Action of embryonic thymus and liver cells on the development of the ascitic form of Ehrlich's carcinoma

The influence of the cells of embryonic thymus and liver on the development of Ehrlich carcinoma was studied. The intraperitoneal injection of the embryonic cells in the adult mice infested by the Ehrlich carcinoma resulted in a marked lengthening of the life time of animals and an increase of the survival percentage. The embryonic cells of thymus and liver inhibited sharply the growth of carcinoma cells in the diffusion chambers as well. In contrast to this, the thymus and bone marrow cells of adult animals, taken in the same concentrations as the embryonic cells, exhibited only a slight inhibiting effect on the growth of tumour cells. On the basis of these data a suggestion is put forward to the effect that the embryonic immunocompetent cells determine the stronger inhibition of tumour growth in the embryos as compared with the adult animals.     

Role of the thymus in generation of lymphocyte functions: II. Serum-mediated inhibition of development of mitogen-reactive lymphocytes in embryonic mouse thymus organ cultures

The thymuses of 14-day-old mouse embryos could be grown in serum-free organ cultures for at least 14 days with development of relatively large numbers of lymphocytes. These also acquired a strong reactivity to the mitogens concanavalin A (Con A) and leucoagglutinin (LA). Supplementing the organ culture medium with serum from calf (CS), fetal calf (FCS), mouse (MS), or fetal mouse (FMS) gave a serum concentration-dependent inhibition of development of mitogen reactivity, without clearly altering the quantitative lymphoid development in the organ cultures. Adult sera were more suppressive than fetal sera. All of nine tested FCS lots were inhibitory and the inhibiting activity was mainly found in the albumin fraction upon Sephadex G-200 gel filtration. Although FCS prevented development of mitogen-reactive cells in organ cultures of thymuses of 14-day-old embryos, it had much less effect on cultures of 15-day-old thymuses. FCS present during the entire organ culture period most efficiently inhibited generation of mitogen reactivity. If present only during the first or second half of the 14-day culture period, the inhibition was still marked but less complete.     

Contribution of neural crest-derived cells in the embryonic and adult thymus

Neural crest (NC)-derived mesenchyme has previously been shown to play an important role in the development of fetal thymus. Using Wnt1-Cre and Sox10-Cre mice crossed to Rosa26(eYfp) reporter mice, we have revealed NC-derived mesenchymal cells in the adult murine thymus. We report that NC-derived cells infiltrate the thymus before day 13.5 of embryonic development (E13.5) and differentiate into cells with characteristics of smooth muscle cells associated with large vessels, and pericytes associated with capillaries. In the adult organ at 3 mo of age, these NC-derived perivascular cells continue to be associated with the vasculature, providing structural support to the blood vessels and possibly regulating endothelial cell function.     

肝窦内皮细胞在肝纤维化发生发展中的作用及其机制

肝窦内皮细胞(liver sinusoidal endothelial cells,LSECs)是肝脏抵御炎症和免疫反应的第一道防线,其独特的窗孔结构及功能特性决定它在肝纤维化发生发展中起重要作用。LSECs主要通过介导肝脏炎症反应、肝窦毛细血管化、活化肝星状细胞(hepatic stellate cells,HSC)、引发细胞外基质(extracellular matrix,ECM)的生成与降解失衡等途径参与肝纤维化的发生发展。靶向LSECs对治疗肝纤维化极具潜力,阐述清楚二者调控关系,将为抗肝纤维化治疗提供新的理论依据。     

Role of endothelial cells in early pancreas and liver development

Liver and pancreas initially develop by budding from the embryonic endoderm. The formation of these organs coincides with the appearance of endothelial cells (ECs) adjacent to the endoderm. ECs either develop in situ in organs, or are recruited by organs and are induced to form blood vessels. Recent reports on liver and pancreas have now shown that ECs also induce essential steps in organ formation such as morphogenesis and cell differentiation. This review summarizes reports on EC signaling during organogenesis and cell differentiation.     

Early fetal hematopoietic development from in vitro differentiated embryonic stem cells.

In this report we describe the efficient hematopoietic differentiation of embryonic stem (ES) cells in vitro. When cultured in semisolid medium two of five ES cell lines efficiently generated embryoid bodies (EBs) containing blood islands in which hematopoietic cells from all six myeloid lineages could be detected. Among a variety of growth factors tested, only erythropoietin significantly increased blood island formation. We directly demonstrate the presence of hematopoietic progenitors in the EBs by employing an in vitro precursor assay. Colony-forming cells (CFC) of all myeloid lineages as well as bi- and multipotent (CFC-MIX) progenitors were readily identified, and a detailed time-course analysis of their appearance was performed. Despite a high frequency of CFC-MIX in vitro, we did not observe any spleen colony-forming cells (CFU-S) in vivo. We conclude that hematopoietic differentiation of ES cells under these conditions reflects formation of the complete range of blood cells found in the yolk sac of the early fetus. Therefore this system provides a unique model in which to study the earliest events of hematopoietic development in vitro.     

Ontogeny of T cells: development of pre-T cells from fetal liver and yolk sac in the thymus microenvironment

The patterns of development of T cells from the very early stem cells that settle in the embryonic thymus have been studied. For this purpose, mouse embryonic thymuses (14 days) depleted of thymocytes were reconstituted with hemopoietic stem cells from fetal liver (FL) and yolk sac (YS) and T-cell development was followed in vitro in organ culture. It was found that cells derived from FL and YS of 10- to 14-day-old embryos were capable of reconstituting depleted thymic explants and exhibiting membrane markers in a pattern similar to that of thymocytes developing in intact thymic explants. Furthermore, these cells responded to concanavalin A in proliferative and cytotoxic assays as measured by limiting-dilution analysis. Thus, lymphohemopoietic stem cells emerging in the embryo prior to thymus lymphoid development are capable of differentiation in the thymus microenvironment into T cells, identified by phenotypic markers and functions that are characteristic of cells developing in the intact embryonic thymus.     

Early embryonic development in vitro by coculture with oviductal epithelial cells in pigs

This experiment was designed to evaluate the ability of three different somatic cell cultures to promote development of early cleavage stage pig embryos. A total of 245 2-cell, 4-cell, 8-cell, and 16-cell pig embryos were cocultured for 5 days with porcine oviductal epithelial cells (POEC), porcine fetal fibroblast monolayer (PEF), a combined POEC and PEF coculture system (PEF-POEC), or Dulbecco's Modified Eagle Medium alone (DMEM). Embryos were collected at slaughter from the reproductive tracts of superovulated prepubertal gilts. Embryos were recovered, evaluated, and randomly placed in one of the four treatment groups. POEC were recovered from oviductal flushes, washed, and placed in 24-well plates. PEF were obtained from 30-day to 60-day fetuses and established in culture. Finally, PEF-POEC consisted of a confluent monolayer of PEF in the bottom of 24-well plates also containing a Costar semipermeable membrane chamber with POEC in it. Embryos were evaluated every 24 h to determine stage of development. More (p less than 0.05) embryos developed to blastocysts in POEC (70% and 54%, respectively) and PEF-POEC (67% and 61%, respectively), than in either DMEM (16% and 2%, respectively) or PEF (27% and 23%, respectively). However, development of embryos did not differ (p less than 0.05) for POEC and PEF-POEC. These data indicate the presence of a primary culture of POEC promotes in vitro development of early cleavage stage pig embryos.     

Hematopoietic stem cells in mice during embryonic development preceding the establishment of liver hematopoiesis]

We studied the time course of appearance of CFUs (7-8 days old) in embryos of (C57B1/6 x CBA)F1 mice from the 8th day of embryonic development. Significant amounts of CFUs could be detected from the 10th day of development, initially in the body of the embryo from the stage of 30-33 pairs of somites, then in the yolk sac and still later, from the stage of about 40 pairs of somites, in liver anlage. CFUs could not be reliably detected until the 9th day of development either in the embryo itself or in the yolk sac. However, after incubation of nine day old embryos for four days in organ culture, such cultures contained CFUs. CFUs could be found in significant levels in embryos explanted from the embryos at the stage no earlier than 24 pairs of somites. When the yolk sac and the embryo were cultivated separately, CFUs could also be detected, however, the removal of liver primordium from the embryo did not influence the amount of CFUs in its body. CFUs were not found in cultures of liver primordium from nine day old embryos. Thus, we can detect pre-CFUs in 9 day old embryos at the stage 25-28 pairs of somites using the system of organ culture; at the same time CFUs cannot be found in intact embryos of the same age. These data provide evidence that before the establishment of liver hemopoiesis precursors of CFUs are located both in the yolk sac and in the embryo outside rudimentary liver. However, our results do not provide any data for the conclusion about the primary source of pre-CFUs in the mouse embryo.     

Retinol improves bovine embryonic development in vitro

Retinoids are recognized as important regulators of vertebrate development, cell differentiation, and tissue function. Previous studies, performed both in vivo and in vitro, indicate that retinoids influence several reproductive events, including follicular development, oocyte maturation and early embryonic development. The present study evaluated in vitro effects of retinol addition to media containing maturing bovine oocytes and developing embryos in both a low oxygen atmosphere (7%) and under atmospheric oxygen conditions (20%). In the first experiment, abbatoir collected bovine oocytes were matured in the presence or absence of varying concentrations of retinol. After a 22–24 hour maturation period the oocytes were fertilized, denuded 18 hours later and cultured in a modified synthetic oviductal fluid (mSOF) in a humidified atmosphere at 38.5 degrees C, 5% CO2, 7% O2 and 88% N2. Cleavage rates did not differ among control and retinol-treated oocytes in all three experiments. Addition of 5 micromolar retinol to the maturation medium (IVM) tended (p < 0.07) to increase blastocyst formation (blastocyst/putative zygote; 26.1% +/- 2.2%) compared to the controls (21.9% +/- 1.9%). Further analysis revealed when blastocyst development rates fell below 20% in the control groups, 5 micromolar retinol treatment dramatically improved embryonic development, measured by blastocyst/putative zygote rate (14.4 +/- 2.1 vs 23.7 +/- 2.5; p < 0.02). The 5 micomolar retinol treatment also enhanced the blastocyst/cleaved rate by nearly 10% (23.7% vs 34.6%; p < 0.02). In the second and third experiments addition of 5 micromolar retinol to the embryo culture medium (IVC) under low oxygen conditions did not significantly improve cleavage or blastocyst rates, but 5 micromolar retinol significantly increased blastocyst development under 20% O2 conditions (p < 0.001). These studies demonstrate that supplementation of 5 micromolar retinol to the maturation medium may improve embryonic development of bovine oocytes indicated by their increased blastocyst rate. A significant improvement in the blastocyst development with the 5 micromolar retinol treatment under atmospheric conditions suggests a beneficial antioxidant effect during embryo culture.     

Sulfate esterifying activity of embryonic chick liver in vitro

A method has been described for the study of tissue sulfate-conjugating systems in vitro. Liver slices from embryonic chicks were maintained in vitro in a medium containing labeled inorganic sulfate and phenol. It was found that more of the sulfate was esterified at 20 °C. than at 37 °C. due to the longer continued activity at the lower temperature. All sulfate-esterifying activity was lost in liver slices maintained at 37 °C. for 30 hr. while those cultures maintained at 20 °C. continued to esterify sulfate for 70 hr.On the basis of our data there would appear to be a change in the thermal stability of the sulfate-esterifying enzyme system of the chick liver upon its transition from the embryonic stage to the stage of the fully developed chick. Data were presented for the chick 4 months ex ovo. We have been unable to detect any analogous temperature effects upon the sulfate-esterifying system in the livers of embryonic and adult rats.     

Evidence for a cyclic renewal of lymphocyte precursor cells in the embryonic chick thymus

Experiments involving sequential transplantations of the chick embryonic thymus at E9 to E12 into a first 3-day host quail embryo and then into a second chick host allowed demonstration of the cyclic periodicity of hemopoietic cell seeding of the embryonic thymus. After a first wave of colonization occurring between E6.5 and E8, the thymus becomes refractory to hemopoietic cell entry for about 4 days. It resumes its capacity to be seeded by a second wave of blood-borne stem cells at E12. After a second period of non receptivity starting at E14, a third wave of incoming cells reaches the thymus around E18. Therefore, with a slightly different periodicity, the same cyclic mechanism regulates the renewal of lymphocytes in chick and quail embryos. Quail hemopoietic cells were immunostained in the chimeric thymuses, with a species specific monoclonal antibody (anti-MB1) which recognizes a common surface antigenic determinant on all endothelial and blood cells of the quail (except erythrocytes). Two steps could thus be distinguished in the seeding process. When the thymus becomes receptive for hemopoietic cells, the latter first accumulate in the intrathymic blood vessels before penetrating massively in the thymic parenchyma. The quail chick-chimera system combined with the use of a species- and cell-type-specific antibody provides a unique tool for studying thymic colonization by lymphocyte precursors.     

Glucocorticoids modulate the in vitro development of the embryonic rat pancreas

The effect of the glucocorticoid analogue, dexamethasone, on the development of the embryonic pancreas was studied in tissue culture. It specifically enhances the accumulation of exocrine enzymes without altering the level of general cell proteins. The enhancement, however, is not symmetrical: the cellular levels of the two major exocrine products, amylase and chymotrypsinogen, are increased about 10- and 2- fold, respectively. Two other zymogens that are present in minor quantities, procarboxypeptidases A and B, are also increased, whereas no effect is seen on lipase A. Coordinate with these effects on synthesis, there is a dramatic change in the morphology of dexamethasone-stimulated acinar cells. Their number of zymogen granules is higher and crystalline arrays are found in the rough endoplasmic reticulum. Dexamethasone also inhibits cell replication, perhaps by selectively inhibiting the last cell divisions of the culture period. At the same time, there is a disproportionate reduction in the insulin content of cultured rudiments. We find that pancreatic development is normal in the absence of dexamethasone and that this glucocorticoid does not precociously induce the appearance of the specific secretory products, but rather enhances by a constant degree their synthesis and accumulation. Therefore, we conclude that glucocorticoids may play a modulatory but not an inductive role in pancreatic development.