DNA Content in Embryonic Extracellular Vesicles Is Independent of the Apoptotic Rate in Bovine Embryos Produced In Vitro.

Pre-implantation embryos release extracellular vesicles containing different molecules, including DNA. The presence of embryonic DNA in E-EVs released into the culture medium during in vitro embryo production could be useful for genetic diagnosis. However, the vesicles containing DNA might be derived from embryos suffering from apoptosis, i.e., embryos of bad quality. This work intended to confirm that embryos release DNA that is useful for genotyping by evaluating the effect of embryonic apoptosis on DNA content in E-EVs. Bovine embryos were produced by parthenogenesis and in vitro fertilization (IVF). On Day 5, morulae were transferred to individual cultures in an EV-depleted SOF medium. On Day 7, embryos were used to evaluate cellular apoptosis, and each culture medium was collected to evaluate E-EV concentration, characterization, and DNA quantification. While no effect of the origin of the embryo on the apoptotic rate was found, arrested morulae had a higher apoptotic rate. E-EVs containing DNA were identified in all samples, and the concentration of those vesicles was not affected by the origin or quality of the embryos. However, the concentration of DNA was higher in EVs released by the arrested parthenogenetic embryos. There was a correlation between the concentration of E-EVs, the concentration of DNA-positive E-EVs, and the concentration of DNA. There was no negative effect of apoptotic rate on DNA-positive E-EVs and DNA concentration; however, embryos of the best quality with a low apoptotic rate still released EVs containing DNA. This study confirms that the presence of DNA in E-EVs is independent of embryo quality. Therefore, E-EVs could be used in liquid biopsy for noninvasive genetic diagnosis.

Disruption of the Mouse Blood-Brain Barrier by Small Extracellular Vesicles from Hypoxic Human Placentas.

Cerebrovascular complications, including cerebral edema and ischemic and hemorrhagic stroke, constitute the leading cause of maternal mortality associated with preeclampsia. The underlying mechanisms of these cerebrovascular complications remain unclear. However, they are linked to placental dysfunction and blood-brain barrier (BBB) disruption. Nevertheless, the connection between these two distant organs is still being determined. Increasing evidence suggests that the placenta releases signaling molecules, including extracellular vesicles, into maternal circulation. Extracellular vesicles are categorized according to their size, with small extracellular vesicles (sEVs smaller than 200 nm in diameter) considered critical signaling particles in both physiological and pathological conditions. In preeclampsia, there is an increased number of circulating sEVs in maternal circulation, the signaling function of which is not well understood. Placental sEVs released in preeclampsia or from normal pregnancy placentas exposed to hypoxia induce brain endothelial dysfunction and disruption of the BBB. In this protocol, we assess whether sEVs isolated from placental explants cultured under hypoxic conditions (modeling one aspect of preeclampsia) disrupt the BBB in vivo.

Reduced Brain Cortex Angiogenesis in the Offspring of the Preeclampsia-Like Syndrome.

BACKGROUND: Children from pregnancies affected by preeclampsia have an increased risk of cognitive and behavioral alterations via unknown pathophysiology. We tested the hypothesis that preeclampsia generated reduced brain cortex angiogenesis in the offspring. METHODS: The preeclampsia-like syndrome (PELS) mouse model was generated by administering the nitric oxide inhibitor NG-nitroarginine methyl ester hydrochloride. Confirmatory experiments were done using 2 additional PELS models. While in vitro analysis used mice and human brain endothelial cells exposed to serum of postnatal day 5 pups or umbilical plasma from preeclamptic pregnancies, respectively. RESULTS: We report significant reduction in the area occupied by blood vessels in the motor and somatosensory brain cortex of offspring (postnatal day 5) from PELS compared with uncomplicated control offspring. These data were confirmed using 2 additional PELS models. Furthermore, circulating levels of critical proangiogenic factors, VEGF (vascular endothelial growth factor), and PlGF (placental growth factor) were lower in postnatal day 5 PELS. Also we found lower VEGF receptor 2 (KDR [kinase insert domain-containing receptor]) levels in mice and human endothelial cells exposed to the serum of postnatal day 5 PELS or fetal plasma of preeclamptic pregnancies, respectively. These changes were associated with lower in vitro angiogenic capacity, diminished cell migration, larger F-actin filaments, lower number of filopodia, and lower protein levels of F-actin polymerization regulators in brain endothelial cells exposed to serum or fetal plasma of offspring from preeclampsia. CONCLUSIONS: Offspring from preeclampsia exhibited diminished brain cortex angiogenesis, associated with lower circulating VEGF/PlGF/KDR protein levels, impaired brain endothelial migration, and dysfunctional assembly of F-actin filaments. These alterations may predispose to structural and functional alterations in long-term brain development.

Brain Vascular Dysfunction in Mothers and Their Children Exposed to Preeclampsia.

Preeclampsia is a maternal syndrome characterized by the new onset of hypertension and proteinuria after 20 weeks of gestation associated with multisystemic complications, including brain alterations. Indeed, brain complications associated with preeclampsia are the leading direct causes of fetal and maternal morbidity and mortality, especially in low- and middle-income countries. In addition to the well-recognized long-term adverse cardiovascular effects of preeclampsia, women who have had preeclampsia have higher risk of stroke, dementia, intracerebral white matter lesions, epilepsy, and perhaps also cognitive decline postpartum. Furthermore, increasing evidence has also associated preeclampsia with similar cognitive and cerebral disorders in the offspring. However, the mechanistic links between these associations remain unresolved. This article summarizes the current knowledge about the cerebrovascular complications elicited by preeclampsia and the potential pathophysiological mechanisms involved, emphasizing the impaired brain vascular function in the mother and their offspring.

Synthesis and Characterisation of ASA-PEEK Composites for Fused Filament Fabrication.

In this paper, a series of polymer composites made from acrylonitrile-styrene-acrylate (ASA) and poly (ether ether ketone) (PEEK) were manufactured. ASA acts as a polymer matrix while PEEK is loaded in the form of micro-particles that act as a reinforcing filler. The composites were compounded by single screw extrusion and then, different specimens were manufactured either via injection moulding (IM) or fused filament fabrication (FFF). Two different types of PEEK (commercial and reused) in different concentrations (3 and 6 wt.%) were tested and their influence in the mechanical, structural, and thermal properties were studied. It was observed that reused PEEK enhanced the stiffness and tensile strength and thermal stability of the composites both, for injected and printed specimens. This evidences the suitability of these composites as potential candidates as novel materials with enhanced properties following an approach of circular economy.

Are Anthelminthic Treatments of Captive Ruminants Necessary?

Anthelmintics are frequently administered to animals to limit fecal egg elimination, so that wild animals in captive breeding programs are treated to maintain a proper health condition. This is effective from a health management perspective, but on the other hand, it could prevent captive animals from developing an effective immunity against parasites that they might encounter when reintroduced into their original geographic areas. The aim of this study was to describe the dynamics of parasite infections in captive Cuvier's gazelles (Gazella cuvieri) not treated with anthelmintics for two years and to evaluate the factors related to their fecal egg shedding. Fifteen one-year-old males were enclosed together and captured monthly to collect feces directly from the rectum. Fecal egg counts were performed, and eggs were classified as strongylid-like, Nematodirus sp., or Trichuris sp. Fecal egg shedding for the three groups of parasites did not vary significantly over the duration of the study. Only precipitation affected the egg-shedding pattern of all parasites, while inbreeding was positively associated with the number of strongylid-like parasites. These findings suggest an equilibrium between hosts and parasites in absence of treatment during the study. The anthelmintic treatment as a systematic prophylaxis method in captive animals should be avoided and replaced by systematic coprological and clinical vigilance, as well as targeted treatment in the case of a significant rise of fecal egg counts.

Zoological institutions as hotspots of gastrointestinal parasites that may affect the success of ungulate reintroduction programmes.

BACKGROUND: Ungulates from zoological institutions are frequently used as founders in reintroduction programmes. These animals are subject to specific parasite management as parasitic infections have previously been associated with failed Bovidae reintroductions. METHODS: Questionnaires to obtain data on how these institutions screen for seasonal parasite presence and the clinical signs they induced in threatened ungulates were sent to 65 institutions involved in European Ex situ Programmes (58.5% response rate). Temperature and relative humidity data were also obtained to categorize each zoological centre. RESULTS: Strongyloides spp. (52.6%), Trichuris spp. (42.1%), Trichostrongylidae family (39.4%) and Eimeria spp. (36.8%) were the most frequently reported parasites in the received questionnaires. Climatic variables did not influence parasite presence. CONCLUSION: Our results suggest that artificial microenvironments created by husbandry practices and enclosure design in zoos could create hotspots for gastrointestinal parasites. To maximise the success of reintroduction projects, we recommend that the influence of microclimates on parasite burdens be evaluated.

Survey of husbandry practices for bovidae in zoos: the importance of parasite management for reintroduction programmes.

Animals from zoological institutions may be used for reintroductions. These individuals are considered healthy, but they are not necessarily free of parasites, despite the minimum husbandry standards required of zoological institutions as described in the European Association of Zoos and Aquaria guidelines. In this sense, parasitism has been identified as the cause of failure, or has added difficulties, in some reintroduction programmes. Here the authors attempt to summarise the risk of parasitism to animals originating from zoological institutions by analysing a questionnaire about parasite prevalence, sampling methods, treatment and control in three ungulates in European zoos. Completed questionnaires were received from 38 institutions (58.5 per cent response rate). Most of the responding institutions (97 per cent) detected the eggs of endoparasites in faeces, but only one reported ectoparasites. Most institutions followed a similar preventive schedule, with ivermectin as the preferred prophylactic treatment for parasites, commonly administered in food every six months. The frequent use of concentrating flotation techniques as the sole method to evaluate the presence of parasite eggs in faecal samples is not recommended because it fails to detect trematode and lung nematode infections, so it would be better to use flotation techniques together with sedimentation procedures or serological and molecular tests. The results suggest that parasite control in zoological institutions can be complicated, indicating the need to implement a specific management schedule for institutions involved in reintroduction projects.

Estimates of direct and indirect effects for early juvenile survival in captive populations maintained for conservation purposes: the case of Cuvier's gazelle.

Together with the avoidance of any negative impact of inbreeding, preservation of genetic variability for life-history traits that could undergo future selective pressure is a major issue in endangered species management programmes. However, most of these programmes ignore that, apart from the direct action of genes on such traits, parents, as contributors of offspring environment, can influence offspring performance through indirect parental effects (when parental genotype and phenotype exerts environmental influences on offspring phenotype independently of additive genetic effects). Using quantitative genetic models, we estimated the additive genetic variance for juvenile survival in a population of the endangered Cuvier's gazelle kept in captivity since 1975. The dataset analyzed included performance recording for 700 calves and a total pedigree of 740 individuals. Results indicated that in this population juvenile survival harbors significant additive genetic variance. The estimates of heritability obtained were in general moderate (0.115-0.457) and not affected by the inclusion of inbreeding in the models. Maternal genetic contribution to juvenile survival seems to be of major importance in this gazelle's population as well. Indirect genetic and indirect environmental effects assigned to mothers (i.e., maternal genetic and maternal permanent environmental effects) roughly explain a quarter of the total variance estimated for the trait analyzed. These findings have major evolutionary consequences for the species as show that offspring phenotypes can evolve strictly through changes in the environment provided by mothers. They are also relevant for the captive breeding programme of the species. To take into account, the contribution that mothers have on offspring phenotype through indirect genetic effects when designing pairing strategies might serve to identify those females with better ability to recruit, and, additionally, to predict reliable responses to selection in the captive population.

Effect of applying positive pressure with or without endotracheal suctioning during extubation: a laboratory study.

BACKGROUND: During invasive mechanical ventilation, secretions accumulate in the subglottic space; consequently, there is a risk of aspiration of these secretions into the airway during cuff deflation and extubation. To minimize this risk, 2 extubation methods are used. The first consists of introducing a suction catheter into the endotracheal tube (ETT) and the trachea. After initiating suctioning, the cuff is deflated and the ETT is removed together with the suction catheter. The second technique involves applying positive pressure to the ETT using a resuscitation bag. Once the manual breath is delivered, the ETT cuff is deflated and the ETT is removed without suction. The aim of this laboratory study is to determine the existence and magnitude of differences in leak volume during cuff deflation and extubation using various combinations of positive pressure with or without endotracheal suctioning. METHODS: An ETT connected to a ventilator was placed in a model trachea. Colored water was instilled in the space above the cuff. To measure the leak volume, a collection chamber was attached to the distal end of the model. Nine procedures were defined, based on the delivery of different positive pressure levels with or without endotracheal suctioning during extubation. The volume of leakage, in milliliters, was the unit of analysis. Procedures yielding values lower than 1 mL were assessed by the Friedman test, and a P value of less than .05 was considered significant. Post hoc comparisons were performed with a Wilcoxon test, followed by a Bonferroni correction. RESULTS: The application of CPAP 15, pressure support ventilation (PSV)15/10, and PSV 20/5 produced 0.4, 0.2, and 0.1 mL of leak volume, respectively. Statistically significant differences were found between CPAP 15 and PSV 15/10 (P = .003) and between CPAP 15 and PSV 20/5 (P = .01), but not between PSV 15/10 and PSV 20/5 (P = .30). The addition of suctioning increased leak volume, with statistically significant differences between CPAP 15 and CPAP 15 + endotracheal suctioning (P = .001) and between PSV 15/10 and PSV 15/10 + endotracheal suctioning (P = .001). CONCLUSIONS: Endotracheal suctioning during cuff deflation and extubation produced greater leakage. Application of CPAP 15, PSV 15/10, and PSV 20/5 resulted in the lowest leak values, with the best results being obtained with the use of PSV 15/10 and PSV 20/5.

Changes in Habitat Structure May Explain Decrease in Reintroduced Mohor Gazelle Population in the Guembeul Fauna Reserve, Senegal.

Reintroduction is a widespread method for saving populations of endangered species from extinction. In spite of recent reviews, it is difficult to reach general conclusions about its value as a conservation tool, as authors are reluctant to publish unsuccessful results. The Mohor gazelle is a North African gazelle, extinct in the wild. Eight individuals were reintroduced in Senegal in 1984. The population grew progressively, albeit slowly, during the first 20 years after release, but then declined dramatically, until the population in 2009 was estimated at no more than 13-15 individuals. This study attempts to determine the likelihood of gazelle-habitat relationships to explain why the size of the gazelle population has diminished. Our results show that the Mohor gazelle in Guembeul is found in open habitats with less developed canopy where the grass is shorter, suggesting the possibility that changes in habitat structure have taken place during the time the gazelles have been in the Reserve, reducing the amount of suitable habitat. Reintroduction design usually concentrates on short-term factors that may affect survival of the released animals and their descendants (short-term achievement), while the key factors for assessing its success may be those that affect the long-term evolution of the population.