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Mouse primordial germ cells (PGCs) are specified in the posterior embryo and migrate through hindgut endoderm and dorsal mesentery to the genital ridge. Although they move individually, PGCs do not migrate synchronously and are not all successful migrants. When migration ends at E11.5, we find approximately 6% of mouse PGCs outside the genital ridge; lacking survival cues, they are poised to undergo apoptosis. What heterogeneities distinguish a successful PGC, able to colonize the gonad, from a failed migrant? Are heterogeneities shared between clonally labeled mother and daughter cells? Our previous data show that mouse PGCs cultured ex vivo mount variable responses to migration cues, suggesting PGC behavioral variation may arise intrinsically; we also showed that proliferation is regulated by the somatic niche, increasing proliferation of PGCs as they progress along the migratory route. These data indicate that migratory speed and PGC-niche interactions may influence PGC gonadal colonization. To characterize intrinsic heterogeneities in migrating mouse PGCs, we conducted single cell RNA sequencing of purified PGCs and surrounding somatic niche for E9.5, E10.5, and E11.5 migratory timepoints. For E9.5 and E10.5, we generated libraries after splitting PGCs into anterior "leading" vs posterior "lagging" populations. Upon bioinformatic integration of all libraries, we analyzed differential gene expression probing positional differences as well as age related differences. We identified an E11.5 subcluster that may represent lagging PGCs transitioning medial/midline aorta-gonad-mesonephros tissues to the gonadal ridge. This cluster expresses imprinting genes. We have shown in vivo through HCR probes that cells expressing Igf2, a top imprinting gene marker of this cluster, are localized in the extragonadal space. We also probed receptor ligand interactions in migratory PGCs, and found several known receptor-ligand pairs, like Cxcl12-Cxcr4 survival signals. Interestingly, we identified potentially novel regulators of migrating germ cells, including ephrins. Additionally, re reanalyzed some human fetal PGCs and found that Slit-Robo signaling was implicated in both mouse and human PGC-somatic cell interactions. Additionally, we investigated whether mitochondrial heteroplasmies might contribute to migratory heterogeneities we observed. Using our single cell libraries, we were able to sequence some regions deeply enough to call variants. In general, we found that at either timepoint, anterior PGCs did not have different levels of heteroplasmy compared to posterior PGCs. However, we did observe that heteroplasmies increased from E9.5 compared to E10.5. In early human PGCs, it is known that nonsynonymous mutations decrease over time; in mouse, however, we saw that nonsynonymous mutations increased at E10.5 compared to E9.5.Finally, using fluorescent lineage labeling, we can test whether these heterogeneities are clonal. Interestingly, cells labeled while in the hindgut still localize near their clonal family in the genital ridge. Studying transcriptional profiles in migratory PGCs across different ages is revealing signaling mechanisms associated with specific niches as well as the extent of heterogeneity between cells.

RESUMEN Se caracteriza la cola en dos especies simpátricas de saurios endémicos de Ecuador. Utilizamos como técnicas de muestreo trampas de goma en ocho transectos de 500 m de distancia y trampas de caída de 80x80x80 cm. Para evaluar la condición de las colas se establecieron las categorías: intacta, ausente y regenerada. Entre 2012 y 2014 capturamos 138 saurios, 125 individuos de Stenocercus guentheri y trece de Pholidobolus montium. Alrededor del 61 % de los individuos tenían la cola intacta. Reportamos por primera vez bifurcación de la cola en Stenocercus guentheri; observamos tres individuos con esta característica. ABSTRACT The tail is characterized in two sympatric species of endemic saurians from Ecuador. We used glue traps in eight transects of 500 m distance and fall traps of 80x80x80 cm. The condition tails intact, absent, and regenerated were studied. Between 2012 and 2014 we captured 138 individuals, 125 individuals belong to Stenocercus guentheri and thirteen to Pholidobolus montium. About 61 % of the individuals had an intact tail. For the first time, we reported tail bifurcation in Stenocercus guentheri; three cases were observed with this condition.

Highly conserved gene regulatory networks are responsible for pattern formation in development. In case of insects that host cellular endosymbionts, challenges are faced for understanding pattern formation because of the interactions between the endosymbiont and intercellular signals of the host developmental gene regulatory network. Studying how these networks are operated in the presence of endosymbionts is critical for understanding of the process of endosymbiosis at the organismal level as well as the evolution of developmental systems.We characterized Dorsal-Ventral (DV) patterning system of the ant, Camponotus floridanus. Since DV patterning depends heavily on Bone Morphogenetic Protein Signaling in many insects, we used molecular techniques such as gene cloning, in situ hybridization, and RNA interference to examine the expression and function of developmental genes that are particularly involved in Bone Morphogenetic Protein Signaling.Here we show the expression and function of core BMP signaling components for DV patterning of ant C. floridanus.

Summary While diet modulates immunity, its impact on B cell ontogeny remains unclear. Using mixture modeling, a large-scale isocaloric dietary cohort mouse study identified carbohydrate as a major driver of B cell development and function. Increasing dietary carbohydrate increased B cell proportions in spleen, mesenteric lymph node and Peyer’s patches, and increased antigen-specific immunoglobulin G production after immunization. This was linked to increased B lymphopoiesis in the bone marrow. Glucose promoted early B lymphopoiesis and higher total B lymphocyte numbers than fructose. It drove B cell development through glycolysis and oxidative phosphorylation, independently of fatty acid oxidation in vitro and reduced B cell apoptosis in early development via mTOR activation, independently of interleukin-7. Ours is the first comprehensive study showing the impact of macronutrients on B cell development and function. It shows the quantitative and qualitative interplay between dietary carbohydrate and B cells and argues for dietary modulation in B cell-targeting strategies., Graphical abstract, Highlights • Dietary carbohydrates dynamically drive B cell lymphopoiesis and increase splenic B cells • Dietary glucose, but not fructose, affects B cell lymphopoesis • Dietary carbohydrates activate mTOR in B cells, particularly in early progenitors • Dietary carbohydrates increase plasma B cells and IgG response in immunized mice, Biological sciences; Immunology; Cell biology; Developmental biology

Excluding outlier plants (biological replicates deviating from the expected distribution on a multi-criteria basis) from phenotypic datasets is necessary to avoid false-positive associations between genome markers and traits., Based on case studies, we discuss the extent to which genome-wide association studies (GWAS) are affected by outlier plants, i.e. those deviating from the expected distribution on a multi-criteria basis. Using a raw dataset consisting of daily measurements of leaf area, biomass, and plant height for thousands of plants, we tested three different cleaning methods for their effects on genetic analyses. No-cleaning resulted in the highest number of dubious quantitative trait loci, especially at loci with highly unbalanced allelic frequencies. A trade-off was identified between the risk of false-positives (with no-cleaning and/or a low threshold for minor allele frequency) and the risk of missing interesting rare alleles. Cleaning can lower the risk of the latter by making it possible to choose a higher threshold in GWAS.

La reproduction des vaches allaitantes présente des particularités par rapport à celle des vaches laitières. Améliorer leur reproduction est un levier pour augmenter leur productivité. Les génisses de race à viande sont pubères lorsqu’elles ont atteint 50-55% du poids adulte vs 40-45% chez les génisses laitières. Les premières peuvent être mises à la reproduction vers 70% du poids adulte contre 60% pour les secondes. Le taux de renouvellement est assez faible en élevage allaitant, de 18 à 30% selon les races. Après le vêlage, la durée de l’anoestrus postpartum est plus longue qu’en race laitière du fait de la tétée du veau. L’expression des chaleurs est bonne chez la vache allaitante : les chaleurs durent entre 6 et 10 h dans les 3 principales races (Charolaise, Limousine et Blonde d’Aquitaine), et l’acceptation du chevauchement est exprimée dans 90 à 100% des oestrus. La monte naturelle domine en élevage allaitant : seuls 3 à 24% des veaux sont issus d’insémination selon les races. La fertilité des vaches allaitantes est actuellement mal connue, la performance de reproduction est plutôt approchée par la fécondité : l’intervalle vêlage-vêlage moyen des troupeaux est compris entre 379 et 444 jours selon les races. La proportion de vaches atteintes d’endométrite après vêlage semble élevée chez la vache allaitante comme chez la vache laitière (environ 20% même au-delà de 50 jours postpartum) mais les effets de cette affection sur la reproduction ont été peu explorés. De nombreuses données françaises concernant les races à viande spécialisées et rustiques sont présentées dans cet article., Reproduction in beef cows is different compared to dairy cows. An increase in reproductive performance is a key factor by which to improve productivity in beef cows. Puberty occurs when beef heifers reach 50-55% of their adult weight vs. 40-45% in dairy breeds. Beef heifers can be bred at 70% of their adult weight vs. 60% for dairy heifers. Replacement rate is low in beef production, from 18 to 30% depending on the breed. Beef cows have a longer anœstrous period after calving compared to dairy cows because the calf suckles its dam. The expression of oestrus is high in beef cows: the length of œstrus expression is between 6 and 10 hours in the 3 specialized French beef breeds (Charolaise, Limousine, Blonde d’Aquitaine) and 90 to 100% of cows express standing behavior at œstrous. Natural service is the most frequent breeding technique used in beef production: only 3 to 24% of calves are produced by artificial insemination depending on the breed. The fertility of beef cows is currently poorly understood. Reproductive efficiency is estimated by calving interval which is between 379 and 444 days depending on the breed. The prevalence of endometritis is high in both beef and dairy cows (levels of 20% have been recorded after 50 days postpartum) but its effect on reproductive performance has rarely been explored in beef cows. The present paper presents a large amount of French data concerning specialized and hardy French beef breeds.

Douglas-fir is a conifer species of major economic importance worldwide, including Western Europe and New Zealand. Herein we describe some characterization and significant refinement of somatic embryogenesis in Douglas-fir, with focus on maturation. The most typical structures observed in the embryonal masses were large polyembryogenic centres (up to 800-1500 μm) with a broad meristem, creating a compact ce ll “package” with suspensor cells. Singulated somatic embryos composed of both a embryonal head (300-400 μm) and long, tightly arranged suspensor were also frequent. Embryo development was enhanced following embryonal mass dispersion on filter paper discs at low density (50-100 mg fresh mass). Moreover, increasing gellan gum concentration in maturation medium (up to 10 g L-1) improved both the quantity and quality of cotyledonary somatic embryos (SEs), which were subsequently able to germinate and develop into plantlets at high frequency. Embryogenic yield was highly variable among the seven embryogenic lines tested (27-1544 SE g-1 fresh mass). Interestingly secondary somatic embryogenesis could be induced from cotyledonary SEs of both low- and highly-productive lines with some useful practical outcomes: secondary lines from low-performance lines (30-39 478 SE g-1) displayed significantly higher embryogenic yield (148-1343 SE g-1). In our best conditions, the total protein content in cotyledonary SEs increased significantly with maturation 41 duration (up to 150 μg mg-1 fresh mass after 7 weeks) but remained below that of mature zygotic embryos (300 μg mg-1). The protein pattern was similar in both somatic and zygotic embryos, with major storage proteins identified as 7S-vicilin- and legumin-like proteins.

Objective According to the Developmental Origin of Health and Disease (DOHaD) concept, maternal obesity and accelerated growth in neonates predispose offspring to white adipose tissue (WAT) accumulation. In rodents, adipogenesis mainly develops during lactation. The mechanisms underlying the phenomenon known as developmental programming remain elusive. We previously reported that adult rat offspring from high-fat diet-fed dams (called HF) exhibited hypertrophic adipocyte, hyperleptinemia and increased leptin mRNA levels in a depot-specific manner. We hypothesized that leptin upregulation occurs via epigenetic malprogramming, which takes place early during development of WAT. Methods As a first step, we identified in silico two potential enhancers located upstream and downstream of the leptin transcription start site that exhibit strong dynamic epigenomic remodeling during adipocyte differentiation. We then focused on epigenetic modifications (methylation, hydroxymethylation, and histone modifications) of the promoter and the two potential enhancers regulating leptin gene expression in perirenal (pWAT) and inguinal (iWAT) fat pads of HF offspring during lactation (postnatal days 12 (PND12) and 21 (PND21)) and in adulthood. Results PND12 is an active period for epigenomic remodeling in both deposits especially in the upstream enhancer, consistent with leptin gene induction during adipogenesis. Unlike iWAT, some of these epigenetic marks were still observable in pWAT of weaned HF offspring. Retained marks were only visible in pWAT of 9-month-old HF rats that showed a persistent “expandable” phenotype. Conclusions Consistent with the DOHaD hypothesis, persistent epigenetic remodeling occurs at regulatory regions especially within intergenic sequences, linked to higher leptin gene expression in adult HF offspring in a depot-specific manner., Highlights • The white adipose tissue is an important target of developmental programming. • Higher leptin gene expression occurs in offspring from obese dams in a depot-specific manner. • Leptin upregulation occurs via epigenetic malprogramming during development of adipose tissue. • Persistent genomic epigenetic remodeling occurs in adipose tissue of offspring from obese dams. • Intergenic regions were more affected than the leptin promoter region in offspring of obese dams.

Thème: Folliculogenesis, oogenesis and superovulationAbstract + poster; The LH surge promotes prostaglandin E2 (PGE2) production within the preovulatory follicle. Oocytemicroenvironment levels of PGE2 affect the developmental competence after fertilization. This studyaimed to characterize the follicular fluid PGE2 enrichment during superovulation treatment. Six heifers(Holstein, 20.3 +/-0.85 months old) received FSH (Stimufol®, Reprobiol, Belgium): half dose, ie, 250 μgof porcine follitropin (pFSH), combined with 50 μg of porcine lutropin (pLH). At the rate of 8 injections,in degressive dose, spread over 4 days. PGF2α (Estrumate®, MSD Santé Animale, France) was injectedat the same time as the 5th injection of Stimufol®.LH peak was assumed to occur between 35 and 40hours after the PGF2α injection. Individual sampling of fluid from antral follicles was performed byovum pick up 12 hours before PGF2α injection and 60 hours after PGF2α injection. This protocol wasdesigned to allow the collection of fluids from ovarian follicle containing either a pre-matured or amatured oocyte. Each heifer was his own control as we took the "pre-matured" follicular fluid on a firstovary and the "matured" follicular fluid on the 2nd ovary, 3 days later. The punctures were repeatedtwice and were cross-checked for the next repetition to evaluate the impact of the ipsi or contralateralside of the corpus luteum (CL) on the follicular fluid composition. The volume of fluid was measured foreach punctured follicle. The PGE2 concentration of the follicular fluid was measured by Elisa (CaymanChemical) to determine the progress of terminal follicular differentiation. An average of 13 +/- 5.06 and28 +/-13.9 follicles were punctured per session/heifer for respectively pre-matured (n=78) and matured(n=169) follicles. The mean collected volume differed between the two groups (pre-matured: 0.229+/-0.213 ml/follicle; matured: 0.575+/-0.379 ml/follicle; two samples t-test, pval

International audience

International audience; It is well known that adipokines are endocrine factors that are mainly secreted by white adipose tissue. Their central role in energy metabolism is currently accepted. More recently, their involvement in fertility regulation and the development of some reproductive disorders has been suggested. Data concerning the role of leptin and adiponectin, the two most studied adipokines, in the control of the reproductive axis are consistent. In recent years, interest has grown about some novel adipokines, chemerin, visfatin, resistin and apelin, which have been found to be strongly associated with obesity and insulin-resistance. Here, we will review their expression and role in male and female reproduction in humans and animal models. According to accumulating evidence, they could regulate the secretion of GnRH (Gonadotropin-Releasing Hormone), gonadotropins and steroids. Furthermore, their expression and that of their receptors (if known), has been demonstrated in the human and animal hypothalamo-pituitary-gonadal axis. Like leptin and adiponectin, these novel adipokines could thus represent metabolic sensors that are able to regulate reproductive functions according to energy balance changes. Therefore, after investigating their role in normal fertility, we will also discuss their possible involvement in some reproductive troubles known to be associated with features of metabolic syndrome, such as polycystic ovary syndrome, gestational diabetes mellitus, preeclampsia and intra-uterine growth retardation in women, and sperm abnormalities and testicular pathologies in men.

Bovine preimplantation embryos at the single cell level: an accurate model for human reproductive approaches? . 2. CellFit Workshop 2019

Background Islets of Langerhans transplantation is a promising therapy for type 1 diabetes mellitus, but this technique is compromised by transplantation stresses including inflammation. In other tissues, co-transplantation with mesenchymal stem cells has been shown to reduce damage by improving anti-inflammatory and anti-oxidant defences. Therefore, we probed the protection afforded by bone marrow mesenchymal stem cells to islets under pro-inflammatory cytokine stress. Methods In order to evaluate the cytoprotective potential of mesenchymal stem cells on rat islets, co-cultures were exposed to the interleukin-1, tumour necrosis factor α and interferon γ cocktail for 24 h. Islet viability and functionality tests were performed. Reactive oxygen species and malondialdehyde were measured. Expression of stress-inducible genes acting as anti-oxidants and detoxifiers, such as superoxide dismutases 1 and 2, NAD(P)H quinone oxidoreductase 1, heme oxygenase-1 and ferritin H, was compared to non-stressed cells, and the corresponding proteins were measured. Data were analysed by a two-way ANOVA followed by a Holm-Sidak post hoc analysis. Results Exposure of rat islets to cytokines induces a reduction in islet viability and functionality concomitant with an oxidative status shift with an increase of cytosolic ROS production. Mesenchymal stem cells did not significantly increase rat islet viability under exposure to cytokines but protected islets from the loss of insulin secretion. A drastic reduction of the antioxidant factors heme oxygenase-1 and ferritin H protein levels was observed in islets exposed to the cytokine cocktail with a prevention of this effect by the presence of mesenchymal stem cells. Conclusions Our data evidenced that MSCs are able to preserve islet insulin secretion through a modulation of the oxidative imbalance mediated by heme and iron via heme oxygenase-1 and ferritin in a context of cytokine exposure. Electronic supplementary material The online version of this article (10.1186/s13287-019-1190-4) contains supplementary material, which is available to authorized users.

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International audience; Root hairs are protrusions from root epidermal cells with crucial roles in plant soil interactions. Although much is known about patterning, polarity and tip growth of root hairs, contributions of membrane trafficking to hair initiation remain poorly understood. Here, we demonstrate that the trans-Golgi network-localized YPT-INTERACTING PROTEIN 4a and YPT-INTERACTING PROTEIN 4b (YIP4a/b) contribute to activation and plasma membrane accumulation of Rho-of-plant (ROP) small GTPases during hair initiation, identifying YIP4a/b as central trafficking components in ROP-dependent root hair formation.

International audience; Of the hepatic cell lines developed for in vitro studies of hepatic functions as alternatives to primary human hepatocytes, many have lost major liver-like functions, but not HepaRG cells. The increasing use of the latter worldwide raises the need for establishing the reference functional status of early biobanked HepaRG cells. Using deep proteome and secretome analyses, the levels of master regulators of the hepatic phenotype and of the structural elements ensuring biliary polarity were found to be close to those in primary hepatocytes. HepaRG cells proved to be highly differentiated, with functional mitochondria, hepatokine secretion abilities, and an adequate response to insulin. Among differences between primary human hepatocytes and HepaRG cells, the factors that possibly support HepaRG transdifferentiation properties are discussed. The HepaRG cell system thus appears as a robust surrogate for primary hepatocytes, which is versatile enough to study not only xenobiotic detoxification, but also the control of hepatic energy metabolism, secretory function and disease-related mechanisms.

While obesity is widely recognized as a risk factor for cancer, survival among patients with cancer is often higher for obese than for lean individuals. Several hypotheses have been proposed to explain this “obesity paradox,” but no consensus has yet emerged. Here, we propose a novel hypothesis to add to this emerging debate which suggests that lean healthy persons present conditions unfavorable to malignant transformation, due to powerful natural defenses, whereby only rare but aggressive neoplasms can emerge and develop. In contrast, obese persons present more favorable conditions for malignant transformation, because of several weight‐associated factors and less efficient natural defenses, leading to a larger quantity of neoplasms comprising both nonaggressive and aggressive ones to regularly emerge and progress. If our hypothesis is correct, testing would require the consideration of the raw quantity, not the relative frequency, of aggressive cancers in obese patients compared with lean ones. We also discuss the possibility that in obese persons, nonaggressive malignancies may prevent the subsequent progression of aggressive cancers through negative competitive interactions between tumors.

International audience; Background : Non-specific Lipid Transfer Proteins (nsLTPs) are widely distributed in the plant kingdom and constitute a superfamily of related proteins. Several hundreds of different nsLTP sequences—and counting—have been characterized so far, but their biological functions remain unclear. It has been clear for years that they present a certain interest for agronomic and nutritional issues. Deciphering their functions means collecting and analyzing a variety of data from gene sequence to protein structure, from cellular localization to the physiological role. As a huge and growing number of new protein sequences are available nowadays, extracting meaningful knowledge from sequence–structure–function relationships calls for the development of new tools and approaches. As nsLTPs show high evolutionary divergence, but a conserved common right handed superhelix structural fold, and as they are involved in a large number of key roles in plant development and defense, they are a stimulating case study for validating such an approach. Methods : In this study, we comprehensively investigated 797 nsLTP protein sequences, including a phylogenetic analysis on canonical protein sequences, three-dimensional structure modeling and functional annotation using several well-established bioinformatics programs. Additionally, two integrative methodologies using original tools were developed. The first was a new method for the detection of (i) conserved amino acid residues involved in structure stabilization and (ii) residues potentially involved in ligand interaction. The second was a structure–function classification based on the evolutionary trace display method using a new tree visualization interface. We also present a new tool for visualizing phylogenetic trees. Results : Following this new protocol, an updated classification of the nsLTP superfamily was established and a new functional hypothesis for key residues is suggested. Lastly, this work allows a better representation of the diversity of plant nsLTPs in terms of sequence, structure and function.

The upper airway epithelium, which is mainly composed of multiciliated, goblet, club and basal cells, ensures proper mucociliary function and can regenerate in response to assaults. In chronic airway diseases, defective repair leads to tissue remodeling. Delineating key drivers of differentiation dynamics can help understand how normal or pathological regeneration occurs. Using single-cell transcriptomics and lineage inference, we have unraveled trajectories from basal to luminal cells, providing novel markers for specific populations. We report that: (1) a precursor subgroup of multiciliated cells, which we have entitled deuterosomal cells, is defined by specific markers, such as DEUP1, FOXN4, YPEL1, HES6 and CDC20B; (2) goblet cells can be precursors of multiciliated cells, thus explaining the presence of hybrid cells that co-express markers of goblet and multiciliated cells; and (3) a repertoire of molecules involved in the regeneration process, such as keratins or components of the Notch, Wnt or BMP/TGFβ pathways, can be identified. Confirmation of our results on fresh human and pig airway samples, and on mouse tracheal cells, extend and confirm our conclusions regarding the molecular and cellular choreography at work during mucociliary epithelial differentiation., Highlighted Article: Single-cell RNAseq data in fresh human airway epithelial tissues and air liquid cultures identifies novel cell populations and offers new insights into the molecular mechanisms occurring during mucociliary epithelium regeneration.

Background Lactation and associated metabolic stresses during the post-partum period have been shown to impair fertility in dairy cows. The oviduct plays key roles in embryo development and the establishment of pregnancy in cattle. The aim of this study was to investigate the effects of lactation and location relative to the corpus luteum (CL) on the transcriptome of the bovine oviduct epithelium. Results An original animal model was used. At 60 days post-partum, Holstein lactating (n = 4) and non-lactating (i.e. never milked after calving; n = 5) cows, as well as control nulliparous heifers (n = 5), were slaughtered on Day 3 following induced estrus, and epithelial samples from the oviductal ampulla and isthmus ipsilateral and contralateral to the corpus luteum (CL) were recovered for RNA sequencing. In the oviduct ipsilateral to the CL, differentially expressed genes (DEGs) were identified between heifers compared with both postpartum cow groups. However, only 15 DEGs were identified between post-partum lactating and non-lactating cows in the ipsilateral isthmus and none were identified in the ipsilateral ampulla. In contrast, 192 and 2583 DEGs were identified between ipsilateral and contralateral ampulla and isthmus, respectively. In both regions, more DEGs were identified between ipsilateral and contralateral oviducts in non-lactating cows and heifers than in lactating cows. Functional annotation of the DEGs associated with comparisons between metabolic groups highlighted a number of over-represented biological functions and cell pathways including immune response and cholesterol/steroid biosynthesis. Conclusions Gene expression in the oviduct epithelium, particularly in the isthmus, was more affected by the location relative to the CL than by lactation at Day 3 post-estrus. Furthermore, the effect of the proximity to the CL was modulated by the metabolic status of the cow. Electronic supplementary material The online version of this article (10.1186/s12864-019-5616-2) contains supplementary material, which is available to authorized users.

International audience; Lipopolysaccharide (LPS) expressed on the surface of Gram-negative bacteria activates pro-inflammatory pathways, dys-regulates the function of endometrial cells and is a key player in the mechanisms involved in endometritis. This study aimed to investigate the effects of LPS on bovine endometrial epithelial cells (bEEC) from whole transcriptome with a special focus on genes involved in embryo-maternal interactions. Following in vitro culture, bEEC from three cows were exposed to 0, 2, and 8 μg/mL LPS for 24h. RNA samples extracted at 0 and 24 hours were analyzed by RNA sequencing (RNA-seq). At 24h, 2035 differentially expressed genes (DEGs) were identified between controls and samples treated with 2 μg/mL LPS. Gene ontology analysis showed that over-expressed DEGs were associated to immune response, response to stress and external stimuli, catalytic activity, and cell cycle. Genes associated with cell membrane and cell adhesion pathways were under-expressed. LPS induced changes in expression of specific genes related to embryo-maternal interactions including under-expression of eight members of the cadherin superfamily, over-expression of six members of the mucin family, and differential expression of a large set of genes binding the above molecules and of more than 20 transcripts coding for cytokines and their receptors. Type I interferon-τ dependent genes were also over-expressed. From a sub-set of 19 genes, (biological replicates of bEEC from cows taken at time 6 (n = 3), 24 (n = 6) and 48 hours (n = 3), and 2 technical replicates per sample) differential gene expression was confirmed by RT2-qPCR (r2 between fold changes at 24 hours by RT2-qPCR and RNA-seq = 0.97). These results indicate that LPS affects the function of bEEC in many ways by differential transcription, glycolytic metabolism and oxidative stress. Many transcriptomic signatures related to implantation and embryo maternal interactions were strongly affected by LPS. These results pave the way for further studies to investigate the duration of these changes and their possible impact on endometrial function and fertility.

International audience

International audience; Olive (Olea europaea L.) is a major fruit crop in the Mediterranean Basin. Ex-situ olive management is essential to ensure optimal use of genetic resources in breeding programs. The Worldwide Olive Germplasm Bank of Córdoba (WOGBC), Spain, and Marrakech (WOGBM), Morocco, are currently the largest existing olive germplasm collections. Characterization, identification, comparison and authentication of all accessions in both collections could thus provide useful information for managing olive germplasm for its preservation, exchange within the scientific community and use in breeding programs. Here we applied 20 microsatellite markers (SSR) and 11 endocarp morphological traits to discriminate and authenticate 1091 olive accessions belonging to WOGBM and WOGBC (554 and 537, respectively). Of all the analyzed accessions, 672 distinct SSR profiles considered as unique genotypes were identified, but only 130 were present in both collections. Combining SSR markers and endocarp traits led to the identification of 535 cultivars (126 in common) and 120 authenticated cultivars. No significant differences were observed between collections regarding the allelic richness and diversity index. We concluded that the genetic diversity level was stable despite marked contrasts in varietal composition between collections, which could be explained by their different collection establishment conditions. This highlights the extent of cultivar variability within WOGBs. Moreover, we detected 192 mislabeling errors, 72 of which were found in WOGBM. A total of 228 genotypes as molecular variants of 74 cultivars, 79 synonyms and 39 homonyms as new cases were identified. Both collections were combined to define the nested core collections of 55, 121 and 150 sample sizes proposed for further studies. This study was a preliminary step towards managing and mining the genetic diversity in both collections while developing collaborations between olive research teams to conduct association mapping studies by exchanging and phenotyping accessions in contrasted environmental sites.

International audience; The nutritional transition that the western population has undergone is increasingly associated with chronic metabolic diseases. In this work, we evaluated a diet rich in saturated fatty acids (hyperlipidic, HL) after weaning of the offspring rats submitted to maternal protein restriction on the hepatic mitochondrial bioenergetics. Wistar rats were mated and during gestation and lactation, mothers received control diets (NP, normal protein content 17%) or low protein (LP, 8% protein). After weaning, rats received either NL (normolipidic) or HL (+59% SFA) diets up to 90 days of life. It was verified that all respiratory states of hepatic mitochondria showed a reduction in the LP group submitted to the post-weaning HL diet. This group also presented greater mitochondrial swelling compared to controls, potentiated after Ca(2+) addition and prevented in the presence of EGTA (calcium chelator) and cyclosporin A (mitochondrial permeability transition pore inhibitor). There was also an increase in liver protein oxidation and lipid peroxidation and reduction in catalase and glutathione peroxidase activities in the LP group fed HL diet after weaning. Our data suggest that adult rats subjected to maternal protein restriction were more susceptible to hepatic mitochondrial damage caused by a diet rich in saturated fatty acids post-weaning.

Context and challenges: Like most other perennial crops, the grapevine needs to undergo a juvenile period before fruiting. Thus, the development of reproductive organs from seedlings is possible only after the second or third vegetative cycle. Each proleptic axis then displays only one to three inflorescences per growing cycle. These biological features and the size of the adult vine are major hindrances to the design of experiments on fruit and plant physiology, and complicate and lengthen the time required for studies in grapevine breeding and genetics.Significance of the review: The microvine is a dwarf phenotype resulting from a mutation in the VviGAI1 gene, which induces miniaturization of all vegetative organs and conversion of tendrils into inflorescences without affecting berry development. The small size of the microvine allows tight control of environmental conditions. Spatial developmental gradients fit well with temporal series of each phytomer position. Thus, kinetic profiles can be inferred from spatial information. In the first part of the paper, we describe the molecular and genetic mechanisms determining microvine phenotypes, reviewing the main biological properties of the microvine model. Subsequently, the results of recent studies in which the model was used for research in grapevine physiology and genetics are summarized. The review focuses on experiments investigating the effects of temperature on vegetative and reproductive organogenesis, berry development, and biomass allocation at the whole-plant level. Furthermore, we discuss and illustrate how the model can be used to identify (QTL) quantitative trait loci in fruit development and adaptive traits that could be useful when selecting genotypes in anticipation of the effects of global warming.

Exposure to fine-particulate air pollution is a major global health concern because it is associated with reduced birth weight and an increased risk of cardiovascular disease. Here we have investigated the potential for exposure to diesel exhaust during pregnancy to influence mammary gland development and milk composition. Female rabbits were therefore exposed by nose-only inhalation to either diluted diesel exhaust fumes (1 mg/m3) or clean air for 2h/day, 5 days/week, from the 3rd to the 27th days of pregnancy. On Day 28 of pregnancy, mammary glands were collected from twelve females (six controls and six diesel-exposed) and assessed for morphological and functional alterations. Milk samples were collected from eighteen dams (nine controls and nine diesel-exposed) during early (days 2 to 4) and established (days 13 to 16) lactation to verify the composition of fatty acids and major proteins and leptin levels. The mammary alveolar lumina contained numerous fat globules, and stearoyl CoA reductase expression was higher in mammary epithelia from diesel exhaust-exposed rabbits, which together suggested increased mammary lipid biosynthesis. Gas chromatography analysis of the composition of milk fatty acids revealed a sharp rise in the total fatty acid content, mainly due to monounsaturated fatty acids. Liquid chromatography-mass spectrometry analysis of milk samples enabled identification and quantification of the main rabbit milk proteins and their main phosphorylated isoforms, and revealed important changes to individual casein and whey protein contents and to their most phosphorylated isoforms during early lactation. Taken together, these findings suggest that repeated daily exposure to diesel exhaust fumes during pregnancy at urban pollution levels can influence lipid metabolism in the mammary gland and the lipid and protein composition of milk. As milk may contribute to metabolic programming, such alterations affecting milk composition should be taken into account from a public health perspective.

Background/Aims Maternal obesity is associated with fertility disorders, obstetric complications, and development of metabolic syndrome in offspring. The recommendation to overweight or obese women is to lose weight before pregnancy, but the impact of these prescriptions on the offspring health is insufficiently studied. Previous results on late-term mice fetuses have shown that maternal obesity leads to fetal growth restriction and to the modification of epigenetic machinery-related gene expression in fetal liver and placenta. Moreover, fetuses from the preconceptional weight loss maternal group normalize their growth, but some fetal genes stay differentially expressed. Method We analysed the metabolic phenotype of offspring born to control (CTRL), obese (OB) or weight loss after diet-induced obesity (WL) mothers. To highlight a possible conditioning effect, offspring were either fed a control (CD) or a high fat diet (HFD). Their metabolism and olfactory behavior were monitored for up to 6 months. Then, we analyzed the metabolome of three tissues involved in food intake and nutrient management (liver, olfactory bulb and hypothalamus) in male adult offspring. Results Multiple Factorial Analysis integration of offspring phenotypic data showed a major influence of the post-weaning diet and pointed a difference in HFD-fed males according to their maternal group (OB vs WL). This confirmed a sex-dependant metabolic conditioning by the maternal environment. The olfactory sensitivity, measured by electro-olfactogram, was reduced in WL male offspring, whatever the postnatal diet. The metabolomics study annotated 278, 258 and 200 metabolites in the liver, olfactory bulb and hypothalamus respectively. Again, the post-weaning diet had a major effect in the three tissues but interestingly, the maternal group also influenced the hepatic metabolome of adult offspring. Conclusions These data, integrating metabolism, olfactory behavior and metabolome, provide new and original information on the effects of preconceptional maternal metabolic status in the offspring health conditioning upon diet challenges.

The vast majority of previous studies on epigenetics in plants have centered on the study of inheritance of DNA methylation patterns in annual plants. In contrast, perennial plants may have the ability to accumulate changes in DNA methylation patterns over numerous years. However, currently little is known about long-lived perennial and clonally reproducing plants that may have evolved different DNA methylation inheritance mechanisms as compared to annual plants. To study the transmission of DNA methylation patterns in a perennial plant, we used apple (Malus domestica) as a model plant. First, we investigated the inheritance of DNA methylation patterns during sexual reproduction in apple by comparing DNA methylation patterns of mature trees to juvenile seedlings resulting from selfing. While we did not observe a drastic genome-wide change in DNA methylation levels, we found clear variations in DNA methylation patterns localized in regions enriched for genes involved in photosynthesis. Using transcriptomics, we also observed that genes involved in this pathway were overexpressed in seedlings. To assess how DNA methylation patterns are transmitted during clonal propagation we then compared global DNA methylation of a newly grafted tree to its mature donor tree. We identified significant, albeit weak DNA methylation changes resulting from grafting. Overall, we found that a majority of DNA methylation patterns from the mature donor tree are transmitted to newly grafted plants, however with detectable specific local differences. Both the epigenomic and transcriptomic data indicate that grafted plants are at an intermediate phase between an adult tree and seedling and inherit part of the epigenomic history of their donor tree.

According to the &ldquo, developmental origins of health and disease&rdquo, (DOHaD) concept, maternal obesity predisposes the offspring to non-communicable diseases in adulthood. While a preconceptional weight loss (WL) is recommended for obese women, its benefits on the offspring have been poorly addressed. We evaluated whether preconceptional WL was able to reverse the adverse effects of maternal obesity in a mouse model, exhibiting a modification of foetal growth and of the expression of genes encoding epigenetic modifiers in liver and placenta. We tracked metabolic and olfactory behavioural trajectories of offspring born to control, obese or WL mothers. After weaning, the offspring were either put on a control diet (CD) or a high-fat (HFD). After only few weeks of HFD, the offspring developed obesity, metabolic alterations and olfactory impairments, independently of maternal context. However, male offspring born to obese mother gained even more weight under HFD than their counterparts born to lean mothers. Preconceptional WL normalized the offspring metabolic phenotypes but had unexpected effects on olfactory performance: a reduction in olfactory sensitivity, along with a lack of fasting-induced, olfactory-based motivation. Our results confirm the benefits of maternal preconceptional WL for male offspring metabolic health but highlight some possible adverse outcomes on olfactory-based behaviours.

Macrophage elastase, or MMP-12, is mainly produced by alveolar macrophages and is believed to play a major role in the development of chronic obstructive pulmonary disease (COPD). The catalytic domain of MMP-12 is unique among MMPs in that it is very highly active on numerous substrates including elastin. However, measuring MMP-12 activity in biological fluids has been hampered by the lack of highly selective substrates. We therefore synthesized four series of fluorogenic peptide substrates based on the sequences of MMP-12 cleavage sites in its known substrates. Human MMP-12 efficiently cleaved peptide substrates containing a Pro at P3 in the sequence Pro-X-X↓Leu but lacked selectivity towards these substrates compared to other MMPs, including MMP-2, MMP-7, MMP-9 and MMP-13. On the contrary, the substrate Abz-RNALAVERTAS-EDDnp derived from the CXCR5 chemokine was the most selective substrate for MMP-12 ever reported. All substrates were cleaved more efficiently by full-length MMP-12 than by its catalytic domain alone, indicating that the C-terminal hemopexin domain influences substrate binding and/or catalysis. Docking experiments revealed unexpected interactions between the peptide substrate Abz-RNALAVERTAS-EDDn and MMP-12 residues. Most of our substrates were poorly cleaved by murine MMP-12 suggesting that human and murine MMP-12 have different substrate specificities despite their structural similarity.

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Diffusible signals are known to orchestrate patterning during embryogenesis, yet diffusion is sensitive to noise. The fact that embryogenesis is remarkably robust suggests that additional layers of regulation reinforce patterning. Here, we demonstrate that geometrical confinement orchestrates the spatial organisation of initially randomly positioned subpopulations of spontaneously differentiating mouse embryonic stem cells. We use micropatterning in combination with pharmacological manipulations and quantitative imaging to dissociate the multiple effects of geometry. We show that the positioning of a pre-streak-like population marked by brachyury (T) is decoupled from the size of its population, and that breaking radial symmetry of patterns imposes polarised patterning. We provide evidence for a model in which the overall level of diffusible signals together with the history of the cell culture define the number of T+ cells, whereas geometrical constraints guide patterning in a multi-step process involving a differential response of the cells to multicellular spatial organisation. Our work provides a framework for investigating robustness of patterning and provides insights into how to guide symmetry-breaking events in aggregates of pluripotent cells., Highlighted Article: Asymmetric geometrical confinement guides polarised patterning and ensures positional precision of a primitive streak-like population of cells in mouse pluripotent cultures.

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National audience; The International Society for Embryo Technologies (IETS) was created in 1974. To meet the needs for specific regulation in terms of health safety, IETS created an Import & Export committee, chaired by R Mapletoft, in the early 80’s. A formal relationship between IETS and the World Organization for Animal Health (OIE) was then set up in 1987. With the development of new embryo technologies and the concern about the safety of animals produced with such technologies entering the food chain, the scope of the committee was expanded in 2000 under the initiative of M Thibier so as to include those issues, resulting in the current IETS Health and Safety Advisory Committee (HASAC, https://www.iets.org/ comm_hasac.asp). HASAC functions to advise the IETS Board members, IETS members and OIE. HASAC is composed of IETS members from academic, regulatory and industry sectors, under the direction of a chair nominated by the IETS Board of Governors (BoG) (current chair: Julie Gard, succeeding to F Fieni, P Chavatte-Palmer and M Thibier). Its aims are: - to review regularly and extensively the literature in order to evaluate potential risks, based on scientific evidence, with regard to international trade of embryos and human consumption of animal or animal products derived from embryo transfer and related technologies. - to provide guidelines such as codes of practice, recommendations and any information pertinent to the safe movement of embryos and safe introduction into the food chain of animal or animal products derived from reproductive biotechnologies without unduly restricting technological advances and commerce.- to communicate to IETS members all its achievements and upon acceptance by IETS BoG, to all the relevant International Agencies. Three HASAC subcommittees meet annually but work electronically throughout the year: 1. The Research subcommittee maintains and revises literature relevant to the animal health implications of current and emerging technologies. A large database, including an abstract, HASAC interpretation and conclusions on all publications examined, is updated each year. Based on these yearly evaluations, science-based recommendations are then handed for evaluation to the Regulatory subcommittee. 2. The Regulatory subcommittee examines new evidence provided by the scientific subcommittee and discusses whether current recommendations for the management of the risks potentially associated with embryos in terms of pathogen-embryo interactions should be modified. Any proposed modifications to alter text with regards to regulations and/or OIE recommendations are put forward to the BoG and, if approved, subsequently formally proposed to the OIE for incorporation into the Terrestrial Animal health Code. Commission members of OIE most often support the proposed amendments from IETS/HASAC before finally being voted on, and mostly accepted, by the OIE General Assembly. 3. The Manual, Forms and Certificates subcommittee develops, elaborates and publishes a code of practice providing guidelines to practitioners for managing their operations so that pertinent quality assurance standards will be met. Additionally, this committee creates and maintains recording and identification systems of embryos to ensure traceability.

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