Large-scale screening for developmental genes in embryonic stem cells and embryoid bodies using retroviral entrapment vectors

Xiong, Jing Wei; Battaglino, Ricardo A.; Leahy, Amy; Stuhlmann, Heidi

doi:10.1002/(sici)1097-0177(199806)212:2<181::aid-aja4>3.0.co;2-d

Cited by 30 publications

(19 citation statements)

References 51 publications

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“…13,15 Its restricted expression during embryogenesis in the vascular endothelium and its precursors in the yolk sac blood islands suggested that Egfl7 plays a role during endothelial lineage determination, and possibly during embryonic vasculogenesis and angiogenesis. Egfl7-specific transcripts were found in most adult organs tested, and in primary and transformed endothelial cell lines.…”

Section: Discussionmentioning

confidence: 99%

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EGFL7 Is a Chemoattractant for Endothelial Cells and Is Up-Regulated in Angiogenesis and Arterial Injury

Campagnolo¹,

Leahy²,

Chitnis³

et al. 2005

The American Journal of Pathology

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The endothelium of the adult vasculature is normally quiescent, with the exception of the vasculature of the female reproductive system. However, in response to appropriate stimuli (ie, wound healing, atherosclerosis, tumor growth and metastasis, arthritis) the vasculature becomes activated and grows new capillaries through angiogenesis. We have recently identified a novel endothelial-restricted gene, Egfl7, that encodes a 41-kd secreted protein (Fitch MJ, Campagnolo L, Kuhnert F, Stuhlmann H: Egfl7, a novel epidermal growth factor-domain gene expressed in endothelial cells. Dev Dyn 2004, 230:316 -324). Egfl7 is expressed at high levels early during mouse embryonic development and is strictly associated with the vascular bed. In this study, we investigated Egfl7 expression in the quiescent adult vasculature, in the pregnant uterus, and in two different models of arterial injury, namely ballooning and ferric chloride injury. By RNA in situ hybridization, Egfl7 expression in the vasculature was found to be restricted to the endothelium of the capillaries and mature vessels. In the pregnant uterus, increased vascularization was accompanied by up-regulation of Egfl7. On arterial injury, Egfl7 expression was up-regulated in the regenerating endothelium, but not in the neointima. Importantly, the EGFL7 protein acted as a chemoattractant for embryonic endothelial cells and fibroblasts in a cell migration assay. Together, these results suggest that Egfl7 functions in the formation and maintenance of endothelial integrity and that its up- In the adult mammalian organism, the vasculature is normally quiescent. Arterial endothelial cells have an extremely low turnover rate (ϳ1 in every 10 5 cells undergoes cell division 1 ). However, adult endothelial cells are not postmitotic and, in response to appropriate stimuli, they can proliferate and form new blood vessels by a process termed angiogenesis. 2-4 Angiogenesis describes the formation of new capillaries and larger vessels by sprouting or splitting from pre-existing vessels. Typically, the sprouting of vessels involves activation of quiescent endothelial cells, proteolytic degradation of the extracellular matrix, chemotactic migration, invasion into the surrounding stroma, proliferation and differentiation of endothelial cells, and formation of a new lumen and maturation of the endothelium. 2,[5][6][7][8] This angiogenic sprouting process occurs under physiological conditions during the female reproductive cycle (ovulation, implantation, pregnancy) and wound healing, as well as under pathological conditions in solid tumors and metastases, rheumatoid arthritis, retinopathies, hemangiomas, and psoriasis. 2,7,9,10 Several of the key players in both embryonic and adult angiogenesis are vascular-specific growth factor ligands

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Section: Discussionmentioning

confidence: 99%

“…In a genetic screen for early circulatory system genes 13,14 we have identified a novel murine gene, epidermal growth factor (EGF)-like domain 7 (Egfl7). 15 Egfl7 encodes a secreted protein with an apparent molecular weight of ϳ41 kd that contains an aminoterminal signal peptide and two centrally located EGF domains.…”

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confidence: 99%

EGFL7 Is a Chemoattractant for Endothelial Cells and Is Up-Regulated in Angiogenesis and Arterial Injury

Campagnolo¹,

Leahy²,

Chitnis³

et al. 2005

The American Journal of Pathology

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123

150

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“…3,36 Briefly, ES cells were dissociated into single cells with 0.25% Trypsin/EDTA (Gibco/BRL). After MEF feeder cells were depleted by incubating cell suspensions for 10 minutes at 37°C, ES cells were suspended and counted by Trypan Blue staining (Bioss, Beijing, China).…”

Section: Es Cell Culture and Differentiationmentioning

confidence: 99%

“…36 Transfected ES cells were placed on neomycin-resistant SNL (STO [mouse embryonic fibroblast cell line] that produces LIF) feeder cells (a gift from Dr K. Choi) in the ES cell medium. After 24 hours, ES cells were selected in the ES medium supplemented with 0.25 mg/mL G418 (Geneticin Selective Antibiotic; Invitrogen, Carlsbad, CA) for 10 days.…”

Section: Generation Of Stable Lycat Transgenic Es Cell Linesmentioning

confidence: 99%

Mouse lysocardiolipin acyltransferase controls the development of hematopoietic and endothelial lineages during in vitro embryonic stem-cell differentiation

Wang

Faloon

Tan

et al. 2007

Blood

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The blast colony-forming cell (BL-CFC) was identified as an equivalent to the hemangioblast during in vitro embryonic stem (ES) cell differentiation. However, the molecular mechanisms underlying the generation of the BL-CFC remain largely unknown. Here we report the isolation of mouse lysocardiolipin acyltransferase (Lycat) based on homology to zebrafish lycat, a candidate gene for the cloche locus. Mouse Lycat is expressed in hematopoietic organs and is enriched in the IntroductionIt was proposed nearly a century ago that a common progenitor generates both the hematopoietic and endothelial lineages. 1,2 Using in vitro mouse embryonic stem (ES) cell differentiation, the blast colony-forming cell (BL-CFC) that clonally generates both endothelial and hematopoietic cells in the presence of vascular endothelial growth factor (VEGF) was characterized. [3][4][5] The BL-CFC was later isolated in vivo from the posterior primitive streak of mid-gastrulation mouse embryos. 6 Flk1 ϩ Scl ϩ and Brachyury ϩ Flk1 ϩ cells are enriched for the hemangioblast. 6,7 Fate mapping in the zebrafish gastrula suggests that hemangioblasts are interspersed with cells that only give rise to either blood cells or endothelial cells in the ventral mesoderm. 8,9 However, the molecular identity and plasticity of the hemangioblast remain largely unknown.The in vitro differentiation of mouse ES cells, along with genetically modified ES cells, has proved to be valuable in deciphering the underlying signaling pathways in hemangioblast development. 10,11 Several pathways are revealed to participate in hemangioblast development, including the Bmp4-Gata2 signaling in embryoid bodies (EBs), the VEGF-Flk1-Plcg1 signaling in mice and EBs, and the transcription factors Scl, Runx1, Mixl1, and Hex in EBs. 7,[12][13][14][15][16][17][18][19][20][21][22][23][24][25] In zebrafish, the cloche (clo) mutant provides genetic evidence that a single gene is required for both endothelial and hematopoietic lineages. 26,27 cloche acts upstream of all known hematopoietic (scl, gata1, gata2) and endothelial (flk1, etsrp, fli1) genes. 26,[28][29][30][31][32][33][34] We have recently cloned the zebrafish lysocardiolipin acyltransferase (lycat) gene from the cloche genetic interval. lycat is required for the generation of both endothelial and hematopoietic lineages and acts upstream of scl and etsrp in zebrafish embryos (J.-W.X., Qingming Yu, Jiaojiao Zhang, and John D. Mably, "An acyltransferase controls the generation of hematopoietic and endothelial lineages in zebrafish," manuscript submitted July 2007). Interestingly, we found that mouse Lycat messenger RNA (mRNA) could partially rescue the cloche mutant phenotype. Therefore the mouse Lycat gene may also play an important role in hemangioblast, endothelial, and hematopoietic cell development.Here we report the isolation of a mouse orthologue of zebrafish lycat and the characterization of mouse Lycat role in the generation of hemangioblasts and endothelial and hematopoietic lineages during in vitro ES cell differentiat...

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“…The ability to differentiate ES cells in vitro allows for their use as model systems for the study of developmental potential and also as valuable reagents for stem cell therapeutic approaches [22][23][24]. The method proposed here is an efficient in vitro system to screen and study effects of small molecules and bioagents.…”

Section: Discussionmentioning

confidence: 99%

Differentiation of neuronal cells using a murine embryonic stem cell-based method

Ezekiel¹

2013

ABB

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The differentiation and screening methodology proposed here is an efficient in vitro system to screen and study effects of small molecules and bioagents and is an alternative to studies that use live animals and embryos. The method is based on engineering a stable murine embryonic stem (ES) cell line expressing lineage-specific promoters that drive selection and reporter genes. Additionally, uniform embryoid bodies (EBs) are used for differentiation studies that allow synchronous differentiation. The reporter and selection marker genes are expressed only in lineages where the promoter is functional. The differentiated cell type can be identified by reporter gene expression and the selection marker can be used for selective enrichment of that particular cell population. The method described here is useful in screening small molecules or bioagents that can differentiate stem cells into particular lineages or cell types. Identified compounds are useful in areas such as stem cell-based regenerative medicine and therapeutics. The method described here has been applied to neuronal cell differentiation.

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Large-scale screening for developmental genes in embryonic stem cells and embryoid bodies using retroviral entrapment vectors

Cited by 30 publications

References 51 publications

EGFL7 Is a Chemoattractant for Endothelial Cells and Is Up-Regulated in Angiogenesis and Arterial Injury

EGFL7 Is a Chemoattractant for Endothelial Cells and Is Up-Regulated in Angiogenesis and Arterial Injury

Mouse lysocardiolipin acyltransferase controls the development of hematopoietic and endothelial lineages during in vitro embryonic stem-cell differentiation

Differentiation of neuronal cells using a murine embryonic stem cell-based method

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