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Yolk sac (YS) CSF1 receptor positive (CSF1R + ) cells are thought to be the progenitors for tissue-resident macrophages present in various tissues. The YS progenitors for tissue-resident macrophages are referred to as erythroid-myeloid progenitors (EMPs). However, diverse types of hematopoietic progenitors are present in the early YS, thus it is not precisely known which type of hematopoietic cell gives rise to the CSF1R + lineage. In this study, an analysis was conducted to determine when CSF1R + progenitors appeared in the early YS. It showed that CSF1R + cells appeared in the YS as early as embryonic day 9 (E9) and that the earliest hematopoietic progenitors that differentiate into CSF1R + cells were found in E8. Since these progenitors possessed the capability to generate primitive erythroid cells, it was likely that primitive erythroid lineages shared progenitors with the CSF1R + lineage. Mutual antagonism appears to work between PU.1 and GATA1 when CSF1R + cells appear in the early YS. One day later (E9), multiple progenitors, including myeloid-restricted progenitors and multipotent progenitors, in the YS could immediately generate CSF1R + cells. These results suggest that EMPs are not an exclusive source for the CSF1R + lineage; rather, multiple hematopoietic cell populations give rise to CSF1R + lineage in the early YS., (© 2022. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Ethnopharmacological Relevance: Liang-Ge-San (LGS), a traditional Chinese medicine (TCM) formula, is usually used in acute inflammatory diseases in China., Aim of the Study: This study aims to detect the optimal combination of anti-inflammatory components from LGS., Materials and Methods: Four mainly representative components (phillyrin, emodin, baicalin, and liquiritin) from LGS were chosen. The optimal combination was investigated by orthogonal design study. Zebrafish inflammation model was established by lipopolysaccharide (LPS)-yolk microinjection, and then the anti-inflammatory activities of different combinations were determined by survival analysis, changes on inflammatory cells infiltration, the MyD88/NF-κB and MAPK pathways and inflammatory cytokines production., Results: The different combinations of bioactive ingredients from LGS significantly protected zebrafish from LPS-induced inflammation, as evidenced by decreased recruitment of macrophages and neutrophils, inhibition of the MyD88/NF-κB and MAPK pathways and down-regulation of TNF-α and IL-6. Among them, the combination group 8 most significantly protected against LPS. The combination of group 8 is: 0.1 μM of emodin, 2 μM of baicalin, 20 μM of phillyrin and 12.5 μM of liquiritin., Conclusion: The optimized combination group 8 exerts the most significant anti-inflammatory activity by inhibiting the recruitment of inflammatory cells, activation of the MyD88/NF-κB and MAPK pathways and the secretion of pro-inflammatory cytokines. This present study provides pharmacological evidences for the further development of new modern Chinese drug from LGS to treat acute inflammatory diseases, but indicated the use of zebrafish in the screening of components from formulas., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Epidermal resident γδ T cells, or dendritic epidermal T cells (DETCs) in mice, are a unique and conserved population of γδ T cells enriched in the epidermis, where they serve as the regulators of immune responses and sense skin injury. Despite the great advances in the understanding of the development, homeostasis, and function of DETCs in the past decades, the origin and the underlying molecular mechanisms remain elusive. Here, we reviewed the recent research progress on DETCs, including their origin and homeostasis in the skin, especially at transcriptional and epigenetic levels, and discuss the involvement of DETCs in skin diseases.

Tissue-resident macrophages are a diverse population of cells that perform specialized functions including sustaining tissue homeostasis and tissue surveillance. Here, we report an interstitial subset of CD169 + lung-resident macrophages that are transcriptionally and developmentally distinct from alveolar macrophages (AMs). They are primarily localized around the airways and are found in close proximity to the sympathetic nerves in the bronchovascular bundle. These nerve- and airway-associated macrophages (NAMs) are tissue resident, yolk sac derived, self-renewing, and do not require CCR2 + monocytes for development or maintenance. Unlike AMs, the development of NAMs requires CSF1 but not GM-CSF. Bulk population and single-cell transcriptome analysis indicated that NAMs are distinct from other lung-resident macrophage subsets and highly express immunoregulatory genes under steady-state and inflammatory conditions. NAMs proliferated robustly after influenza infection and activation with the TLR3 ligand poly(I:C), and in their absence, the inflammatory response was augmented, resulting in excessive production of inflammatory cytokines and innate immune cell infiltration. Overall, our study provides insights into a distinct subset of airway-associated pulmonary macrophages that function to maintain immune and tissue homeostasis., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

The chicken yolk sac (YS) plays an important role in nutrient absorption and immune function for the developing embryo. The avian β-defensins (AvBD) are cationic peptides that are important members of the innate immune system. The objective of this study was to profile AvBD mRNA expression patterns and distribution of cells expressing AvBD mRNA in the chicken YS. Expression of AvBD1, 2, 7, and 10 mRNA was low at embryonic day 7 (e7), increased to e9 through e13 and then declined to e19. Using in situ hybridization, AvBD10 mRNA was found to be expressed in endodermal epithelial cells, while AvBD1, 2, and 7 mRNA were expressed in heterophils. The developmental expression pattern and distribution of AvBD mRNA in the YS reveals the importance of these genes to protection of the developing chick embryo., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Distinct macrophage subsets populate the developing embryo and fetus in distinct waves. However little is known about the functional differences between in utero macrophage populations or how they might contribute to fetal and neonatal immunity. Here we tested the innate immune response of mouse macrophages derived from the embryonic yolk sac and from fetal liver. When isolated from liver or lung, CD11b HI fetal liver derived macrophages responded to the TLR4 agonist LPS by expressing and releasing inflammatory cytokines. However F4/80 HI macrophages from the yolk sac did not respond to LPS treatment. While differences in TLR4 expression did not appear to explain these data, F4/80 HI macrophages had much lower NLRP3 inflammasome expression compared to CD11b HI macrophages. Gene expression profiling also demonstrated LPS-induced expression of inflammatory genes in CD11b HI macrophages, but not in F4/80 HI cells. Genes expressed in LPS-treated CD11b HI macrophages were more likely to contain predicted NF-κB binding sites in their promoter regions. Our data show that CD11b HI macrophages derived from fetal liver are the major pro-inflammatory cells in the developing fetus. These findings could have important implications in better understanding the fetal inflammatory response and the unique features of neonatal immunity.

The murine epidermis harbors two immune cell lineages, Langerhans cells (LCs) and γδ T cells known as dendritic epidermal T cells (DETCs). LCs develop from both early yolk sac (YS) progenitors and fetal liver monocytes before locally self-renewing in the adult. For DETCs, the mechanisms of homeostatic maintenance and their hematopoietic origin are largely unknown. Here, we exploited multicolor fate mapping systems to reveal that DETCs slowly turn over at steady state. Like for LCs, homeostatic maintenance of DETCs is achieved by clonal expansion of tissue-resident cells assembled in proliferative units. The same mechanism, albeit accelerated, facilitates DETC replenishment upon injury. Hematopoietic lineage tracing uncovered that DETCs are established independently of definitive hematopoietic stem cells and instead originate from YS hematopoiesis, again reminiscent of LCs. DETCs thus resemble LCs concerning their maintenance, replenishment mechanisms, and hematopoietic development, suggesting that the epidermal microenvironment exerts a lineage-independent influence on the initial seeding and homeostatic maintenance of its resident immune cells., (© 2018 Gentek et al.)

Monocytes and macrophages are critical effectors and regulators of innate immune response. They not only play crucial and distinctive roles in homeostasis, but also contribute to some pathologic processes. The heterogeneity of the macrophage lineage has been widely recognized and, in part, is a result of the specialization of resident macrophages in particular tissue microenvironments. Monocytes are usually known to originate in the bone marrow from a common myeloid progenitor that is shared with neutrophils, and they are then released into the peripheral blood. However, the origin of tissue-resident macrophages, crucial for homeostasis and immunity, has remained controversial until recently. During embryonic organogenesis, macrophages derived from yolk sac and fetal liver precursors are seeded throughout tissues, persisting in the adulthood as resident, self-maintaining populations. After birth, bone marrow-derived monocytes can replenish tissue resident macrophages following injury, infection and inflammation. In this review, we will mainly summarize our current understanding on the origin, ontogeny and fates of tissue macrophages and will briefly discuss the molecular regulation of resident macrophage homeostasis in physiological situation., (© 2018 Wiley Periodicals, Inc.)

Omphalitis or yolk sac infection (YSI) and colibacillosis are the most common infectious diseases that lead to high rates of early chick mortalities (ECMs) in young chicks. Out of numerous microbial causes, avian pathogenic Escherichia coli (APEC) or extraintestinal pathogenic E. coli infections are considered the most common cause of these conditions. YSI causes deterioration and decomposition of yolk, leading to deficiency of necessary nutrients and maternal antibodies, retarded growth, poor carcass quality, and increased susceptibility to other infections, including omphalitis, colibacillosis, and respiratory tract infection. Presently, in ovo injection of antibiotics, heavy culling, or after hatch use of antibiotics is practiced to manage ECM. However, increased antibiotic resistance and emergence of "super bugs" associated with use or misuse of antibiotics in the animal industry have raised serious concerns. These concerns urgently require a focus on host-driven nonantibiotic approaches for stimulation of protective antimicrobial immunity. Using an experimental YSI model in newborn chicks, we evaluated the prophylactic potential of three in ovo-administered innate immune stimulants and immune adjuvants for protection from ECM due to YSI. Our data have shown >80%, 65%, and 60% survival with in ovo use of cytosine-phosphodiester-guanine (CpG) oligodeoxynucleotides (ODN), polyinosinic:polycytidylic acid, and polyphosphazene, respectively. In conclusion, data from these studies suggest that in ovo administration of CpG ODN may serve as a potential candidate for replacement of antibiotics for the prevention and control of ECM due to YSI in young chicks.

Preterm birth is a major risk factor for adverse neurological outcomes in ex-preterm children, including motor, cognitive, and behavioral disabilities. N-acetyl-L-cysteine therapy has been used in clinical studies; however, it requires doses that cause significant side effects. In this study, we explore the effect of low dose N-acetyl-L-cysteine therapy, delivered using a targeted, systemic, maternal, dendrimer nanoparticle (DNAC), in a mouse model of intrauterine inflammation. Our results demonstrated that intraperitoneal maternal DNAC administration significantly reduced the preterm birth rate and altered placental immune profile with decreased CD8 + T-cell infiltration. Furthermore, we demonstrated that DNAC improved neurobehavioral outcomes and reduced fetal neuroinflammation and long-term microglial activation in offspring. Our study is the first to provide evidence for the role of CD8 + T-cell in the maternal-fetal interface during inflammation and further support the efficacy of DNAC in preventing preterm birth and prematurity-related outcomes.

Macrophages are key components of the innate immune system and play pivotal roles in immune response, organ development, and tissue homeostasis. Studies in mice and zebrafish have shown that tissue-resident macrophages derived from different hematopoietic origins manifest distinct developmental kinetics and colonization potential, yet the genetic programs controlling the development of macrophages of different origins remain incompletely defined. In this study, we use zebrafish, where tissue-resident macrophages arise from the rostral blood island (RBI) and ventral wall of dorsal aorta (VDA), the zebrafish hematopoietic tissue equivalents to the mouse yolk sac and aorta-gonad-mesonephros for myelopoiesis, to address this issue. We show that RBI- and VDA-born macrophages are orchestrated by distinctive regulatory networks formed by the E-twenty-six (Ets) transcription factors Pu.1 and Spi-b, the zebrafish ortholog of mouse spleen focus forming virus proviral integration oncogene B (SPI-B), and the helix-turn-helix DNA-binding domain containing protein Irf8. Epistatic studies document that during RBI macrophage development, Pu.1 acts upstream of Spi-b, which, upon induction by Pu.1, partially compensates the function of Pu.1. In contrast, Pu.1 and Spi-b act in parallel and cooperatively to regulate the development of VDA-derived macrophages. Interestingly, these two distinct regulatory networks orchestrate the RBI- and VDA-born macrophage development largely by regulating a common downstream gene, Irf8. Our study indicates that macrophages derived from different origins are governed by distinct genetic networks formed by the same repertoire of myeloid-specific transcription factors., (© 2017 by The American Society of Hematology.)

Tissue-resident macrophages can derive from yolk sac macrophages (YS-Macs), fetal liver monocytes (FL-MOs), or adult bone-marrow monocytes (BM-MOs). The relative capacity of these precursors to colonize a niche, self-maintain, and perform tissue-specific functions is unknown. We simultaneously transferred traceable YS-Macs, FL-MOs, and BM-MOs into the empty alveolar macrophage (AM) niche of neonatal Csf2rb(-/-) mice. All subsets produced AMs, but in competition preferential outgrowth of FL-MOs was observed, correlating with their superior granulocyte macrophage-colony stimulating factor (GM-CSF) reactivity and proliferation capacity. When transferred separately, however, all precursors efficiently colonized the alveolar niche and generated AMs that were transcriptionally almost identical, self-maintained, and durably prevented alveolar proteinosis. Mature liver, peritoneal, or colon macrophages could not efficiently colonize the empty AM niche, whereas mature AMs could. Thus, precursor origin does not affect the development of functional self-maintaining tissue-resident macrophages and the plasticity of the mononuclear phagocyte system is largest at the precursor stage., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Although classified as hematopoietic cells, tissue-resident macrophages (MFs) arise from embryonic precursors that seed the tissues prior to birth to generate a self-renewing population, which is maintained independently of adult hematopoiesis. Here we reveal the identity of these embryonic precursors using an in utero MF-depletion strategy and fate-mapping of yolk sac (YS) and fetal liver (FL) hematopoiesis. We show that YS MFs are the main precursors of microglia, while most other MFs derive from fetal monocytes (MOs). Both YS MFs and fetal MOs arise from erythro-myeloid progenitors (EMPs) generated in the YS. In the YS, EMPs gave rise to MFs without monocytic intermediates, while EMP seeding the FL upon the establishment of blood circulation acquired c-Myb expression and gave rise to fetal MOs that then seeded embryonic tissues and differentiated into MFs. Thus, adult tissue-resident MFs established from hematopoietic stem cell-independent embryonic precursors arise from two distinct developmental programs., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: Embryo resorption is a major problem in human medicine, agricultural animal production and in conservation breeding programs. Underlying mechanisms have been investigated in the well characterised mouse model. However, post mortem studies are limited by the rapid disintegration of embryonic structures. A method to reliably identify embryo resorption in alive animals has not been established yet. In our study we aim to detect embryos undergoing resorption in vivo at the earliest possible stage by ultra-high frequency ultrasound., Methods: In a longitudinal study, we monitored 30 pregnancies of wild type C57BI/6 mice using ultra-high frequency ultrasound (30-70 MHz), so called ultrasound biomicroscopy (UBM). We compared the sonoembryology of mouse conceptuses under spontaneous resorption and neighbouring healthy conceptuses and correlated the live ultrasound data with the respective histology., Results: The process of embryo resorption comprised of four stages: first, the conceptus exhibited growth retardation, second, bradycardia and pericardial edema were observed, third, further development ceased and the embryo died, and finally embryo remnants were resorbed by maternal immune cells. In early gestation (day 7 and 8), growth retardation was characterized by a small embryonic cavity. The embryo and its membranes were ill defined or did not develop at all. The echodensity of the embryonic fluid increased and within one to two days, the embryo and its cavity disappeared and was transformed into echodense tissue surrounded by fluid filled caverns. In corresponding histologic preparations, fibrinoid material interspersed with maternal granulocytes and lacunae filled with maternal blood were observed. In later stages (day 9-11) resorption prone embryos were one day behind in their development compared to their normal siblings. The space between Reichert's membrane and inner yolk sac membrane was enlarged The growth retarded embryos exhibited bradycardia and ultimately cessation of heart beat. Corresponding histology showed apoptotic cells in the embryo while the placenta was still intact. In the subsequent resorption process first the embryo and then its membranes disappeared., Conclusions: Our results provide a temporal time course of embryo resorption. With this method, animals exhibiting embryo resorption can be targeted, enabling the investigation of underlying mechanisms before the onset of total embryo disintegration.

The extra-embryonic yolk sac (YS) is the first hematopoietic site in the mouse embryo and is thought to generate only primitive erythroid and myeloerythroid progenitor cells before definitive HSC emergence within the embryo on E10.5. Here, we have shown the existence of T cell-restricted progenitors in the E9.5 YS that directly engraft in recipient immunodeficient mice. T-cell progenitors were also produced in vitro from both YS and para-aortic splanchnopleura hemogenic endothelial cells, and these T-cell progenitors repopulated the thymus and differentiated into mature T-cell subsets in vivo on transplantation. Our data confirm that the YS produces T-lineage-restricted progenitors that are available to colonize the thymus and provide new insight into the YS as a definitive hematopoietic site in the mouse embryo.

During the first week of the posthatching period, before the immune system is mature enough to produce its own B lymphocytes, a chick's humoral immunity depends on maternal antibodies (IgY) received from the egg yolk. During incubation and after hatching, the yolk sac (YS) membrane transfers nutrients (including IgY) from the egg yolk to the developing embryo or newly hatched chick. The objective of this study was to determine the effects of breeder flock age on the total IgY content of egg yolks and chick YS from a commercial broiler breeder strain. Hatching eggs from the same broiler breeder flock were collected at 32, 40, and 55 wk of age. One group of eggs per flock age was used to determine the egg yolk total IgY content. Another group of eggs was incubated for 21.5 d, and upon hatching, the YS of newly hatched chicks were collected to determine the total IgY content. Egg and egg yolk weight increased with flock age, but YS weights did not reflect egg yolk weight. The total IgY content per gram of egg yolk increased with flock age; this fact plus the observed yolk weight increase with flock age notably increased the total IgY contained in yolks of eggs laid by 55-wk-old breeders. However, chicks hatching from 55-wk-old breeders had less IgY per gram of YS than chicks from 32- and 40-wk-old breeders. Whether there are differences in the rates of YS absorption between chicks of different breeder ages is unknown. This research provided total IgY values for broiler breeder egg yolk and chick YS of a commonly used meat-type chicken strain. Differences in egg yolk and YS total IgY contents due to flock age in this type of bird had not been previously reported. Research on the physiological consequences of YS absorption rates in chicks from different breeder ages is advised.

Fc receptors transport maternal antibodies across epithelial cell barriers to passively immunize newborns. FcRY, the functional counterpart of mammalian FcRn (a major histocompatibility complex homolog), transfers IgY across the avian yolk sac, and represents a new class of Fc receptor related to the mammalian mannose receptor family. FcRY and FcRn bind immunoglobulins at pH ≤6.5, but not pH ≥7, allowing receptor-ligand association inside intracellular vesicles and release at the pH of blood. We obtained structures of monomeric and dimeric FcRY and an FcRY-IgY complex and explored FcRY's pH-dependent binding mechanism using electron cryomicroscopy (cryoEM) and small-angle X-ray scattering. The cryoEM structure of FcRY at pH 6 revealed a compact double-ring "head," in which the N-terminal cysteine-rich and fibronectin II domains were folded back to contact C-type lectin-like domains 1-6, and a "tail" comprising C-type lectin-like domains 7-8. Conformational changes at pH 8 created a more elongated structure that cannot bind IgY. CryoEM reconstruction of FcRY dimers at pH 6 and small-angle X-ray scattering analysis at both pH values confirmed both structures. The cryoEM structure of the FcRY-IgY revealed symmetric binding of two FcRY heads to the dimeric FcY, each head contacting the C(H)4 domain of one FcY chain. FcRY shares structural properties with mannose receptor family members, including a head and tail domain organization, multimerization that may regulate ligand binding, and pH-dependent conformational changes. Our results facilitate understanding of immune recognition by the structurally related mannose receptor family and comparison of diverse methods of Ig transport across evolution.

Langerhans cells (LCs) are myeloid cells of the epidermis, featured in immunology textbooks as bone marrow-derived antigen-presenting dendritic cells (DCs). A new picture of LC origin, homeostasis and function has emerged, however, after genetic labelling and conditional cell ablation models in mice. LC precursors are recruited into the fetal epidermis, where they differentiate and proliferate in situ. In adults, LCs proliferate at steady state, and during inflammation, in response to signals from neighbouring cells. Here we review the experimental evidence that support either extra-embryonic yolk sac (YS) macrophages or hematopoietic stem cells (HSCs) as the origin of LCs. Beyond LC biology, we propose that YS and HSCs can contribute to the development of distinct subsets of macrophages and DCs., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

In Egypt, efforts to control highly pathogenic H5N1 avian influenza virus in poultry and in humans have failed despite increased biosecurity, quarantine, and vaccination at poultry farms. The ongoing circulation of HP H5N1 avian influenza in Egypt has caused >100 human infections and remains an unresolved threat to veterinary and public health. Here, we describe that the failure of commercially available H5 poultry vaccines in Egypt may be caused in part by the passive transfer of maternal H5N1 antibodies to chicks, inhibiting their immune response to vaccination. We propose that the induction of a protective immune response to H5N1 is suppressed for an extended period in young chickens. This issue, among others, must be resolved and additional steps must be taken before the outbreaks in Egypt can be controlled.

Prolonged interference or suppression of maternal antibodies of the humoral immune response of newly hatched chicks to active immunization has been documented; however, the immunological mechanisms responsible for such suppression are still unclear. Laying hens were immunized with dinitrophenyl-keyhole limpet hemocyanin (DNP-KLH). Purified maternal anti-DNP or non-specific IgY antibodies were transferred by yolk sac inoculation to newly hatched chicks, and they were immunized with DNP-KLH or rabbit serum albumen (RSA) at 1 and 4 weeks of age. The concentrations of anti-DNP and anti-RSA antibodies in serum samples of these chicks were measured using an enzyme-linked immunosorbent assay (ELISA). The immune responses of the chicks that received a high dose of maternal anti-DNP antibodies and were immunized with an appropriate dose of DNP-KLH were suppressed. However, those of the chicks that received the same high dose of maternal non-specific IgY antibodies and were immunized with an appropriate dose of DNP-KLH and those of the chicks that received a high dose of maternal anti-DNP antibodies and were immunized with RSA were not suppressed. On the other hand, suppression of anti-DNP antibody production would not be induced if the chicks received a high dose of antigen specific maternal antibodies and were immunized with a high dose of the same antigen. These results revealed that the immune suppressive effect of maternal antibodies on the immune response of the newly hatched chicks was antigen specific and depended mainly on the ratio of antigen/maternal antibody at the time of immunization.

In adults, the nonclassical MHC class I molecule, FcRn, binds both IgG and albumin and rescues both from a degradative fate, endowing both proteins with high plasma concentrations. FcRn also transports IgG from mother to young during gestation. Anticipating that a detailed understanding of gestational IgG transport in the mouse may give us a useful model to understand FcRn function in the human placenta, we have studied FcRn in the mouse yolk sac placenta in detail. Analyzing day 19-20 fetuses of the three FcRn genotypes resulting from matings of FcRn(+/-) parents, we found that FcRn(-/-) fetuses showed negligible IgG concentrations (1.5 microg/ml), whereas IgG concentrations in FcRn(+/-) fetuses were about a half (176 microg/ml) that of FcRn(+/+) fetuses (336 microg/ml), indicating that FcRn is responsible for virtually all IgG transport from mother to fetus. Immunofluorescence and immunoblotting studies indicated that FcRn is expressed in the endoderm of the yolk sac placenta but not in other cells of the yolk sac placenta or in the chorioallantoic placenta. IgG was found in the endoderm of both FcRn(+/+) and FcRn(-/-) yolk sac placentas and in the mesenchyme of FcRn(+/+) but was missing from the mesenchyme of FcRn(-/-) yolk sac placentas, indicating that IgG enters the endoderm constitutively but is moved out of the endoderm by FcRn. The similarities of these results to human placental FcRn expression and function are striking.

We report that angiotensin-converting enzyme (ACE), a critical physiologic regulator of blood pressure, angiogenesis, and inflammation, is a novel marker for identifying hemangioblasts differentiating from human embryonic stem cells (hESC). We demonstrate that ACE+CD45-CD34+/- hemangioblasts are common yolk sac (YS)-like progenitors for not only endothelium but also both primitive and definitive human lymphohematopoietic cells. Thrombopoietin and basic fibroblast growth factor are identified as critical factors for the proliferation of human hemangioblasts. The developmental sequence of human embryoid body hematopoiesis is remarkably congruent to the timeline of normal human YS development, which occurs during weeks 2 to 6 of human gestation. Furthermore, ACE and the renin-angiotensin system (RAS) directly regulate hemangioblast expansion and differentiation via signaling through the angiotensin II receptors AGTR1 and AGTR2. ACE enzymatic activity is required for hemangioblast expansion, and differentiation toward either endothelium or multipotent hematopoietic progenitors is dramatically augmented after manipulation of angiotensin II signaling with either AGTR1- or AGTR2-specific inhibitors. The RAS can therefore be exploited to direct the hematopoietic or endothelial fate of hESC-derived hemangioblasts, thus providing novel opportunities for human tissue engineering. Moreover, the initial events of human hematoendotheliogenesis can be delineated in a manner previously impossible because of inaccessibility to early human embryonic tissues.

Passerine birds have played an important role in the establishment, maintenance, and spread of West Nile virus (WNV) in North America, and some are susceptible to WNV-associated mortality. Characterization of passive transfer of anti-WNV antibodies in passerines is important to understanding transmission and demographic effects of WNV on wild birds. We showed passively acquired maternal antibodies to WNV in the house sparrow (Passer domesticus). Although all seropositive females (N = 18) produced antibody-positive egg yolks, only 20% of seropositive mothers (3/15) produced seropositive chicks. The estimated average half-life of maternal antibodies in chick sera was 3 days, and no antibodies were detected after 9 days post-hatch (DPH). Maternal antibodies failed to provide protection against viremia in chicks at 21-25 DPH. Although the observed duration of persistence of passively inherited anti-WNV antibodies in house sparrows differs from some non-passerine birds, it remains unknown whether similar patterns occur in other passerines.

In mammals the transfer of passive immunity from mother to young is mediated by the MHC-related receptor FcRn, which transports maternal IgG across epithelial cell barriers. In birds, maternal IgY in egg yolk is transferred across the yolk sac to passively immunize chicks during gestation and early independent life. The chicken yolk sac IgY receptor (FcRY) is the ortholog of the mammalian phospholipase A2 receptor, a mannose receptor family member, rather than an FcRn or MHC homolog. FcRn and FcRY both exhibit ligand binding at the acidic pH of endosomes and ligand release at the slightly basic pH of blood. Here we show that FcRY expressed in polarized mammalian epithelial cells functioned in endocytosis, bidirectional transcytosis, and recycling of chicken FcY/IgY. Confocal immunofluorescence studies demonstrated that IgY binding and endocytosis occurred at acidic but not basic pH, mimicking pH-dependent uptake of IgG by FcRn. Colocalization studies showed FcRY-mediated internalization via clathrin-coated pits and transport involving early and recycling endosomes. Disruption of microtubules partially inhibited apical-to-basolateral and basolateral-to-apical transcytosis, but not recycling, suggesting the use of different trafficking machinery. Our results represent the first cell biological evidence of functional equivalence between FcRY and FcRn and provide an intriguing example of how evolution can give rise to systems in which similar biological requirements in different species are satisfied utilizing distinct protein folds.

Antinuclear antibodies, ANA, were usually detected with antigen of somatic cell nucleus. It has not been reported to detect ANA with egg cell nucleus as antigen. Enzyme linked immuosorbent assay, ELISA, coated with yolk was developed to detect ANA in our laboratory. A quality control test, cross absorption test, and cross antibody-induced test with yolk were performed. Results showed a good agreement between our method and IFA through measurement of the same samples from patients suspected of having rheumatic connective tissue diseases (Kappa=0.668, P=0.000). The results were not influenced by the RF and different sources of egg. CVs of inter-assay, were less than 10%. The cross absorption test was negative, as well; the ANA to somatic cell nucleus could be induced with egg cell nucleus. It is implied that there were both cross as well as overlapped Egg-ANA and Somatic-ANA. As egg nucleus, its volume was large, its purification was simple, so the better method might be established.

The Atlantic sharpnose shark, Rhizoprionodon terraenovae, is viviparous species that forms a yolk sac placenta to facilitate exchange between mother and embryo. However, very little is known about the immunological aspects of this organ in sharks. To begin to understand this, we used histology, histochemistry and immunohistochemistry to investigate the sharpnose shark placenta throughout gestation. We report the presence of lymphoid aggregates in the maternal portion of the placenta during all stages of gestation, and their increasing size and vascularity near term. Immunoglobulin is found in the maternal tissues of the placenta, but its presence in embryonic tissue and potential transfer from maternal circulation remains unclear. Placental cells resembling mammalian uterine NK cells and melanomacrophages of lower vertebrates are described for the first time. Similarities with mammalian placentae point to shared aspects in the co-evolution of reproductive and immune systems, even between two phylogenetically diverse groups in which placentation arose by convergent evolution.

ACTH and ACTH receptor-like molecules were found at the examined stages of development (2, 4, 8, 12, 18, and 24 days post-hatching) in yolk sac, pronephros tubules, interrenal tissue, thymus, liver, spleen, cardinal veins, and skin of the teleost fish Dicentrarchus labrax. ACTH and the related receptor-like molecules show a similar distribution. LPS treatment at two different stages (8 and 24 days post-hatching) provoked both a release and an induction of ACTH-like molecules, suggesting an important role of this peptide to control the modifications in body homeostasis during the first period of the sea bass' life, i.e., 30 days post-hatching, before the lymphoid cells have reached complete maturation.

In the midgestation murine embryo, several major vascular tissues contain hematopoietic stem cell (HSC) activity. These include the aorta-gonad-mesonephros region (AGM), yolk sac, and fetal liver. Recently, the placenta was demonstrated to harbor hematopoietic progenitors, but it was not examined for HSC activity. We demonstrate here that the placenta also harbors adult-repopulating HSCs. Placental HSCs begin to be detected at embryonic day (E) 11, and HSC numbers increase dramatically between E11 and E12, exceeding the numbers in the circulating embryonic blood. Furthermore, all placental HSC activity is restricted to the GFP+ fraction of cells in Ly-6A (Sca-1) GFP transgenic embryos. Cells coexpressing GFP and endothelial markers CD34 and CD31 are found in the embryonic vasculature of the placental labyrinth. Moreover, placental cell expression of other HSC markers and transcription factors suggests that HSC emergence may occur in the placenta, as has been proposed for other embryonic hematopoietic sites.

In mammals, IgG is transferred from mother to young by the MHC-related receptor FcRn, which binds IgG in acidic endosomes and releases it at basic pH into blood. Maternal IgY, the avian counterpart of IgG, is transferred to embryos across yolk sac membranes. We affinity-purified the chicken yolk sac IgY receptor (FcRY) and sequenced its gene. FcRY is unrelated to MHC molecules but is a homolog of the mammalian phospholipase A(2) receptor. Analytical ultracentrifugation and truncation experiments suggest that FcRY forms a compact structure containing an IgY binding site at acidic pH but undergoes a conformational change at basic pH that disrupts the site. FcRY is thus unrelated to mammalian FcRn in both its structure and mechanism for pH-dependent binding, illustrating distinct routes utilized by evolution to transfer antibodies.

Problem: A variety of reproductive impairments have been reported in the context of the antiphospholipid syndrome (APS). APS is associated with the presence of antibodies to negatively charged phospholipids that may affect the outcome of pregnancy., Method of Study: Rat embryos were cultured within their yolk sacs. The effects of two antiphosphatidylserine monoclonal aPS antibodies (HL5B, RR7F) regarding their influence on growth and apoptotic events of the yolk sacs, as well as on growth and the morphology of the embryos, were studied., Results: Exposure of rat embryos and their yolk sacs to aPS inhibited yolk sac growth. Moreover, increased number of apoptotic events of giant cells in the aPS-exposed ectoplacental cone was found in comparison with control IgG-exposed giant cells (P < 0.05). No significant damage was observed in the embryos., Conclusions: The results suggest that aPS affect growth and apoptosis of rat ectoplacental cone.

Plasma cells play a pivotal role in the immune system and are responsible for the synthesis and release of immunoglobulins. Numerous in vitro culture experiments on the yolk sac demonstrated the generation of mature cells of the myeloid and lymphoid lineages under appropriate culture conditions. However, there are no reports describing the development of mature lymphoid cells in the yolk sac so far. For this reason, we undertook this study to investigate the development of antibody-containing plasma cells during early yolk sac haematopoiesis. Immunohistochemistry and immunoelectron microscopy were employed in the study. Results of this work demonstrated very weak immune staining for the intracytoplasmic IgA, IgG, and IgM at days 10 and 11 of embryonic life, while dark staining was obtained at 12 days. Positive staining was localized to the endodermal cell layer. Electron microscopic examinations revealed the existence of cells with the typical characteristics of plasma cells inside the endodermal cell layer, which may suggest their endodermal origin. To further verify the nature of these cells, intracytoplasmic immunoglobulins were demonstrated by immunoelectron microscopy. The present study demonstrated emergence of mature functioning plasma cells in early rat yolk sac. In a previous work we hypothesized the possibility of endodermal origin of yolk sac macrophages. This study adds additional evidence to support that hypothesis. The possible role of plasma cells in the yolk sac is discussed.

Renal cell carcinoma antigen is a rather specific marker for normal and neoplastic renal tissue. We investigated the expression of this antigen in 34 gonadal and extragonadal germ cell tumors, including 8 pure yolk sac carcinomas and 26 embryonal carcinomas, 15 of which were combined with teratomas, seminomas, and dysgerminomas. Renal cell carcinoma antigen was demonstrated in all 8 yolk sac tumors and 21 of 26 embryonal carcinomas (81%). In yolk sac tumors, renal cell carcinoma antigen reactivity was diffusely present throughout the tumors. In embryonal carcinomas, this marker was identified only in yolk sac components. Both intracytoplasmic and membranous staining patterns were present. No reactivity was noticed in embryonal carcinoma cells, seminoma, dysgerminoma, and other components of teratomas. The study suggests an antigenic similarity between renal tubules and yolk sac tumors. Furthermore, the renal cell carcinoma antigen may be used as an addition to the panel of immunocytochemical markers for yolk sac carcinomas.

B cell lymphogenesis in mammals occurs in various tissues during development but it is generally accepted that it operates by the same mechanism in all tissues. We show that in swine, the frequency of in-frame (IF) VDJ rearrangements differs among yolk sac, fetal liver, spleen, early thymus, bone marrow, and late thymus. All VDJ rearrangements recovered and analyzed on the 20th day of gestation (DG20) from the yolk sac were 100% IF. Those recovered at DG30 in the fetal liver were >90% IF, and this predominance of cells with apparently a single IF rearrangement continued in all organs until approximately DG45, which corresponds to the time when lymphopoiesis begins in the bone marrow. Thereafter, the proportion of IF rearrangements drops to approximately 71%, i.e., the value predicted whether VDJ rearrangement is random and both chromosomes were involved. Unlike other tissues, VDJs recovered from thymus after DG50 display a pattern suggesting no selection for IF rearrangements. Regardless of differences in the proportion of IF rearrangements, we observed no significant age- or tissue-dependent changes in CDR3 diversity, N region additions, or other characteristics of fetal VDJs during ontogeny. These findings indicate there are multiple sites of B cell lymphogenesis in fetal piglets and differences in the frequency of productive VDJ rearrangements at various sites. We propose the latter to result from differential selection or a developmentally dependent change in the intrinsic mechanism of VDJ rearrangement.

1. Early granulocytic response was evaluated in chick embryos inoculated with herpesvirus of turkeys (HVT). 2. Fifty 10-d-old specific pathogen-free embryos were divided into two groups, inoculated via yolk sac. Group 1 were inoculated with a complete dose of HVT and group 2 with vaccinediluent only. 3. Samples were taken for histological evaluation of yolk sac, liver, chorioallantoic membrane, brain and heart from 5 embryos per group on days 12, 14, 16, 18 and 20 of embryonic life. 4. Increases in numbers of granulocytes were detected on days 14 and 16 in the yolk sac, and on d 14 and 20 in the liver of in embryos, which received HVT. In addition, the chorioallantoic membrane was infiltrated with granulocytes. 5. The results confirm that granulopoiesis in the yolk sac is stimulated in the early stages of incubation if a viral antigen is present. The virus also appears to trigger the presence of granulocytes in embryonic liver and chorioallantoic membranes.

Although N-cadherin is necessary for organ formation originating in the endoderm, the expression of N-cadherin in gastric carcinoma and its role has not yet been reported. The present study was conducted to determine the pattern of immunohistochemical expression of E-cadherin and N-cadherin, using formalin-fixed, paraffin-embedded tissues from 97 primary gastric carcinomas, including 17 which were producing alpha-fetoprotein (AFP). Samples were subdivided into 50 tubular adenocarcinomas and 47 poorly differentiated adenocarcinomas. Results showed that E-cadherin was expressed in varying degrees in areas of cell adhesion between tumor cells, in 94 out of 97 cases studied. Three cases which showed no expression of E-cadherin were diagnosed as AFP-producing tumors by immunohistochemistry. Expression of N-cadherin was observed in varying degrees in the intercellular spaces between tumor cells in 11 tubular adenocarcinomas and in six poorly differentiated adenocarcinomas, including E-cadherin-negative cases, all of which were AFP positive. The present findings suggest a possible role for N-cadherin in gastric carcinoma.

At birth the tissues of marsupial immune system are underdeveloped. The young animal is not immunocompetent. Histological and immunohistochemical studies of pouch young epithelial tissues provide a clear picture of tissue development but the timing of onset of immunocompetence awaits definition. The survival of the neonatal marsupial in a microbially rich environment is dependent on maternal strategies, including immunoglobulin transfer via milk and, in some species, prenatally via the yolk sac placenta. It is also likely that pouch secretions play a role. This review summarizes our current knowledge of the pathway of immunological development in marsupials and the protection and threats afforded by the pouch environment.

Background: Heterologous antiserum to the visceral yolk sac (AVYS) is teratogenic, inducing a spectrum of malformations in vivo and producing similar effects in vitro. Numerous studies support the concept that AVYS-induced malformations result from embryonic nutritional deficiency, without affecting the maternal nutritional status. This has provided a useful model with which to investigate the nutritional requirements of the early embryo, as well as the role of various nutrients in the etiology of congenital defects., Methods: In the current investigation, we examined the effects of methionine and other nutrients on AVYS-induced embryotoxicity in vitro. For these experiments, we cultured rat embryos (9.5 p.c) for 48 hr with AVYS and/or methionine at several concentration levels., Results: The addition of L-methionine to AVYS-exposed cultures reduced dysmorphology and open neural tube; this effect was concentration dependent. AVYS-induced dysmorphology was completely prevented at a concentration of L-methionine corresponding to 50-fold the basal serum concentration. Utilization of D-methionine, L-leucine, or folic acid (5-methyltetrahydrofolate, MTHF) instead of L-methionine had no protective effects., Conclusions: These results suggest that, although AVYS limits the supply of all amino acids to the embryo, embryopathy largely results from a deficiency of methionine. Furthermore, although endocytosis and degradation of proteins by the VYS supplies most amino acids to the embryo, free amino acids may be compensatory when this source is reduced. These results support those of previous investigations that suggest methionine is required for normal NT closure and that methionine is a limiting nutrient for embryonic development., (Copyright 2000 Wiley-Liss, Inc.)

The antigen specificity of a rat monoclonal antibody TER-119 was investigated. In adult mice, TER-119 reacted with mature erythrocytes, 20-25% of bone marrow cells and 2-3% of spleen cells but not with thymocytes nor lymph node cells. In fetal haematopoietic tissues, 30-40% of d 10 yolk sac cells, 80-90% of d 14 fetal liver cells and 40-50% of newborn liver cells were reactive with TER-119. TER-119+ cells in adult bone marrow expressed significant levels of CD45 but not myeloid (Mac-1, Gr-1) or B-cell (B220) markers. Morphological examination and haematopoietic colony-forming assays for isolated TER-119+ cells revealed that TER-119 reacts with erythroid cells at differentiation stages from early proerythroblast to mature erythrocyte, but not with cells showing typical erythroid blast-forming unit (BFU-E) and erythroid colony-forming unit (CFU-E) activities. Erythroleukaemia cell lines do not express the TER-119 antigen even after stimulation with dimethylsulphoxide. TER-119 immunoprecipitated protein bands with molecular masses of 110 kDa, 60 kDa, 52 kDa and 32 kDa from erythrocyte membrane, whereas only a 52-kDa band was detected by TER-119 in Western blot analysis. Further molecular and cellular analyses indicated that the TER-119 antigen is a molecule associated with cell-surface glycophorin A but not with glycophorin A itself.

We describe a unique, stable pre-pro-B cell line (YS-PPB) derived from AA4.1+ yolk sac cells from day 10 mouse embryos. This cell line, discovered fortuitously during the course of studies of in vitro B cell differentiation, is independent of IL-7 supplementation for long term expansion in vitro. YS-PPB cells as well as clonal sublines expressed AA4.1, CD43, B220, Sca-1, CD19, heat stable antigen, MHC class I, IL-7R, and FcyR, but did not express cytoplasmic mu-chain, surface IgM (sIgM), or MHC class II molecules. PCR analysis showed that the cells expressed TdT, lambda5, and RAG-1 genes, but that their Ig genes were still in germline configuration. The cell line was dependent on direct contact with S17 stromal cells for growth, but, in contrast to bone marrow stem cells, required no additional growth factors for maintenance and expansion. When stimulated with IL-7 and LPS, YS-PPB cells and cells from all tested clonal sublines differentiated into sIgM+ B cells in vitro. Irradiated mice reconstituted with YS-PPB cells yielded spleens containing 38% sIgM+ donor-derived B cells, demonstrating that YS-PPB cells, although stably arrested in development at the boundary between pre-pro-B and pro-B stages of B cell differentiation, still retain their competence to differentiate into mature, Ig-producing B cells when transferred to a normal host environment. Thus, this new cell line can provide a reproducible source of B cell precursors arrested at that critical time in B cell differentiation when the machinery for Ig gene rearrangement is in place but rearrangement has not yet occurred.

Murine fetal liver (FL) and adult bone marrow (BM) hematopoietic stem cells (HSCs) are characterized by cell surface expression of CD38 and c-kit. Because murine yolk sac (YS) HSC activity precedes the initiation of FL hematopoiesis, we investigated whether YS-derived HSCs also expressed c-kit and CD38. c-Kit+ CD38+ lineage- cells derived from day 9 YS as well as adult BM were found to be enriched in high proliferative potential colony-forming cells. c-Kit+ CD38+ lineage- YS or adult BM cells were capable of long-term reconstitution (>6 months) of busulfan-conditioned newborn or lethally irradiated adult mice, respectively. In contrast, c-kit+ CD38- lineage- populations from both tissues were enriched in lineage-committed progenitors and had no long-term HSC activity. We concluded that c-kit and CD38 are cell surface markers of HSCs expressed throughout murine ontogeny.

In the chicken, maternal antibodies are transferred into the egg and subsequently transported into the developing embryo. IgG is the primary immunoglobulin isotype of the egg yolk, while IgM and IgA are mainly found in the albumen. However, considerable amounts of IgM and IgA of unknown origin are found one day prior to hatching in the yolk sac. These antibodies are not synthesized de novo by the embryo proper, thus pointing to a transfer from the albumen into the egg yolk during development. To further address this question, 125I labelled chicken IgA was injected into the albumen of freshly laid eggs. On day 21 of embryonic development, 125I-IgA was found in the yolk sac content. On average (n = 6) 36.2 +/- 7.2% of the injected radioactivity was recovered from this compartment and shown to be associated with IgA. Comparison of total IgA in the albumen of freshly laid eggs with the amounts of IgA in yolk sac content showed similar results with a 44% transfer rate. An increase of the IgA concentration in the yolk sac was first detectable between days 14 and 16 of embryonic development. These data clearly show that IgA is transferred from the albumen into the yolk sac, most likely by a transport across the yolk sac membrane.

The processes of angiogenesis and hematopoiesis require a high degree of coordination during embryogenesis. Whereas much is understood about the development of the vascular system in avian embryos, little information has been attained in mammals, predominantly because there are no specific markers for either blood vessels or hematopoietic cells in any developing mammalian system. We have recently shown that murine CD34 (mCD34) is expressed on the vascular endothelium in all organs and tissues of the adult mouse as well as on a small percentage of presumably hematopoietic stem cells in the bone marrow and fetal liver. Here we show that mCD34 is also expressed on the endothelium of blood vessels and on a subset of hematopoietic-like cells throughout murine development. mCD34 is first observed on the yolk sac endothelium of day 7.5 embryos and on a subset of hematopoietic cells within these yolk sacs. mCD34 expression is maintained on vessels and hematopoietic cells in all organs and tissues throughout embryogenesis. In addition, mCD34 is localized on growth conelike filopodial processes that appear at the budding edge of newly sprouted capillaries. Double staining of capillaries for mCD34 and laminin shows that these growth conelike processes seem to be free of laminin, whereas the formed capillaries seem to be coated with this extracellular matrix protein. Analysis of vessels in developing brain shows that these filopodial processes seem to be directed toward the ventricular epithelium, a previously described site of vascular endothelial growth factor synthesis. Finally, we show that the vascular structures of developing murine embryoid bodies also express mCD34. These data suggest that mCD34 is a useful marker for the analysis of the development of the blood vascular system in murine embryos.

It has been shown previously that erythroblast antigen (AG-EB) is an interspecies antigen expressed on the surface of nucleated red cells and reticulocytes. We have recently produced the monoclonal antibody MAE 15 that reacted specifically with murine epitope of AG-EB. Using MAE15 and immunocytochemical techniques on the light and electron microscopic levels the expression of AG-EB was studied in mouse embryonal and adult tissues. In mouse embryos AG-EB was revealed in erythroid cells of the yolk sac, liver and spleen, in epithelial cells of the intestine, salivary and thyroid glands as well as in the skin. The adult pregnant mice displayed AG-EB in cells of the yolk sac, decidua and trophoblast. The normal adult mice definitely expressed AG-EB not only in nucleated erythroid cells of the bone marrow and spleen but also in cells of non-hemopoietic organs and tissues: in epithelium of intestine, some glands and tubules of the kidney and testis; in endothelium of blood brain capillaries; in cells of the glandular layer of the epidermis; in insertion disks of myofibrils. Our findings suggest a possible transport role of AG-EB in different "barrier" cells.

Anti-Plasmodium spp. antibody titers of mating pairs of adult, captive-reared, African black-footed penguins (Spheniscus demersus) and their chicks were determined using the enzyme-linked-immunosorbent assay (ELISA). Two Plasmodium falciparum antigens were used for the ELISA: R32tet32 (sporozoite antigen), and crude red blood cell extract (CRBCE). Eighteen chicks were bled weekly for ten weeks starting with their day of hatching. The yolk sacs of two penguin eggs were biopsed for ELISA-detectable maternal antibodies (MAB). None of the 28 adult penguins were parasitemic by Giemsa-stained thin blood smear; however, all had anti-Plasmodium spp. immunoglobulins reacting with P. falciparum antigens. All 18 newly hatched chicks had anti-Plasmodium spp. MAB while housed in a mosquito-free environment. The level of MAB in the newly hatched chicks was correlated significantly (P < 0.001) with antibody level detected in their female parents (R32tet32: r = 0.87, CRBCE: r = 0.89). No correlation was found between antibody titers of the newly hatched chicks and their male parents. The level of maternal-fetal antibodies was regressed significantly (P < 0.001) over the 10-week period. Penguin chicks over 10 weeks of age had no anti-Plasmodium spp. MAB. Egg-yolk samples had significantly (P < 0.03) higher MAB titers than female parents that laid these eggs.

Turkey poults from hens vaccinated against avian group A rotavirus were examined to study the transfer of maternally derived anti-rotavirus IgG (rIgG) to the mucosal surfaces (intestinal and tracheal), serum, yolk, and bile. During the first week of life, maternal rIgG titers in intestinal mucosal washings were 200-to-500-fold less than rIgG titers in the circulation, as determined by enzyme-linked immunosorbent assay (ELISA). The intestinal titers in 10- and 13-day-old poults were negligible. A moderate linear correlation (r = 0.6) was present between rIgG titer in the blood circulation and the intestines, with a serum cutoff level of 10,000 ELISA units. Maternal rIgG was detected in tracheal washings only during the first 3 days of life. Biliary rIgG titers were fourfold higher than intestinal titers at day of hatch but had declined considerably in 1-day-old poults. Yolk had relatively high rIgG titers at hatching. Maternal rIgG titer in the small intestine was determined after in situ ligation of the individual segments; it was highest in the duodenum, followed by the ileum and jejunum. There was evidence that rIgG in the intestine was transferred from the blood and not directly from the yolk sac. Bidirectional movement of rIgG between circulation and intestine was also detected. Maternal rIgG was not detected in the intestinal washings of progeny from hens naturally infected with rotavirus.

A rabbit anti-rat yolk sac antiserum, administered by intraperitoneal injection at 0.25 mL/100 g bodyweight into 8.5-day pregnant rats, resulted in a resorption incidence of 20%, and growth retardation and malformation of all surviving fetuses at term. In all subsequent experiments, pregnant rats either received this same dose of antiserum at 8.5 days or were untreated. When 9.5-day rat conceptuses were cultured for 48 h in a medium containing serum from a 9.5-day pregnant rat that had been treated with antiserum, development was severely abnormal, regardless of whether conceptuses were explanted from antiserum-treated or untreated dams. In contrast, culture for 48 h in serum from untreated 9.5-day pregnant rats resulted in normal growth and development of conceptuses explanted from untreated dams, and in slight growth retardation and dysmorphogenesis in 9.5-day conceptuses explanted from antiserum-treated dams. In the former, development was similar to that attained by 11.5-day conceptuses from untreated dams; in the latter, development was appreciably better than in 11.5-day conceptuses from antiserum-treated dams. These results indicate that the critical period of exposure of embryos in utero to teratogenic antiserum after administration to the 8.5-day pregnant rat is longer than 24 h and that a significant insult is delivered to the conceptus before 9.5 days ea. Using 9.4-day ea conceptuses in culture, pinocytosis (uptake of 125I-labelled polyvinylpyrrolidone) by the yolk sac, and its inhibition by antiserum, was demonstrated. No pinocytic activity was evident in the embryo itself. These data support the hypothesis that anti-yolk sac antiserum exerts its teratogenic action by inhibiting the nutritional function of the yolk sac during early organogenesis.

The route of transfer of anti-Borrelia duttonii antibody subclasses from mother to young and their role in protection against borrelial challenge infection in ddY mice were investigated. Offspring from infected and noninfected mice were segregated and nursed by noninfected or infected mothers. Enzyme-linked immunosorbent assay analysis of antibodies of the cross-suckled offspring revealed that anti-B. duttonii immunoglobulin G1 (IgG1) is transferred exclusively in milk and that IgG2a is transferred mainly in milk but also slightly through the yolk sac route. On the other hand, IgG3 is transferred mainly through the yolk sac route but also slightly in milk, whereas IgG2b is transferred through both routes but to a lesser extent. Anti-borrelial IgM was not detected in any offspring. The protective role of transferred IgG subclasses was examined by challenge infection with B. duttonii. Offspring from noninfected mice fed by infected mothers had IgG1, IgG2a, and IgG3 at challenge and were completely protected against the challenge infection. On the other hand, offspring from infected mice fed by noninfected mothers had only IgG3, and 8 of 10 were completely protected from challenge infection whereas the other 2 contracted slight and transient spirochetemia. These findings suggested that anti-borrelial IgG3 alone has considerable protective activity and that IgG1, IgG2a, or both, either by themselves or together with IgG3, have a complete protective activity against borrelial infection.

An indirect immunoperoxidase monolayer assay was used to measure the location of virus and class-specific antibodies to Newcastle disease virus (NDV) of chickens. The intra-ocular vaccination of chicks with the Ulster and Hitchner B1 strains of NDV resulted in the highest titre of virus being recovered from the Harderian gland (HG) at 10(6) iu/0.1 g tissue without any accompanying virus in the faeces. Maternal IgG antibody which was detected in the lacrimal fluid at 1/50 of serum levels of IgG antibody could prevent this replication. Lacrimal IgA antibody to NDV was shown to be absolutely dependent on local virus replication for its production because it reached titres of 10(3) after local intra-ocular infection compared with < 10(1.5) after two intravenous inoculations of inactivated virus. Biliary IgA, by contrast, reached 10(3.5) after either immunisation schedule. Lacrimal IgG and IgM antibody to NDV occurred at 1-9% of the titres of serum antibody to NDV after immunisation with inactivated virus or the passive transfer of NDV-immune serum between chickens. This percentage increased to 13-33% of serum titres after intra-ocular infection with NDV as if the local replication of NDV in the HG stimulated lacrimal antibody of all classes. This is the first report of NDV replication in the HG and of virus-specific IgM in the lacrimal fluid.

The tissue distribution, molecular weight, and biochemical nature of an antigen detected by a mouse monoclonal antibody designated 283D3 and raised against rabbit visceral yolk sac endodermal cells, has been investigated. The antigen is located on the luminal side of apical tubules and large sub-apical vesicles in rabbit yolk sac endoderm and proximal kidney tubule epithelial cells. It is expressed in a similar polarised fashion in epithelial cells lining the epididymis. Western blotting showed the antigen to comprise proteins of molecular weight 330-380 kDa. The antigen has been affinity purified from yolk sac and kidney and is predominantly protein in nature with a small percentage of N-linked carbohydrate. In terms of tissue distribution and molecular weight it has close similarity to Heymann nephritis antigen but differs in not being confined to coated pits. Its function is not known, but the association with endocytic elements implies a possible role in non-specific protein absorption.