Endothelial cells are a specialized type of epithelial cell which forms the inner layer of blood vessels.
These cells play a key role in angiogenesis, the development of new blood vessels from pre-existing vessels. Angiogenesis is a multi-step process that is important for both physiological and pathological development. During angiogenesis, endothelial cells are activated and express matrix metalloproteinases (MMPs), which degrade the vascular basement membrane. In response to environmental cues, endothelial cells secrete MMPs and then invade through the basement membrane to form new capillary networks.
Endothelial cells are tested in a variety of assays for functions that contribute to the angiogenesis process. Collagen I coated surfaces are suitable for culturing endothelial cells such as fetal bovine heart endothelial cells (FBHECs) and human umbilical vein endothelial cells (HUVECs) (Figure 1). In vitro assays of endothelial cell function include cell migration1, invasion2, and tubule formation3-9. Both the BD BioCoat™ Angiogenesis System: Endothelial Cell Invasion and the BD BioCoat Angiogenesis System: Endothelial Cell Migration allow for rapid data collection without multiple handling steps. These quantitative assays utilize BD FluoroBlok™ microporous polyethylene terephthalate (PET) membranes (3 μm pore size) which effectively block the fluorescence signal from labeled cells that have not invaded or migrated through the membrane, respectively, thereby allowing the selective detection of cells that reside on the underside of the membrane (Figure 2). To perform fluorescence detection, cells may be pre-labeled or post-labeled with a fluorescent dye (Figure 3). The pre-labeling technique enables real-time kinetic measurements of cell migration or invasion. Endothelial cells must be able to migrate and enzymatically degrade the basement membrane in order for angiogenesis to occur. The wells of the BD BioCoat Angiogenesis System: Endothelial Cell Invasion are evenly coated with BD Matrigel™ Matrix, which allows researchers to examine the ability of endothelial cells to invade through reconstituted basement membrane in response to chemoattractants, such as VEGF, in the presence or absence of anti-angiogenic agents (Figure 4).
BD BioCoat Angiogenesis System: Endothelial Cell Migration consists of BD FluoroBlok inserts evenly coated with human fibronectin (Figure 5). Studies conducted using the post-labeling technique demonstrated that BD™ HUVEC-2 cells migrate towards VEGF in a concentration dependent manner (Figure 6).
During angiogenesis, endothelial cells form capillaries once they have invaded through the basement membrane. The correct culture surface is critical for successful endothelial cell tube formation in vitro. Both primary endothelial cells and endothelial cell lines have been demonstrated to form tubules on the BD BioCoat Angiogenesis System: Endothelial Cell Tube Formation (Figures 7-9) which is comprised of a 3D gel of BD Matrigel Matrix. The BD BioCoat Angiogenesis Systems are available in 24- and 96-Multiwell formats, which can be used for moderate to high throughput compound screening. BD Matrigel™ Matrix has also been extensively used to study in vivo angiogenesis4-5, 10-12 as a less technically challenging alternative to the corneal implantation model. A "plug" of material is placed subcutaneously, followed by histological quantification 7-10 days later. These in vitro and in vivo assays give researchers multiple options for exploring endothelial cell functions that are essential during angiogenesis.
* BD BioCoat Angiogenesis System: Endothelial Cell Tube Formation offers a standardized and robust assay for studying endothelial cell tubulogenesis. For customers interested in establishing an assay for tube formation using vialed BD Matrigel Matrix, we recommend pre-testing lots to ensure optimal performance.
Nakamura K, Taguchi E, Miura T, Yamamoto A, Takahashi K, Bichat F, Guilbaud N, Hasegawa K, Kubo K, Fujiwara Y, Suzuki R, Kubo K, Shibuya M, Isoe T. (2006) KRN951, a highly potent inhibitor of vascular endothelial growth factor receptor tyrosine kinases, has antitumor activities and affects functional vascular properties. Cancer Res. 66(18):9134.
Steinle JJ, Booz GW, Meininger CJ, Day JNE, Granger HJ. (2003) β3- adrenergic receptors regulate retinal endothelial cell migration and proliferation. J Biol Chem. 278(23):20681.
Di Simone N, De Santis M, Tamburrini E, Di Nicuolo F, Lucia MB, Riccardi P, D’Ippolito S, Cauda R, , Caruso A. (2007) Effects of antiretroviral therapy on tube-like network formation of human endothelial cells. Biol Pharm Bull. 30(5):982.
Kong D, Li Y, Wang Z, Banerjee S, Sarkar FH. (2007) Inhibition of angiogenesis and invasion by 3,3’-diindolylmethane is mediated by the NF-κB downstream target genes MMP-9 and uPA that regulated bioavailability of VEGF in prostate cancer. Cancer Res. 67(7):3310.
Michaud-Levesque J, Demeule M, Beliveau R. (2007) In vivo inhibition of angiogenesis by a soluble form of melanotransferrin. Carcinogenesis. 28(2):280.
Nishiyama K, Takaji K, Uchijima Y, Kurihara Y, Asano T, Yoshimura M, Ogawa H, Kurihara H. (2007) Protein kinase A-regulated nucleocytoplasmic shuttling of Id1 during angiogenesis. J Biol Chem. 282(23):17200.
Takeda Y, Kazarov AR, Butterfield CE, Hopkins BD, Benjamin LE, Kaipainen A, Hemler ME. (2007) Deletion of tetraspanin Cd151 results in decreased pathologic angiogenesis in vivo and in vitro. Blood. 109(4):1524.
Di Simone N , Di Nicuolo F, Sanguinetti M, Castellani R, D’Asta M, Caforio L, Caruso A. (2006) Resistin regulates human choriocarcinoma cell invasive behaviour and endothelial cell angiogenic processes. J Endocrinol. 189:691.
Folkman J and Haudenshschild C. (1980) Angiogenesis in vitro. Nature. 288(5791):551.
Birdsey GM, Dryden NH, Amsellem V, Gebhardt F, Sahnan K, Haskard DO, Dejana E, Mason JC, Rand AM. (2008) Transcription factor Erg regulates angiogenesis and endothelial apoptosis through VE-cadherin. Blood. 111(7):3498.
Murphy EZ, Majeti BK, Barnes LA, Makale M, Weis SM, Lutu-Fuga K, Wrasidlo W, Cheresh DA. (2008) Nanoparticle-mediated drug delivery to tumor vasculature suppresses metastasis. Proc Natl Acad Sci. 105(27):9343.
Kisuck J, Butterfield CE, Duda DG, Eichenberger SC, Saffaripour S, Ware J, Ruggeri ZM, Jain RK, Folkman J, Wagner DD. (2006) Platelets and platelet adhesion support angiogenesis while preventing excessive hemorrhage. Proc Natl Acad Sci. 103(4):855.
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