Days Of Embryonic Life Research Articles

The eighth International Forum on Stem Cells: virtual meeting, October 20-21, 2022.

The eighth International Forum on Stem Cells: virtual meeting, October 20-21, 2022.

Late diagnosis of Neural Tube Defects: About two uncommon cases

Background: Neural tube defects are pathologies of the central nervous system occurring between the 23rd and 27th day of embryonic life. Classified into two groups: cephalic pole anomaly (anencephaly) and spinal pole anomaly (spina bifida), these are uncommon anomalies representing one case per 1000 births, often resulting in therapeutic termination of pregnancy. The etiopathogenesis is not yet known but an interaction of genetic, environmental and drug factors is implicated. Folic acid supplementation is a means of prevention. Ultrasound is an essential and increasingly effective means of antenatal screening. Our objective is to report two uncommon cases of neural tube closure anomalies diagnosed at birth due to unmonitored pregnancies.

Pentalogy of Cantrell with encephalocele – A case report with review of literature

Pentalogy of Cantrell is a rare structural disorder. It is a combination of congenital defects in the midline comprising of supraumbilical abdominal wall defect, defect in the lower sternum, deficient anterior diaphragm, defect in diaphragmatic pericardium and intracardiac defects. It is proposed to result from an insult occurring as early as 14 to 17 day of embryonic life. It involves the ventral midline due to disruption of midline mesenchyme. Etio-pathogenesis is still largely unknown. The insult could be environmental or disruption of a single gene, either autosomal or X linked. Very rarely Cantrell’s syndrome is associated with neural tube disorders. Here, we report one such rare case of Pentalogy of Cantrell.

Distinct kinetic mechanisms of H3K4 methylation catalyzed by MLL3 and MLL4 core complexes

The methyltransferases MLL3 and MLL4 primarily catalyze the monomethylation of histone H3 lysine 4 (H3K4) on enhancers to regulate cell-type-specific gene expression and cell fate transition. MLL3 and MLL4 share almost identical binding partners and biochemical activities, but perform specific and nonredundant functions. The features and functions that distinguish MLL3 and MLL4 remain elusive. Here, we characterize the kinetic mechanisms of MLL3 and MLL4 ternary complexes containing the catalytic SET domain from MLL3 or MLL4 (MLL3SET or MLL4SET), the SPRY domain of ASH2L (ASH2LSPRY), and a short fragment of RBBP5 (RBBP5AS-ABM) to search for possible explanations. Steady-state kinetic analyses and inhibition studies reveal that the MLL3 complex catalyzes methylation in a random sequential bi–bi mechanism. In contrast, the MLL4 complex adopts an ordered sequential bi–bi mechanism, in which the cofactor S-adenosylmethionine (AdoMet) binds to the enzyme prior to the H3 peptide, and the methylated H3 peptide dissociates from the enzyme before S-adenosylhomocysteine (AdoHcy) detaches after methylation. Substrate-binding assays using fluorescence polarization (FP) confirm that AdoMet binding is a prerequisite for H3 binding for the MLL4 complex but not for the MLL3 complex. Molecular dynamic simulations reveal that the binding of AdoMet exclusively induces conformational constraints on the AdoMet-binding groove and the H3 substrate-binding pocket of MLL4, therefore stabilizing a specific active conformation to ease entry of the substrate H3. The distinct kinetic mechanisms and conformational plasticities provide important insights into the differential functions of MLL3 and MLL4 and may also guide the development of selective inhibitors targeting MLL3 or MLL4.

Circulation Index as a Quantitative Indicator of Angiogenesis in Chorioallantoic Membrane of Chicken Broilers

Summary Chorioallantoic membrane (CAM) is an extraembryonic membrane very frequently used for in vivo studies in various researches. Since researches require a fast method for quantifying the CAM angiogenic response, there is a need to develop a new precise and unbiased method of quantification of angiogenesis in CAM, which would be easy to perform and suitable for analysis of a large number of samples. The objective of this paper is to apply a new method of quantification of angiogenesis in investigation of the development of blood vessels in the CAM, in particular days of embryonic life considered essential for CAM development. The present research included 75 fertilized eggs of heavy hybrid Ross 308. CAM sampling for stereological analyses was in key phases of embryonic development, namely on the 12th, 15th and 19th day. The results of the present investigation show that the increase in embryonic age results in increase in circulation index, which is also an indicator of angiogenic processes developing in CAM. The lowest value of circulation index (0.1952) was recorded on the first sampling day (E12), while the highest value (0.2666) was recorded on the last sampling day (E19). This method may be applied in researching different factors which affect angiogenesis in CAM.

Does the Primo Vascular System Originate from the Polar Body

Background: Polar bodies degenerate on the first day of embryonic life, and there is no known or specific role for them in the human embryo yet. However, it is not logical to have useless cells. Our previous reports indicated a role for the primo vascular system (PVS) in human beings. There is a possible link between polar bodies and the PVS in embryonic life. Methods: The following databases were searched for peer-reviewed articles in English: Cochrane Collaboration, PubMed, MEDLINE, CINAHL, AMED, Age line, and Social Services s. Our search included the following key words individually or in combination: alternative medicine(s), integrated medicine, holistic care, complementary alternative medicine, embryology, traditional Chinese medicine, traditional medicine, PVS, Bong Han duct in relation to polar bodies. Results: There are no reports relating polar bodies to our keywords. Conclusion: The PVS has been suggested to be responsible for embryonic development. This system contains some pluripotent cells and proteins, some of which are not even present in the blood circulation in chicken embryos. The system becomes only visible after staining with trypan blue. Accordingly, this report tries to present the PVS as the ruler of embryonic cell division and development, which regulates all complicated events during that period of life. To the best of our knowledge, this is the first report suggesting that embryonic development is controlled via the PVS, which originates from polar bodies. i 2014 S. Karger AG, Basel

Mouse thymic mast cells in normal state and after stress-induced atrophy

Background. Studying of thymic mast cells population in normal state and after stressinduced atrophy. Methods. The study was performed on 80 thymus of white outbred mice with using of histochemistry and immunohistochemistry methods. Sections of embryonic thymus were stained with toluidine blue. Adult mice were given a single injection of 2,5 mg of hydrocortisone for induction of thymic accidental transformation; sections were stained with toluidine blue and alcian blue-safranin. Within of immunohistochemical research paraffin sections of adult thymus were stained with polyclonal antibodies to synaptophysin and tyrosine hydroxylase with alcian blue stain. Results. Mast cells (MCs) appeared in thymus on 19th day of embryonic life and demonstrated mainly a medullar location. In adult animals MCs were observed only in the connective tissue of the capsule, interlobular septa, subcortex and perivascular space. Mast cells matured in thymus after accidental transformation. The localization of developing mast cells was changing from medullar and cortical to capsular. A morphological proximity between nerve terminals and mast cells has been observed in normal adult thymus. Some of these nerves were catecholaminergic. Conclusion. Possible important role of thymic mast cells and mast cellsnerves interaction in normal state and after accidental transformation is discussed.

A case of intraocular yolk sac tumor in a child and its pathogenesis

While yolk sac tumor is one of the most common malignant germ cell tumors occurring in young children, it is rarely found in extragonadal sites. We report a case of intraocular yolk sac tumor in a 4-year-old boy. The diagnosis was confirmed by histologic examination and by the rapid normalization of serum alpha-fetoprotein level following enucleation. We propose that yolk sac cells can potentially migrate into the eye at 22 days of embryonic life during neural tube formation, when the head and tail of the neuropore open contemporaneously and communicate with the amniotic cavity.

The in Vitro and in Ovo Effects of Environmental Illumination and Temperature on the Melatonin Secretion from the Embryonic Chicken Pineal Gland

Pineal glands of chicken embryos were placed into a perifusion system for 4 days. The pineal glands were illuminated or exposed to elevated temperature for 8 or 12 h during the in vitro experiment and/or in ovo. Both daily illumination and repeated elevations of environmental temperature transitionally inhibited melatonin release before, and controlled the phase of melatonin rhythm after, the 17th day of embryonic life (E17). In addition, the in ovo rhythmic illumination applied before E13 advances the development of the circadian hormone synthesis.

Embryo‐maternal Communication during the First Days of Embryonic Life

The mechanisms of embryo‐maternal communication during the first days of embryonic life are largely unknown. Using the bovine as a model, the aims of our study were to morphologically characterize the interaction between the pre‐implantation embryo and the epithelium of the maternal ampulla, isthmus and uterotubal junction by light and scanning electron microscopy. For this purpose, oviducts were removed from cows revealing a functional corpus luteum on day 3 after insemination. These were compared to oviducts removed on day 3 (metestrus) of the estrous cycle. Three days after insemination, the majority of the epithelial cells in the ampulla were secretory cells distinctly protruding into the oviductal lumen. Contrary the ampulla of cows on day 3 of the cycle predominantly revealed ciliated cells in the oviductal epithelium. As shown by Periodic Acid Schiff reaction (PAS) with and without amylase digestion, the secretory cells of the ampulla synthesized merely glycoproteins during metestrus, but large amounts of glycogen during pregnancy. In the isthmus no morphological differences were seen between pregnant and cyclic cows. The most conspicuous finding during pregnancy was seen in the uterotubal junction: Vital cumulus cells embedded in between epithelial cells had developed short cytoplasmic processes intensely contacting the epithelial uterine cells. The embryos obtained ex vivo were regularly covered with a thick layer of homogenous extracellular matrix. Contrary embryos produced in vitro– both with and without coculture with oviductal cells –revealed a clearly visible zona pellucida with spongy appearance and numerous pores. Our results imply that already during the first days of life there is intense interaction of the pre‐implantation embryo and the maternal genital tract part of which may be mediated by cumulus cells.

Expression of the rat connexin 39 (rCx39) gene in myoblasts and myotubes in developing and regenerating skeletal muscles: an in situ hybridization study

We report a detailed analysis of the expression pattern of the recently identified rat connexin gene, named rat connexin 39 (rCx39), both during embryonic development and in adult life. Qualitative and quantitative reverse transcription/polymerase chain reaction analysis showed intense expression of rCx39 restricted to differentiating skeletal muscles, with a peak of expression detected at 18 days of embryonic life, followed by a rapid decline to undetectable levels within the first week of postnatal life. A combination of the in situ hybridization technique for the detection of rCx39 mRNA and immunohistochemistry for myogenin, a myoblast-specific marker, allowed us to establish that the mRNA for this connexin was expressed in myogenin-positive myoblasts and early myotubes but disappeared in mature myotubes. Moreover, in adult animals, rCx39 mRNA was expressed in myogenic cells involved in skeletal myofiber regeneration following a crush injury. This is the first case of a connexin being mainly expressed in the myogenic cell lineage. The information presented should pave the way to novel molecular approaches in studies on the role of connexin-based gap-junctional communication in skeletal muscle differentiation and regeneration.

Pregnancy outcome in non-obstructive azoospermia is as effective as in obstructed azoospermia when sperm is retrieved

Evaluation of the outcome of in vitro fertilization rate (FR), number of high quality embryos on the third day of embryonic life (D3) and pregnancy rate (PR) obtained from male partners with obstructive azoospermia (OA) and non-obstructive azoospermia (NOA).

The unique transcriptome through day 3 of human preimplantation development

Successful human development is dependent upon a cascade of events following fertilization. Unfortunately, knowledge of these critical events in humans is remarkably incomplete. Although hundreds of thousands of human embryos are cultured yearly at infertility centers worldwide, the vast majority fail to develop in culture or following transfer to the uterus. In this study, we sought to characterize global patterns of gene expression in individual, normal embryos during the first three days of embryonic life using microarrays; we then compared gene expression between normally growing and growth-arrested embryos using quantitative PCR. Our results documented several novel findings. First, we found that a complex pattern of gene expression exists; most genes that are transcriptionally modulated during the first three days following fertilization are not upregulated, as was previously thought, but are downregulated. Second, we observed that the majority of genes exhibiting differential expression during preimplantation development are of unknown identity and/or function. Third, we show that embryonic transcriptional programs are clearly established by day 3 following fertilization, even in embryos that arrested prematurely with 2-, 3- or 4-cells. This indicates that failure to activate transcription is not associated with the majority of human preimplantation embryo loss. Finally, taken together, these results provide the first global analysis of the human preimplantation embryo transcriptome, and demonstrate that RNA can be amplified from single oocytes and embryos for analysis by cDNA microarray technology, thus lending credence to additional studies of genetic regulation in these cell types, as well as in other small biological samples.

Ontogeny of plasma cells in the early rat yolk sac.

The present study investigated the development of plasma cells in the early rat yolk sac (days 10-16 of gestation) by light microscopy, transmission electron microscopy, immunoelectron microscopy, and indirect immunofluoresce techniques. Cells delineating the morphology of plasma cells in the yolk sac were observed as early as 12 days of embryonic life. As for positive immune staining for the intra-cytoplasmic immunoglobulin (Ig) production (IgA, IgM and IgG), the intensity of the immune staining was very weak on days 10 and 11 of gestation, while it turned very dense on day 12 of gestation. At 14 days of gestation, the number of positive cells was markedly reduced. Immunoelectron microscopy visualized products of the immune reaction in cisterns of the rough endoplasmic reticulum. Conventional electron microscopic examination of 12, 13, and 16-day yolk sacs confirmed the development and differentiation of plasma cells with their well-known ultrastructural features, making this the first study to demonstrate these in the early rat yolk sac. The development of plasma cells in the early yolk sac implies the ability of the yolk sac to effect a humoral immune response at this stage of fetal life. The probable role of plasma cells in the yolk sac is also discussed.

Epidermal growth factor regulates amino acid transport in chick embryo hepatocytes via protein kinase C

System A-mediated amino acid transport, activation of different steps of signal transduction and involvement of different isoforms of protein kinase C (PKC) have been investigated in chick embryo hepatocytes after epidermal growth factor (EGF) stimulation. EGF rapidly (10 min) increased the rate of aminoisobutyric acid (AIB) uptake in chick embryo hepatocytes freshly isolated on the 19th day of embryonic life, while no change was detectable at other embryonal stages. The growth factor stimulation was abolished by PKC and tyrosine kinase inhibitors and was mimicked by 4-phorbol-12-myristate-13-acetate, dimethyl-2 (PMA). EGF treatment did not modify the phosphorylation of the γ isoform of phospholipase C (PLC-γ), and inositol trisphosphate (IP3) and intracellular calcium levels, but it induced an increase in PKC activity. Our data show that EGF regulates amino acid uptake, via PKC and without PLC-γ activation, only in the last period of chick embryo hepatocyte development. The effects of growth factor on PKC activity suggest the involvement of PKC-α and -ε isoforms in EGF modulation of amino acid transport.

Epidermal growth factor regulates amino acid transport in chick embryo hepatocytes via protein kinase C.

System A-mediated amino acid transport, activation of different steps of signal transduction and involvement of different isoforms of protein kinase C (PKC) have been investigated in chick embryo hepatocytes after epidermal growth factor (EGF) stimulation. EGF rapidly (10 min) increased the rate of aminoisobutyric acid (AIB) uptake in chick embryo hepatocytes freshly isolated on the 19th day of embryonic life, while no change was detectable at other embryonal stages. The growth factor stimulation was abolished by PKC and tyrosine kinase inhibitors and was mimicked by 4-phorbol-12-myristate-13-acetate, dimethyl-2 (PMA). EGF treatment did not modify the phosphorylation of the isoform of phospholipase C (PLC-), and inositol trisphosphate (IP3) and intracellular calcium levels, but it induced an increase in PKC activity. Our data show that EGF regulates amino acid uptake, via PKC and without PLC- activation, only in the last period of chick embryo hepatocyte development. The effects of growth factor on PKC activity suggest the involvement of PKC- and - isoforms in EGF modulation of amino acid transport.

Developmental expression and localization of glycogen synthase kinase-3β in rat brain

Glycogen synthase kinase (GSK)-3beta is a protein kinase in the wingless/wnt pathway and as such is involved in the regulation of growth and development of the neural tissue in Drosophila and in vertebrates. This enzyme is also abundantly expressed in the mammal adult brain, where it might play a role in the regulation of several substrates. The expression and the neuroanatomical distribution of GSK-3beta immunoreactivity in the rat brain from embryonic up to adult stages has been studied. GSK-3beta was expressed in the developing brain with the highest expression observed from 18 days of embryonic life up to 10 days of postnatal life. Its expression decreased thereafter and was lowest in the adult. GSK-3beta was strongly expressed in developing neurons but only weakly expressed in layers containing neuroblasts. In the adult and during development, GSK-3beta was detected in the pericarya and proximal part of dendrites. In the embryo, an intense GSK-3beta immunoreactivity was also observed in axonal tracts. This axonal immunoreactivity had markedly decreased by 10 days of postnatal life and was absent at 20 days of postnatal life and in the adult. No GSK-3beta immunoreactivity was detected in astrocytes. The GSK-3beta immunoreactivity was found in most brain regions, although significant local variations of GSK-3beta expression were observed. The developmental evolution of GSK-3beta compartmentalization in neurons parallels that of phosphorylated tau, a protein considered to be a physiological substrate for the kinase.

Apoptosis and Involution of Bursa of Fabricius in Chickens.

Apoptosis was studied in chickens with reference to bursal involution. Morphological and biochemical apoptosis occurred first on day 18 of incubation in bursa of normal embryos. Such phenomena were never detected before 18 th day of embryonic life. Therefore, apoptotic induction was stage specific in chicken bursa. Embryos treated with testosterone propionate (TP) showed DNA fragmentation on day 18, 8 days after the TP injection; these apoptotic responses were negligible on other experimental days. Furthermore, apoptotic cells were occasionally seen among the bursal lymphocytes or epithelial cells in the TPtreated embryos. Embryos injected with half dose of TP showed lower levels of DNA fragmentation than the normals, and the cells undergoing apoptosis were low in number. After hatching, apoptosis was detected morphologically and biochemically in normal bursa, especially after the stage of sexual maturation. These findings demonstrate an apparent correlation between apoptosis and normal involution of bursa after maximal stage of tissue maturation and are suggesting for an apoptosis-independent process of bursal involution introduced by TP treatment.

Involvement of lot cells in guidance of mitral cell axons

The olfactory marker protein, a protein specific to the olfactory sensory neurons, has been studied in mouse during embryogenesis and in the postnatal period up to 30 days, with the unlabeled antibody enzyme method of immunohistochemistry. Olfactory neurons, which are morphologically detectable in 10-day-old embryos, do not contain olfactory marker protein. The protein is present in the olfactory neuroepithelium at embryonic day 14 and its appearance coincides with the establishment of sensory synapses in the olfactory bulb. Neurons containing the protein increase in number up to 30 days after birth. At 15 days of embryonic life, immunostaining was observed in sensory axons at the rostral tip of the olfactory bulb, and by embryonic day 17 a plexus of stained fibres has covered the bulbar surface. Between embryonic day 15 and postnatal day 1, olfactory axons have been observed to reach the mitral cell layer. In the vomeronasal system the olfactory marker protein is present at later stages and both the receptors' perikarya and their axons and axon terminals in the accessory olfactory bulb show a lower level of staining than the olfactory system proper.This study of the olfactory marker protein has allowed us to correlate its appearance with significant developmental phenomena.

Microglia development in the quail cerebellum

We used the QH1 antibody to study changes in the morphological features and distribution of microglial cells throughout development in the quail cerebellum. Few microglial precursors were present in the cerebellar anlage before the ninth incubation day (E9), whereas many precursors apparently entered the cerebellum from the meninges in the basal region of the cerebellar peduncles between E9 and E16. From this point of entry into the nervous parenchyma, they spread through the cerebellar white matter, forming a 'stream' of labeled cells that could be seen until hatching (E16). The number of microglial cells in the cerebellar cortex increased during the last days of embryonic life and first posthatching week, whereas microglial density within the white matter decreased after hatching. As a consequence, the differences in microglial cell density observed in the cerebellar cortex and the white matter during embryonic life diminished after hatching, and microglia showed a nearly homogeneous pattern of distribution in adult cerebella. Ameboid and poorly ramified microglial cells were found in developing stages, whereas only mature microglia appeared in adult cerebella. Our observations suggest that microglial precursors enter the cerebellar anlage mainly by traversing the pial surface at the basal region of the peduncles, then migrate along the white matter, and finally move radially to the different cortical layers. Differentiation occurs after the microglial cells have reached their final position. In other brain regions the development of microglia follows similar stages, suggesting that these steps are general rules of microglial development in the central nervous system.