Unraveling the distinct distributions of VE- and N-cadherins in endothelial cells: A key role for p120-catenin

Endothelial cells express two different classical cadherins, vascular endothelial (VE) cadherin and neural (N) cadherin, having distinct functions in the vascular system. VE-cadherin is specific to endothelial adherens junctions and is strictly necessary for vascular morphogenesis. On the contrary, N-cadherin shows diffuse localization on the cell surface and interacts with mural cells for vessel stabilization. In this study, we sought to clarify the cellular mechanisms leading to the distinct cellular locations and functions of the two cadherins in the endothelium. VE-cadherin has been shown to be responsible for the junctional exclusion of N-cadherin. Using several endothelial models, we demonstrate that this property is dependent on VE-cadherin binding to p120 catenin (p120^ctn). Moreover, although in the absence of VE-cadherin N-cadherin can localize to cell contacts, angiogenesis remains impaired, demonstrating that endothelial junction formation is not sufficient for normal vessel development. Interestingly, we show that VE-cadherin, but not N-cadherin, is partially associated with cholesterol-enriched microdomains. Lipid raft-associated-VE-cadherin is characterized by a very high level of p120^ctn association, and this association is necessary for VE-cadherin recruitment into lipid rafts. Altogether, our results indicate a critical role for p120^ctn in regulating the membrane distribution of endothelial cadherins with functional consequences in terms of cadherin stabilization and intracellular signaling.     

利用胚胎干细胞分化的拟胚体研究小鼠早期胚胎发育过程

小鼠胚胎干细胞(ES细胞)具有分化的全能性已经得到广泛共识。ES细胞在体外分化所形成的拟胚体在结构上能够模仿早期胚胎发育过程,包括在内细胞团表面形成内胚层、柱状上皮细胞的分化,以及中央空腔的形成。本文介绍利用拟胚体研究小鼠早期胚胎发育过程中各个胚胎阶段的发育、细胞程序性死亡的发生及TGF-β信号在胚胎发育过程中的作用。     

Establishment and characterization of a pluripotent stem cell line derived from human amniotic membranes and initiation of germ layers in vitro

OBJECTIVES: Pluripotent stem cells are proposed to be used in regenerative therapy and may exist in the human amniotic membrane. The present article is aimed at establishing a pluripotent stem cell line from human placenta. METHODS: HAM-1 (stem cell line derived from human amniotic membranes) was established by the colonial cloning technique using aMEM culture medium containing 10 ng/ml of EGF, 10 ng/ml of hLIF and 10% fetal bovine serum. RESULTS: HAM-1 cells appeared to maintain a normal karyotype indefinitely in vitro and expressed markers characteristic of stem cells from mice and human, namely alkaline phosphatase. Also, these cells contributed to the formation of chimeric mouse embryoid bodies and gave rise to cells of all germ layers in vitro. CONCLUSIONS: This study demonstrates that human amniotic membranes derived stem cells have a wide developmental capability and might be utilized to regenerate different types of cells or tissues for transplantation therapy.     

Pluripotent differentiation in vitro of murine Es-D3 embryonic stem cells

Although the ES-D3 murine embryonic stem cell line was one of the first derived, little information exists on the in vitro differentiation potential of these cells. We have used immunocytochemical and flow cytometric methods to monitor ES-D3 embryoid body differentiation in vitro during a 21-d period. Spontaneous differentiation of embryoid body cells was induced by leukemia inhibitory factor withdrawal in the absence of feeder cells. The pluripotent stem cell markers Oct-3/4, SSEA-1, and EMA-1 were found to persist for at least 7 d, whereas the primitive endoderm marker cytokeratin endo-A was expressed at increasing levels from day 6. The localization of these antigens within the embryoid bodies suggested that embryonic ectoderm- and primitive endoderm-derived tissues were segregated. Localized expression of class III beta-tubulin and sarcomeric myosin also was detected, indicating that representatives of all three embryonic germ layers were present after induction of differentiation in vitro.     

High Frequency Induction of Somatic Embryoid of Rice in Suspension Culture

On MS and B5 basic media the mature seed of rice (Oryza sativa L.) was induced to produce somatic embryoid. The results showed that the rate of occurrence of somatic embryoid varied with the variety of rice and the concentration of 2,4-D was important to the occurrence of somatic embryoid. For those varieties of rice suitable for induction, the induction factors such as KT, zeatin, BAP, ABA, osmotic pressure and so forth were stringently required at different stages of somatic embryoid development, and they also influenced the formation of somatic embryoid. The results of various layered cultures of embryonic calli proved that the somatic embryoid was derived from the surface layer callus containing embryonic cells. At the induction stage no somatic embryoid was formed in callus. The rate of somatic embryoid formation was over seventy per cent on liquid differentiation medium I.     

Enhanced cardiomyogenesis of human embryonic stem cells by a small molecular inhibitor of p38 MAPK

Abstract Human embryonic stem cells (hESC) can differentiate to cardiomyocytes in vitro but with generally poor efficiency. Here, we describe a novel method for the efficient generation of cardiomyocytes from hESC in a scalable suspension culture process. Differentiation in serum-free medium conditioned by the cell line END2 (END2-CM) readily resulted in differentiated cell populations with more than 10% cardiomyocytes without further enrichment. By screening candidate molecules, we have identified SB203580, a specific p38 MAP kinase inhibitor, as a potent promoter of hESC-cardiogenesis. SB203580 at concentrations <10 μM, induced more than 20% of differentiated cells to become cardiomyocytes and increased total cell numbers, so that the overall cardiomyocyte yield was approximately 2.5-fold higher than controls. Gene expression indicated that early mesoderm formation was favored in the presence of SB203580. Accordingly, transient addition of the inhibitor at the onset of differentiation only was sufficient to determine the hESC fate. Patch clamp electrophysiology showed that the distribution of cardiomyocyte phenotypes in the population was unchanged by the compound. Interestingly, cardiomyogenesis was strongly inhibited at SB203580 concentrations ≥15 μM. Thus, modulation of the p38MAP kinase pathway, in combination with factors released by END2 cells, plays an essential role in early lineage determination in hESC and the efficiency of cardiomyogenesis. Our findings contribute to transforming human cardiomyocyte generation from hESC into a robust and scalable process.