Spontaneous transformation and immortalization of human endothelial cells

Takahashi, Kimiko; Sawasaki, Yoshio; Hata, Junichi; Mukai, Kiyoshi; Goto, Teppei

doi:10.1007/bf02624456

Cited by 404 publications

(243 citation statements)

References 32 publications

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“…In order to assess cell viability, an MTT cytotoxicity assay consisting of tetrazolium salt reduced to formazan by mitochondrial enzymes of viable cells was used proportionally to its dehydrogenase activity and defined as a 3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyl-tetrazolium bromide colorimetric assay (11).…”

Section: Methodsmentioning

confidence: 99%

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Cytotoxicity of root canal sealers on endothelial cell cultures

et al. 2013

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This study evaluated, in vitro, the cytotoxicity of six root canal sealers after 12, 24 and 72 h of contact time, using an endothelial ECV-304 cell line. The MTT assay was used for analysis of cell viability. Twelve specimens of each sealer were prepared and randomly assigned to 6 groups according to the commercial brands (n=4/time). A control group was also formed, which was not subjected to the contact with sealers. To assess the effects of sealers on endothelial cells, the specimens were placed in culture plate wells and incubated at 37°C with 5% CO 2 and 100% humidity. MTT assays were performed in quadruplicate after 12, 24 and 72 h of contact of the sealer specimens with monolayers. Statistical analysis was performed by two-way ANOVA with Bonferroni post-hoc test at a significance level of 5%. Analysis of absorbance in the experimental groups showed that GuttaFlow presented the lowest cytotoxicity, with a mean absorbance of 0.048, followed by Pulp Canal Sealer (0.038), Sealer 26 (0.038), Endo Densell (0.036) and Pulp Fill (0.035). The control group had a mean absorbance of 0.098. Based on the results, Endofill and GuttaFlow were the most and the least cytotoxic sealers, respectively.

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Section: Methodsmentioning

confidence: 99%

“…ECV304 endothelial cells are obtained from the human umbilical cord vein. This line is characterized by a monolayered growth pattern, high proliferative potential, with no growth factor-specific requirement (11).…”

Section: Introdutionmentioning

confidence: 99%

Cytotoxicity of root canal sealers on endothelial cell cultures

et al. 2013

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“…ECV cells (spontaneously transformed human umbilical vein endothelial cells; Takahashi et al, 1990) were maintained in RPMI-1640, 10% FCS, 50 IU ml À1 penicillin, 50 mg ml À1 streptomycin and split twice weekly 1:10. ECV cells display the following endothelial cell markers: factor VIII-related antigen (negative), UEA-I and PHM5 (positive) and ultrastructurally endothelium-speci®c Weibel± Palade bodies (Takahashi et al, 1990). Epithelial cell markers, such as cytokeratin, and two other epithelial cell markers are not found (Takahashi et al, 1990).…”

Section: Cell Culturementioning

confidence: 99%

Fibronectin-binding protein acts as Staphylococcus aureus invasin via fibronectin bridging to integrin alpha5beta1

Sinha¹,

François²,

Nüße³

et al. 1999

Cell Microbiol

507

563

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SummaryThe ability of Staphylococcus aureus to invade mammalian cells may explain its capacity to colonize mucosa and to persist in tissues after bacteraemia. To date, the underlying molecular mechanisms of cellular invasion by S. aureus are unknown, despite its high prevalence and dif®culties in treatment. Here, we show cellular invasion as a novel function for an S. aureus adhesin, previously implicated solely in attachment. S. aureus, but not S. epidermidis, invaded epithelial 293 cells in a temperature-and F-actindependent manner. Formaldehyde-®xed and live bacteria were equally invasive, suggesting that no active bacterial process was involved. All clinical S. aureus isolates analysed, but only a subset of laboratory strains, were invasive. Fibronectin-binding proteins (FnBPs) acted as S. aureus invasins, because: (i) FnBP deletion mutants of invasive laboratory strains lost invasiveness; (ii) expression of FnBPs in noninvasive strains conferred invasiveness; and (iii) the soluble isolated ®bronectin-binding domain of FnBP (D1±D4) completely blocked invasion. Integrin a 5 b 1 served as host cell receptor, which interacted with staphylococcal FnBPs through cellular or soluble ®bronectin. FnBP-de®cient mutants lost invasiveness for epithelial cells, endothelial cells and ®broblasts. Thus, ®bronectin-dependent bridging between S. aureus FnBPs and host cell integrin a 5 b 1 is a conserved mechanism for S. aureus invasion of human cells. This may prove useful in developing new therapeutic and vaccine strategies for S. aureus infections.

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“…HeLa-H2B is a clone from the cervical carcinoma model cell line (HeLa) stably expressing the histone protein H2B fused to GFP (31). ECV-304 was originally described as an immortalized endothelial cell line (32). However, more recent evidence has suggested that ECV-304 is a derivative of the human bladder carcinoma cell line T24 (33).…”

Section: Cell Culture and Transfectionmentioning

confidence: 99%

Controlled light exposure microscopy reveals dynamic telomere microterritories throughout the cell cycle

et al. 2008

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Telomeres are complex end structures that confer functional integrity and positional stability to human chromosomes. Despite their critical importance, there is no clear view on telomere organization in cycling human cells and their dynamic behavior throughout the cell cycle. We investigated spatiotemporal organization of telomeres in living human ECV-304 cells stably expressing telomere binding proteins TRF1 and TRF2 fused to mCitrine using four dimensional microscopy. We thereby made use of controlled light exposure microscopy (CLEM), a novel technology that strongly reduces photodamage by limiting excitation in parts of the image where full exposure is not needed. We found that telomeres share small territories where they dynamically associate. These territories are preferentially positioned at the interface of chromatin domains. TRF1 and TRF2 are abundantly present in these territories but not firmly bound. At the onset of mitosis, the bulk of TRF protein dissociates from telomere regions, territories disintegrate and individual telomeres become faintly visible. The combination of stable cell lines, CLEM and cytometry proved essential in providing novel insights in compartment-based nuclear organization and may serve as a model approach for investigating telomere-driven genome-instability and studying long-term nuclear dynamics. ' 2008 International Society for Advancement of Cytometry Key termstelomere; TRF1; TRF2; nuclear organization; chromatin dynamics; CLEM; live cell imaging TELOMERES are the natural ends of linear chromosomes. A mammalian telomere consists of a double stranded array of simple TTAGGG repeats ending in a single stranded overhang that folds back to form a T-loop structure (1). In combination with sufficient telomere repeats, a complex of indirect and direct telomere binding proteins, dubbed shelterin, assures proper telomere structure and function. The specificity of shelterin for telomeric DNA is due to the recognition of TTAGGG repeats by three of its components: the homodimers telomeric repeat binding factor 1 and 2 (TRF1 and TRF2) that bind the duplex part of telomeres, and protection of telomeres 1 that binds to the single stranded TTAGGG repeats present at the three-overhang and in the D loop of the T-loop configuration (2-5).The nucleus has a meticulous organization with functional relevance to the cell assuring proper gene expression, replication and genome stability (6-8). Telomeres are considered to play an important role in spatial genome organization. Whereas yeast telomeres are known to form few large silencing clusters at the heterochromatic nuclear periphery (9,10), there is no consensus on how telomeres are organized in the mammalian nucleus. From an architectural point of view, human telomeres have been proposed to be anchored to a nuclear scaffold, thereby possibly providing struc-

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Spontaneous transformation and immortalization of human endothelial cells

Cited by 404 publications

References 32 publications

Cytotoxicity of root canal sealers on endothelial cell cultures

Cytotoxicity of root canal sealers on endothelial cell cultures

Fibronectin-binding protein acts as Staphylococcus aureus invasin via fibronectin bridging to integrin alpha5beta1

Controlled light exposure microscopy reveals dynamic telomere microterritories throughout the cell cycle

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