Decorin developmental expression and function in the early avian embryo

Decorin, a proteoglycan, interacts with extracellular matrix proteins, growth factors and receptors. Decorin expression and spatio-temporal distribution were studied by RT-PCR and immunofluorescence, while decorin function was examined by blocking antibodies in the early chick embryo. Decorin was first detectable at stage XIII (late blastula). During gastrulation (stage HH3-4), decorin fluorescence was intense in epiblast cells immediately adjacent to the streak, and in migrating cells. Decorin fluorescence was intense in endoderm and strong at mesoderm-neural plate surfaces at stage HH5-6 (neurula). At stage HH10-11 (12 somites), decorin fluorescence was intense in myelencephalon and then showed distinct expression patterns along the myelencephalon axes by stage HH17. Decorin fluorescence was intense in neural crest cells, dorsal aorta, heart, somite and neuroepithelial cells apposing the somite, nephrotome, gut and in pancreatic and liver primordia. Antibody-mediated inhibition of decorin function affected the head-to-tail embryonic axis extension, indicating that decorin is essential for convergent extension cell movements during avian gastrulation. Decorin was also essential for retinal progenitor cell polarization, neural crest migration, somite boundary formation and cell polarization, mesenchymal cell polarization and primary endoderm displacement to the embryo periphery. The embryonic blood vessels were deformed, the dorsal mesocardium was thinned and the cardiac jelly was abnormally thickened in the heart. Decorin is known to modulate collagen fibrillogenesis, a key mechanism of matrix assembly, and cell proliferation. Decorin also appears to be essential for the coordination of cell and tissue polarization, which is an important feature in organ patterning of the embryo.     

Entangled lives: Implications of the developmental origins of health and disease hypothesis for bioarchaeology and the life course

Epidemiological research since the 1980s has highlighted the consequences of early life adversity, particularly during gestation and early infancy, for adult health (the “Barker hypothesis”). The fast‐evolving field of molecular epigenetics is providing explanatory mechanisms concerning phenotypic plasticity in response to developmental stressors and the accumulation of disease risk throughout life. In addition, there is now evidence for the heritability of poor health across generations via epigenetic modifications. This research has the potential to invoke a paradigmatic shift in how we interpret factors such as growth insults and immune response in past skeletal remains. It demonstrates that health cannot be understood in terms of immediate environmental circumstances alone. Furthermore, it requires both a theoretical and practical re‐evaluation of disease biographies and the life course more generally. Individual life courses can no longer be regarded as discrete, bounded, life histories, with clearly defined beginning and end points. If socioeconomic circumstances can have intergenerational effects, including disease susceptibility and growth stunting, then individual biographies should be viewed as nested or “embedded” within the lives of others. This commingling of life courses may prove problematic to unravel; nevertheless, this review aims to consider the potential consequences for bioarchaeological interpretations. These include a greater consideration of: the temporal power of human skeletons and a life course approach to past health; infant health and the implications for maternal well‐being; and the impact of non‐proximate stressors (e.g., early life and ancestral environments) on the presence of health indicators. Am J Phys Anthropol 158:530–540, 2015. © 2015 Wiley Periodicals, Inc.     

Novel phenotypes and developmental arrest in early embryo specific mutants of maize

Embryo specific (emb) mutants exhibit aberrant embryo development without deleterious effects on endosperm development. We have analyzed five emb mutants of maize, which, based on their developmental profiles can be divided into two groups: mutants arrested at early stages and mutants with novel phenotypes. The members of the first group resemble wild-type proembryos and never reach other developmental stages. In the second group the tube-shaped mutants emb*-8522 and emb*-8535 completely lack apical-basal differentiation, while in mutant emb*-8516 a second embryo-like structure arises from the suspensor. The five emb mutations analyzed are non-allelic and two of the mutations are very likely caused by insertion of the transposon mutator, opening the door for their molecular analysis. Received: 10 February 1999 / Accepted: 7 July 1999     

The developmental origins of obesity and related health disorders--prenatal and perinatal factors

Obesity, and its health-related sequelae (the metabolic syndrome), have recently emerged as a global health crisis. The prevalence of childhood and adult obesity in economically developed and developing countries world-wide has more than doubled over the past decade. While genetic factors, increasingly sedentary lifestyles, and overnutrition have all been cited as important components of the obesity crisis, recent epidemiological and experimental evidence suggests that developmental factors--especially those that occur in utero and during early postnatal life--play a significant role in the pandemic. Research into the 'developmental origins of health and disease' (DOHaD) has now firmly established that pre- and perinatal developmental perturbations which predispose to obesity in adult life can result from a variety of factors, including both nutritional surplus and deficiency, and there is growing evidence that these physiological traits can be passed on epigenetically to subsequent generations. Anthropological perspectives regarding the developmental origins of obesity and its related health problems cannot only shed further light on contemporary ethnic human health disparities, but can offer unique insights into the relevance of the developmental origins of disease to community-based public health interventions.     

兔早期胚胎发育相关新基因IFRG的克隆和表达

近来的研究表明干扰素在哺乳动物早期胚胎发育中有重要的作用。我们首次克隆了兔早期胚胎发育相关新基因IFRG（干扰素应答基因）的全长cDNA序列（AJ584672）,根据该cDNA序列以兔卵巢cDNA为模板经PCR扩增后克隆了兔IFRG cDNA的完整开放阅读框(396bp)。将其克隆到原核表达载体pGEX-4T-2上,在大肠杆菌BL21中进行了GST-IFRG融合蛋白的表达。 经IPTG诱导培养后,SDS-PAGE电泳检测结果显示在41kDa处有特异性表达蛋白,回收融合蛋白作为抗原免疫小鼠,制备多克隆抗体,通过Western杂交表明该融合蛋白具有生物免疫活性。     

A provisional epithelium in leech embryo: Cellular origins and influence on a developmental equivalence group

Segmental tissues of glossiphoniid leeches arise from rostrocaudally arrayed columns (bandlets) of segmental founder cells (primary m, n, o, p, and q blast cells) which undergo stereotyped sublineages to generate identifiable subsets of definitive progeny. The bandlets lie at the surface of the embryo beneath the squamous epithelium of a transient embryonic covering called the provisional integument. This "provisional epithelium" derives from microsomes produced during the early cleavage divisions. Previous experiments have shown that the primary o and p blast cells constitute an equivalence group, i.e., are initially developmentally equipotent and undergo hierarchical interactions which cause them to assume distinct O and P fates. Here, we examine the role of the provisional epithelium in determining the fates of the underlying o and p blast cells. Experiments entailing the microinjection of individual micromeres with cell lineage tracers show that, at stages 7-8 of normal development, the epithelium comprises coherent and relatively stereotyped domains derived from particular micromeres. Upon photoablating domains of epithelium labeled with photosensitizing lineage tracer, the normal assignment of O fates is disturbed; o blast cells divide symmetrically (as p blast cells do) and some supernumerary definitive progeny expressing P fates arise within the O lineage. We therefore conclude that the epithelium is essential for generation and/or reception of signal(s) by which the o and p blast cells' normally determine their fates. Finally, a new tracer substance, biotinylated fixable dextran (BFD), is described which was essential for this study by virtue of its superior resistance to photobleaching and which offers several other advantages as well.     

Modelling the origins of legume domestication and cultivation

Ladizinsky’s (1987) mathematical model of the domestication of lentils and other Near Eastern legumes is invalid. Ladizinsky believed that the model predicts that high rates of seed harvesting would lead to fixation, in wild populations, of genes conferring lack of seed dormancy (a domesticated trait). In fact, however, the model gives rapid fixation of alleles for non-dormancy underall circumstances; gathering by humans has no effect on allelic frequencies. It appears that in these species cultivation must have preceded the morphological changes that distinguish domesticated from wild plants. The addition of more realistic assumptions to the model does not alter this conclusion. I suggest several scenarios that could explain Near Eastern legume domestication: the most plausible of these postulates that cultivation of cereals led to scheduling conflicts which necessitated the abandonment of harvesting of wild legumes, and hence the initiation of their cultivation. Mathematical modelling may be able to contribute to our understanding of agricultural origins, but it must be carried out with greater rigor and closer attention to the theoretical literature.     

Molecular and chemical genetic approaches to developmental origins of aging and disease in zebrafish

The incidence of diseases increases rapidly with age, accompanied by progressive deteriorations of physiological functions in organisms. Aging-associated diseases are sporadic but mostly inevitable complications arising from senescence. Senescence is often considered the antithesis of early development, but yet there may be factors and mechanisms in common between these two phenomena over the dynamic process of aging. The association between early development and late-onset disease with advancing age is thought to come from a consequence of developmental plasticity, the phenomenon by which one genotype can give rise to a range of physiologically and/or morphologically adaptive states in response to different environmental or genetic perturbations. On the one hand, we hypothesized that the future aging process can be predictive based on adaptivity during the early developmental period. Modulating the thresholds of adaptive plasticity by chemical genetic approaches, we have been investigating whether any relationship exists between the regulatory mechanisms that function in early development and in senescence using the zebrafish (Danio rerio), a small freshwater fish and a useful model animal for genetic studies. We have successfully conducted experiments to isolate zebrafish mutants expressing apparently altered senescence phenotypes during embryogenesis (“embryonic senescence”), subsequently showing shortened lifespan in adulthoods. We anticipate that previously uncharacterized developmental genes may mediate the aging process and play a pivotal role in senescence. On the other hand, unexpected senescence-related genes might also be involved in the early developmental process and regulation. The ease of manipulation using the zebrafish system allows us to conduct an exhaustive exploration of novel genes and small molecular compounds that can be linked to the senescence phenotype, and thereby facilitates searching for the evolutionary and developmental origins of aging in vertebrates. This article is part of a Special Issue entitled: Animal Models of Disease.     

The developmental origins of well-being

Low birthweight is now known to be associated with increased rates of coronary heart disease and the related disorders, stroke, hypertension and adult-onset diabetes. These associations have been extensively replicated in studies in different countries and are not the result of confounding variables. They extend across the normal range of birthweight and depend on lower birthweights in relation to the duration of gestation rather than the effects of premature birth. The associations are thought to be consequences of developmental plasticity, the phenomenon by which one genotype can give rise to a range of different physiological or morphological states in response to different environmental conditions during development. Recent observations have shown that impaired growth in infancy and rapid childhood weight gain exacerbate the effects of impaired prenatal growth. A new vision of optimal early human development is emerging, which takes account of health and well-being throughout life.     

Response characteristics of the mitochondrial DNA genome in developmental health and disease

This review focuses on mitochondrial biology in mammalian development; specifically, the dynamics of information transfer from nucleus to mitochondrion in the regulation of mitochondrial DNA genomic expression, and the reverse signaling of mitochondrion to nucleus as an adaptive response to the environment. Data from recent studies suggest that the capacity of embryonic cells to react to oxygenation involves a tradeoff between factors that influence prenatal growth/development and postnatal growth/function. For example, mitochondrial DNA replication and metabolic set points in nematodes may be determined by mitochondrial activity early in life. The mitochondrial drug PK11195, a ligand of the peripheral benzodiazepine receptor, has antiteratogenic and antidisease action in several developmental contexts in mice. Protein malnutrition during early life in rats can program mitochondrial DNA levels in adult tissues and, in humans, epidemiological data suggest an association between impaired fetal growth and insulin resistance. Taken together, these findings raise the provocative hypothesis that environmental programming of mitochondrial status during early life may be linked with diseases that manifest during adulthood. Genetic defects that affect mitochondrial function may involve the mitochondrial DNA genome directly (maternal inheritance) or indirectly (Mendelian inheritance) through nuclear-coded mitochondrial proteins. In a growing number of cases, the depletion of, or deletion in, mitochondrial DNA is seen to be secondary to mutation of key nuclear-coded mitochondrial proteins that affect mitochondrial DNA replication, expression, or stability. These defects of intergenomic regulation may disrupt the normal cross-talk or structural compartmentation of signals that ultimately regulate mitochondrial DNA integrity and copy number, leading to depletion of mitochondrial DNA.     

Developmental plasticity and developmental origins of non-communicable disease: theoretical considerations and epigenetic mechanisms

There is now evidence that developmental influences have lifelong effects on cardiovascular and metabolic function and that elements of the heritable or familial component of susceptibility to cardiovascular disease, obesity and other non-communicable diseases (NCD) can be transmitted across generations by non-genomic means. In animals the developmental environment induces altered phenotypes through genetic, physiological (especially endocrine) and epigenetic mechanisms. The latter include DNA methylation, covalent modifications of histones and non-coding RNAs. Such ‘tuning’ of phenotype has potential adaptive value and may confer Darwinian fitness advantage because it either adjusts the phenotype to current circumstances and/or attempts to match an individual’s responses to the environment predicted to be experienced later. When the phenotype is mismatched to the later environment, e.g. from inaccurate nutritional cues from the mother or placenta before birth, or from rapid environmental change through improved socio-economic conditions, risk of NCD increases. Such mechanisms are also thought to play roles in ageing and early onset of puberty, reinforcing a life-course perspective on such adaptive responses, especially the detrimental later effects of trade-offs. Epigenetic changes induced during development are highly gene-specific and function at the level of individual CpG dinucleotides in both gene promoter and intergenic regions. Evidence is accruing that endocrine or nutritional interventions during early postnatal life can reverse epigenetic and phenotypic changes induced, for example, by unbalanced maternal diet during pregnancy. Elucidation of epigenetic processes may permit perinatal identification of individuals most at risk of later NCD and enable early intervention strategies to reduce such risk.     

Dual origins of the prechordal cranium in the chicken embryo

The prechordal cranium, or the anterior half of the neurocranial base, is a key structure for understanding the development and evolution of the vertebrate cranium, but its embryonic configuration is not well understood. It arises initially as a pair of cartilaginous rods, the trabeculae, which have been thought to fuse later into a single central stem called the trabecula communis (TC). Involvement of another element, the intertrabecula, has also been suggested to occur rostral to the trabecular rods and form the medial region of the prechordal cranium. Here, we examined the origin of the avian prechordal cranium, especially the TC, by observing the craniogenic and precraniogenic stages of chicken embryos using molecular markers, and by focal labeling of the ectomesenchyme forming the prechordal cranium. Subsequent to formation of the paired trabeculae, a cartilaginous mass appeared at the midline to connect their anterior ends. During this midline cartilage formation, we did not observe any progressive medial expansion of the trabeculae. The cartilages consisted of premandibular ectomesenchyme derived from the cranial neural crest. This was further divided anteroposteriorly into two portions, derived from two neural crest cell streams rostral and caudal to the optic vesicle, called preoptic and postoptic neural crest cells, respectively. Fate-mapping analysis elucidated that the postoptic neural crest cells were distributed exclusively in the lateroposterior part of the prechordal cranium corresponding to the trabeculae, whereas the preoptic stream of cells occupied the middle anterior part, differentiating into a cartilage mass corresponding to the intertrabecula. These results suggest that the central stem of the prechordal cranium of gnathostomes is composed of two kinds of distinct cartilaginous modules: a pair of trabeculae and a median intertrabecula, each derived from neural crest cells populating distinct places of the craniofacial primordia through specific migratory pathways.     

Modelling infectious disease to support human health

During the current COVID-19 pandemic, there has been renewed scientific and public focus on understanding the pathogenesis of infectious diseases and investigating vaccines and therapies to combat them. In addition to the tragic toll of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), we also recognize increased threats from antibiotic-resistant bacterial strains, the effects of climate change on the prevalence and spread of human pathogens, and the recalcitrance of other infectious diseases – including tuberculosis, malaria, human immunodeficiency virus (HIV) and fungal infections – that continue to cause millions of deaths annually. Large amounts of funding have rightly been redirected toward vaccine development and clinical trials for COVID-19, but we must continue to pursue fundamental and translational research on other pathogens and host immunity. Now more than ever, we need to support the next generation of researchers to develop and utilize models of infectious disease that serve as engines of discovery, innovation and therapy.

Summary: This Editorial considers how knowledge from animal and other models of infectious disease can impact our understanding of human biology and potential therapies, focusing largely on zebrafish. It also highlights ways in which DMM is supporting these areas.

As an Editor at Disease Models & Mechanisms (DMM) and an academic researcher using zebrafish as a model to study tuberculosis, it is especially exciting to read and publish research in zebrafish to obtain, in a whole, live vertebrate, insights into infectious diseases and therapies (Box 1). Indeed, zebrafish provide a remarkable vertebrate model for many questions related to infectious disease. Embryos and larvae are optically transparent, enabling microscopy of both pathogen and host that would be more challenging or cumbersome in other systems (Fig. 1). Knock-in of fluorescent tags at endogenous loci allows direct and detailed in vivo visualization of the host immune response (Cronan et al., 2018). Both forward and reverse genetic approaches for understanding infection are straightforward and are buttressed by the high-throughput capabilities of this model, in which a single tank of adult zebrafish can produce hundreds of embryos per week. Furthermore, chemical biology screens and interventions using intact, living animals are uniquely accessible to researchers, as zebrafish larvae and embryos are permeable to diverse small molecules and fit within a single well of 96-well and 384-well plates (Patton et al., 2021). Open in a separate windowFig. 1.Zebrafish larva infected with fluorescent Mycobacterium abscessus expressing TdTomato, shown in red. Image courtesy of Matt Johansen (Johansen et al., 2021).Box 1. DMM highlights zebrafish advancing knowledge in infectious diseaseRecent publications in DMM show the potential of the zebrafish model system to provide new or fuller insights into infectious diseases and therapies. One question being addressed is how basic cell-autonomous immune processes function in the context of a full organism. Various Reviews have highlighted what we have learned about the role of pyroptosis in host defence against bacterial infections (Brokatzky and Mostowy, 2022), as well as advances in understanding the diverse roles that macrophages and neutrophils play during the initial response to a variety of infectious and inflammatory stimuli (Rosowski, 2020). Zebrafish can also provide models for parasitic diseases that are relatively neglected, and we were pleased to publish a zebrafish model that provides insight into Toxoplasma pathogenesis, particularly the in vivo interactions of Toxoplasma with macrophages (Yoshida et al., 2020). Research dissecting the role of host immune cells in pathogen responses can be further potentiated by new tools, such as those developed by the Lieschke laboratory using macrophage and neutrophil-specific Cas9 driver lines to allow cell-specific genetic perturbation (Isiaku et al., 2021).Non-tuberculous mycobacteria causing pulmonary disease are a growing threat worldwide, with an antibiotic resistance profile that makes them very difficult to treat (Stout et al., 2016; Vinnard et al., 2016). An exploration of phage therapy for non-tuberculous mycobacteria in the zebrafish provides new insights into exciting clinical work that bookends this publication (Johansen et al., 2021). Engineered bacteriophages targeting specific strains of Mycobacterium abscessus have now been used clinically in cases of advanced lung disease (Dedrick et al., 2019; Nick et al., 2022). In other work, Habjan et al. employed the zebrafish mycobacterial infection model as an early screening step for anti-tuberculosis hits from in vitro screens that might have the best chance for in vivo translation. Following up on a screen for novel in vitro activity against Mycobacterium tuberculosis that identified ∼240 compounds, they identified 14 compounds with good in vivo activity. Impressively, they went on to identify the target of the strongest in vivo hit as being a mycobacterial aspartyl-tRNA synthase through screening for resistant mutants in both Mycobacterium marinum and Mycobacterium tuberculosis (Habjan et al., 2021).Drug screens, like those discussed above, are possible due to the permeability of the zebrafish to small molecules, which also allows creative ways to control the induction of host cytokines. DMM published an approach that enables drug-inducible, tissue-specific, titratable expression of different cytokines (Ibrahim et al., 2020). Harnessing this permeability in zebrafish can also enable detailed exploration of the effects of drugs, such as broadly used glucocorticoids, on specific innate immune cell types (Xie et al., 2019).The zebrafish has also been used as a model to understand infectious disease therapies targeting the pathogen directly. A recent paper describes the in vivo efficacy of nanoparticle-based delivery of lipophilic antibiotics, as well as use of the zebrafish to screen different formulations (Knudsen Dal et al., 2022). Finally, in the adult zebrafish sphere, a recent Review focused on how zebrafish can inform vaccine development strategies (Saralahti et al., 2020).These recent publications highlight some of the strengths of the zebrafish model for infectious disease research. DMM aims to be at the forefront in encouraging scientists and clinicians to leverage these insights for future therapies.Although efforts in zebrafish are often recognized and valued within the model organism community and beyond, it can sometimes be hard to break through to the world of clinical research. I vividly remember the excitement of being invited to present my work as a starting assistant professor at an early-career researcher lunch with a prominent visiting scientist, only to have my research and plans dismissed with some variation of “Well, why don''t you try to figure out what''s actually going on in people?”.Indeed, this is what many zebrafish researchers are ultimately trying to do by a different route. The goal of harnessing the knowledge we generate in models to impact human biology and therapies is an important part of the scientific enterprise. Many of us want and expect our findings to be relevant beyond the context of a model system. In my field, it has been exciting to see work in the zebrafish emerge that has led to the discovery of fundamentally conserved features of tuberculosis and host immunity – from zebrafish to humans – and has since translated to ongoing clinical trials.However, although we might hope that our work will be inherently understood and utilized in the clinical context, maximizing the potential of this research requires community advocates and communicators to help place the work in context. This can be achieved through ongoing dialogue among researchers, clinicians and patients to understand medical needs and perspectives. For example, DMM and The Company of Biologists have been long-time supporters of societies, such as the Zebrafish Disease Models Society, which focuses on the translational potential of zebrafish for understanding human disease and for developing new therapies, including some being investigated in clinical trials.Thus, it is useful to consider the following three broad themes when using model organisms in infectious disease research:

1. Conserved host–pathogen interactions in model systems. Although we all recognize, even at a strictly visual level, the many differences between the biology of a model organism and human biology, there is fundamentally conserved biology to be explored. Immune signalling pathways and underlying principles, as well as molecular and cellular details, first discovered and dissected in worms, flies, fish, mice and other model organisms, have translated remarkably well to human biology in many cases.
2. Model diversity. Divergent biology – in addition to being fascinating and important for the sake of knowledge itself – also leads to vital new insights and therapeutic approaches. As just one example, bacteriophages were instrumental in the discovery of fundamental aspects of gene regulation, have been used to facilitate genetic manipulation of seemingly genetically intractable pathogens, and are now being engineered and deployed therapeutically. And the study of bacterial–bacteriophage interactions of course led to all the advances made possible by CRISPR. These and many other examples from models that diverge from humans all support open-mindedness in science and emphasize the strength of laboratories taking diverse approaches and using diverse models. Pressing questions and opportunities in this realm are many, including investigation of how some non-human immune systems – those of bats, as just one example – permit asymptomatic tolerance of viruses that may be pathogenic in humans (Hayman, 2019). Which animal species restrict human pathogens via immune mechanisms that might eventually be harnessed therapeutically? Some of these topics will be prominent in a 2023 meeting organized by DMM entitled ‘Infectious Diseases Through an Evolutionary Lens’, which will take place in London at the British Medical Association House (Fig. 2).Open in a separate windowFig. 2.DMM''s 2023 meeting is entitled ‘Infectious Diseases Through an Evolutionary Lens’ and will take place in London at the British Medical Association House. Register your interest here: https://www.biologists.com/infectious-diseases-through-an-evolutionary-lens-contact-form/.
3. Engineering preclinical and predictive models of infectious disease. With advances in gene editing and the ability to make specific base edits, it is possible to precisely model human variants in an in vivo context during infection. Organisms like the zebrafish can provide useful models to delve into the specific consequences of these variants. Orthogonal approaches include mammalian animal models and advanced human cell models (Leist et al., 2020; van der Vaart et al., 2021). Discussion between scientists doing this preclinical work and clinical collaborators will be needed to determine to what degree the model recapitulates human disease and how these models can be used to advance new therapies. Recently, we have seen some of the landscape for clinical trials change, and in public health emergencies, collaborations would ideally accelerate the time from discovery to clinic. Again, this will require dialogue with and buy-in from clinical researchers to put together rigorous clinical trials.

DMM seeks to create and contribute to the ongoing conversations among and between basic scientists, clinical researchers and clinicians, with insights and criticisms from each of these domains. By highlighting rigorous, high-quality science in these areas, we hope to contribute to improved understanding of infectious diseases and new approaches to treatment.