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PubMed1999

New reproductive technologies.

The ethical calculus is described. There are four entities that have a stake in all ethical decisions. These are the individual, family, community and society at large. Values assigned to these entities can be modified by experience, cultural background, religious authority and the law. In the simple case of in vitro fertilization (IVF), there seems to be general international acceptance, except for the official position of the Roman Catholic Church, the origin of which dissent is extensively discussed. There is concern about the number of pre-zygotes/pre-embryos to be transferred, the disposition of any unfertilized egg and the reporting of results. Clear ethical considerations arise as IVF is applied in more complex situations, such as unmarried couples, the use of donor gametes and donor pre-embryos, the use of cryopreservation, intracytoplasmic sperm injection (ICSI), prenatal genetic diagnosis, surrogate gestational motherhood and research on the pre-embryo. The ethical, religious and legal acceptance of these depends on the weight given to elements of the ethical calculus as described in the chapter.

Ethics, Medical↗
PubMed1995

Physiologically based pharmacokinetic models applicable to organogenesis: extrapolation between species and potential use in prenatal toxicity risk assessments.

A physiologically based pharmacokinetic (PBPK) model describing the disposition of 2-methoxyacetic acid (2-MAA; the proximate toxicant derived from oxidation of the ethylene glycol ether, 2-methoxyethanol) was developed in pregnant rodents. This model was validated with pharmacokinetic (PK) data from dams and embryos during major organogenesis. A physiological model of human pregnancy was then combined with the PBPK model and linked to an empirical 2-MAA PK model with 2 maternal compartments and a single or multiple conceptus compartment, depending on the developmental stage. This approach is intended to allow more realistic human pregnancy risk assessments by refining the reference dose calculations via uncertainty factors. It will be possible to eliminate an uncertainty factor of 10 for interspecies extrapolations in the 2-methoxyethanol risk assessment if the PBPK model described here is used.

Acetates↗
PubMed1991

Segmental origin and migration of neural crest cells in the hindbrain region of the chick embryo.

A vital dye analysis of cranial neural crest migration in the chick embryo has provided a positional fate map of greater resolution than has been possible using labelled graft techniques. Focal injections of the fluorescent membrane probe DiI were made into the cranial neural folds at stages between 3 and 16 somites. Groups of neuroepithelial cells, including the premigratory neural crest, were labelled by the vital dye. Analysis of whole-mount embryos after 1-2 days further development, using conventional and intensified video fluorescence microscopy, revealed the pathways of crest cells migrating from mesencephalic and rhombencephalic levels of the neuraxis into the subjacent branchial region. The patterns of crest emergence and emigration correlate with the segmented disposition of the rhombencephalon. Branchial arches 1, 2 and 3 are filled by crest cells migrating from rhombomeres 2, 4 and 6 respectively, in register with the cranial nerve entry/exit points in these segments. The three streams of ventrally migrating cells are separated by alternating regions, rhombomeres 3 and 5, which release no crest cells. Rostrally, rhombomere 1 and the caudal mesencephalon also contribute crest to the first arch, primarily to its upper (maxillary) component. Both r3 and r5 are associated with enhanced levels of cell death amongst cells of the dorsal midline, suggesting that crest may form at these levels but is then eliminated. Organisation of the branchial region is thus related by the dynamic process of neural crest immigration to the intrinsic mechanisms that segment the neuraxis.

Animals↗
PubMed1985

Distributions of vimentin and desmin in developing chick myotubes in vivo. II. Immunoelectron microscopic study.

The distribution of the intermediate filament proteins vimentin and desmin in developing and mature myotubes in vivo was studied by single and double immunoelectron microscopic labeling of ultrathin frozen sections of iliotibialis muscle in 7-21-d-old chick embryos, and neonatal and 1-d-old postnatal chicks. This work is an extension of our previous immunofluorescence studies of the same system (Tokuyasu, K. T., P. A. Maher and S. J. Singer, 1984, J. Cell Biol., 98:1961-1972). In immature myotubes of 7-11-d embryos, significant labeling for desmin and vimentin was found only in intermediate filaments, and these proteins coexisted in the same individual filaments. Each of the two proteins was present in irregular clusters along the entire length of a filament. No exclusively vimentin- or desmin-containing filaments were observed at this stage. In the early myotubes, the intermediate filaments were essentially all longitudinally oriented, even when they contained three times as much desmin as vimentin. No special relationship was recognized between the dispositions of the filaments and the organization of the myofibrils. Occasionally, several myofibrils were already aligned in lateral registry at this early stage, but labeling for desmin and vimentin was largely absent at the level of the Z bands. Instead, the Z bands appeared to be covered by elements of the sarcoplasmic reticulum. The confinement of intermediate filaments to the level of the Z bands occurred in the myotubes of later embryos after the extensive lateral registry of the Z bands. Thus, intermediate filaments are unlikely to play a primary role in producing the lateral registration of myofibrils during myogenesis, but may be important in determining the polarization of the early myotube and the alignment of its organelles. Throughout the development of myotubes, desmin and vimentin remained in the form of intermediate filaments, although the number of filaments per unit volume of myotube appeared to be reduced as myofibrils increased in number in maturing myotubes. This observation indicated that the transverse orientation of intermediate filaments in mature myotubes does not result from the de novo polymerization of subunits from Z band to Z band, but a continuous shifting of the positions and directions of intact filaments.

Animals↗
PubMed1992

Fate mapping the neural plate and the intraembryonic mesoblast in the upper layer of the chicken blastoderm with xenografting and time-lapse videography.

The disposition of the Anlage fields of the neural plate and the intraembryonic mesoblast in the upper layer of the chicken blastoderm was studied at the primitive streak stage prior to the regression of Hensen's node (stages 5V to 6V, L. Vakaet (1970) Arch. Biol. 81, 387-426). Chicken blastoderms were cultured by New's technique on a mixture of thin egg white and agar. The anterior half of the deep layer was reflected with a tungsten needle. A circular fragment of the upper layer was punched out with a pulled out Pasteur pipette and discarded. It was replaced with an isotopic and isopolar piece of quail upper layer that was punched out with the same pipette. The deep layer was replaced and the chimeras were reincubated for 24 hours. The xenografts were followed with time-lapse videography. After fixation, the quail cells were located using Le Douarin's quail nucleolar marker technique. Integrating the observations with time-lapse videography and the results of Feulgen stained sections, we have drawn a new fate map of the disposition of the Anlage fields in the upper layer of the chicken blastoderm at stages prior to the regression of Hensen's node (stages 5V to 6V). The disposition of the neural plate and of the notochord, somites, nephrotome and lateral plates was therefore determined before the Anlage fields are morphologically discernible. The pathway of the fields in the upper layer towards their disposition was documented with time-lapse videography in chimeric chicken blastoderms that developed normally.

Animals↗
PubMed1983

Changes in nuclear ribonucleoprotein constituents and chromatin disposition during neuronal differentiation and maturation.

Morphological and quantitative changes of ribonucleoproteic (RNP) structures and chromatin are studied in the nuclei of undifferentiated cells, neuroblasts and neurons in several degrees of maturation, in order to relate them to the drastic modifications in transcription and/or RNA processing taking place during cell differentiation. Undifferentiated (matrix) cells of 2-day embryos differ from that of 4-day embryos in their nucleolar volume and in the amount of compact chromatin. These differences are interpreted as the earliest signs of neuroblast differentiation. All over the process of matrix cell-bipolar-multipolar neuroblast differentiation there is a large spreading of compact chromatin well before any important change in RNP structures or in nuclear volume. The most remarkable increase in nuclear volume and in the amount of RNP particles occurs during the differentiation of multipolar neuroblasts to immature neurons, which is characterized by large synaptogenic activity. The interpretation of these changes is discussed in connection, on one hand with the metabolic effects of synapses, and on the other hand with the variations of gene expression taking place in cell differentiation and under other natural and experimental situations.

Animals↗
PubMed1980

Intracellular filament bundles in whole mounts of chick and human myoblasts extracted with triton X-100.

The method of Triton X-100 extraction and critical point drying of whole mounts of cultured chick and human myoblasts was used to study the presence of intracellular bundles of filaments within these cells. Observation by means of transmission and scanning electron microscopy demonstrated a complex system of filament bundles which appeared morphologically and spatially heterogeneous. Most obvious were long dense bundles of cables traversing along the ventral surface of developing myoblasts, presumably the 'stress fibers' seen in light microscopy. Other bundle types occurred which were composed of loose aggregates of filaments coursing through the remnant cell body. A prominent accumulation of filaments was also seen at the lateral edges of these myoblasts. These lateral edge cables were thicker and denser than any other type of filament bundle observed in the myoblasts. Reaction of unextracted myoblasts directly to human antiplatelet myosin conjugated to rhodamine demonstrated that the most intense reaction also occurred along the lateral edges of both human and chick myoblasts. During development of chick myoblasts the filament bundles became oriented parallel to the cell axis giving the cell a fusiform morphology. It is possible that the various filament bundle structures and their differing structural and spatial dispositions could be related to functional differences among the diverse population of intracellular bundles of filaments.

Animals↗
PubMed2002

[Reproductive development of initial sporophytes in Pinus sylvestris L].

The reproductive development of Pinus sylvestris from generative primordia to sporophyte initial (embryo tier of 16-celled proembryo) was studied. In the reproductive cycle, three types of stages are distinguished and described in association with: 1) hereditary program formation; 2) a new reproductive initial origin, and 3) increase in propogative energy. The layout of disposition of these stages in the reproductive cycle is given.

Pinus↗
PubMed1990

Prenatal diagnosis of sex in bovine fetuses by amniocentesis.

The sex of 1, 056 bovine fetuses was diagnosed by cytogenetic analysis of amniotic cells collected surgically from cows made pregnant as a result of embryo transfer. The fetuses ranged in gestational age from 7 to 22 wk at the time of amniocentesis. Amniotic cells were cultured for 5 to 30 d to obtain a sufficient number of cells for cytogenetic analysis. The growth rate of 819 samples was examined in detail. On average, amniotic cells from pregnancies that were 7 or 8 wks old required about 13 d to reach a cell concentration sufficient for analysis, whereas those from pregnancies that were 10 wk old or more required only 10 d or less to reach that concentration. Final disposition of 325 pregnancies subjected to amniocentesis was unequivocally confirmed. Of these, the sex of 302 fetuses (93%) was determined with a male: female ratio of 51:49. The gender of 213 of 220 fetuses (96.8%) was correctly dignosed, as verified by examination of either 33 intentionally induced abortuses or of 187 live calves. Thirteen percent (29) of 216 pregnancies that were allowed to proceed to term ended in spontaneous abortion, a rate only about 5% higher than that reported to occur normally in embryo transfer pregnancies. The remaining 53 pregnancies were induced to abort, but it was not possible to recover and verify the sex of those fetuses. The capability to diagnose fetal sex in utero yielded other useful information as well. For example, the sex ratio of 1,056 fetuses during development from 7 to 15+ wk of gestation was found to be the same (51:49) as it was at calving. Finally, amniocentesis to determine prenatal sex provides time to alter the sex ratio of a calf crop. It may ultimately prove valuable as an adjunct to genetic engineering of cattle to identify presumptive transgenic calves in utero.

Journal Article↗
PubMedDate not supplied

[Teratogenic and metamorphosis inhibiting activity of retinoic acid in Ciona intestinalis].

Exposure of embryos of the sea squirt, Ciona intestinalis, to all-trans-retinoic acid (between 10(-5) and 10(-7) M) causes specific malformations of the larvae and suppression of settlement and metamorphosis. Whether the vitamin A derivative was administered at the 2-cell stage, or at the early gastrula stage did not affect the nature of the extent of the ensuing anomalies. Malformations include a dorsal bulge of the body, an irregular disposition of ocellar pigment, and a twisted tail. Treated larvae have no statocyst pigment. There is also a reduction in size of the body, compared to control larvae. Retinoic acid retarded hatching, or even blocked it (at 10(-5) M). Obviously retinoic acid interferes with the induction of metamorphosis, possibly through neutralizing one or more factors responsible for settlement and metamorphosis. The possible role of thyroxin in these processes is being critically evaluated.

Animals↗
PubMed1997

Disposition of netobimin, albendazole, and its metabolites in the pregnant rat: developmental toxicity.

Netobimin (NTB), a benzimidazole prodrug with a good anthelmintic spectrum, was administered orally to female rats at a dose of 59.5 mg NTB/kg, to study its pharmacokinetic behavior and the disposition of its most important metabolites, albendazole (ABZ), albendazole sulfoxide (ABZSO), and albendazole sulfone (ABZSO2). ABZ was found in plasma after 6 hr. Peak plasma concentrations (Cmax) and areas under curves (AUC) of ABZSO were eight- and fourfold higher, respectively, than those of ABZSO2. To study NTB disposition in pregnant rats, three different drug doses (50, 59.5, and 70.7 mg/kg) were given. No significant differences were found between plasma concentrations for each metabolite at the three doses studied. Only ABZ concentrations rose slightly as dose increased. ABZ, ABZSO, and ABZSO2 were found in amniotic sacs and embryos at concentrations that were higher than plasma at the same times. The fetuses obtained after administration of each of the doses of NTB were studied to detect developmental toxicity. A significant correlation was found between rate of developmental toxicity and metabolite concentration. ABZSO embryo concentrations could be the main factor accounting for toxicity.

Albendazole↗
PubMed1986

Identification of the chick neural retina cell surface N-acetylgalactosaminyltransferase using monoclonal antibodies.

Intact embryonic chick neural retina cells have at their surface an N-acetylgalactosaminyltransferase which catalyzes the incorporation of N-acetylgalactosamine from UDP-N-acetylgalactosamine into endogenous macromolecular acceptors. The enzyme along with its endogenous acceptors can be isolated as a particulate complex following treatment of membrane-enriched fractions with Triton X-100. In this paper we report on two separate fusions generating monoclonal antibodies: one using as immunogen the particulate complex and the second using as immunogen a soluble N-acetylgalactosaminyltransferase found in tissue-culture-conditioned medium which lacks endogenous acceptor activity. Antibodies from both fusions recognize an antigen which is tightly associated with the particulate transferase/acceptor complex and a soluble antigen having N-acetylgalactosaminyltransferase activity toward exogenously added acceptors. The antibodies recognize a component of ca Mr 220,000, which shows N-acetylgalactosaminyltransferase activity after SDS-gel electrophoresis and transfer to nitrocellulose. This component comigrates on two-dimensional gel electrophoresis with an iodinatable cell surface component whose presence at the cell surface correlates with endogenous transferase activity. We conclude that the antibodies recognize the transferase enzyme itself. Immunohistochemical analysis shows that the enzyme is initially localized throughout the embryonic neural retina in a pattern indicative of a cell surface disposition but becomes restricted to the outer plexiform layer and to outer segments in the adult.

Animals↗
PubMed1988

[Exogenous harmful substances in pregnancy and their effects on the embryo and fetus].

When using drugs during pregnancy the advantages and risks of a therapy have to be considered carefully. In order to assess the risk of possible teratogenic effects under the influence of external harmful substances the following factors have to be taken into account: the type of the drug administered, kind, dose and duration of the medication, impaired metabolism of drugs owing to diseases of te mother, placenta transfer, time of the exposition during pregnancy, genetic disposition. As in other cases in pharmacology and toxicology there are clear relations of dose and effect as a result of an exposition. High doses, administered for a short time, are extremely dangerous as they may affect the fetus. A deleterious effect on the fetus in the first trimester results in abortions and malformations, in the second and third trimester in general retardations of the fetal development, postnatal impairments of the functional development, increasing perinatal mortality, premature deliveries, stillbirths, transplacental carcinogesis. A long-term exposition or the administration of high doses of different substances at the end of pregnancy may produce syndromes which may possibly endanger the newborn. Following drugs are among these pharmacons: Benzodiazepines, Neuroleptics, tricyclic antidepressants, Lithium, Beta-receptor-blocking agents, Beta-sympathomimetics using as tocolytics, Captopril. Withdrawal syndromes of the newborns have been described after a long-term therapy with certain drugs or due to drug addiction of the mother. In every pregnancy there is a certain risk of giving birth to an abnormal child. Malformations observed at birth or immediately afterwards happen according to the definition with a frequency of 2 to 4% (FRG), according to sources from other countries with a frequency of 2.5 to 6.5%. If you add anomalies diagnosed within first year of life a deformity rate of 10% can be worked out, approximately 4 to 5% of which can be traced back to the exposition to drugs and environmental chemicals. A teratogenic influence would always have to work together with other factors to cause a faulty development of the fetus.

Abnormalities, Drug-Induced↗
PubMed1993

[Differentiational profiles of skeletal muscle internal membrane systems directly related to excitation-contraction coupling].

The arrangements of internal membrane systems (transverse (T) tubules, sarcoplasmic reticulum (SR) and T-SR junction (triad)) are matched to contractile profiles in adult muscle fibers. The developmental profiles of the these internal membrane systems directly related to the excitation-contraction (EC) coupling were investigated in the chicken anterior (ALD, slow) and posterior (PLD, fast) latissimus dorsi muscles in ovo (14 to 21 days incubation: E 14-E 21) and after hatching (1 to 30 days after hatching: D 1-D 30). These internal membrane systems were visualized using confocal and electron microscopes. ALD and PLD have similar contraction time in early developmental stage (E 14-E 16), however, PLD suddenly becomes faster at E 17-E 18. Following the differentiation of faster contraction properties (E 18-E 19), density of triads increased significantly in PLD. Early development of T-tubules, on the other hand, is quite similar in both ALD and PLD. Peripherally located, longitudinally oriented T tubules, and the first T networks crossing the fiber center appear earlier in ALD (E 14-E 16) than PLD, but have similar dispositions. The final fiber type specific disposition of T-tubules is established after hatching, and three major components of the E-C coupling pathway are in place as soon as myofibrils develop.

Animals↗
PubMed2006

Enhanced tumorigenesis in p53 knockout mice exposed in utero to high-dose vitamin E.

The limited antioxidative capacity of the embryo and fetus may increase their risk for cancer initiation and/or promotion by reactive oxygen species (ROS)-mediated oxidative DNA damage and/or signaling. To determine if cancer can originate in utero, a high dietary dose of the antioxidant vitamin E (VE) (10% dl-alpha-tocopherol-acetate) was given to cancer-prone p53 knockout mice throughout pregnancy. Although reducing fetal death (P < 0.05), in utero exposure to VE enhanced postnatal tumorigenesis in both +/- (P < 0.04) and -/- (P < 0.0008) p53-deficient offspring. VE did not alter maternal weights, offspring p53 genotypic distribution or tumor spectrum. Constitutive embryonic DNA oxidation in untreated -/- p53 embryos [gestational day (GD) 13] was higher than in +/- and +/+ p53 littermates (P < 0.05). VE reduced DNA oxidation in -/- p53 embryos (P < 0.05) without affecting +/- and +/+ p53 littermates. VE had contrasting, tissue-dependent effects on fetal (GD 19) DNA oxidation, with reductions in -/- and +/- p53-deficient fetal brains (P < 0.01), increases in skin (P < 0.05) and no effect in liver and thymus. The 250-fold increase in dietary VE levels produced only 1.6-6.3-fold, tissue-dependent increases in tissue concentrations. The greatest increase, in fetal skin, correlated with increased DNA oxidation in that tissue in -/- and +/- p53-deficient fetuses and enhanced tumorigenesis in these genotypes. These results show that some cancers may originate in utero and the risk can be enhanced by embryonic and fetal exposure to high dietary levels of VE. The elevated DNA oxidation in some tissues of untreated -/- p53 offspring suggests that ROS may contribute to their higher baseline tumor incidence. The limited and tissue-dependent disposition of VE indicates substantial conceptal regulation. The similarly selective and contrasting effects of VE on DNA oxidation may contribute to its controversial protective efficacy and suggest that its effects on tumorigenesis are cell-specific, possibly in high doses involving a pro-oxidative mechanism.

Animals↗
PubMed1986

Toxicological aspects of cyclamate and cyclohexylamine.

In the late 1960s the artificial sweetener cyclamate was implicated as a bladder carcinogen in rats. This finding and other concerns about its safety ultimately led to a ban on cyclamate in the U.S. and restrictions on its use in many other countries. Since that time, the carcinogenic potential of cyclamate and cyclohexylamine, its principal metabolite, has been reevaluated in a group of well-controlled, well-designed bioassays that have failed to substantiate the earlier findings. This review of the published and unpublished literature on cyclamate attempts to evaluate the carcinogenicity question and other important aspects of the toxicity of cyclamate and cyclohexylamine, including their effects on various organ systems, their genotoxic potential, and their effects on reproduction. In addition, the physiological disposition of cyclamate is reviewed, with particular attention directed toward the site and extent of its conversion to cyclohexylamine.

Abnormalities, Drug-Induced↗
PubMed1981

Development of a sensory afferent projection in the grasshopper embryo. I. Growth of peripheral pioneer axons within the central nervous system.

The grasshopper's cercal nerve is established early in embryogenesis by an identified pair of peripheral neurons called the cercal pioneers. Like the peripheral pioneer neurons in other insect appendages, these two cells send their axons from the periphery to the rudimentary CNS and thus lay the foundation for a nerve that will later be followed by a large number of sensory axons. In this paper, cobalt fills of the primordial cercal nerve were used to characterize the disposition of these peripheral pioneer axons within the embryonic CNS. The pioneer axons stained by this technique terminate in ellipsoidal growth cones which have filopodia radiating from the leading edge and a single long terminal filament pointing along the path the axon is taking. The growing axons also bear filopodia along their sides, but these structures disappear as the cells mature. The pioneer axons of the cercal nerve make an abrupt turn where they first enter the ganglion rudiment and join the axons of the primary longitudinal tract. The pioneers then grow along this tract for several hundred microns without forming secondary growth cones or branches. This prolonged absence of central arborization distinguishes the peripheral pioneer axons from the axons of later-arising epidermal sensory neurons.

Animals↗
PubMed1995

Development of a physiologically based pharmacokinetic model describing 2-methoxyacetic acid disposition in the pregnant mouse.

Using the potent developmental toxicant 2-methoxyethanol (2-ME) as a prototypical compound, a physiologically based pharmacokinetic (PBPK) model was developed to describe the disposition of its primary metabolite and proximate toxicant 2-methoxyacetic acid (2-MAA) in the pregnant CD-1 mouse. Data were collected during early, mid, and late organogenesis, specifically Gestation Days (GD) 8, 11, and 13 (GD 0 = plug-positive date). Pharmacokinetics and tissue partition coefficients for 2-MAA were determined in maternal plasma and conceptus on GD 8 and in maternal plasma, embryo, and extraembryonic/amniotic fluid (EAF) on GD 11 and 13. For simulation of GD 8 data, the conceptus was described as a single compartment, combining the yolk sac placenta, embryo, EAF, and decidua. For GD 11 and 13, the placenta, embryo, and EAF were explicitly described. Several hypotheses were tested for their ability to predict 2-MAA dosimetry. These hypotheses were encoded as alternative models having (a) blood flow-limited delivery of 2-MAA to model compartments, (b) pH trapping of ionized 2-MAA within compartments, (c) active transport of 2-MAA into compartments, and (d) reversible binding of 2-MAA within compartments. While the flow-limited description adequately predicted GD 8 dosimetry, the best simulations of the pharmacokinetic data collected on GD 11 and 13 were obtained with the active transport models. Since the mechanism by which 2-MAA accumulates into the embryo and EAF has not yet been elucidated, these mathematical descriptions are empirical. Further development of this PBPK model for 2-MAA in pregnant mice, in particular its scale-up to humans, will facilitate more realistic human risk assessments for the developmental toxicity of 2-ME and related compounds.

Acetates↗