12-day Embryos Research Articles

A prospective study of the proteome of equine pre-implantation embryo.

The present study was conducted to investigate the global proteome of 8-day-old equine blastocysts. Follicular dynamics of eight adult mares were monitored by ultrasonography and inseminated 24 h after the detection of a preovulatory follicle. Four expanded blastocysts were recovered, pooled, and subjected to protein extraction and mass spectrometry. Protein identification was conducted based on four database searches (PEAKS, Proteome Discoverer software, SearchGUI software, and PepExplorer). Enrichment analysis was performed using g:Profiler, Panther, and String platforms. After the elimination of identification redundancies among search tools (at three levels, based on identifiers, peptides, and cross-database mapping), 1977 proteins were reliably identified in the samples of equine embryos. Proteomic analysis unveiled robust metabolic activity in the 8-day equine embryo, highlighted by an abundance of proteins engaged in key metabolic pathways like the TCA cycle, ATP biosynthesis, and glycolysis. The prevalence of chaperones among highly abundant proteins suggests that regulation of protein folding, and degradation is a key process during embryo development. These findings pave the way for developing new strategies to improve equine embryo media and optimize invitro fertilization techniques.

Transcriptomic Comparative Analysis of Two Breeds of Mongolian Sheep at 16-day Embryos

Transcriptomic Comparative Analysis of Two Breeds of Mongolian Sheep at 16-day Embryos

Molecular signature of the ontogenic development of the prawn Macrobrachium tenellum

The prawn Macrobrachium tenellum shows aquaculture potential due to its well-defined reproductive cycle linked to female nutritional requirements. Significant changes occur in egg composition during the 16 to 17-day embryo development. Understanding the ontogenic proteins is crucial for developmental insights and controlled reproduction. We employed free-label quantitative proteomics to analyze egg peptides at the initial and final stages of wild females. Using the emPAI protocol and Proteome Discoverer 2.0, we identified 89 differentially expressed proteins in M. tenellum eggs. Of these, 27 were exclusive to early-stage development and three to late-stage. Abundant proteins included Vitellogenin, glyceraldehyde-3-phosphate dehydrogenase, histone 4, beta-actin, and hemocyanin. Gene Ontology analysis revealed 518 terms across molecular functions, biological processes, and cellular components using the GoRetriever tool of AgBase and the CateGOrizer tool of the Animal Genome Research Program. Carbohydrate metabolism was significant in early-stage development, with glyceraldehyde-3-phosphate dehydrogenase being the second most abundant protein. Proteins involved in ATP synthesis and cytoplasmic proteins associated with catalytic and binding activities related to primary metabolism were also detected. Our study elucidates the role of Vitellogenin in lipid transport activity and its potential involvement in the juvenile hormone feedback pathway. This pathway includes farnesoic acid O-methyltransferase and juvenile hormone epoxide oxidase, regulating protein biosynthesis, molt cycles (including chitinase activity), and potentially influencing controlled reproduction. Our proteomic analysis provides insights into the molecular mechanisms driving Ontogenic development in Macrobrachium tenellum, with implications for controlled reproduction strategies and advancements in aquaculture practices.

Successful mule embryo transfer to a recipient mule during late diestrus

Mating between a donkey (Equus asinus) and a mare (Equus caballus) results in a mule foal, which is an infertile hybrid due to the different number of chromosomes in the parent species. However, mules have been successfully used as recipients of donkey embryos, horse embryos and, recently, mule embryos. In non-cyclic recipient mules, progesterone supplementation is needed to sustain pregnancy, but some mules can develop pre-ovulatory follicles which ovulate and form active CLs that produce enough progesterone to sustain pregnancy. In spite of this, it is possible that the induction of additional CL formation to increase progesterone concentrations prior to embryo transfer could increase the chances of successful pregnancy. In this study an 8-day old mule embryo produced by the insemination of a mare with fresh donkey semen was transferred to a cyclic mule that had been induced to form a secondary CL by the injection of 1.8mg of deslorelin acetate on day 8 post-ovulation, when a spontaneously developed follicle reached 38 mm of diameter. Progesterone concentrations at that time confirmed that the primary CL was active. Secondary ovulation occurred 2 days later and a secondary CLformed during the following days, resulting in a significant increase in blood progesterone concentrations. The 8-day old embryo was transferred nine days after the secondary ovulation (19 days after the primary ovulation), when both the primary and secondary CL were fully active as indicated by sustained progesterone concentrations and by their diameters of 37 and 35.5 mm, respectively. Pregnancy diagnosis was made 4 days after transfer of the embryo, that is, 23 days after the primary ovulation and 13 days after the secondary ovulation. At that time, the 12-day old embryo was within a 15mm embryonic vesicle. On days 55 and 58 of pregnancy two new supplementary CL formed spontaneously, reaching diameters of 36 and 29mm. This report of a successful mule-in-mule pregnancy using as a recipient a mule with both a primary and an artificially induced secondary CL suggests the possibility of improving the chances of successful pregnancy in recipient mules by increasing endogenous progesterone production through induction of secondary ovulations prior to embryo transfer. Acknowledgements. This study was supported by grant PAPIIT IN 223920, DGAPA-UNAM.

Proteomic characterization of eight-day-old equine blastocysts produced in vivo

Early embryo development is dependent on molecular signatures of the gametes, gene expression and maternal-embryo interactions. The proteome of the embryo reflects its developmental state and defines its fate. The present study aimed to describe the proteome of 8-day old equine blastocysts produced in vivo. Follicular dynamics of eight mares were monitored by ultrasonography. When a preovulatory follicle was confirmed, mares received GnRH and were inseminated. After eight days, four embryos were recovered. Proteins were extracted from the embryos, pooled and subjected to mass spectrometry. Four bioinformatic platforms were used to increase the accuracy of the protein identification as they have different capacities and coverages. Overall, there was a list of 4,014 proteins, but there were overlaps among the bioinformatic search engines. After elimination of redundancies (at 3 levels, based on identifiers, peptides and cross-bank mapping), a final count of 1,977 proteins were identified reliably in the samples of equine embryos. There were 1,413 proteins identified by the Peaks program, 316 by Proteome Discoverer, 165 by SearchGUI, and 83 proteins using the PepExplorer program. The low number of overlaps observed for PepExploreris due to the nature of that particular search, which used peptides sequenced but not identified by the Peaks program. Biological processes of the embryo proteins identified included cellular and metabolic processes. Cellular components involved the cell, organelle and protein-containing complex. Molecular functions of embryo proteins included binding, catalytic activities and structural molecule activity. Six pathways relevant to embryo development (FDR < 0.05, Panther platform) related to the TCA cycle, pyruvate metabolism, glycolysis, purine metabolism, and the pentose phosphate pathway. Proteins of the 8-day old equine embryos were associated with pathways of cell-cell communication and extracellular matrix remodeling and pathways related to intracellular remodeling, such as cytoskeleton regulation by Rho GTPase. In summary, the current approach based on mass spectrometry and a combination of search engines allowed the identification of 1,977 proteins in 8-day-old equine embryos. This is, to date, the largest atlas of proteins from embryos of the equine species. Knowledge of protein expression by the embryo helps to understand its physiology, metabolic events and future identification of biomarkers for health.

Influence of an equine viral arteritis (EVA) outbreak on embryo recovery, pregnancy and live foal rates at an insemination and embryo transfer center in Northern Germany 2020

Equine viral arteritis (EVA) is a contagious disease caused by the equine arteritis virus (EAV). Although EVA outbreaks occur infrequently, EAV is present in different horse populations in many countries and can cause high economic loss. EVA is of special concern to horse breeders because it can cause abortion in pregnant mares, death in very young foals and infected stallions might become permanent carriers of the virus. Most horses exposed to EAV will develop no signs of the disease. If clinical illness does occur, EVA can be difficult to diagnose because it is clinically similar to several other equine diseases. Incubation period varies between 2 – 14 days, depending on the route of exposure. The interval can be 2 – 3 days after respiratory exposure and 6 – 8 days after venereal transmission of the virus. (Balasuriya.et.al. Equine Infectious Diseases (Second Edition). 2014; 169-181). In total, 72 horses (64 mares, 8 foals) were based at the stud at the time of confirmed outbreak. The first clinical signs of fever of unknown origin began on 1st June 2020. Blood samples of all horses were taken on 5th June 2020 and showed a 42.0% seroconversion via ELISA and VNT. PCR testing was performed on nasal swabs and viral DNA was detected in 8.0% of horses. A consecutive blood sample two weeks later showed88.0% seroconversion. Embryo collection was carried out using a closed system. An experienced team of veterinarians transferred all embryos non-surgically using the Wilsher-technique (Wilsher.et.al. Equine Vet Edu 2004; 16:1:39-44). Retrospective data analysis of embryo recovery rate, 7-day post embryo transfer pregnancy rate- and live foal rate were calculated for Period One (P1, before possible incubation period, 13th February 2020 – 30th April 2020) and Period Two (P2, incubation period and outbreak, 1st May 2020 – 8th June 2020). Statistical analysis was performed using Chi-Square-Test. A total of 100 embryo flushes were performed in P1 and 116 in P2. Embryo recovery rates were 66.0% for P1 and 63.8% for P2 (n.s.), pregnancy rates after transfer were 90.9% for P1 and 71.6% for P2 (n.s.). Live foal rates after a positive 7-day-scan were 93.3% for P1 and 77.4% for P2 (p<0.05). Foal survival rate was 69.8% in P2 compared to 91.7% in P1, when comparing the number of live foals at 14 days post-partum with the number of positive pregnancies (p<0.01). Apart from the possible economic impact, EVA seems to have a great effect on the live foal rate and foal survival, but not on the actual positive embryo recovery rate and initial 7-day-post-embryo transfer pregnancy rate.

Neuroprotective effect of thyroid hormones on methamphetamine-induced neurotoxicity via cell surface receptors

Thyroid hormones (THs) have an essential role in normal brain development and function. Methamphetamine (MA) is a widely abused psychostimulant that induces irreversible damages to neuronal cells. In the current study, we used rat primary hippocampal neurons (PHNs) to investigate the neuroprotective effect of THs against MA neurotoxicity. PHNs were prepared from 18-day rat embryos and cell viability was assessed using MTT assay, following treatment with various concentrations of MA, T3, T4 or tetrac, an integrin αvβ3 cell surface receptor antagonist. Our results showed that 7 mM MA induced an approximately 50 % reduction in the PHNs viability. Treatment with 800 nM T3 or 8 μM T4 protected PHNs against MA toxicity, an effect which was blocked in the presence of tetrac. These findings suggest that THs protect PHNs against MA-induced cell death by the activation of integrin αvβ3 cell surface receptors. So, targeting integrin αvβ3 receptors or using THs can be considered as promising therapeutic strategies to overcome MA neurotoxicity.

The Amphibian Short-Term Assay: Evaluation of a New Ecotoxicological Method for Amphibians Using Two Organophosphate Pesticides Commonly Found in Nature-Assessment of Biochemical, Morphological, and Life-History Traits.

Amphibia is the most threatened class among vertebrates, with >40% of the species threatened with extinction. Pollution is thought to alter amphibian population dynamics. With the growing interest in behavioral ecotoxicology, the neurotoxic organophosphate pesticides are of special concern. Understanding how exposure to neurotoxics leads to behavioral alterations is of crucial importance, and mechanistic endpoints should be included in ecotoxicological methods. In the present study, we tested an 8-day assay to evaluate the toxicity of two organophosphates, diazinon and chlorpyrifos, on Xenopus laevis, that is, on biochemical, morphological, and life-history traits related to locomotion capacities. The method involves measuring biomarkers such as glutathione-S-transferase (GST) and ethoxyresorufin-O-deethylase (EROD; two indicators of the detoxifying system) in the 8-day-old larvae as well as acetylcholinesterase (AChE) activity (involved in the nervous system) in 4-day-old embryos and 8-day-old larvae. Snout-to-vent length and snout-to-tail length of 4-day-old embryos and 8-day larvae were recorded as well as the corresponding growth rate. Fin and tail muscle widths were measured as well for testing changes in tail shape. Both tests showed effects of both organophosphates on AChE activity; however, no changes were observed in GST and EROD. Furthermore, exposure to chlorpyrifos demonstrated impacts on morphological and life-history traits, presaging alteration of locomotor traits. In addition, the results suggest a lower sensitivity to chlorpyrifos of 4-day-old embryos compared to 8-day-old larvae. Tests on other organophosphates are needed to test the validity of this method for the whole organophosphate group. Environ Toxicol Chem 2022;41:2688-2699. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Transcriptome Integration Analysis at Different Embryonic Ages Reveals Key lncRNAs and mRNAs for Chicken Skeletal Muscle.

The growth and development of skeletal muscle at embryonic stages are vital and it directly affects the growth performance of chickens. Long non-coding RNA (lncRNA) plays an important role in this process. In the experiment, we collected the leg muscles of fast- and slow-growing Bian chickens both at 14- and 20-day embryo ages (14E and 20E) for RNA-seq. Finally, 292 and 347 differentially expressed (DE) lncRNAs were identified in F14vsF20 and S14vsS20, and 1,295 and 1,560 DE mRNAs were also screened, respectively. Then we constructed lncRNA-mRNA networks for the two groups, respectively, and found that 6 of the top 10 lncRNAs ranked with degree are same. GO analysis showed that 12 of the top 20 terms were same in the two comparison groups and most of them were related to energy metabolisms, such as cellular respiration and aerobic respiration. KEGG enrichment revealed that up to 16 pathways of the top 20 in F14vsF20 were same as that of S14vsS20 and most of them were related to growth, including citrate cycle (TCA cycle) and oxidative phosphorylation. Further analysis showed that there were 602 and 102 same DE mRNAs and DE lncRNAs between the two comparison groups. We then identified 442 lncRNA-mRNA pairs, including 201 mRNAs and 32 lncRNAs. Protein-Protein Interactions (PPI) network was predicted for the 201 mRNAs and three core networks were obtained using the plug-in MCODE of Cytoscape. Then the function of genes in the three core networks was further analyzed with ClueGo and they were mainly enriched in six groups of biological processes. On this basis, combined with KEGG pathways and lncRNA-mRNA networks, we identified several candidate lncRNAs and mRNAs. Among them, lncRNAs mainly include TCONS_00061389, TCONS_00025495, TCONS_00017622, TCONS_00216258 and TCONS_00084223, and mRNAs include PLK1, BUB1, TTK, NDUFS7 NDUFAB1, PDHA1, CDK1, SDHA, ACO2 and MDH1. The results would provide a foundation for further experiments on the role of lncRNAs in the regulation of muscle development. And it could also contribute to further clarify the regulatory mechanism of chicken skeletal muscle.

Identification of crucial circRNAs in skeletal muscle during chicken embryonic development

BackgroundChicken provides humans with a large amount of animal protein every year, in which skeletal muscle plays a leading role. The embryonic skeletal muscle development determines the number of muscle fibers and will affect the muscle production of chickens. CircRNAs are involved in a variety of important biological processes, including muscle development. However, studies on circRNAs in the chicken embryo muscle development are still lacking.ResultsIn the study, we collected chicken leg muscles at 14 and 20-day embryo ages both in the fast- and slow-growing groups for RNA-seq. We identified 245 and 440 differentially expressed (DE) circRNAs in the comparison group F14vsF20 and S14vsS20 respectively. GO enrichment analysis for the host genes of DE circRNAs showed that biological process (BP) terms in the top 20 related to growth in F14vsF20 were found such as positive regulation of transcription involved in G1/S phase of mitotic cell cycle, multicellular organismal macromolecule metabolic process, and multicellular organismal metabolic process. In group S14vsS20, we also found some BP terms associated with growth in the top 20 including actomyosin structure organization, actin cytoskeleton organization and myofibril assembly. A total of 7 significantly enriched pathways were obtained, containing Adherens junction and Tight junction. Further analysis of those pathways found three crucial host genes MYH9, YBX3, IGF1R in both fast- and slow-growing groups, three important host genes CTNNA3, AFDN and CREBBP only in the fast-growing group, and six host genes FGFR2, ACTN2, COL1A2, CDC42, DOCK1 and MYL3 only in the slow-growing group. In addition, circRNA-miRNA network also revealed some key regulation pairs such as novel_circ_0007646-miR-1625-5p, novel_circ_0007646-miR-1680-5p, novel_circ_0008913-miR-148b-5p, novel_circ_0008906-miR-148b-5p and novel_circ_0001640-miR-1759-3p.ConclusionsComprehensive analysis of circRNAs and their targets would contribute to a better understanding of the molecular mechanisms in poultry skeletal muscle and it also plays an important guiding role in the next research.

Identifying Positive Cases in an Avian Model of Induced-Thyrotoxicosis

Thyrotoxicosis is a condition of excess circulating thyroid hormones. Maternal thyrotoxicosis has been related to the development of craniosynostosis (CS) in approximately 3% of births. CS occurs when the cranial sutures of the skull fuse prematurely. However, the mechanism for thyroid hormone-induced CS in those cases is unclear. Currently, we are establishing an avian model of thyroxine-induced CS to study these mechanisms. Two groups of fertilized chicken eggs were injected with saline or 25 ng T4 (thyroxine) on embryonic days 11 and 15 and collected on 19 prior to hatching. Data were collected from each group including: body mass and length, tibia and femur length, liver and heart mass, and circulating T4 concentrations. Our study aimed to characterize all possible physiological changes associated with our treatment and determine the subset of positive cases. Following thyroxine exposure, 19-day old embryos were 4% shorter (p&lt;0.05) with a corresponding 8% decrease in tibia length (p&lt;0.01) and 9% decrease in femur length (p&lt;0.01) compared to the controls. Measured T4 concentrations were 88% higher in serum samples following thyroxine-exposure compared to the controls (p&lt;0.01). Body, heart, and liver mass were not different between groups. We were then able to isolate positive cases from correlation plots based on clustering observed in anticipated regions. In conclusion these results support that there is a subset of samples within the treatment group that display the expected phenotypes. It is in this subgroup of data that our focus can be shifted to study the mechanism of thyroxine-induced CS.&#x0D; Acknowledgment of NASA West Virginia Space Grant Consortium (Grant #80NSSC20M0055) and the Genomics Core Facility and WV-INBRE (NIH grant P20GM103434).

Comparative Analysis of miRNA Expression Profiles in Skeletal Muscle of Bian Chickens at Different Embryonic Ages.

Simple SummaryThe development of skeletal muscle during the embryonic period has an important impact on the meat production of poultry, and miRNAs are vital regulators of skeletal muscle development. In this study, through the RNA-seq analysis of leg muscles from 14- and 20-day Bian chicken embryos, we screened out 127 and 131 differentially expressed miRNAs (DEMs) in a fast-growing group (F14 vs. F20) and a slow-growing group (S14 vs. S20), respectively. Based on functional enrichment analysis and interaction network analysis, we further screened out 13 miRNAs and 5 hub genes, which potentially play important roles in skeletal muscle development. This study provides data support for future studies on the regulatory mechanisms of miRNAs in the skeletal muscle development of Bian chickens.MicroRNAs (miRNAs) are widely involved in the growth and development of skeletal muscle through the negative regulation of target genes. In order to screen out the differentially expressed miRNAs (DEMs) associated with skeletal muscle development of Bian chickens at different embryonic ages, we used the leg muscles of fast-growing and slow-growing Bian chickens at the 14th and 20th embryonic ages (F14, F20, S14 and S20) for RNA-seq. A total of 836 known miRNAs were identified, and 121 novel miRNAs were predicted. In the F14 vs. F20 comparison group, 127 DEMs were screened, targeting a total of 2871 genes, with 61 miRNAs significantly upregulated and 66 miRNAs significantly downregulated. In the S14 vs. S20 comparison group, 131 DEMs were screened, targeting a total of 3236 genes, with 60 miRNAs significantly upregulated and 71 miRNAs significantly downregulated. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that the predicted target genes were significantly enriched in 706 GO terms and 6 KEGG pathways in the F14 vs. F20 group and 677 GO terms and 5 KEGG pathways in the S14 vs. S20 group. According to the interaction network analysis, we screened five coexpressed DEMs (gga-miR-146a-3p, gga-miR-2954, gga-miR-34a-5p, gga-miR-1625-5p and gga-miR-18b-3p) with the highest connectivity degree with predicted target genes between the two comparison groups, and five hub genes (HSPA5, PKM2, Notch1, Notch2 and RBPJ) related to muscle development were obtained as well. Subsequently, we further identified nine DEMs (gga-let-7g-3p, gga-miR-490-3p, gga-miR-6660-3p, gga-miR-12223-5p, novel-miR-327, gga-miR-18a-5p, gga-miR-18b-5p, gga-miR-34a-5p and gga-miR-1677-3p) with a targeting relationship to the hub genes, suggesting that they may play important roles in the muscle development of Bian chickens. This study reveals the miRNA differences in skeletal muscle development between 14- and 20-day embryos of Bian chickens from fast- and slow-growing groups and provides a miRNA database for further studies on the molecular mechanisms of the skeletal muscle development in Bian chickens.

Conventional biotyping revealed Mycoplasma and antimicrobial resistant Escherichia coli and Salmonella isolates during disease outbreak in broiler farms and hatcheries in Leyte, Philippines

A respiratory disease outbreak investigation was carried out from October to December 2015 in three broiler farms and two hatcheries in Leyte. Three- to seven-day-old chicks from farms were initially brought to CVM diagnostic laboratory for necropsy and microbial analyses. Subsequent visits to hatcheries and farms were made for random swabbing and sampling of physical facilities, equipment, vehicle, unhatched eggs, 18-day old eggs, eggshells, and day-old chicks. Chick necropsy, embryo examination, and bacteriological method for Mycoplasma, Escherichia coli, Salmonella, and Aspergillus were performed. Antimicrobial sensitivity testing was conducted for Salmonella and E. coli isolates. The main clinical sign observed in chicks was dyspnea. Necropsy revealed consistent bilateral fibrinous airsacculitis and perihepatitis with few cases of pericarditis, wherein sampled organs consistently showed the presence of E. coli. Out of 83 samples collected, 19 (22.89%) Salmonella, 40 (48.19%) E. coli, and no Aspergillus were isolated. Most E. coli were isolated from chick embryos and necropsied chickens, while Salmonella isolates were mostly from the environment. Mycoplasma was isolated from 63.64% (7/11) of sampled lungs, air sacs, and liver from unhatched embryos (100%), 18-day old embryos (50%), and day-old chicks (33.33%). The majority of Salmonella and E. coli isolates showed multidrug resistance against amoxicillin, chloramphenicol, erythromycin, trimethoprim/sulfamethoxazole (TMPS), and tetracycline. Few isolates were sensitive to ciprofloxacin. Our results should raise awareness on the occurrence of antimicrobial-resistant E. coli and Salmonella, and coinfection with Mycoplasma among broilers in Leyte. Therefore, proper selection and usage of antibiotics should be advocated in all poultry farms.

Protective effect of mesenchymal stem cell-derived exosomal treatment of hippocampal neurons against oxygen-glucose deprivation/reperfusion-induced injury.

Individuals who survive a cardiac arrest often sustain cognitive impairments due to ischemia-reperfusion injury. Mesenchymal stem cell (MSC) transplantation is used to reduce tissue damage, but exosomes are more stable and highly conserved than MSCs. This study was conducted to investigate the therapeutic effects of MSC-derived exosomes (MSC-Exo) on cerebral ischemia-reperfusion injury in an in vitro model of oxygen-glucose deprivation/reperfusion (OGD/R), and to explore the underlying mechanisms. Primary hippocampal neurons obtained from 18-day Sprague-Dawley rat embryos were subjected to OGD/R treatment, with or without MSC-Exo treatment. Exosomal integration, cell viability, mitochondrial membrane potential, and generation of reactive oxygen species (ROS) were examined. Terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick-end labeling (TUNEL) staining was performed to detect neuronal apoptosis. Moreover, mitochondrial function-associated gene expression, Nrf2 translocation, and expression of downstream antioxidant proteins were determined. MSC-Exo attenuated OGD/R-induced neuronal apoptosis and decreased ROS generation (P<0.05). The exosomes reduced OGD/R-induced Nrf2 translocation into the nucleus (2.14±0.65 vs. 5.48±1.09, P<0.01) and increased the intracellular expression of antioxidative proteins, including superoxide dismutase and glutathione peroxidase (17.18±0.97 vs. 14.40±0.62, and 20.65±2.23 vs. 16.44±2.05, respectively; P<0.05 for both). OGD/R significantly impaired the mitochondrial membrane potential and modulated the expression of mitochondrial function-associated genes, such as PINK, DJ1, LRRK2, Mfn-1, Mfn-2, and OPA1. The abovementioned changes were partially reversed by exosomal treatment of the hippocampal neurons. MSC-Exo treatment can alleviate OGD/R-induced oxidative stress and dysregulation of mitochondrial function-associated genes in hippocampal neurons. Therefore, MSC-Exo might be a potential therapeutic strategy to prevent OGD/R-induced neuronal injury.

Norepinephrine regulates osteogenesis in the embryonic period of development

Sympathetic and sensory nerve fibers regulate osteosynthesis and osteoresorption processes throughout life. Fundamental research and clinical data confirm the existence of functional interactions between neurons and bone tissue cells and indicate the catabolic and anabolic effect of sympathetic nervous system mediators on bone tissue. There is practically no information about the regulation of osteoremodeling in embryogenesis. Objective: to study the effect of norepinephrine on the growth of bone tissue explants in the embryonic period of development. The studies were performed on the bone tissue explants of 12-day old chicken embryos. Norepinephrine (10-10 M - 10-4 M), propranolol (10-10 M), atenolol (10-4 M), urapidil (10-6 M) were added to the experimental Petri dishes. Norepinephrine (10-6 M) stimulates the growth of the bone tissue explants through α1-adrenoreceptors. The osteotoxic effect of high doses of the drug is realized through β2-adrenoreceptors. Embryonic osteogenesis is regulated by norepinephrine in dose-dependent manner. Physiological effect of the substance depends on the interaction with certain types of adrenoreceptors.

USE OF INVOCELL INTRAVAGINAL EMBRYO CULTURE SYSTEM FOR FIVE DAYS OF INCUBATION: A PROSPECTIVE STUDY OF SAFETY AND TOLERABILITY

The purpose of this single-center study was to prospectively evaluate the INVOcell culture device with its retention device in planned day-5 embryo transfer cycles, as to its retention, comfort, and possible vaginal mucosal irritation. INVOcell is presently approved in the US for up to 3 days. The study was prospectively approved by WCG IRB (#1295533). Women scheduled for oocyte retrieval planning blastocyst transfer at a single center (N=31) were invited to participate in the study; all gave written informed consent. Two failed screening; 29 were enrolled in the study aged 26-41 (median 36) years with body mass index of 20.1-39.5 (median 30.5) kg/m2. A vaginal speculum exam was done at the completion of each retrieval, and after INVOcell removal at 5 days; before examination of INVOcell contents, all women completed the study questionnaire. The primary endpoint of the study was device retention, determined by patient report. Secondary endpoints of patient comfort and vaginal tissue reaction to the INVOcell system were assessed after its removal by patient questionnaire and speculum exam. The INVOcell system was retained by 100% of participants. One woman felt that the system was dislodged; she was able to readjust the system to maintain it in position. A conservative analytic approach thus gives a 96% rate of successful retention. All women completed the questionnaire, which assessed overall subject comfort and satisfaction with wearing the INVOcell system. Only two subjects reported any discomfort, and that was at a tolerable level not requiring medical intervention. The overall Likert rating for discomfort of 0.17 (pain scale of 0-10 with 0 being no pain/discomfort and 10 being severe pain/discomfort) demonstrates the favorable comfort level for the INVOcell and retention device during 5-day incubation. Whereas bleeding was noted immediately after oocyte retrieval in the majority of women and considered to be a normal sequela of this procedure, at Day-5 vaginal speculum exam no lesions, ulcerations, erythema, or bleeding were noted. In conclusion, our study demonstrates that the INVOcell system can be safely used for a 5-day embryo incubation without causing vaginal irritation or mucosal injury. Our results also verify that the system is well tolerated.

Developmental morphological analyses on the preglottal salivary gland in Japanese quails (Coturnix japonica).

To understand the development of the mucous preglottal salivary gland in Coturnix japonica (Japanese quail), morphological and histochemical studies were performed on 20 healthy Japanese quail embryos (aging from 10th to 17th incubation days) and 25 healthy quail chicks (aging from 0th to 60th days). The primordia of preglottal salivary gland were observed as an epithelial bud at the early embryonic stage, which then elongated and differentiated into secretory units by the end of this stage. In Japanese quails, the preglottal salivary gland was a mucous polystomatic tubuloalveolar unpaired gland composed of two lateral portions and a middle one embedded into the submucosa of the lingual root. The gland openings accompanied taste pore (8.17 μm) of taste buds associated salivary glands type; some skeletal muscle fibers embedded among secretory lobules extended from muscle cricohyoideus at 14th day-old quail chick. Also, both herbts corpuscles and secretory motor plexus could be detected among secretory lobules. Based on our investigations, the development of the preglottal salivary gland could clearly be distinguished in the embryonic stage into pre bud and bud stages at 10th day old, cord and branching stages ended by cavitation at 11th day old, canalization stage at 13th day old, lobulation and secretory stages by the 17th day old. The secretory materials showed different histochemical reactions ended with highly alcinophilic mucous indicated highly sialomucin (acidic) content. Myoepithelial cells could be demonstrated at a 17-day old quail embryo and thereafter surrounded the secretory endpieces of the preglottal salivary gland.

Transcriptome Analysis Reveals the Profile of Long Non-coding RNAs During Chicken Muscle Development.

The developmental complexity of muscle arises from elaborate gene regulation. Long non-coding RNAs (lncRNAs) play critical roles in muscle development through the regulation of transcription and post-transcriptional gene expression. In chickens, previous studies have focused on the lncRNA profile during the embryonic periods, but there are no studies that explore the profile from the embryonic to post-hatching period. Here, we reconstructed 14,793 lncRNA transcripts and identified 2,858 differentially expressed lncRNA transcripts and 4,282 mRNAs from 12-day embryos (E12), 17-day embryos (E17), 1-day post-hatch chicks (D1), 14-day post-hatch chicks (D14), 56-day post-hatch chicks (D56), and 98-day post-hatch chicks (D98), based on our published RNA-seq datasets. We performed co-expression analysis for the differentially expressed lncRNAs and mRNAs, using STEM, and identified two profiles with opposite expression trends: profile 4 with a downregulated pattern and profile 21 with an upregulated pattern. The cis- and trans-regulatory interactions between the lncRNAs and mRNAs were predicted within each profile. Functional analysis of the lncRNA targets showed that lncRNAs in profile 4 contributed to the cell proliferation process, while lncRNAs in profile 21 were mainly involved in metabolism. Our work highlights the lncRNA profiles involved in the development of chicken breast muscle and provides a foundation for further experiments on the role of lncRNAs in the regulation of muscle development.

Innervation of Meissner's corpuscles and Merkel -cells by transplantation of embryonic dorsal root ganglion cells after peripheral nerve section in rats.

Transplantation of embryonic motor neurons has been shown to improve motor neuron survival and innervation of neuromuscular junctions in peripheral nerves. However, there have been no reports regarding transplantation of sensory neurons and innervation of sensory receptors. Therefore, we hypothesized that the transplantation of embryonic sensory neurons may improve sensory neurons in the skin and innervate Merkel cells and Meissner's corpuscles. We obtained sensory neurons from dorsal root ganglia of 14-day rat embryos. We generated a rat model of Wallerian-degeneration by performing sciatic nerve transection and waiting for one week after. Six months after cell transplantation, we performed histological and electrophysiological examinations in naïve control, surgical control, and cell transplantation groups. The number of nerve fibers in the papillary dermis and epidermal-dermal interface was significantly greater in the cell transplantation than in the surgical control group. The percent of Merkel cells with nerve terminals, as well as the average number of Meissner corpuscles with nerve terminals, were higher in the cell transplantation than in the surgical control group, but differences were not significant between the two groups. Moreover, the amplitude and latency of sensory conduction velocity were evoked in rats of the cell transplantation group. We demonstrated that the transplantation of embryonic dorsal root ganglion cells improved sensory nerve fiber number and innervation of Merkel cells and Meissner's corpuscles in peripheral nerves.

Teratogenic Effect of High Dose of Syzygium guineense (Myrtaceae) Leaves on Wistar Albino Rat Embryos and Fetuses.

Syzygium guineense is an important medicinal plant effective against hypertension, diabetes mellitus, and cancer but with no evidence of its teratogenicity. This study was planned to investigate the teratogenic potential of S. guineense leaves on rat embryos and fetuses. Five groups of Wistar albino rats, each consisting of ten pregnant rats, were used as experimental animals. Groups I-III rats were treated with 250, 500, and 1000 mg/kg of hydroethanolic extract of S. guineense leaves, and groups IV and V were control and ad libitum control, respectively. Rats were treated during day 6–12 of gestation. Embryos and fetuses were retrieved at day 12 and day 20 of gestation, respectively. The embryos were assessed for developmental delays and growth retardation. The fetuses were examined for gross external, skeletal, and visceral anomalies. In 12-day old rat embryos, crown-rump length, number of somites, and morphological scores were significantly reduced by the treatment of 1000 mg/kg of the extract. The external morphological and visceral examinations of rat fetuses did not reveal any detectable structural malformations in the cranial, nasal, oral cavities, and visceral organs. The ossification centers of fetal skull, vertebrae, hyoid, forelimb, and hindlimb bones were not significantly varied across all groups. However, even if not statistically significant, high-dose treated rat fetuses had a reduced number of ossification centers in the sternum, caudal vertebrae, metatarsal, metacarpal, and phalanges. Treatment with the hydroethanolic extract of S. guineense leaves produced no significant skeletal and soft tissue malformations. The plant extract did not produce significant teratogenic effects on rat embryos/fetuses up to 500 mg/kg doses but retarded the growth of embryos at high dose (1000 mg/kg) as evidenced by decreased crown-rump length, number of somites, and morphological scores. Therefore, it is not advisable to take large doses of the plant during pregnancy.