Course Of Embryogenesis Research Articles

Effects of host genetics and environment on egg-associated microbiotas in brown trout (Salmo trutta).

Recent studies found fish egg-specific bacterial communities that changed over the course of embryogenesis, suggesting an interaction between the developing host and its microbiota. Indeed, single-strain infections demonstrated that the virulence of opportunistic bacteria is influenced by environmental factors and host immune genes. However, the interplay between a fish embryo host and its microbiota has not been studied yet at the community level. To test whether host genetics affects the assemblage of egg-associated bacteria, adult brown trout (Salmo trutta) were sampled from a natural population. Their gametes were used for full-factorial invitro fertilizations to separate sire from dam effects. In total, 2520 embryos were singly raised under experimental conditions that differently support microbial growth. High-throughput 16S rRNA amplicon sequencing was applied to characterize bacterial communities on milt and fertilized eggs across treatments. Dam and sire identity influenced embryo mortality, time until hatching and composition of egg-associated microbiotas, but no link between bacterial communities on milt and on fertilized eggs could be found. Elevated resources increased embryo mortality and modified bacterial communities with a shift in their putative functional potential. Resource availability did not significantly affect any parental effects on embryo performance. Sire identity affected bacterial diversity that turned out to be a significant predictor of hatching time: embryos associated with high bacterial diversity hatched later. We conclude that both host genetics and the availability of resources define diversity and composition of egg-associated bacterial communities that then affect the life history of their hosts.

Diverse and pervasive subcellular distributions for both coding and long noncoding RNAs.

In a previous analysis of 2300 mRNAs via whole-mount fluorescent in situ hybridization in cellularizing Drosophila embryos, we found that 70% of the transcripts exhibited some form of subcellular localization. To see whether this prevalence is unique to early Drosophila embryos, we examined ∼8000 transcripts over the full course of embryogenesis and ∼800 transcripts in late third instar larval tissues. The numbers and varieties of new subcellular localization patterns are both striking and revealing. In the much larger cells of the third instar larva, virtually all transcripts observed showed subcellular localization in at least one tissue. We also examined the prevalence and variety of localization mechanisms for >100 long noncoding RNAs. All of these were also found to be expressed and subcellularly localized. Thus, subcellular RNA localization appears to be the norm rather than the exception for both coding and noncoding RNAs. These results, which have been annotated and made available on a recompiled database, provide a rich and unique resource for functional gene analyses, some examples of which are provided.

Epigenetic control of the immune system: a lesson from Kabuki syndrome

Kabuki syndrome (KS) is a rare multi-systemic disorder characterized by a distinct face, postnatal growth deficiency, mild-to-moderate intellectual disability, skeletal and visceral (mainly cardiovascular, renal, and skeletal) malformations, dermatoglyphic abnormalities. Its cause is related to mutations of two genes: KMT2D (histone-lysine N-methyltransferase 2D) and KDM6A (lysine-specific demethylase 6A), both functioning as epigenetic modulators through histone modifications in the course of embryogenesis and in several biological processes. Epigenetic regulation is defined as the complex of hereditable modifications to DNA and histone proteins that modulates gene expression in the absence of DNA nucleotide sequence changes. Different human disorders are caused by mutations of genes involved in the epigenetic regulation, and not surprisingly, all these share developmental defects, disturbed growth (in excess or defect), multiple congenital organ malformations, and also hematological and immunological defects. In particular, most KS patients show increased susceptibility to infections and have reduced serum immunoglobulin levels, while some suffer also from autoimmune manifestations, such as idiopathic thrombocytopenic purpura, hemolytic anemia, autoimmune thyroiditis, and vitiligo. Herein we review the immunological aspects of KS and propose a novel model to account for the immune dysfunction observed in this condition.

Embryogenesis of angiosperms: ultrastructural transformations of plastids

Electron microscopic study of <em>Alcea rosea</em> and <em>Valeriana officinalis</em> embryogenesis showed the ultrastructural changes in the embryo plastidome. It is assumed that plastids of the mature angiosperms zygote are of one genetic type. Tissue ditferentiation and changes of plastid functions in the course of embryogenesis and plant development underlie the histological differentiation of plastids and their ontogenic transformations (metamorphosis).

Rape embryogenesis. V. Accumulation of lipid bodies

In embryo of winter rape var. Górczański lipid bodies have been observed in the light microscope starting from two-celled proembryo. Their number and size increase in the course of embryogenesis, especially since greening of endosperm and embryo. Lipid bodies, either single or in clusters, are present in all embryo cells, the clusters occur in various parts of the cytoplasm. During embryo maturation lipid bodies move and locate in a number of layers by the cell wall. At the same time their appearance change. Typical storage lipids originate. Lipid bodies are unevenly distributed within the embryo: their number and size decrease along the embryo axis from its part below cotyledons towards root apex. Moreover, they are histologically diversified: the biggest are located in epidermis and cortex, whereas the smallest -in central cylinder. In columella there are fewest lipid bodies.

Identification of the Precise Kairomone-sensitive Period and Histological Characterization of Necktooth Formation in Predator-induced Polyphenism inDaphnia pulex

Many organisms have the ability to alter their development in the presence of predators, leading to predator-induced defenses that reduce vulnerability to predation. In the water flea Daphnia pulex, small protuberances called 'neckteeth' form in the dorsal neck region in response to kairomone(s) released by predatory phantom midges (Chaoborus larvae). Although previous studies suggested that kairomone sensitivity begins when chemoreceptors begin to function during embryogenesis, the exact critical period was unknown to date. In this study, we investigated the period of kairomone sensitivity and the process of necktooth formation in D. pulex through extensive treatments with pulses of kairomone(s). First, we described the time course of embryogenesis, which we suggest should be used as the standard in future studies. We found the kairomone-sensitive period to be relatively short, extending from embryonic stage 4 to postembryonic first instar. We observed cell proliferation and changes in cell structure in response to the kairomone treatment, and propose a model for necktooth formation. Preliminary LiCl treatment suggests the Wnt signaling pathway involved in crest formation as a candidate for the molecular mechanism underlying this process. Our study provides basic insight toward understanding the mechanisms underlying adaptive polyphenism in D. pulex.

Cell Proliferation Kinetics and Branching Stochastic Processes

The main purpose of this work is to present some new ideas and results obtained in modeling of cell proliferation kinetics. Recent advances in experimental techniques of flow cytometry have made it possible to collect a wealth of information about the status of individual cells isolated from dissociated tissues. When one is interested in modeling tissue development starting from the earliest embryonic stages it is reasonable to begin $% 0$ cells because these cells appear only in the course of embryogenesis. New type cells (immigrants) of age zero arrive in the population of cells in accordance with a non-homogeneous Poisson process with arrival rate $r(t)$. Upon arrival, these immigrants are assumed to be of age zero. Upon completion of its lifespan, every cell either divides into two new cells, or it goes out of the process of proliferation (differentiation or death). These two events occur with probability $p$ and $q=1-p$, respectively. The time to division or differentiation of any cell is described by a non-negative random variable $\tau $ with c.d.f. $G(x)=\mathbf{P}\{\eta \leq x\}$. Cells are assumed to evolve independently of each other. Motivated by the above example, we investigate properties of a class of Markov branching processes with non-homogeneous Poisson immigration. We consider a more general process than the one presented above so that the scope of our work does not remain limited to the study of oligodendrocyte generation. Limiting distributions are obtained in the supercritical case and among them an analogue of the LLN and of the CLT.

The ventral proximal nail fold: stem cell niche of the nail and equivalent to the follicular bulge – a study on developing human skin

Although the bulge is well characterized as a stem cell niche of the hair follicle, comparatively little is known about the location of stem cells in the nail. Herein, we describe the spatiotemporal expression pattern of six stem cell markers in the developing human nail and compared it with the embryonic and fetal human hair follicle. The areas of proliferative activity were additionally examined using labeling with Ki-67. We examined immunohistochemically samples from embryonic and fetal human nail, hair and skin for the expression of cytokeratin 15 (CK15, two clones), cytokeratin 19 (CK19), PHLDA1, CD200, nestin and Ki-67 using standard techniques. CK15 (clone LHK15), CK19 and PHLDA1 are negative in the nail and hair matrix but positive in the ventral proximal nail fold and in the follicular bulge. Over the course of embryogenesis they display a highly specific spatiotemporal expression pattern both in the nail and in the hair follicle. We propose that at least during embryogenesis the proximal ventral nail fold represents the niche for the nail stem cells. In contrast to animal experiments, autoradiographic pulse-chasing studies cannot be performed in human, and immunohistochemical studies are a valid alternative although they have their limitations. Further studies on adult human nail units are suggested.

Molecular Characterization of Zebrafish Embryogenesis via DNA Microarrays and Multiplatform Time Course Metabolomics Studies

One of the greatest strengths of "-omics" technologies is their ability to capture a molecular snapshot of multiple cellular processes simultaneously. Transcriptomics, proteomics, and metabolomics have, individually, been used in wide-ranging studies involving cell lines, tissues, model organisms, and human subjects. Nonetheless, despite the fact that their power lies in the global acquisition of parallel data streams, these methods continue to be employed separately. We highlight work done to merge transcriptomics and metabolomics technologies to study zebrafish (Danio rerio) embryogenesis. We combine information from three bioanalytical platforms, that is, DNA microarrays, (1)H nuclear magnetic resonance ((1)H NMR), and mass spectrometry (MS)-based metabolomics, to identify and provide insights into the organism's developmental regulators. We apply a customized approach to the analysis of such time-ordered measurements to provide temporal profiles that depict the modulation of metabolites and gene transcription. Initially, the three data sets were analyzed individually but later they were fused to highlight the advantages gained through such an integrated approach. Unique challenges posed by fusion of such data are discussed given differences in the measurement error structures, the wide dynamic range for the molecular species, and the analytical platforms used to measure them (i.e., fluorescence ratios, NMR, and MS intensities). Our data analysis reveals that changes in transcript levels at specific developmental stages correlate with previously published data with over 90% accuracy. In addition, transcript profiles exhibited trends that were similar to the accumulation of metabolites over time. Profiles for metabolites such as choline-like compounds (Trimethylamine-N-oxide, phosphocholine, betaine), creatinine/creatine, and other metabolites involved in energy metabolism exhibited a steady increase from 15 hours post fertilization (hpf) to 48 hpf. Other metabolite and transcript profiles were transiently rising and then falling back to baseline. The "house keeping" metabolites such as branched chain amino acids exhibited a steady presence throughout embryogenesis. Although the transcript profiling corresponds to only 16 384 genes, a subset of the total number of genes in the zebrafish genome, we identified examples where gene transcript and metabolite profiles correlate with one another, reflective of a relationship between gene and metabolite regulation over the course of embryogenesis.

Notch and delta mRNAs in early‐stage and mid‐stage drosophila embryos exhibit complementary patterns of protein‐producing potentials

Notch and Delta proteins generate Notch signaling that specifies cell fates during animal development. There is an intriguing phenomenon in Drosophila embryogenesis that has not received much attention and whose significance to embryogenesis is unknown. Notch and Delta mRNAs expressed in early-stage embryos are shorter than their counterparts in mid-stage embryos. We show here that the difference in sizes is due to mRNA 3' processing at alternate polyadenylation sites. While the early-stage Notch mRNA has a lower protein-producing potential than the mid-stage Notch mRNA, the early-stage Delta mRNA has a higher protein-producing potential than the mid-stage Delta mRNA. Our data can explain the complementary patterns of Notch and Delta protein levels in early- and mid-stage embryos. Our data also raise the possibility that the manner and regulation of Notch signaling change in the course of embryogenesis and that this change is effected by 3' UTR and mRNA 3' processing factors.

Actions speak louder

A fluorescence resonance energy transfer (FRET)-based biosensor helps scientists monitor the activation of an essential signaling protein over the course of embryogenesis in Drosophila melanogaster.

Novel crosstalk to BMP signalling: cGMP-dependent kinase I modulates BMP receptor and Smad activity

Integration of multiple signals into the canonical BMP/Smad pathway poses a big challenge during the course of embryogenesis and tissue homeostasis. Here, we show that cyclic guanosine 3',5'-monophosphate (cGMP)-dependent kinase I (cGKI) modulates BMP receptors and Smads, providing a novel mechanism enhancing BMP signalling. cGKI, a key mediator of vasodilation and hypertension diseases, interacts with and phosphorylates the BMP type II receptor (BMPRII). In response to BMP-2, cGKI then dissociates from the receptors, associates with activated Smads, and undergoes nuclear translocation. In the nucleus, cGKI binds with Smad1 and the general transcription factor TFII-I to promoters of BMP target genes such as Id1 to enhance transcriptional activation. Accordingly, cGKI has a dual function in BMP signalling: (1) it modulates BMP receptor/Smad activity at the plasma membrane and (2) after redistribution to the nucleus, it further regulates transcription as a nuclear co-factor for Smads. Consequently, cellular defects caused by mutations in BMPRII, found in pulmonary arterial hypertension patients, were compensated through cGKI, supporting the positive action of cGKI on BMP-induced Smad signalling downstream of the receptors.

Germline Competent Embryonic Stem Cells Derived from Rat Blastocysts

Rats have important advantages over mice as an experimental system for physiological and pharmacological investigations. The lack of rat embryonic stem (ES) cells has restricted the availability of transgenic technologies to create genetic models in this species. Here, we show that rat ES cells can be efficiently derived, propagated, and genetically manipulated in the presence of small molecules that specifically inhibit GSK3, MEK, and FGF receptor tyrosine kinases. These rat ES cells express pluripotency markers and retain the capacity to differentiate into derivatives of all three germ layers. Most importantly, they can produce high rates of chimerism when reintroduced into early stage embryos and can transmit through the germline. Establishment of authentic rat ES cells will make possible sophisticated genetic manipulation to create models for the study of human diseases.

Epigenetic inactivation of the RB1 gene as a factor of genomic instability: A possible contribution to etiology of chromosomal mosaicism during human embryo development

The methylation status of the cell cycle control gene RB1 has been studied in placental tissues of spontaneous abortions of the first trimester of pregnancy with mosaic variants of numerical chromosomal abnormalities verified by a molecular genetic examination. Aberrant methylation of the gene promoter region has been revealed for the first time in 20% of embryos with chromosomal mosaicism that died in utero. A maximum frequency of epimutations was recorded in a group of embryos with a low level of abnormal cells for which mitotic errors are most likely to determine the formation of mosaic aneuploidy in primary euploid zygotes. It has been suggested that aberrant epigenetic genomic modifications at early stages of human embryonic development can be one of the mechanisms promoting genomic instability realized in the form of mosaic abnormalities of the karyotype that are incompatible with the normal course of embryogenesis.

Abortive embryogenesis in hybrid swarm populations of Pinus sylvestris L. and Pinus muga Turra

Comparative study on fertilization process in Pinus sylvestris, Pinus mugo and in their putative hybrid swarm individuals was done involving pre-zygotic and post-zygotic stages. The amount of surviving ovules from open pollination reflecting the mode of interaction between pollen grains and nucellar tissue of an ovule averaged at 8.1 of sound ovules per conelet in Pinus sylvestris, 7.3 ovules in the hybrid swarm population and at 4.9 ovules in Pinus mugo. A strong correlation was observed between the number of surviving ovules and the proportion of germinating seeds in the compared species and hybrids. Normal course of embryogenesis in Pinus sylvestris and Pinus mugo contrasted with increased frequency of disturbances observed in the hybrid swarm individuals. The differential survival rates of the ovules and deviations from typical pattern of embryogenesis are discussed from the standpoint of cross-ability relationship between Pinus sylvestris and Pinus mugo.

The Role of Phospholipid Hydroperoxide Glutathione Peroxidase Isoforms in Murine Embryogenesis

Phospholipid hydroperoxide glutathione peroxidase (GPx4) is a selenocysteine-containing enzyme, and three different isoforms (cytosolic, mitochondrial, and nuclear) originate from the GPx4 gene. Homozygous GPx4-deficient mice die in utero at midgestation, since they fail to initiate gastrulation and do not develop embryonic cavities. To investigate the biological basis for embryonic lethality, we first explored expression of the GPx4 in adult murine brain and found expression of the protein in cerebral neurons. Next, we profiled mRNA expression during the time course of embryogenesis (embryonic days 6.5-17.5 (E6.5-17.5)) and detected mitochondrial and cytosolic mRNA species at high concentrations. In contrast, the nuclear isoform was only expressed in small amounts. Cytosolic GPx4 mRNA was present at constant levels (about 100 copies per 1000 copies of glyceraldehyde-3-phosphate dehydrogenase mRNA), whereas nuclear and mitochondrial isoforms were down-regulated between E14.5 and E17.5. In situ hybridization indicated expression of GPx4 isoforms in all developing germ layers during gastrulation and in the somite stage in the developing central nervous system and in the heart. When we silenced expression of GPx4 isoforms during in vitro embryogenesis using short interfering RNA technology, we observed that knockdown of mitochondrial GPx4 strongly impaired segmentation of rhombomeres 5 and 6 during hindbrain development and induced cerebral apoptosis. In contrast, silencing expression of the nuclear isoform led to retardations in atrium formation. Taken together, our data indicate specific expression of GPx4 isoforms in embryonic brain and heart and strongly suggest a role of this enzyme in organogenesis. These findings may explain in part intrauterine lethality of GPx4 knock-out mice.

A homologue of the vertebrate SET domain and zinc finger protein Blimp-1 regulates terminal differentiation of the tracheal system in the Drosophila embryo

The B-lymphocyte-inducing maturation protein (Blimp-1) gene encodes a zinc finger and SET/PR domain-containing transcriptional factor. A number of functional studies in a variety of vertebrate species have demonstrated that Blimp-1 is a master regulator of cell fate determination and cell differentiation in a wide diversity of developmental contexts. Despite all of this significance, the role, if any, of a homologue of Blimp-1 in directing morphogenetic events during embryonic development of invertebrates has so far remained completely unexplored. In this report, we describe the identification of a Drosophila homologue of Blimp-1 and show that the gene is expressed in diverse cell types during the course of embryogenesis. Further, using genetic analysis, we demonstrate that its wild-type activity is critically required for the maturation of the tracheal system into properly differentiated tubes.

Unexpected Difficult Intubation due to Lingual Thyroid: A case report

Lingual thyroid is a rare clinical entity that is due to the failure of the thyroid gland to descend early in the course of embryogenesis. It may be present with symptoms of dysphagia and upper airway obstruction. We report here on the case of a 63-year-old female who was scheduled for an operation for lumbar disc herniation and she could not be intubated. The cause of the airway obstruction was an ectopic thyroid at the base of the tongue, which made visualization of the glottis impossible. Several attempts at endotracheal intubation were unsuccessful. Fortunately, the patient was mask ventilated. We awakened the patient and consulted an otolaryngologist, and she was diagnosed with lingual thyroid. After a week, she was operated on using spinal anesthesia. (Korean J Anesthesiol 2006; 51: 236~8)

Specification of midbrain territory.

The vertebrate neural plate is subdivided into four distinct territories comprising the presumptive forebrain, midbrain, hindbrain, and the spinal cord, shortly after gastrulation. Initially, this subdivision relies on a defined pattern of expression of distinct transcription and secreted factors within the newly formed neuroectoderm, even before morphological partitioning is evident. Subdivision of the neural plate into distinct territories is a complex process, which is also known as patterning or regionalisation, and involves both planar and vertical signals coming from within the neuroectoderm and from neighbouring non-neural tissues. During the course of embryogenesis, this gross subdivision of the neural plate is progressively refined by a variety of mechanisms, leading to the establishment of various subdomains that ultimately give rise to specific cell populations characteristic for the corresponding brain and spinal cord regions. Once again, a prominent feature of these later processes is the defined expression of specific genes within the developing neural tube. In the present review, we will concentrate on the genes active in the progressive refinement of the midbrain territory as a distinct subdivision of the brain. We will also give an outlook on genes that are active during early induction of the anterior neural plate and genetic mechanisms that control the generation of specific cell populations of the ventral midbrain, with special focus on the mesencephalic dopaminergic neurons.

Stimulatory Effect of Insulin and Epidermal Growth Factor on Activity of Protein Kinase A, Glucose-6-Phosphate Dehydrogenase, and Glycogen Synthase in Skeletal Muscles of Chick Embryos and Chickens

Insulin is able to produce two types of regulatory effects on muscles—metabolic and growth stimulating. Study of the effect of insulin and epidermal growth factor (EGF) on activity of cAMP-dependent protein kinase (PKA), glucose-6-phosphate dehydrogenase (G-6PDH), a starting enzyme of pentosephosphate pathway (PPP), and glycogen synthase (GS), a key enzyme of the glycogen synthesis, has shown that both types of the insulin effects do not arise simultaneously in the course of embryogenesis. The growth-stimulating effect mediated by adenylyl cyclase—cAMP is revealed since the 10th embryonal day. It was established for the first time that insulin could participate in growth stimulation by activating PKA in vivo and in vitro in muscles of the 10–14-day old embryos and the 8–10-day old chickens. The stimulating effect of insulin on G-6PDH activity is revealed since the same embryonal period and gradually increases. Insulin in vivo and in vitro simulates the glycogen synthase activity by increasing its conversion from non-active to active (GS-I) form only in the 15-day old embryos and in chickens. The stimulating effects of insulin and EGF on both G-6PDH activity (in embryos and chickens) and GS (in chickens) was shown to blocked by selective inhibitors of tyrosine kinases, thyrphostin 47 and genestein, in the dose-dependent manner, which indicates involvement of receptor of the tyrosine kinase type in these effects. The complex of the established facts permits concluding that insulin at early embryonal stages stimulates in the chicken muscles the PKA and G-6PDH activities involved in action of this hormone on growth, which is especially pronounced at the stage of myoblast proliferation. Meanwhile, the metabolic insulin effect (stimulation of the glycogen synthase system) appears in the second half of embryonal period and coincides in time with the period of muscle cell differentiation and active muscle contractures.