The effect of the humoral action of the cells from chick embryo hematopoietic organs on the colony-forming capacity of normal mouse bone-marrow cells and in activated erythropoiesis

The cells of chick embryo hematopoietic organs were cultivated together with mouse bone marrow cells in bicameral diffusion chambers under normal conditions and in activated erythropoiesis. The yolk sac cells inhibited colony formation on the 2nd, 4th days of development, the hepatic cells--on the 8th, 12th days, the splenic cells--on the 13th, 17th days, and the bone marrow cells--on the 15th day. The cells of the yolk sac sharply intensify the formation of colonies by the marrow mononuclears, in particular, on the 6th developmental day in activated erythropoiesis. The yolk sac (on the 4th, 6th days of development), the liver, and the spleen produce humoral factors which facilitate the formation of erythroid colonies by the marrow cells of mice. The cells of a 4-day-old yolk sac inhibit the formation of granulocytic colonies, and the synthesis of inhibitors ceases in a 6-day-old yolk sac. It is suggested that the synthesis of local humoral inhibitors of the proliferation of colony-forming units controls the intensity of hematopoiesis in the organ and, by reducing proliferation, facilitates the emergency of cells into differentiation independently, according to the law or under the effect of local humoral poetins.     

Erythropoiesis in the yolk sac of the bat Tadarida brasiliensis cynocephala

The process of erythropoiesis and vasculogenesis in the yolk sac of the bat (Tadarida brasiliensis cynocephala) has been studied through the use of both light and electron microscopy. Stem cells arise from the leading edge of the migrating splanchnic mesoderm and transform into primitive erythroblasts. Differentiation involves either contact or association with the endodermal cells, since all erythropoietic activity occurs on the endodermal side of the expanding vascular bed, and many of the cells are in close apposition to the lateral or basal plasma membranes of the endodermal cells. Endodermal cells also phagocytize developing primitive erythroblasts during the later stage of the process when erythropoiesis is subsiding in the yolk sac. Cells destined to become the endothelium of the expanding vascular bed also arise from the leading edge of the migrating splanchnic mesoderm. Their process of differentiation involves the development of cytoplasmic extensions that may surround a group of differentiating erythroblasts, enclosing them in the newly formed lumen of the blood vessel. The cytoplasmic extensions make contact and develop junctional complexes with similar processes from other cells to complete the lumen of the lengthening vascular bed. Cells of the granulocyte series or megakaryocytes are not observed in the yolk sac of the bat as has been described in certain other species.     

Erythropoiesis in the yolk sac of the bat Tadarida brasiliensis cynocephala.

The process of erythropoiesis and vasculogenesis in the yolk sac of the bat (Tadarida brasiliensis cynocephala) has been studied through the use of both light and electron microscopy. Stem cells arise from the leading edge of the migrating splanchnic mesoderm and transform into primitive erythroblasts. Differentiation involves either contact or association with the endodermal cells, since all erythropoietic activity occurs on the endodermal side of the expanding vascular bed, and many of the cells are in close apposition to the lateral or basal plasma membranes of the endodermal cells. Endodermal cells also phagocytize developing primitive erythroblasts during the later stage of the process when erythropoiesis is subsiding in the yolk sac. Cells destined to become the endothelium of the expanding vascular bed also arise from the leading edge of the migrating splanchnic mesoderm. Their process of differentiation involves the development of cytoplasmic extensions that may surround a group of differentiating erythroblasts, enclosing them in the newly formed lumen of the blood vessel. The cytoplasmic extensions make contact and develop junctional complexes with similar processes from other cells to complete the lumen of the lengthening vascular bed. Cells of the granulocyte series or megakaryocytes are not observed in the yolk sac of the bat as has been described in certain other species.     

RP59, a marker for osteoblast recruitment, is also detected in primitive mesenchymal cells, erythroid cells, and megakaryocytes.

We recently described a novel protein in bone marrow of rats, RP59, as a marker for cells with the capacity to differentiate into osteoblasts. In this work, its expression pattern was further investigated to learn about the origin and biological relevance of RP59 expressing marrow cells. As revealed by in situ hybridization and by immunohistochemistry of yolk sac embryos, RP59 was found in the cells of the primitive ectoderm and primitive streak as well as in blood islands and extraembryonal mesoderm. Later, RP59 occurred in fetal liver cells and in circulating blood. From the time around birth, it was found in bone marrow and spleen cells. In addition, in vitro-formed blood vessels contained RP59-positive cells in the lumen. Endothelial cells and the vast majority of cells outside the blood vessels were not labeled. Concerning more mature hematopoietic cell types, RP59 was observed in megakaryocytes and nucleated erythroblasts, but absent from lymphoid cells. In conclusion, RP59 was induced in early mesoderm. It was maintained in the erythroid and megakaryotic lineages and, as earlier described, in young osteoblasts.     

A light and electron microscopic study of the mouse visceral yolk sac endodermal cells in the middle and late embryonic periods, showing the possibility of definitive erythropoiesis

Hematological studies have revealed the importance of the visceral yolk sac (VYS) in the primitive erythropoiesis of mouse embryos at an early stage before day 12. We examined the possibility of the occurrence of extra-embryonic erythropoiesis at a stage later than embryonic day 12 by light and electron microscopic analyses. Surprisingly, a novel structure in the form of erythrocyte-like globules was observed in the VYS endodermal cells. They were consistently present in the VYS endodermal cells from embryonic day 12 until day 18 (birth is day 19), by immunocytochemical and enzyme histochemical analyses. They were immuno-positive for mouse erythrocyte antibody and also positive for the benzidine reaction showing the presence of hemoglobin. The erythrocyte-like globules were shown to be the erythrocytes present in the cytoplasm. These results indicated that erythropoiesis in the VYS endodermal cells continues from the early embryonic stage, as primitive erythropoiesis, until the late stage.     

Hemopoietic cells in the liver and spleen of the embryonic and early postnatal mouse: a karyometrical observation

Hemopoietic cells in the liver and spleen of embryonal and early neonatal mice were karyometrically examined by light microscopy. Hepatic hemopoiesis started at 11 days gestation, and proerythroblasts and less differentiated cells with nuclei larger than 7 micron in diameter, appeared in the primitive hepatic cords. After 12 days, the liver contained a number of mature erythroblasts. The immature large hemopoietic cells disappeared from the liver at 16 days gestation. Splenic hemopoiesis began at 16 days gestation. Hemopoietic cells with nuclei measuring larger in diameter than 8 micron appeared in the splenic cords, and the immature cells could be seen even at 2 days after birth. Development of embryonic hemopoiesis from the liver to the spleen is discussed in relation to yolk sac and bone marrow hemopoiesis.     

The ontogenesis of the hematopoietic system revisited

The ontogenesis of hematopoiesis is classically described as a series of successive steps: the first takes in the yolk sac where blood islands differentiate. Then, cells deriving from these structures migrate and populate the transient hematopoietic organs such as the liver and the spleen. At last, the eventual migration allows the establishment of bone marrow hematopoiesis. This theory described in almost all the textbooks of Human Embryology does not fit with recent experimental data. Indeed, the construction of quail-chick chimeras shows that the yolk sac does not contribute to the adult hematopoiesis in birds. Adult hematopoietic cells arise from a population located on the ventral side of the aorta both in birds and mammals. The aortic population derives from the para-aortic splanchnopleura and its derivative, the so-called AGM (Aorta-Gonad-Mesonephros). These new data provide new concepts to understand the process of ontogenesis of the hematopoietic system in vertebrates.     

ELECTRON MICROSCOPIC STUDIES OF MACROPHAGES IN EARLY HUMAN YOLK SACS

Distribution and fine structure of macrophages were studied in 10 human embryos in the 6th and 7th week of gestation, 5.5 to 12 mm in crown-rump length. The yolk sac macrophages were found in the extravascular mesenchymal tissues and intravascular spaces long before the first appearance of bone marrow and lymphatic tissues in the embryos. In addition to the macrophages, the fibroblastic cells and the cells of erythropoietic series were also present in the extravascular space. The macrophages showed a variety of cellular structures suggesting transition from immature cell type with no heterophagolysosomes to mature cell type in phagocytosis. The mature macrophages avidly phagocytized the primitive erythroblasts and occasionally platelets. They were positively stained for lysosomal enzymes and were characterized by numerous pleomorphic heterophagolysosomes which exhibited various stages of digestion of phagocytized blood cells. The origin of intravascular macrophages may be in either migrated extravascular macrophages or phagocytic endothelial cells. The phagocytosis and degradation of erythroblasts appear to be one of the main functions of yolk sac macrophages. The presence of the macrophages in mitosis indicates their proliferation in situ.     

Electron microscopic studies of macrophages in early human yolk sacs

Distribution and fine structure of macrophages were studied in 10 human embryos in the 6th and 7th week of gestation, 5.5 to 12 mm in crown-rump length. The yolk sac macrophages were found in the extravascular mesenchymal tissues and intravascular spaces long before the first appearance of bone marrow and lymphatic tissues in the embryos. In addition to the macrophages, the fibroblastic cells and the cells of erythropoietic series were also present in the extravascular space. The macrophages showed a variety of cellular structures suggesting transition from immature cell type with no heterophagolysosomes to mature cell type in phagocytosis. The mature macrophages avidly phagocytized the primitive erythroblasts and occasionally platelets. They were positively stained for lysosomal enzymes and were characterized by numerous pleomorphic heterophagolysosomes which exhibited various stages of digestion of phagocytized blood cells. The origin of intravascular macrophages may be in either migrated extravascular macrophages or phagocytic endothelial cells. The phagocytosis and degradation of erythroblasts appear to be one of the main functions of yolk sac macrophages. The presence of the macrophages in mitosis indicates their proliferation in situ.     

The microtubule marginal band of mammalian red blood cells

Summary A microtubule marginal band consisting of up to 50 microtubules is present in the primitive, circulating, nucleated, disk-shaped blood cells of fetal rats, mice and man. This marginal band is associated with vesicles of endoplasmic reticulum. Bundles of microtubules are also present in erythroblasts of the definitive line in fetal liver parenchyma and bone marrow of adults, but a typical and distinctly marginal localization is not seen in these cells. The presence of microtubule marginal bands in the primitive erythrocytes of mammals is considered as an ontogenetic recapitulation of vertebrate erythropoiesis.     

Early complete hydatidiform moles contain inner cell mass derivatives

In four cases of early complete hydatidiform moles, confirmed to be androgenetic in origin by DNA studies, we have identified non-chorionic inner cell mass derived structures which are not commonly observed in specimens of later gestational age. These structures include nucleated red blood cells, endothelial cells, stromal macrophages, amnion and yolk sac. The latter four structures were confirmed by specific immunocytochemical stains. Recognition that such structures can accompany complete hydatidiform moles has both theoretical and practical significance. From a theoretical perspective, it demonstrates that the maternal genome is not required for the initiation of amniogenesis, development of the yolk sac, vasculogenesis, or hematopoiesis. From a practical perspective it emphasizes that complete hydatidiform moles, with their markedly increased risk of subsequent choriocarcinoma, cannot be excluded based on the finding of “fetal structures.” Am. J. Med. Genet. 70:273–277, 1997. © 1997 Wiley-Liss, Inc.     

Splenic erythropoiesis in rats under hypoxic and post-hypoxic conditions

Summary In rats chronic hypoxia causes extramedullary haematopoiesis mainly localized in the spleen. It is not known how splenic erythropoiesis develops and how it regresses after termination of hypoxia. In this study the spleen of rats exposed to chronic hypoxia was studied by light and electron microscopy; the findings were compared to relevant peripheral blood values.Splenic erythropoiesis begins almost immediately after exposure to hypoxia and reaches its maximum after 24 weeks. It occurs mainly in the splenic cords drawing upon local erythroblasts and is accompanied by an increase in splenic weight as well as a decrease in splenic iron stores.After termination of hypoxia marked phagocytosis by splenic cord macrophages diminishes the number of erythroblasts and of erythrocytes with a concomitant increase in splenic iron stores. Thus, splenic erythropoiesis appears to be inhibited as part of a rebound phenomenon and returned to normal by phagocytosis of erythroid cells within 4 weeks after cessation of the hypoxic stimulus.Dedicated to Professor K. Lennert on the occasion of his 65th birthday     

Notch/Delta4 interaction in human embryonic liver CD34+ CD38- cells: positive influence on BFU-E production and LTC-IC potential maintenance

We investigated whether Notch signaling pathways have a role in human developmental hematopoiesis. In situ histochemistry analysis revealed that Notch1, 2, and 4 and Notch ligand (Delta1-4, and Jagged1) proteins were not expressed in the yolk sac blood islands, the para-aortic splanchnopleure, the hematopoietic aortic clusters, and at the early stages of embryonic liver hematopoiesis. Notch1-2, and Delta4 were eventually detected in the embryonic liver, from 34 until 38 days postconception. Fluorescence-activated cell sorter analysis showed that first-trimester embryonic liver CD34(+)CD38(low) cells expressed both Notch1 and Notch2. When these cells were cultured on S17 stroma stably expressing Delta4, a 2.6-fold increase in BFU-E number was observed at day 7, as compared with cultures with control stroma, and this effect was maintained for 2 weeks. Importantly, exposure of these cells to Delta4 under these conditions maintained the original frequency and quality of long-term culture-initiating cells (LTC-ICs), while control cultures quickly resulted in the extinction of this LTC-IC potential. Furthermore, short-term exposure of embryonic liver adherent cells to erythropoietin resulted in a dose-dependent increase in Delta4 expression, almost doubling the expression observed with untreated stroma. This suggests that Delta4 has a role in the regulation of hematopoiesis after a hypoxic stress in the fetus.     

大鼠脾脏的组织发生

应用光镜、扫描电镜和透射电镜对大鼠脾脏的组织发生进行了形态学观察。脾原基发生在妊娠第16天,继则血管出现,血窦形成,脾脏造血开始。妊娠第19天,网状细胞围绕着发生中的小动脉形成动脉周围鞘。随之淋巴细胞聚集于小动脉周围。出生后,各种血细胞造血相继终止,红、白髓不断发育成熟,生后第5天观察到淋巴树突细胞。边缘带、边缘窦和脾小结相继出现。生后第25天观察到原始生发中心。第40天后,脾脏各部结构基本发育完善。     

Experimental study of hematopoiesis in rats with exchange replacement of blood by perfluorotributylamine emulsion

Exchange blood replacement with perfluorotributylamine (PFTBA) emulsion was carried out in experiments on unanesthetized rats. During the period of blood exchange and for the next 5 days the rats were maintained on an increased oxygen concentration. On the sixth day the animals were returned to ordinary atmospheric conditions, and systematic observation commenced on that day and continued for 1 year. On the sixth day after exchange blood replacement, marked activation of erythropoiesis was found in the bone marrow of the rats and reticulocytosis in their peripheral blood. Meanwhile a decrease in the hemoglobin concentration and an increase in the number of lymphocytes and mature and immature neutrophils were found in the blood; immature erythropoietic cells-polychromatophilic erythroblasts and normoblasts-were present. The normal peripheral blood indices were completely restored by the 13th day and the morphological composition of the bone marrow by the 28th day; for the next 12 months hematopoiesis in the rats was maintained without appreciable deviations. It can be concluded from these results that the PFTBA emulsion has no adverse effect on hematopoiesis in animals and does not disturb the normal process of regulation of hematopoiesis under these experimental conditions.Laboratory of Pathological Physiology and Laboratory of New Blood Substitutes, Central Institute of Hematology and Blood Transfusion, Ministry of Health of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR N. A. Fedorov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 87, No. 6, pp. 531–533, June, 1979.     

Migration of Prebursal Stem Cells from the Early Chicken Embryo to the Yolk Sac

Allogeneic yolk sac—embryo chimaeras were constructed by association of S¹⁵B¹⁵ yolk sac and B²B² embryo on day 2 of incubation. Five days later yolk sac cells from the chimaeras were injected intravenously into 14-day-old irradiated embryos, using recipients of B²B² and B¹⁵B¹⁵ genotypes. One week after hatching, cells in the bursa of Fabricius and peripheral blood erythrocytes were studied for la-like antigens and B alloantigens, respectively, to determine whether they were derived from the embryo or yolk sac part of the chimaera. The results obtained demonstrate that prebursal and erythropoietic stem cells migrate from the early embryo to the yolk sac during the 2nd to the 7th day of incubation. They also exclude the de novo generation of prebursal stem cells in the yolk sac.     

Development of B-lymphocyte colony-forming cells in foetal mouse tissues.

In CBA and (CBA X C57B1)F1 mice, cells forming B-lymphocyte colonies in agar culture were first detected in the 17-day foetal liver and the following day in the spleen, bone marrow and peripheral blood. Colony-forming cells were not detected in the yolk sac or foetal thymus. Adult levels of colony-forming cells were achieved within 3 days of birth. In organ cultures of 15-day foetal liver or spleen, B-lymphocyte colony-forming cells developed during a 5-day incubation period, indicating that both organs can function as bursal analogues. Foetal liver colony-forming cells were of small size and generated colonies of cells with a pattern of membrane immunoglobulin similar to colony cells generated by cells from adult animals.     

超声检测早孕期卵黄囊的临床意义

卵黄囊是孕囊内非常重要的结构,在胚胎早期发育中发挥着造血、营养、生殖、代谢、免疫、排泄和内分泌等功能。卵黄囊容易受多种因素影响,在超声上表现为直径过大或过小、形态不规则、回声异常、钙化等,卵黄囊的形态改变和妊娠结局相关,形态异常易引起早期自然流产,卵黄囊直径过大和染色体异常密切相关,多篇文献报道卵黄囊和羊膜囊数目的非一致性,说明卵黄囊不能确定羊膜囊性。最新研究发现巨噬细胞来源于卵黄囊,参与免疫调节。卵黄囊和早产以及糖尿病的关系还在表面的推测上,缺乏大量临床数据及理论依据来进一步证实。     

The amniotic, allantoic and yolk sac epithelia of the cat: SEM and TEM studies

Summary The epithelial layers of the feline allantoamnion and yolk sac between the 23rd day and full term were studied using SEM, TEM, and light microscopical histochemistry. Amniotic, allantoic and yolk sac fluid were analysed. The attenuated and relatively inert amniotic epithelium sloughs off completely around the 54th day, the amniotic cavity is then lined by fibrous connective tissue only. The exocoelomic epithelium is regarded as the source of the abundant macrophages, rich in lysosomal enzymes, that are seen in the allantoamniotic membrane after obliteration of the exocoelomic cleft. The allantoic epithelium has been mistaken for the amniotic one by many authors. Its cuboidal, glycogen-rich cells develop coral-like luminal outgrowths after the 45th day, which contain almost all kinds of organelles. The basal cell poles form large interdigitations, and the nuclei become heavily lobulated. The changes are accompanied by drastic alterations in the ionic composition of the allantoic fluid. The yolk sac endoderm differs greatly from the allantoic endoderm. It is the most active epithelium of the accessory fetal membranes. The yolk sac mesothelium retains many long microvilli over the whole gestational period. The nutritional value of the yolk sac fluid is only higher than that of the amniotic fluid in its glycerol and cholesterol values.