Embryological Research Research Articles

Traversing birth: continuity and contingency in research on development in nineteenth-century life and human sciences.

In the history of life sciences, it has often been argued that the individual organism emerged, around 1800, as a four-dimensional entity--a temporalized entity. Against this backdrop, the article asks how research on development contributed to structuring the time of the organism in terms of a historical process, that is, by understanding a given phenomenon as brought forth by what preceded it and as establishing conditions for what will follow, thus relating the past, the present and the future in a specific way. To shed light on this conceptualization, we must take into account not only embryological research on morphogenesis but also physiological research on the genesis of vital functions and the causation of congenital anomalies. Three layers of structuring time in such research may be discerned: the making of a trans-natal continuity of the developing organism; a conceptualization of birth as a threshold of past and future that paradigmatically reveals the historical understanding of such developmental continuity; and an approach to intergenerational transmission that confronts developmental continuity with historical contingency. My contribution focuses on the work of William T. Preyer and Charles Féré but, in a genealogical vein, situates their work in the larger context of nineteenth-century research on development.

Observing temporal order in living processes: on the role of time in embryology on the cell level in the 1870s and post-2000.

The article analyses the role of time in the visual culture of two phases in embryological research: at the end of the nineteenth century, and in the years around 2000. The first case study involves microscopical cytology, the second reproductive genetics. In the 1870s we observe the first of a series of abstractions in research methodology on conception and development, moving from a method propagated as the observation of the "real" living object to the production of stained and fixated objects that are then aligned in temporal order. This process of abstraction ultimately fosters a dissociation between space and time in the research phenomenon, which after 2000 is problematized and explicitly tackled in embryology. Mass data computing made it possible partially to re-include temporal complexity in reproductive genetics in certain, though not all, fields of reproductive genetics. Here research question, instrument and modelling interact in ways that produce very different temporal relationships. Specifically, this article suggests that the different techniques in the late nineteenth century and around 2000 were employed in order to align the time of the researcher with that of the phenomenon and to economize the researcher's work in interaction with the research material's own temporal challenges.

The Work Surfaces of Morphogenesis: The Role of the Morphogenetic Field

How biological form is generated remains one of the most fascinating but elusive challenges for science. Moreover, it is widely documented in contemporary literature that development is tightly coordinated. The idea that such development is governed by a coordinating field of force, the morphogenetic field, and its position in embryology research paradigms, is traced in this article. Empirical evidences for field phenomena are described, ranging from bioelectromagnetic effects, morphology, transplantation, regeneration, and other data. Applications of medical potential including treatment of cancer, birth defects, and wound healing are highlighted. The article hypothesizes that distinct morphological forms may have distinct field parameters. Experimentally tractable field parameters may thus provide an exciting research program for probing morphogenesis and phylogenetic diversity.

Embryological processes in ovules of Rudbeckia laciniata L. (Asteraceae) from Poland

Embryological researches of an invasive plant Rudbeckia laciniata focused on the process of megasporogenesis, female gametophyte formation, and fertilization. A disturbed first meiotic division results in the formation of restitution nucleus. In some megaspore mother cells micronuclei or vacuoles were observed near the restitution nucleus. Different course of megasporogenesis led to the development of unreduced embryo sacs with various organization which is reflected in the embryo formation and capacity of seed germination. Our research confirmed the occurrence of hemigamy in the Polish representatives of R. laciniata. In spite of a reduced germination capacity of R. laciniata seeds, their production allows a long-distance dispersal and expansion of the species.

“No interest in human anatomy as such”: Frederic Wood Jones dissects anatomical investigation in the United States in the 1920s

In 1926, Frederic Wood Jones, professor of Anatomy at the University of Adelaide and a leading figure in the British anatomical world, took a Rockefeller Foundation funded trip to the United States in order to inspect anatomy programmes and medical museums and to meet leading figures in the anatomical and anthropological world. His later reflections paint a picture of a discipline in transition. Physical anthropology and gross anatomy were coming to a crisis point in the United States, increasingly displaced by research in histology, embryology and radiological anatomy. Meanwhile, in Britain and its colonial outposts, anatomists such as Wood Jones were attempting to re-invigorate the discipline in the field, studying biological specimens as functional and active agents in their particular milieus, but with human dissection at the core. Thus, an examination of this trip allows us to see how the interaction between two traditions in anatomy informed the process of the development of human biology in this critical period.

Restoring the resting potential brings maturity to induced stem cell–derived human cardiac myocytes

Reprogramming of mature human cells to produce the equivalent of embryonic stem cells offers exciting possibilities for therapeutic tissue repair and artificial organs. A more immediate benefit is the possibility of testing potential new drugs and other therapies in human cells of any type that can easily be created in vitro without the use of human blastocysts and with a minimum of invasive procedures in patients. Fundamental research in embryology lead to the identification of growth factors that could be used to force cultured embryonic stem cells to differentiate into mature adult cells including atrial and ventricular cardiac myocytes. This work grew out of the initial approach of allowing the harvesting and subculturing regions of embryos growing in vitro that showed spontaneous beating. A spectacular advance came in 2006 when Takahashi and Yamanaka 1 showed that mouse or human adult fibroblasts could revert to pluripotent stem cells when exposed to 4 factors: Oct3/4, Sox2, c-Myc, and Klf4 under embryonic stem cell culture conditions. In 2012, Yamanaka received the Nobel Prize in Medicine or Physiology for this work, which he shared with Gurdon, who showed in l962 that the specialization of mature cells can be reversed by transplantation of a frog intestinal cell nucleus into a frog egg. By the time of this award, substantial work by others had shown what additional factors or conditions are needed to cause the so-called induced pluripotent stem (IPS) cells to differentiate into adult cardiac myocytes or other cellular components including vascular and endothelial cells. 2 In the heart, embryonic development and tissue differentiation is accompanied by important changes in the electrical activity that alter the cardiac action potential and underlying ionic currents. While the genome must contain sequence information for all ion channels in the heart and other tissues, the expression of the ion channels varies with the stage of embryonic development and with anatomic location. In the embryonic hearts of vertebrates, the action potential of the immature ventricle is similar to that found in the adult sinoatrial node. Specifically, there is spontaneous diastolic depolarization (the pacemaker potential) with a relatively low maximum diastolic potential in immature atrial and ventricular cells, which disappears during development. The upstroke of the action potential is mediated by calcium channels in the early embryo and by sodium channels in the late embryo and adult. The action potential upstroke becomes increasingly rapid during embryonic development, as demonstrated by Nathan and DeHaan 3 in the chick heart. Mature atrial and ventricular cells have a resting potential that is due to predominant permeability to potassium and is approximately � 90 mV, which is the potassium equilibrium

Occupational egg allergy in an embryological research facility

Hen and quail eggs are commonly used in embryological research. While immunoglobin E (IgE)-associated allergy to hens' egg proteins is recognized in employees in the food industry, there are no previous reports from workers in embryological research. Two newly identified cases prompted us to examine the extent of this problem in a university laboratory. To determine the prevalence and determinants of sensitization to egg among a group of workers in an embryological research laboratory. Following the identification of egg sensitization in two research workers, we surveyed 116 employees from a single embryology research laboratory in 2005. Sensitization to egg was assessed by skin prick tests and/or specific IgE measurement and examined in relation to a number of potential determinants, including the extent of appropriate control measures. Four employees were sensitized to egg, each with digestive symptoms of egg allergy. All had worked directly with eggs, giving a prevalence of specific sensitization in that group of 6.1% (95% confidence interval (CI) [1.7%, 14.8%]). There was a statistically significant trend for those sensitized to report a shorter duration of exposure to eggs and less frequent use of hygiene control measures to reduce exposure. Sensitization to eggs among those occupationally exposed to egg protein in research work occurs more commonly than in the general population. The presence of digestive symptoms after eating eggs may be indicative of sensitization to egg and should be incorporated into health evaluation of exposed workers.

FISH reanalysis of inner cell mass and trophectoderm samples of previously array-CGH screened blastocysts shows high accuracy of diagnosis and no major diagnostic impact of mosaicism at the blastocyst stage

Does comprehensive chromosome screening (CCS) of cells sampled from the blastocyst trophectoderm (TE) accurately predict the chromosome complement of the inner cell mass (ICM)? Comprehensive chromosome screening of a TE sample is unlikely to be confounded by mosaicism and has the potential for high diagnostic accuracy. The effectiveness of chromosome aneuploidy screening is limited by the technologies available and chromosome mosaicism in the embryo. Combined with improving methods for cryopreservation and blastocyst culture, TE biopsy and CCS is considered to be a promising approach to select diploid embryos for transfer. The study was performed between January 2011 and August 2011. In the first part, a new ICM isolation method was developed and tested on 20 good morphology blastocysts. In the main phase of the study, fluorescence in situ hybridization (FISH) was used to reanalyse the ICMs and TEs separated from 70 embryos obtained from 26 patients undergoing blastocyst stage array comparative genome hybridization (aCGH) PGS cycles. The isolated ICM and TE fractions were characterized by immunostaining for KRT18. Then, non-transferrable cryopreserved embryos were selected for the FISH reanalysis based on previous genetic diagnosis obtained by TE aCGH analysis. Blastocysts either diploid for chromosome copy number (20) or diagnosed as single- (40) or double aneuploid (10) were included after preparing the embryo into one ICM and three equal-sized TE sections. Accuracy of the aCGH was measured based on FISH reanalysis. Chromosomal segregations resulting in diploid/aneuploid mosaicism were classified as 'low-', 'medium-' and 'high-' grade and categorized with respect to their distribution (1TE, 2TE, 3TE, ICM or ALL embryo). Linear regression model was used to test the relationship between the distributions and the proportion of aneuploid cells across the four embryo sections. Fisher's exact test was used to test for random allocation of aneuploid cells between TE and ICM. All ICM biopsy procedures displayed ICM cells in the recovered fraction with a mean number of ICM cells of 26.2 and a mean TE cell contamination rate of 2%. By FISH reanalysis of previously aCGH-screened blastocysts, a total of 66 aneuploidies were scored, 52 (78.8%) observed in all cells and 14 (21.2%) mosaic. Overall, mosaic chromosomal errors were observed only in 11 out of 70 blastocysts (15.7%) but only 2 cases were classified as mosaic diploid/aneuploid (2.9%). Sensitivity and specificity of aCGH on TE clinical biopsies were 98.0 and 100% per embryo and 95.2 and 99.8% per chromosome, respectively. Linear regression analysis performed on the 11 mosaic diploid/aneuploid chromosomal segregations showed a significant positive correlation between the distribution and the proportion of aneuploid cells across the four-blastocyst sections (P < 0.01). In addition, regression analysis revealed that both the grade and the distribution of mosaic abnormal cells were significantly correlated with the likelihood of being diagnosed by aCGH performed on clinical TE biopsies (P = 0.019 and P < 0.01, respectively). Fisher's exact test for the 66 aneuploidies recorded showed no preferential allocation of abnormal cells between ICM and TE (P = 0.33). The study is limited to non-transferable embryos, reanalyzed for only nine chromosomes and excludes segmental imbalance and uniparental disomy. The prevalence of aneuploidy in the study group is likely to be higher than in the general population of clinical PGD embryos. This study showed high accuracy of diagnosis achievable during blastocyst stage PGS cycles coupled with 24-chromosomes molecular karyotyping analysis. The new ICM isolation strategy developed may open new possibilities for basic research in embryology and for clinical grade derivation of human embryonic stem cells. No specific funding was sought or obtained for this study.

Efficient Embryo Transfer in the Common Marmoset Monkey (Callithrix jacchus) with a Reduced Transfer Volume: A Non-Surgical Approach with Cryopreserved Late-Stage Embryos1

Among primates, the common marmoset is suitable for primate embryology research. Its small body size, however, has delayed the technical development of efficient embryo transfer. Furthermore, three factors have been determined to adversely affect the performance of marmoset embryo transfer: nonsurgical approaches, the use of cryopreserved embryos, and the use of late-stage embryos. Here we performed embryo transfer under conditions that included the above three factors and using either a small (1 μl or less) or a large volume (2-3 μl) of medium. The pregnancy and birth rates were 50% (5/10) and 27% (3/11), respectively, when using the large volume, and 80% (8/10) and 75% (9/12), respectively, when using the small volume. The latter scores exceed those of previous reports using comparable conditions. Thus, it appears that these three previously considered factors could be overcome, and we propose that reducing the transfer volume to 1 μl or less is essential for successful marmoset embryo transfer.

Frozen-thawed spermatozoa from oligozoospermic ejaculates are susceptible to in situ DNA fragmentation in polyvinylpyrrolidone-based sperm-immobilization medium

To elucidate the effect of sperm immobilization media that are and are not based on polyvinylpyrrolidone (PVP) on the DNA integrity of fresh and frozen-thawed spermatozoa during standard intracytoplasmic sperm injection (ICSI) conditions. Experimental prospective study. Embryology research laboratory. Forty-six ejaculates from normozoospermic and oligozoospermic men. Assessment of sperm DNA fragmentation by single-cell gel electrophoresis assay. DNA integrity of fresh and frozen-thawed spermatozoa from normozoospermic and oligozoospermic ejaculates exposed to PVP-based and non-PVP-based media. Exposure of fresh and frozen thawed spermatozoa from normozoospermic and oligozoospermic ejaculates to PVP-based medium in an ICSI dish for 30 minutes statistically significantly increased the DNA fragmentation. In contrast, the extent of DNA fragmentation in non-PVP-based medium did not statistically significantly differ from control. A PVP-based medium can induce a statistically significant amount of sperm DNA fragmentation in an ICSI dish, and frozen-thawed sperm from oligozoospermic ejaculates are more susceptible to in situ DNA fragmentation.

Cellular and molecular changes associated with somatic embryogenesis induction in Agave tequilana

In spite of the importance of somatic embryogenesis for basic research in plant embryology as well as for crop improvement and plant propagation, it is still unclear which mechanisms and cell signals are involved in acquiring embryogenic competence by a somatic cell. The aim of this work was to study cellular and molecular changes involved in the induction stage in calli of Agave tequilana Weber cultivar azul in order to gain more information on the initial stages of somatic embryogenesis in this species. Cytochemical and immunocytochemical techniques were used to identify differences between embryogenic and non-embryogenic cells from several genotypes. Presence of granular structures was detected after somatic embryogenesis induction in embryogenic cells; composition of these structures as well as changes in protein and polysaccharide distribution was studied using Coomassie brilliant blue and Periodic Acid-Schiff stains. Distribution of arabinogalactan proteins (AGPs) and pectins was investigated in embryogenic and non-embryogenic cells by immunolabelling using anti-AGP monoclonal antibodies (JIM4, JIM8 and JIM13) as well as an anti-methyl-esterified pectin-antibody (JIM7), in order to evaluate major modifications in cell wall composition in the initial stages of somatic embryogenesis. Our observations pointed out that induction of somatic embryogenesis produced accumulation of proteins and polysaccharides in embryogenic cells. Presence of JIM8, JIM13 and JIM7 epitopes were detected exclusively in embryogenic cells, which supports the idea that specific changes in cell wall are involved in the acquisition of embryogenic competence of A. tequilana.

Chironomid midges: a forgotten model of developmental biology research

AbstractFor more than a century, embryologists have been exploring various model systems to gain insights into developmental processes. This article presents an overview of the role of chironomid midges in embryology research since their introduction as model organisms in the 19th century. We present the vestiges of bibliography since the days of Weismann (1834–1914), who raised preliminary queries to unravel many unique features of insect embryogenesis using midges as a crucible. Unfortunately, over the years, chironomid midges got lost into obscurity as a model for developmental biology, which is evident from the paucity of developmental biology–related literature on midges in the past decades. Through this essay, the authors intend to share reminiscences of the heydays of chironomid research with the wider community of zoologists with an aim of reviving chironomid embryology. Midges not only possess the basic qualities essential for an ideal model system, but being one of the ancestral dipteran stocks, they can also prove an excellent test system for evo‐devo, transgenetic, and embryogenomic investigations that utilize methodologies at the interface of developmental biology and high‐throughput molecular genetic and genomics approach. An introspection of re‐introducing chironomid midgesas model system will be rewarding for the contemporary developmental biologists.

Between Biochemists and Embryologists – The Biochemical Study of Embryonic Induction in the 1930s

The discovery by Hans Spemann of the "organizer" tissue and its ability to induce the formation of the amphibian embryo's neural tube inspired leading embryologists to attempt to elucidate embryonic inductions' underlying mechanism. Joseph Needham, who during the 1930s conducted research in biochemical embryology, proposed that embryonic induction is mediated by a specific chemical entity embedded in the inducing tissue, surmising that chemical to be a hormone of sterol-like structure. Along with embryologist Conrad H. Waddington, they conducted research aimed at the isolation and functional characterization of the underlying agent. As historians clearly pointed out, embryologists came to question Needham's biochemical approach; he failed to locate the hormone he sought and eventually abandoned his quest. Yet, this study finds that the difficulties he ran into resulted primarily from the limited conditions for conducting his experiments at his institute. In addition, Needham's research reflected the interests of leading biochemists in hormone and cancer research, because it offered novel theoretical models and experimental methods for engaging with the function of the hormones and carcinogens they isolated. Needham and Waddington were deterred neither by the mounting challenges nor by the limited experimental infrastructure. Like their colleagues in hormone and cancer research, they anticipated difficulties in attempting to establish causal links between complex biological phenomena and simple chemical triggering.

The Rabbit(Oryctolagus cuniculus):A Model for Mammalian Reproduction and Early Embryology

INTRODUCTIONThe rabbit is a valuable animal model for a variety of biomedical research areas including in vitro fertilization, early embryology and organogenesis, neurophysiology, ophthalmology, and cardiovascular research. The rabbit is also used as a model for toxicology studies and analyses of drug effects on embryo and fetal development, as well as for research involving the immune system (e.g., host/pathogen interactions, asthma, arthritis, systemic lupus erythematosus). The rabbit’s most prominent use as a laboratory animal is antibody production; most biomedical research institutions thus provide the infrastructure and veterinary expertise for keeping rabbits. This article discusses current advances in applying modern life science research methods to basic and applied embryological research using the rabbit as a model for early intrauterine development in mammals, including state-of-the-art genomic tools and the sequencing data available (and planned) for the rabbit.

The Human Fertilisation and Embryology Act 2008: Tinkering at the Margins

This note suggests that, viewed from a feminist perspective, the reforms contained in the Human Fertilisation and Embryology Act 2008 represent a missed opportunity to re-think the appropriate model of regulation to govern fertility treatment and embryology research in the UK. It argues that reform of the legislation was driven largely by the government’s desire to avoid re-igniting controversies over the legal status of the embryo and abortion and to maintain Britain’s position at the forefront of embryo research and related biotechnologies. It also highlights the importance of media debates, which were highly selective, to the reform process, and suggests that in order to inject feminist values into the process of legislative reform, feminists need to become more media savvy. In the short term, it suggests that there is little prospect of a radical re-thinking of the appropriate ethico-legal response to the wide variety of family forms that reproductive technologies potentially enable, much less of considering our ethical obligations to the new forms of embryos that are now permitted by the 2008 Act. In the meantime, however, it argues that these issues provide productive opportunities for feminist legal theorists to address questions that have been erased or obscured in the course of the 2008 reforms.

Effect of varying equilibration time in a two-step vitrification method on the post-warming DNA integrity of mouse blastocysts

To assess the effect of equilibration time on the DNA integrity of vitrified-warmed mouse blastocysts. Prospective in vitro study. Embryology research laboratory. Mouse nonexpanded blastocysts (NEB) (n = 54) and expanded blastocysts (EB) (n = 56) were vitrified using cryotips. Blastocysts were vitrified using a two-step media protocol, with a first (equilibration) step of 4 minutes [NEB (n = 20), EB (n = 24)], 8 minutes [NEB (n = 17), EB (n = 16)], or 15 minutes: [NEB (n = 17), EB (n = 16)]. Control samples were fresh NEB (n = 17) and EB (n = 18) blastocysts. DNA integrity index (DII) using terminal deoxynucleotide transferase-mediated dUTP nick-end labeling, confocal imaging, and viability. 1) The DII of the vitrified warmed NEB and EB improved significantly with 8-minute equilibration protocol compared with 4 minutes. 2) The DII of the EB significantly decreased with 15-minute equilibration protocol compared with 4 or 8 minutes. 3) The DII in the NEB significantly improved with 8- and 15-minute equilibration protocols compared with 4 minutes, with no significant difference between 8 or 15 minutes. Vitrification with an 8-minute equilibration step improved DII of the NEB and EB after warming.

Transgenerational changes in somatic and germ line genetic integrity of first-generation offspring derived from the DNA damaged sperm

To report a more quantitative approach to study the influence of varying levels of sperm DNA damage on transgenerational changes in genomic instability in a mouse model. Experimental prospective study. Embryology research laboratory. Swiss albino mice. The sperm DNA damage was induced by different doses of gamma-irradiation to male mice followed by mating with healthy female mice. Genomic integrity in embryos, fetus, and spermatozoa of F1 mice derived from the DNA-damaged sperm. The transgenerational changes in genetic integrity were attributed by a dose-dependent increase in the frequency of micronuclei in preimplantation embryos and a concomitant increase in genomic instability in fetal liver cells and sperm chromatin modifications in F1 males. A strong positive correlation was observed between the extent of sperm DNA damage and somatic and germ-line genomic instability. Sperm-mediated transgenerational genomic instability is dependent on the amount of DNA damage present in the sire's sperm at the time of fertilization.

Zebrafish Embryology and Cartilage Staining Protocols for High School Students

The Life Sciences-Howard Hughes Medical Institute Outreach Program at Harvard University supports high school science education by offering an on-campus program for students and their teachers to participate in investigative, hands-on laboratory sessions. The outreach program has recently designed and launched a successful zebrafish embryology protocol that we present here. The main objectives of this protocol are to introduce students to zebrafish as a model research organism and to provide students with direct experience with current techniques used in embryological research. The content of the lab is designed to generate discussions on embryology, genetics, fertilization, natural selection, and animal adaptation. The protocol produces reliable results in a time-efficient manner using a minimum of reagents. The protocol presented here consists of three sections: observations of live zebrafish larvae at different developmental stages, cartilage staining of zebrafish larvae, and a mutant hunt involving identification of two zebrafish mutants (nacre and chokh). Here, we describe the protocol, show the results obtained for each section, and suggest possible alternatives for different lab settings.

Embryogenesis and laboratory maintenance of the foam‐nesting túngara frogs, genus Engystomops (= Physalaemus)

The vast majority of embryological research on amphibians focuses on just a single genus of frogs, Xenopus. To attain a more comprehensive understanding of amphibian development, experimentation on non-model frogs will be essential. Here, we report on the early development, rearing, and embryological analysis of túngara frogs (genus Engystomops, also called Physalaemus). The frogs Engystomops pustulosus, Engystomops coloradorum, and Engystomops randi construct floating foam-nests with small eggs. We define a table of 23 stages for the developmental period in the foam-nest. Embryos were immunostained against Lim1, neural, and somite-specific proteins and the expression pattern of RetinoBlastoma Binding Protein 6 (RBBP6) was analyzed by in situ hybridization. Due to their brief life-cycle, frogs belonging to the genus Engystomops are attractive for comparative and genetic studies of development. Developmental Dynamics 238:1444-1454, 2009. (c) 2009 Wiley-Liss, Inc.

Commentary on Domestic Animals in Agricultural and Biomedical Research: An Endangered Enterprise

Commentary on Domestic Animals in Agricultural and Biomedical Research: An Endangered Enterprise