Practical aspects of equine embryo transfer

This article focuses on practical aspects and most specific points of equine embryo transfer (EET). Induction of polyovylations in donor mares is not possible. Only the repetition of embryo collections can increase the number of foals from the same mares. At each oestrus, follow follicular growth and artificial inseminations are conducted in the same way as for any brood mare. It is essential to know the number and timing of ovulations at least a 12-hours accurency. Induction of ovulation is frequently required, GnRH agonists are used to do that. Embryos are flushed from the donor mare’s uterus, typically on day 7 or 8 after ovulation using a long (~1.5 m) and large (diameter 8 mm) Foley-type balloon-tipped catheter introduced by vaginal route just in front to cervix. Some practical details are important to EET practitioners, firstly, the embryos are easily located under stereo-microscope after filtration of flushing medium, and secondly most equine embryos are immediately transferred or after cooling at 4°C less than 24 hours. To have a recipient mare in the right stage of her oestrous cycle (D5 to D8 after ovulation) is the biggest and the most difficult challenge for EET practitioners because the synchronisation of oestrous and ovulations is very difficult to obtain. Therefore, embryos are transferred transcervically to the uterus of a recipient mare. Recently, a modified method using a speculum and a specific forceps seems to give a much better success rate than the conventional technique which requires a large training. The EET has regulatory constraints: regulations of different breeds allowing or not allowing the embryo transfer and sanitary European regulation concerning intra-community trade of equine embryos (Annex D to Council directive 92/65 EEC) and, in some countries, additional national regulation. The possible sanitary risks of ET in equine have been still little studied, and without International or European official registrations of EET activity, it is difficult to know the extent of the use of the technique around the world. The average success rate of embryo recovery is of the order of 40 to 50% depending on the fertility of the donor and the type of sperm used, the transfer success rate can reach 70 to 80% if the technique is well mastered by ET practitioners

Law of corresponding states for osmotic swelling of vesicles

As solute molecules permeate into a vesicle due to a concentration difference across its membrane, the vesicle swells through osmosis. The swelling can be divided into two stages: (a) an "ironing" stage, where the volume-to-area ratio of the vesicle increases without a significant change in its area; (b) a stretching stage, where the vesicle grows while remaining essentially spherical, until it ruptures. We show that the crossover between these two stages can be represented as a broadened continuous phase transition. Consequently, the swelling curves for different vesicles and different permeating solutes can be rescaled into a single, theoretically predicted, universal curve. Such a data collapse is demonstrated for giant unilamellar POPC vesicles, osmotically swollen due to the permeation of urea, glycerol, or ethylene glycol. We thereby gain a sensitive measurement of the solutes' membrane permeability coefficients, finding a concentration-independent coefficient for urea, while those of glycerol and ethylene glycol are found to increase with solute concentration. In addition, we use the width of the transition, as extracted from the data collapse, to infer the number of independent bending modes that affect the thermodynamics of the vesicle in the transition region.Comment: 10 page

Extracellular Reactive Oxygen Species (ROS) Production in Fresh Donkey Sperm Exposed to Reductive Stress, Oxidative Stress and NETosis

Altres ajuts: Generalitat de Catalunya 2017-SGR-1229Jenny shows a large endometrial reaction after semen influx to the uterus with a large amount of polymorphonuclear neutrophils (PMN) migrating into the uterine lumen. PMN act as a sperm selection mechanism through phagocytosis and NETosis (DNA extrudes and, together with proteins, trap spermatozoa). While a reduced percentage of spermatozoa are phagocytosed by PMN, most are found to be attached to neutrophil extracellular traps (NETs). This selection process together with sperm metabolism produces a large amount of reactive oxygen species (ROS) that influence the reproductive success. The present study aimed to determine the extracellular ROS production in both sperm and PMN. With this purpose, (1) donkey sperm were exposed to reductive and oxidative stresses, through adding different concentrations of reduced glutathione (GSH) and hydrogen peroxide (HO), respectively; and (2) PMN were subjected to NETosis in the presence of the whole semen, sperm, seminal plasma (SP) or other activators such as formyl-methionyl-leucyl-phenylalanine (FMLP). Extracellular ROS production (measured as HO levels) was determined with the Amplex ® Red Hydrogen Peroxide/Peroxidase Assay Kit. Donkey sperm showed more resilience to oxidative stress than to the reductive one, and GSH treatments led to greater HO extracellular production. Moreover, not only did SP appear to be the main inducer of NETosis in PMN, but it was also able to maintain the extracellular HO levels produced by sperm and NETosis

ProAKAP4 Semen Concentrations as a Valuable Marker Protein of Post-Thawed Semen Quality and Bull Fertility : a Retrospective Study

Functional sperm quality markers to predict bull fertility have been actively investigated. Among them, proAKAP4, which is the precursor of AKAP4, the main structural protein in the fibrous sheath of spermatozoa; appears to be promising, especially since spermatozoa lacking AKAP4 expression were shown to be immotile, abnormal, and infertile. In this study, the objective was to evaluate proAKAP4 concentration values with the classic sperm motility descriptors and fertility outcomes (NRR at 90 days) in post-thawed conditions of 10 bulls' semen. ProAKAP4 expression was confirmed by Western blotting and proAKAP4 concentrations were determined by ELISA. Variations in proAKAP4 concentrations were observed independently of the motility sperm descriptors measured using computer-assisted semen analysis (CASA). A ProAKAP4 concentration of 38.67 ± 8.55 ng/10 million spermatozoa was obtained as a statistical mean of all samples. Threshold values of proAKAP4 were then determined between 19.96 to 96.95 ng/10 million spermatozoa. ProAKAP4 concentrations were positively correlated with progressive motility and the linearity coefficient. The sperm showing the lowest progressive motility were the samples exhibiting proAKAP4 concentrations below 20 ng/10 million spermatozoa. Furthermore, proAKAP4 concentrations were significantly higher in bulls with a higher NRR in the field. Our results demonstrate a correlation between the semen concentration of proAKAP4 and NRR-90d (p = 0.05) in post-thawed bull semen, highlighting the potential of proAKAP4 as a predictive marker of bull fertility

Evaluation of physiological characteristics as selection criteria for drought tolerance in maize inbred lines and their hybrids

Improvement for maize drought tolerance has always been a significantobjective for breeders and plant physiologists. Nowadays, climate change sets new challenges to major crop adaptation at stressful environments. For such a purpose, the measurement of physiological traits related to maize response to drought might prove to be useful indices. The objective of the present study was to establish whether the physiological traits can be used as reliable physiological markers to evaluate the performance of parental genotypes and their hybrids under both dry and normally watered conditions, and under two densities an ultra-low density (ULD) and a normal dense stand (DS). Thirty (30) maize inbred lines and 30 single-crosses among them were evaluated across three diverse locations in Greece. The ULD was 0.74 plants/m-2, while the DS comprised 4.44 plants m-2 in the water deficitregime, and 6.67 and 7.84 plants m-2 in the normal water treatment for lines and hybrids, respectively. There was a very good association between the physiological characteristics studied and grain yield under the ultra-low density and especially for inbred lines. It was shown that the physiological characteristics can facilitate the selection of stress-adaptive genotypes under the low-density conditions and may permit modern maize to be grown at a wider range of environments. At the normal densities such a possibility was not evidenced since physiological parameters and yield did not correlate for either parents or hybrid

Wheat Landraces Are Better Qualified as Potential Gene Pools at Ultraspaced rather than Densely Grown Conditions

The negative relationship between the yield potential of a genotype and its competitive ability may constitute an obstacle to recognize outstanding genotypes within heterogeneous populations. This issue was investigated by growing six heterogeneous wheat landraces along with a pure-line commercial cultivar under both dense and widely spaced conditions. The performance of two landraces showed a perfect match to the above relationship. Although they lagged behind the cultivar by 64 and 38% at the dense stand, the reverse was true with spaced plants where they succeeded in out-yielding the cultivar by 58 and 73%, respectively. It was concluded that dense stand might undervalue a landrace as potential gene pool in order to apply single-plant selection targeting pure-line cultivars, attributable to inability of plants representing high yielding genotypes to exhibit their capacity due to competitive disadvantage. On the other side, the yield expression of individuals is optimized when density is low enough to preclude interplant competition. Therefore, the latter condition appears ideal to identify the most promising landrace for breeding and subsequently recognize the individuals representing the most outstanding genotypes

The structure of a class 3 nonsymbiotic plant haemoglobin from<i>Arabidopsis thaliana</i>reveals a novel N-terminal helical extension

Plant nonsymbiotic haemoglobins fall into three classes, each with distinct properties but all with largely unresolved physiological functions. Here, the first crystal structure of a class 3 nonsymbiotic plant haemoglobin, that fromArabidopsis thaliana, is reported to 1.77 Å resolution. The protein forms a homodimer, with each monomer containing a two-over-two α-helical domain similar to that observed in bacterial truncated haemoglobins. A novel N-terminal extension comprising two α-helices plays a major role in the dimer interface, which occupies the periphery of the dimer–dimer face, surrounding an open central cavity. The haem pocket contains a proximal histidine ligand and an open sixth iron-coordination site with potential for a ligand, in this structure hydroxide, to form hydrogen bonds to a tyrosine or a tryptophan residue. The haem pocket appears to be unusually open to the external environment, with another cavity spanning the entrance of the two haem pockets. The final 23 residues of the C-terminal domain are disordered in the structure; however, these domains in the functional dimer are adjacent and include the only two cysteine residues in the protein sequence. It is likely that these residues form disulfide bondsin vitroand it is conceivable that this C-terminal region may act in a putative complex with a partner moleculein vivo.</jats:p