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PubMed1990

On the possible effects of homeostatic shifts in human embryonic development.

Possible etiological factors of congenital malformations as well as of human trisomies are considered in the framework of the repressor hypothesis. In this approach gene expression is envisaged from the point of view of the functional variations of the total activation energy for normal gene expression in homeostatic equilibrium. We restrict our attention to variations of the total activation energy under the effect of temperature gradients. We discuss the evidence that hyperthermia may be an etiological factor for trisomies in humans.

Animals↗

PubMed1988

[Comparative expression of 2 intermediate filament proteins, peripherin and the 68 kDa neurofilament protein, during embryonal development of the rat].

Peripherin, an intermediate filament protein, was originally detected by biochemical methods in the neurons of the peripheral nervous system. We now studied its expression and cellular localization by immunocytochemical methods in the developing rat embryo, and compared them with the expression and localization of the 68 kDa neurofilament protein. It appears that peripherin is expressed not only in the neurons of the peripheral nervous system, but also in some well defined neuronal populations of the central nervous system. These results focus on the questions of the phylogenetic origin and of the function of peripherin.

Animals↗

PubMed1988

Evaluation of human serum as a culture medium for studies of rat embryonic development in vitro.

The normal growth of rat embryos cultured in human serum supplemented with glucose and 10% rat serum is of potential use in the assessment of teratogenic risk in man. Before this technique can be widely applied, it is necessary to know whether the stages of the menstrual cycle at which a serum sample is tested materially affect the results obtained. Moreover, in order to achieve reproducible conditions we have found it necessary to add a minimal amount of rat serum to the human serum used for culture, but unnecessarily high levels of rat serum supplementation could over-compensate for any growth factor deficiency in human serum. Here we report that culture of rat conceptuses gives similar results irrespective of whether human first, second, or third trimester pregnancy serum, postnatal serum, or serum at various stages of the menstrual cycle is used. We also report that addition of 2% rat serum supplement is sufficient to achieve reproducible rat embryonic growth and differentiation in human serum.

Animals↗

PubMed2000

Regional and cellular specificity of the expression of TPRD, the tetratricopeptide Down syndrome gene, during human embryonic development.

The TPRD gene (tetratricopeptide (TPR) containing Down syndrome gene) is one of the candidate genes in the Down syndrome chromosomal region-1. Duplication of this gene may be the cause of major phenotypic features of Down syndrome. Here we show that the TPRD expression is developmentally regulated during human embryogenesis. At the earliest stages of development (Carnegie 8-12) TPRD expression is ubiquitous. At later developmental stages (Carnegie stages 14, 16 and 18), it becomes restricted to the nervous system, as is the case for the mtprd gene during mouse development. We extended our analysis of TPRD expression during fetal development of the human nervous system (13, 22 and 24 weeks). A new oblique illumination technique was used to compare signal intensity and cell density. Some regions of the nervous system such as the external cortical layers of the brain, and the inner neuroblastic layer of the eye, strongly express the TPRD gene.

Cerebral Cortex↗

PubMed1998

Glycocoding as an information management system in embryonic development.

The past 5 years have seen a burgeoning in the amount of data emerging from laboratories studying the early stages of embryogenesis. Much of this data implicates various aspects of glycobiology in the initiation and regulation of these processes. The level of analytical detail coming from these investigations has surpassed our ability to fully understand its overall significance within the context of the interactive/dynamic developmental process. This review proposes a pause in this seemingly endless quest for more detail so that we may take stock of our goals and objectives. The proposed goals include a mechanistic understanding of the process(es) involved in information managed at various 'transition points' during early embryogenesis, and an understanding of the mechanism(s) by which the spatial/temporal regulation of early development are managed. The 'transitions' which occur during cell to cell cluster, cell cluster to early organ architecture, and early organ architecture to functional organ development are fundamental and mirror the evolutionary process of biological information management. All of these 'transitions' involve increasing complexity, the development of hierarchies of information management systems (integrated bidirectionally), and spatial/temporal regulation which relies on historical events to map future structure and function. This review focuses on a relatively small number of studies which highlight these aspects of early development. A mechanism which involves glycocoding, an extension of the Roseman Hypothesis, and its direct use as an information management system is proposed and some supporting experimental evidence is presented. This extension of an existing hypothesis is related to several recent investigations, and is designed to broaden the experimental design of future studies so that these important process issues can be addressed.

Animals↗

PubMedDate not supplied

The leukemia-associated gene Mllt1/ENL: characterization of a murine homolog and demonstration of an essential role in embryonic development.

MLLT1 (ENL/LTG19) is one of a number of fusion gene partners with the MLL oncogene involved in 11q23 translocations in human leukemia and encodes a transcriptional regulator of unknown function. Leukemias bearing MLL translocations may be myeloid or lymphoid or bear mixed lineage properties; however, those bearing MLL/MLLT1 translocations are predominantly lymphoid, suggesting that MLLT1 may influence the leukemic phenotype. The murine homolog Mllt1 exhibits 86% amino acid sequence identity with the human gene and is broadly expressed in murine tissues and cell lines, with the exception of liver and myeloid cell lines. We have mapped Mllt1 to mouse chromosome 17 band E2 using FISH analysis. The genomic structure and 5' regulatory sequence of Mllt1 are highly conserved between mouse and human. There is also conservation of the genomic structure, but not the promoter, between MLLT1 and MLLT3/AF9, a homologous gene that is also an MLL translocation partner in human leukemias with a predominant myeloid phenotype. Targeted disruption of Mllt1 in mice leads to embryonic lethality prior to 8.5 dpc. These studies indicate that MLLT1 is involved in essential developmental processes and suggest that expression patterns of MLL fusion partners may influence the lineage of MLL-associated leukemias.

Amino Acid Sequence↗

PubMed2003

Prenatal exposure to corticosterone impairs embryonic development and increases fluctuating asymmetry in chickens (Gallus gallus domesticus).

1. The level of corticosterone in fertilised eggs from hens (Gallus gallus domesticus) was manipulated experimentally to elucidate whether stress in laying hens is harmful to the chicks, as manifested by impaired survival and reduced growth, and whether bilateral asymmetry may represent an indicator of environmental stress in poultry. 2. Three hundred and fifty eggs were randomly divided into 4 groups; 1. untreated, 2. control, 3. 10 ng corticosterone/ml and 4. 20 ng corticosterone/ml. Each of the eggs in groups 2, 3 and 4 were injected with 100 microl ethanol-saline solution (25% ethanol in saline) containing 0, 0.6 and 1.2 microg corticosterone, respectively. After the injections, the final concentration of ethanol in the egg (albumen and yolk) was 0.03%, and the concentration of added corticosterone was 0, 10 and 20 ng/ml, respectively, in groups 2, 3 and 4. All the eggs were treated on developmental d 1. 3. Corticosterone injections resulted in greater embryonic mortality, earlier termination of foetal development and reduced growth. Moreover, chicks developing in eggs with an elevated concentration of corticosterone displayed reduced developmental stability as evidenced by increased fluctuating asymmetry (FA) in tarsus length. 4. In conclusion, an increased concentration of corticosterone in the egg was detrimental to survival and growth of the chicks. Prenatal stress also generated bilateral asymmetry, and illustrates the potential application of FA as an indicator of environmental stress in poultry.

Agriculture↗

PubMed1994

Expression and role of c-myc protooncogene in murine preimplantation embryonic development.

PURPOSE: The present study was conducted to investigate the expression and possible role of the c-myc protooncogene in preimplantation embryos by using reverse-transcriptase/polymerase chain reaction (RT-PCR) technique and microinjection of synthetic antisense c-myc oligonucleotide probe, respectively. Total RNA was extracted from oocytes and two cell-, four cell-, early morula-, late morula-, early blastocyst-, and late blastocyst-stage embryos, and cDNA was constructed using MMLV reverse transcriptase. Sense primer (P1) and antisense primer (P2) used were based on the c-myc gene sequence bp 1609-1629 and bp 3279-3299, respectively, that span a 1.37-kb intron. PCR-amplified products of cDNA from oocyte-, two cell-, four cell-, early and late morula-, and blastocyst-stage embryos demonstrated the expected 313-bp product in Southern blot hybridization using a c-myc specific DNA probe, with an indication of lower levels in oocytes and early morulae. RESULTS: Cytoplasmic injection of the antisense c-myc oligonucleotide probe (P2) and not the sense probe (P1) into pronuclear-stage zygotes caused a significant (P = 0.02 to 0.0001) inhibition of development to blastocysts in a concentration-dependent manner, with a maximal inhibition at the first cleavage of zygotes to two cell-stage embryos. There was no effect on the P2 antisense injection on pronucleus formation. CONCLUSION: These results indicate that the c-myc protooncogene is expressed in preimplantation embryos and may have an essential role in normal embryogenesis in mice.

Animals↗

PubMed2000

Enzyme activities involved in lipid metabolism during embryonic development of Macrobrachium borellii.

The activities of the enzymatic systems involved in the activation and degradation of fatty acids, and in the synthesis of triacylglycerols and phospholipids were studied in vitro using total cellular homogenate and subcellular fractions of eggs of the shrimp Macrobrachium borellii at different developing stages. Egg development was divided into seven stages based on morphological features of the embryo. Palmitoyl-CoA ligase activity increased as the embryo developed and showed its maximum at stage V. An increase in the synthesis of triacylglycerols and diacylglycerols was also observed at this stage. Diacylglycerylethers were synthesized more actively during the first stages of development. The higher specific activity observed in total homogenate than in microsomal fraction suggested that their synthesis was not exclusively microsomal. Phospholipid synthesis was very active all along development, reflecting active membrane biosynthesis. The highest activity of the cytosolic triacylglycerol lipase was observed at stage V. Fatty acid degradation, measured as mitochondrial beta-oxidation activity, did not vary significantly during development. We conclude that both the anabolic and catabolic processes concerning lipid metabolism are very active, with values similar to those described for adult hepatopancreas, revealing the major role of lipids during shrimp embryogenesis energetics, and that the highest activities of lipid synthesis-hydrolysis take place at stage V when embryos are under active organogenesis. J. Exp. Zool. 286:231-237, 2000.

Animals↗

PubMed1997

Two populations of luteinizing hormone-releasing hormone neurons in the forebrain of the rhesus macaque during embryonic development.

To investigate the possibility that a second luteinizing hormone-releasing hormone (LHRH) population appears during development in primates, embryos and fetal brains of rhesus monkeys were immunostained with antisera specific to different LHRH forms. Two LHRH cell populations were discernible by immunoreactivity to antisera LR-1 and GF-6. Because one LHRH cell type migrated out from the olfactory placode several days earlier than the other, they were referred to as "early" and "late" LHRH cells, respectively. Although late LHRH neurons were immunoreactive to all anti-mammalian LHRH antisera tested, early LHRH neurons were only detected by antiserum GF-6. Early LHRH neurons (approximately 10 x 7 microm) were smaller than late LHRH neurons (approximately 18 x 7 microm). Early LHRH neurons were first found around the olfactory placode, in the nasal mesenchyme, and in the rostroventral forebrain on embryonic day 30 (E30), whereas late LHRH neurons were first seen in the olfactory pit on E32. Early LHRH cells were located throughout the basal forebrain on E32-E42, whereas late LHRH cells were found in the olfactory pit and along the terminal nerve on E34-E36 and were not seen in the forebrain until E38. By E51-E62, late LHRH neurons reached into the basal hypothalamus in a distribution resembling that in the older brain, while early LHRH neurons were found in the septum, preoptic region, stria terminalis, medial amygdala, claustrum, internal capsule, and globus pallidus. Based on the distribution pattern of immunopositive cells with antiserum LR-1, late LHRH cells are bona fide LHRH neurons that regulate the pituitary-gonadal axis. In contrast, the molecular form of early LHRH cells is unclear, although it is plausible that early LHRH cells may contain the molecule in which the C-terminal epitope of LHRH is modified or absent. It is concluded that in primates there is a second population of LHRH neurons that originates from the embryonic olfactory placode before the origin of mammalian LHRH-like neurons, and that these two populations of LHRH-immunopositive neurons have different morphologic features and different final distributions in the brain.

Animals↗

PubMed1986

Keratan sulfate proteoglycan during embryonic development of the chicken cornea.

Antibodies to corneal keratan sulfate proteoglycan (KSPG) were used to characterize the pattern of KSPG accumulation during differentiation of neural crest cells in the stroma of embryonic chick cornea. Immunohistochemistry with monoclonal antibody I22 to keratan sulfate found this KSPG antigen localized inside stromal cells at stage 29 (Day 6), ca. 12 hr after migration into the primary stroma. A 2- to 3-day lag then occurred before appearance of extracellular keratan sulfate, first seen on Day 9 (Stage 35) in the posterior stroma. Keratan sulfate antigen accumulated in a posterior to anterior direction during subsequent development. Uniform staining of the stroma for keratan sulfate did not occur until after Day 16. Among several tissues, only corneal stroma contained an extracellular matrix which stained for keratan sulfate, though intracellular staining of some cartilage cells was observed. Accumulation of KSPG antigens in developing cornea was measured in unfractionated guanidine extracts with a quantitative ELISA using three different antibodies against KSPG. Increases were first detected after Day 9 using monoclonal I22, and somewhat later with the other two antibodies. Assays with all three antibodies detected a sustained, exponential increase of KSPG throughout the 5 days prior to hatching. Keratan sulfate continued to accumulate after hatching, but an antibody with specificity to KSPG core protein, detected no relative increase in antigen after hatching. This suggests a modulation of KSPG primary structure late in development and after hatching. Overt differentiation of individual neural crest cells thus appears to begin ca. 12 hr after their arrival in the primary stroma; a lag of 2-3 days precedes active secretion of KSPG.

Age Factors↗

PubMed2003

Effect of supplementation of leukemia inhibitory factor and epidermal growth factor on murine embryonic development in vitro, implantation, and outcome of offspring.

OBJECTIVES: To evaluate the supplementation of leukemia inhibitory factor (LIF) or epidermal growth factor (EGF) in a single culture system (IVF50) and a sequential culture system (IVF50-S2) to determine whether they had any additional benefit on embryo development and implantation. DESIGN: Prospective controlled animal study. SETTING: University research laboratory. SUBJECT(S): Two-cell embryos from F1(CBA x C57BL) mice. INTERVENTION(S): Embryos were randomly cultured in different growth factor-treated or untreated in vitro systems. MAIN OUTCOME MEASURE(S): Blastocyst formation and morphology, total cell numbers of day 5 blastocysts, birth rates, and characteristics of the offspring. RESULT(S): The beneficial effects of LIF or EGF on blastocyst development and morphology were observed only in IVF50 medium but not in sequential IVF50-S2 media. In addition, blastocysts generated from the LIF-supplemented cultures had higher total cell numbers and higher total birth rates after transfer compared to those from IVF50 medium alone. No significant effect on fetal development was observed but pups born after treatment in LIF-supplemented sequential cultures had prolonged gestation and increased birth weights. CONCLUSION(S): The beneficial effects of LIF and EGF on in vitro blastocyst development are mainly seen under suboptimal culture conditions (simple medium only) and their effects are masked in an improved in vitro system (sequential culture media). Supplementation of culture media with such factors, however, is still an attractive approach in enhancing embryo viability through increased total cell numbers. The beneficial effect of LIF in improving implantation and subsequent increased birth rates should be further explored.

Animals↗

PubMed1991

In vitro maturation of ovarian oocytes from unstimulated rhesus monkeys: assessment of cytoplasmic maturity by embryonic development after in vitro fertilization.

One-hundred and sixty-six cumulus-enclosed oocytes, obtained from ovaries of unstimulated rhesus monkeys, were subjected to six different treatments in vitro--two types of media (simple = TALP; complex = CMRL) x three levels of gonadotropins (none, FSH, FSH + hCG)--to assess their ability to undergo maturation, fertilization, and embryo development. A summary of development in culture for all experimental treatments is as follows: 58% of oocytes underwent germinal vesicle breakdown; 37% extruded a first polar body; 17% had more than one pronucleus and/or two polar bodies after insemination (i.e., were activated/fertilized); and 12% cleaved (i.e., developed) to at least the 2-cell stage in vitro. Of 45 oocytes incubated only in medium (either simple or complex) without gonadotropins, only 3 were activated/fertilized (6.7%), and only one embryo developed to at least the 2-4-cell stage (2.2%). There were no differences between oocytes incubated with only FSH and oocytes incubated with FSH + hCG. Activation/fertilization (20.7% vs. 6.7%) and embryo development (greater than or equal to 2 cells; 15.7% vs. 2.2%) were significantly higher in treatments with than without gonadotropin supplementation. There were no statistically significant differences attributable to incubation in different media during oocyte maturation. Cumulus-enclosed oocytes recovered from unstimulated ovaries of rhesus monkeys can resume maturation during culture in vitro, as shown by their ability to be fertilized and by the cleavage in vitro of the resultant zygotes.

Animals↗

PubMed1993

Embryonic development of rat sympathetic preganglionic neurons: possible migratory substrates.

Spinal somatic and autonomic (sympathetic preganglionic) motor neurons are generated synchronously and, subsequently, migrate from the ventricular zone together to form a common primitive motor column. However, these two subsets of motor neurons ultimately express several phenotypic differences, including somal size, peripheral targets, and spinal cord locations. While somatic motor neurons remain ventrally, autonomic motor neurons (AMNs) move both dorsally and medially between embryonic days 14 and 18, when they approximate their final locations in spinal cord. The goal of the present investigation was to determine the potential guidance substrates available to AMNs during these movements. The dorsal translocation was studied in developing upper thoracic spinal cord, because, at this level, the majority of AMNs are located dorsolaterally. Sections were double-labeled by ChAT (choline acetyltransferase) and SNAP/TAG-1 (stage-specific neurite associated protein/transiently expressed axonal surface glycoprotein) immunocytochemistry to visualize motor neurons and the axons of early forming circumferential interneurons, respectively. Results showed that during the developmental stage when AMNs translocated dorsally, SNAP/TAG-1 immunoreactive lateral circumferential axons were physically located along the borders of the AMN region, as well as among its constituent cells. These findings indicate that lateral circumferential axons, as well as the SNAP/TAG-1 molecules contained upon their surfaces, are in the correct spatial and temporal position to serve as guidance substrates for AMNs. The medial translocation was studied in developing lower thoracic-upper lumbar spinal cord, because, at this level, more than half of the AMNs are medially located. Sections were double-labeled by ChAT and vimentin immunocytochemistry to visualize motor neurons and radial glial fibers, respectively. Observations on consecutive developmental days of the medial translocation revealed that AMNs were aligned with parallel arrays of radial glial fibers. Thus, the glial processes could serve as guides for the AMN medial movement. Future experimental analyses will examine whether circumferential axons and radial glial fibers are in fact functioning as migratory guides during AMN development, and, if so, whether specific surface molecules on these guides trigger the subsequent differentiation of AMNs.

Animals↗

PubMedDate not supplied

[Embryonic development of Taenia pisiformis and T. hydatigena (Cestoidea, Cyclophyllidea)].

A description of the consecutive stages of ripening of ovary cells of early fission and gastrulation of T. pisiformis and T. hydatigena, peculiar to the common type of fission of Cyclophyllidea is given. The fission has signs of double spiral fission which takes place in primitive Turbellaria. The primitiveness of early embryogenesis is reflected in early determination of macromeres, in the absence of strict regulation in the location of blastomeres, in the formation of blastula and unipolar gastrulation with the absence of morphological differentiation of ectoderm and mesenchyme.

Animals↗

PubMed1996

Early embryonic development and evaluation of equine embryo viability.

Tremendous progress has been made in the development of assisted reproductive techniques that may enhance the reproductive efficiency of the horse. However, techniques that involve the manipulation of oocytes and/or embryos may themselves be detrimental to embryo viability and subsequent development. Therefore, an objective method of assessing viability of embryos before and/or after oocyte/embryo manipulation is desirable. At this time, morphologic evaluation is the most widely used method of determining the viability of equine embryos. Although morphologic assessment of embryo quality will not always accurately predict the survival of individual embryos, it is very useful for predicting the survival of groups of embryos. Other tests that have been used to evaluate equine embryo viability include (1) development during culture in vitro; (2) quantitating metabolism of the fluorescent substrate fluorescein diacetate; (3) quantitating uptake of the fluorescent stain DAPI; and (4) quantitating embryo metabolism. Although these tests offer potential advantages over morphologic assessment alone, their current limitations have prevented their routine use for embryo evaluation. It is likely that as improvements are made in these evaluation methods, they may offer advantages for use alone or in combination to more accurately assess the viability of equine embryos.

Animals↗

PubMed1985

Comparative effects of cathepsin inhibitors on rat embryonic development in vitro. Evidence that cathepsin D is unimportant in the proteolytic function of yolk sac.

The effects of two proteinase inhibitors, leupeptin and pepstatin on the development of 9.5-day rat conceptuses in vitro has been studied. All cultures were of 48 h duration and the inhibitors were present throughout the entire period. When pepstatin was added to the culture medium (5-25 micrograms/ml) conceptuses developed and grew to an extent that did not differ from untreated controls. However, leupeptin (1-4 micrograms/ml) caused severe growth retardation and abnormal development of conceptuses. The effects of the two inhibitors on the hydrolysis of 125I-labelled BSA and haemoglobin by homogenates of 10.5-day yolk sac indicated the biochemical basis for the differential toxic effects of the two inhibitors on development. Leupeptin was highly inhibitory of the degration of both substrates whereas pepstatin caused no inhibition of 125I-labelled BSA hydrolysis, and only a slight inhibition of haemoglobin hydrolysis. These observations demonstrate that cathepsin D, a lysosomal aspartic proteinase that is specifically inhibited by pepstatin is not involved in yolk-sac-mediated protein utilization by early organogenesis-phase conceptuses and that lysosomal cysteine proteinases, specifically inhibited by leupeptin, are of paramount importance in this yolk sac function.

Animals↗

PubMed1996

Effects of follicular fluid during in vitro maturation of bovine oocytes on in vitro fertilization and early embryonic development.

In vivo, oocytes mature while bathed in preovulatory follicular fluid, so we studied maturation of oocytes in vitro in media supplemented with such fluid. Bovine oocytes were obtained from 2-to 6-mm follicles from slaughterhouse ovaries and were matured for 24 h in tissue culture medium 199 + 10% estrous cow serum supplemented with bovine follicular fluid (BFF) obtained from a preovulatory follicle either before (0 h) or 8 or 20 h after a GnRH-induced LH surge. After in vitro fertilization, oocytes matured in 10% or 40% 20-h BFF had higher cleavage rates (p < 0.05) than those matured in 10% or 40% 0-h BFF (80% and 86% vs. 46% and 51%, respectively); oocytes matured in 8-h BFF were intermediate. A greater percentage of oocytes showed cumulus expansion with 20-h BFF than with 0-h BFF (p < 0.05). In experiment 2, more (p < 0.05) embryos from oocytes matured in 20% or 40% 20-h BFF developed to blastocysts after culture of embryos compared to those matured in 20% or 40% 0-h BFF (29% and 29% vs. 7% and 14%, respectively) or those matured in no BFF (12%). Embryos from oocytes matured in a mixture of 0-h and 20-h BFF developed to the blastocyst stage similarly (27%) to those matured in 20-h BFF, suggesting that positive factors in 20-h BFF overcame any inhibitory effect of 0-h BFF. To summarize, bovine oocytes matured with BFF collected 20 h after LH surge result in higher cleavage rates after in vitro fertilization and more blastocysts than those matured in 0-h BFF. When the factors responsible are identified, they might be used to enhance maturation of oocytes in vitro.

Animals↗