal aerosol chemotherapy, named PIPAC, is now used in many centers around the world and as an intraperitoneal drug delivery system for treatment of peritoneal carcinomatosis. Recently, many of us have encountered problems during PIPAC procedures due to changes in material and production features of the original PIPAC nebulizer. Concomitantly, new PIPAC nebulizers proposed by other manufacturers are being launched on the market; which claim that they are the same as the original device in delivering PIPAC. However, these new devices are all different in terms of materials, technical characteristics and costs. We have considered that, to maintain the acquired results of PIPAC, we must ensure that the new systems are equivalent. The characteristics deemed essential by the expert group are as follows: 1: The nebulizer must be able to create droplets through an injector pressure between 10 and 20 bars, 2: The mean droplet size must be 3 micrometers, with 95% of the droplets between 0 and 10 micrometers, 3: The diffusion angle must be 70 degrees, which is the minimum.

Pregnancy and placental development in horses: an update.

Horses have been domesticated by man and historical information mostly associates horses with men. Nowadays, however, horse riding is essentially by women. Women are also very much involved in equine sciences, with a large contribution to the understanding of fetoplacental development. While highlighting the work of female scientists, this review describes the recent advances in equine fetoplacental studies, focusing on data obtained by new generation sequencing and progress on the understanding of the role of placental progesterone metabolites throughout gestation. A second emphasis is made on fetal programming, a currently very active field, where the importance of maternal nutrition, mare management or the use of embryo technologies has been shown to induce long term effects in the offspring that might affect progeny's performance. Finally, new perspectives for the study of equine pregnancy are drawn, that will rely on new methodologies applied to molecular explorations and imaging.

Estimation of milk production in suckling mares and factors influencing their milk yield.

Research about mare's milk is mainly focused on quality and information about quantity is incomplete partly due to the lack of a consensus on the method of measuring milk yield. The live weight, body condition at foaling and age of mares are factors influencing milk yield. The influence of mare parity, however, remains unclear. Over a period of 2 years (2018-2019), milk yield was evaluated on 65 mares (51 multiparous and 13 primiparous). Mares and foals were kept in a group at pasture. One method of milk yield measurement and one proxy method were applied; milking and weight-suckle-weight (WSW), respectively. The procedure was performed at five timepoints during the lactation period (3-30-60-90 and 180 days) without repetition. The relevance of WSW was addressed by studying the correlation between the two methods on 23 individuals. Factors influencing milk yield, through milking data, were studied on 57 individuals. Data was divided into two subsets. The first was an explanatory matrix containing the live weight of mares 24 h after parturition, parity, age, year of lactation and foal gender. The second was a response matrix containing data from milking at the five timepoints of the lactation. A correlation was found (RV = 0.41) between milking and WSW at day 3, however no correlation was found for other timepoints (RV ≤ 0.15). The live weight of the mare 24 h after foaling, age and parity appeared to have a significant impact on milk production (P < 0.05). Thus, older or multiparous mares showed a higher milk yield than younger or primiparous mares. In addition, mares with a higher live weight after foaling produced more milk than those with a lower live weight. Overall, results can lead us to two main conclusions. First, the WSW method performed at five different timepoints of the lactation, but without repeated measurements, is not an efficient way to estimate the milk yield of mares. Secondly, results concerning the live weight and age of mares were in accordance with previous studies. The influence of parity was also highlighted, confirming trends showed by other authors. Age and parity are closely related in our population, making it difficult to differentially assess their effects. Being able to identify the impact of both factors independently would benefit several sectors of the horse industry from sport to mare milk producers.

Assessment of placental perfusion using contrast-enhanced ultrasound: A longitudinal study in pregnant rabbit.

OBJECTIVES: The main aim of the study was to quantitatively characterize placental perfusion using contrast-enhanced ultrasound in a longitudinal study performed in pregnant rabbits. As a secondary objective, we qualitatively assessed the placental perfusion. METHODS: Three pregnant rabbits were used. Contrast-enhanced ultrasound was performed twice for each dam, either on embryo development (ED) day 14, 21 or 28 of pregnancy, under general anesthesia. Both qualitative and quantitative analyses were performed based on ultrasound assessment. RESULTS: Altogether, data from 29 fetuses were analyzed: 10 at ED-14; 12 at ED-21, and 7 at ED-28. Placental vascularization was mainly peripheral at the earliest gestational age and became gradually homogeneous within the whole placenta as pregnancy progressed. Intensity parameters and relative blood flows significantly increased with gestational age. The wash-in and wash-out rates were correlated and a significant increase in wash-in compared to in wash-out rates was observed with increased gestational age. CONCLUSION: Contrast-enhanced ultrasound is feasible in animal models and allows qualitative and quantitative analysis of placental vascularization during pregnancy.

Correlation between uteroplacental three-dimensional power Doppler indices and true uterine blood flow: evaluation in a pregnant sheep model.

OBJECTIVES: Three-dimensional (3D) Doppler quantification within the uteroplacental unit could be of great help in understanding and screening for pre-eclampsia and intrauterine growth restriction. Yet the correlation between 3D Doppler indices and true blood flow has not been confirmed in vivo. The aim of this study was to evaluate this correlation in a pregnant sheep model. METHODS: A blood flow quantitative sensor and a controllable vascular occlusion system were placed around the common uterine artery in seven sheep in late pregnancy, while all the other arterial supplies were ligated. Several occlusion levels were applied, from 0 to 100%, simultaneously with 3D Doppler acquisitions of several placentomes, using standardized settings. Each placentome was analyzed using VOCAL™ (Virtual Organ Computer-aided AnaLysis) software. The correlation between true blood flow and Doppler indices (vascularization index (VI), flow index (FI) and vascularization flow index (VFI)) was evaluated, together with measurement reproducibility. RESULTS: Forty-eight acquisitions were analyzed. All 3D Doppler indices were significantly correlated with true blood flow. Higher correlations were observed for VI and VFI (r = 0.81 (0.74-0.87), P < 0.0001 and r = 0.75 (0.67-0.82), P < 0.0001) compared with FI (r = 0.53 (0.38-0.64) P < 0.0001). Both intra- and interobserver reproducibility were high, with intraclass correlation coefficients of at least 0.799. CONCLUSION: This is the first in-vivo experimental study confirming a significant correlation between true blood perfusion and quantitative 3D Doppler indices measured within the uteroplacental unit. These results confirm the potential usefulness of 3D Doppler ultrasound for the assessment of placental vascular insufficiency both in clinical cases and in a research setting.

Radiofrequency ablation of retained placenta accreta after conservative management: preliminary evaluation in the pregnant ewe and in normal human placenta in vitro.

OBJECTIVE: To evaluate radiofrequency (RF) efficiency and safety for the ablation of retained placenta in humans, using a pregnant sheep model. DESIGN: Experimental study. SETTING: Laboratory of Surgery School, Nancy, France. POPULATION/SAMPLE: Three pregnant ewes/ten human placentas. METHODS: Various RF procedures were tested in pregnant ewes on 50 placentomes (individual placental units). Reproducibility of the best procedure was then evaluated in a further 20 placentomes and on ten human term placentas in vitro after delivery. MAIN OUTCOME MEASURES: Placental tissues destruction, lesions' size, myometrial lesions. RESULTS: Low power (100 W) and low target temperatures (60 degrees C) lead to homogenous tissue destruction, without myometrial lesion. No significant difference was observed in terms of lesion size and procedure duration for in the placentomes of pregnant ewe in vivo and in human placentas in vitro. The diameter of the ablation could be correlated with the tines deployment. CONCLUSION: The placental tissue structure is very permissive to RF energy, which suggests that RF could be used for the ablation of retained placenta, providing optimal control of tissue destruction. These results call for further experimental evaluations.

[Effects of omega 3 supplementation in pregnant women]. / Influence d'une supplémentation en omégas 3 au cours de la grossesse.

Maternal nutrition is a major environmental factor, which can be modified and can affect fetal growth and development with potential long-term consequences. There is currently a strong mediatic pressure for supplementing diets with omega 3 fatty acids. Nevertheless, if beneficial effects seem to be confirmed in adults and in animal models, the evidence for favourable effects of omega 3 supplementation in pregnant women are less obvious. Indeed, there is a trend showing a positive effect on cerebral development, but long term effects have not been demonstrated and both the quantity of omega 3 and the omega 3:omega 6 ratios are not precisely determined. Numerous studies are needed, both in pregnant animal models and in patients, to unravel these effects.

In utero characterisation of fetal growth by ultrasound scanning in the rabbit.

Fetal development is an important factor influencing the susceptibility of adults to metabolic diseases. In order to study the influence of fetal growth on further development in animal models like the rabbit, methods of measurement of fetal and placental size and viability must be established and validated. In this study, 42 New Zealand does bred naturally (N=12) or transferred with in vivo produced embryos (2, 4 or 6 embryos/doe) have been scanned every 2-3 days with a 7.5 MHz transabdominal probe from Day 7 post-coitum until term to measure fetal and placental growth. Vesicle, placental, fetal length and head size have thus been determined according to number of fetuses and time. In late gestation, the fetuses that were transferred in limited numbers to the uterus of does were significantly larger than their natural breeding counterparts probably due to reduced litter size.

[Maternal nutrition: effects on offspring fertility and importance of the periconceptional period on long-term development]. / Nutrition maternelle: incidence sur la fertilité de la descendance et importance de la période périconceptionelle pour le long terme.

The Developmental origins of human adult diseases (DOHAD) has initially emphasised the effects of maternal undernutrition during foetal development on long-term outcomes in the adult offspring, including effects on fertility. More recent work has provided evidence that preconceptional nutritional conditions and periconceptional environment also play a major role in programming the offspring susceptibility to disease. Epigenetic mechanisms, which may be mediated by macro- and micro-nutriments, endocrine status and oxidative stress, are the focus of the mechanistic studies aimed at understanding the processes involved in these effects. This article details available data in the area, using examples from numerous animal studies.

Review: Epigenetics, developmental programming and nutrition in herbivores.

Epidemiological studies in humans and animal models (including ruminants and horses) have highlighted the critical role of nutrition on developmental programming. Indeed, it has been demonstrated that the nutritional environment during the periconceptional period and foetal development can altered the postnatal performance of the resultant offspring. This nutritional programming can be exerted by maternal and paternal lineages and can affect offspring beyond the F1 generation. Alterations in epigenetic mechanisms have been proposed as the causative link behind the programming trajectories observed in the offspring. Although a clear cause-effect relationship between epigenetic modifications during early development and later offspring phenotype has not been demonstrated in livestock species, strong associations have been reported for some epigenetic marks (e.g. messenger RNA) that are worth exploring as possible predictors of future offspring phenotype. In this review, we shortly describe the main epigenetic mechanisms studied so far in mammals (i.e. mainly in the mouse) thought to be associated with developmental programming, and discuss the few studies available in mammalian herbivores (e.g. cattle) showing the effect of nutrition on epigenetic marks and the associated phenotype. Clearly, there is a need to develop research on nutritional strategies capable of modulating the epigenetic machinery with positive influence on the phenotype of livestock herbivores. This type of research is needed to alleviate the challenges currently faced by the livestock industry (e.g. impaired fertility of high-yielding dairy cows). This in turn will have a positive influence on animal welfare and productivity of livestock enterprises.

Placental structure and function in different breeds in horses.

Ponies and sometimes draft horses are often used as experimental models for horses although size and metabolic parameters are known to vary between horse breeds. So far, there is little information about differences of placental structure and no information about differences of placental function between breeds. The aim of this study was to investigate differences in placental size, structure and function at birth in relation to foal size and weight in ponies, Saddlebred and draft horses. Pony, Saddlebred and draft horse pregnancies were obtained by artificial insemination over 2 successive breeding seasons. Foals and total fetal membranes (TFM) were weighed and placentas measured for surface area at term. Placentas were sampled above the umbilical cord insertion. Surface density and volume fraction of the different cellular components of the placenta were measured on histological sections using stereology. The expression of genes involved in growth and development, nutrient transfer and vascularization was compared between groups. Foals and TFM were lighter at birth in ponies than Saddlebred horses, and both were lighter compared to draft horses. The surface density and volume fraction of microcotyledonary vessels was increased in pony compared to Saddlebred placentas. The relative expression of genes involved in growth and development was different between breeds and increased with maternal, fetal and placental weight. Primiparous dams produced lighter foals and smaller placentas, associated with a decreased volume fraction of microcotyledonary vessels and genes involved in growth and development and vascularization. Foal sex had little effect on placental structure and function as the expression of only one gene differed according to sex, with EGFR expression being decreased in placentas of females compared to males. In conclusion, foal and placental weight, as well as placental expression of genes involved in growth and development were correlated with maternal size. Placental structure also differed between breeds, with a stronger difference between ponies and both breeds of horses.

Placental alterations in structure and function in intra-uterine growth-retarded horses.

BACKGROUND: Following embryo transfer (ET), the size and breed of the recipient mare can affect fetal development and subsequent post natal growth rate and insulin sensitivity in foals. OBJECTIVES: To investigate placental adaptation in pregnancies where increased or restricted fetal growth was induced through ET between Pony, Saddlebred and Draught horses. STUDY DESIGN: In vivo experiment. METHODS: Control Pony (P, n = 21) and Saddlebred (S, n = 28) pregnancies were obtained by artificial insemination. Increased pregnancies were obtained by transferring Pony (P-D, n = 6) and Saddlebred (S-D, n = 8) embryos into Draught mares. Restricted pregnancies were obtained by transferring Saddlebred embryos into Pony mares (S-P, n = 6). Placental weight and surface were recorded and samples collected for stereology and analysis of expression of genes involved in placental growth, vascularisation and nutrient transport. Data were analysed by linear model. RESULTS: S-P foals were growth retarded when compared with controls despite increased gestational length. Placental weight was reduced but placental surface density and volume fraction were increased. Placental expression of genes involved in growth and development and nutrient transfer was strongly reduced. In contrast, placental size and weight were increased in enhanced growth P-D and S-D foals. The trophoblastic surface density and the allantoic vessels surface density were decreased in P-D and S-D, respectively, both with very few modifications in gene expression. MAIN LIMITATIONS: Control embryos were produced by artificial insemination whereas experimental embryos were produced by ET. CONCLUSIONS: Placental structure and gene expression are modified after ET into a smaller or larger breed than that of the embryo. These adaptations contribute to the observed phenotype of foal growth restriction or enhanced growth at birth.

Impact of a gestational exposure to diesel exhaust on offspring gonadal development: experimental study in the rabbit.

The aim of the present work was to address experimentally the possible impact of exposure to air pollution during gestation on the differentiation and function of the gonads of the offspring using a rabbit model. Rabbits were exposed daily to diluted diesel exhaust gas or filtered air from the 3rd until the 27th day of gestation, during which time germ cells migrate in genital ridges and divide, and fetal sex is determined. Offspring gonads were collected shortly before birth (28th day of gestation) or after puberty (7.5 months after birth). The structure of the gonads was analyzed by histological and immunohistological methods. Serum concentrations of testosterone and anti-Müllerian hormone were determined using ELISA. The morphology and the endocrine function of the gonads collected just at the arrest of the exposure were similar in polluted and control animals in both sexes. No differences were observed as well in gonads collected after puberty. Sperm was collected at the head of the epididymis in adults. Sperm motility and DNA fragmentation were measured. Among all parameters analyzed, only the sperm DNA fragmentation rate was increased three-fold in exposed males. Mechanisms responsible for these modifications and their physiological consequences are to be further clarified.

Maternal parity affects placental development, growth and metabolism of foals until 1 year and a half.

Primiparous mares are known to produce smaller foals than multiparous mares. This difference seems to be partly explained by the reduced exchange surface and volume of the placental villi in primiparous compared to multiparous placentas. The effect of maternal parity on foals' post-natal growth, metabolism and sexual maturation, however, has been given little consideration. The objectives of this work were to analyse placental biometry and structure at term, growth of foals and yearlings, their metabolism and testicular maturation at one year of age. Twenty multiparous mares (M), aged over 6 years and 12 primiparous mares (P), aged up to 5 years were artificially inseminated with the same stallion and monitored the same way until foaling. At birth, foals and placentas were measured and placentas were sampled above at the umbilical cord insertion, as well as in the pregnant and the non-pregnant horn to perform stereological analyses. Foals were weighed and measured until 540 days of age. At 120 and 360 days of age, an Intravenous Glucose Tolerance Test was performed on foals and yearlings. At 360 days of age, the males were castrated and testicular maturation analysed by RT-qPCR. At birth, P dams produced lighter and smaller foals and placentas. The foal birth weight to placental surface ratio was lower in the P compared to the M group. P Foals remained lighter than M foals until 360 days of age and smaller until at least 540 days of age. At 120 days of age, P foals had a higher glucose tolerance than M foals, and then may be less mature than M foals in terms of the control of their glucose homeostasis. At 360 days of age, the testicles of prepubertal P stallions were less mature in the P vs the M group. In conclusion, primiparous dams produce intrauterine growth restricted, less mature and smaller foals compared to multiparous dams with altered metabolism and growth until at least 540 days of age. These differences could affect the sport career of these foals, especially if it begins at an early age.

Placental function and structure at term is altered in broodmares fed with cereals from mid-gestation.

INTRODUCTION: Feeding pregnant broodmares with cereal concentrates has been shown to increase maternal insulin resistance and affect foal metabolism in the short and long-term. These effects are likely to be mediated by the placenta. Here, we investigated feto-placental biometry and placental structure and function at term in mares fed with or without cereals concentrates. MATERIAL AND METHODS: From 7 months of gestation, 22 multiparous mares were fed forage only (group F (n = 12)) or received forage and cracked barley (group B (n = 10)) until foaling. Foals and placentas were weighed and placental samples were collected above the umbilical cord insertion at birth. Placental histological structure was studied by stereology. A RNAseq analysis was performed on 9 placentas of each group. Enrichment of gene sets was analysed using the Gene Set Enrichment Analysis (GSEA) software using the KEGG and GO databases. RESULTS: No difference in feto-placental biometry was observed between groups. The volume of microcotyledonary vessels was decreased in B placentas and the vascular wall of allantoic arterioles was thickened. Gene sets involved in neutral amino acids, folate and anions transport and fatty acids, cholesterol and folate degradation were down-regulated while gene sets involved in RNA expression, inflammation and vascularisation were up-regulated in B placentas. CONCLUSION: Feeding pregnant mares with concentrates from mid-gestation alters the placental function and structure as observed in other species in cases of maternal insulin resistance.

Maternal obesity increases insulin resistance, low-grade inflammation and osteochondrosis lesions in foals and yearlings until 18 months of age.

INTRODUCTION: Obesity is a growing concern in horses. The effects of maternal obesity on maternal metabolism and low-grade inflammation during pregnancy, as well as offspring growth, metabolism, low-grade inflammation, testicular maturation and osteochondrotic lesions until 18 months of age were investigated. MATERIAL AND METHODS: Twenty-four mares were used and separated into two groups at insemination according to body condition score (BCS): Normal (N, n = 10, BCS ≤4) and Obese (O, n = 14, BCS ≥4.25). BCS and plasma glucose, insulin, triglyceride, urea, non-esterified fatty acid, serum amyloid A (SAA), leptin and adiponectin concentrations were monitored throughout gestation. At 300 days of gestation, a Frequently Sampled Intravenous Glucose Tolerance Test (FSIGT) was performed. After parturition, foals' weight and size were monitored until 18 months of age with plasma SAA, leptin, adiponectin, triiodothyronine (T3), thyroxine (T4) and cortisol concentrations measured at regular intervals. At 6, 12 and 18 months of age, FSIGT and osteoarticular examinations were performed. Males were gelded at one year and expression of genes involved in testicular maturation analysed by RT-qPCR. RESULTS: Throughout the experiment, maternal BCS was higher in O versus N mares. During gestation, plasma urea and adiponectin were decreased and SAA and leptin increased in O versus N mares. O mares were also more insulin resistant than N mares with a higher glucose effectiveness. Postnatally, there was no difference in offspring growth between groups. Nevertheless, plasma SAA concentrations were increased in O versus N foals until 6 months, with O foals being consistently more insulin resistant with a higher glucose effectiveness. At 12 months of age, O foals were significantly more affected by osteochondrosis than N foals. All other parameters were not different between groups. CONCLUSION: In conclusion, maternal obesity altered metabolism and increased low-grade inflammation in both dams and foals. The risk of developing osteochondrosis at 12 months of age was also higher in foals born to obese dams.