Transformation of cultured human vascular endothelium by SV40 DNA

Gimbrone, Michael A.; Fareed, George C.

doi:10.1016/0092-8674(76)90132-x

Cited by 105 publications

(45 citation statements)

References 35 publications

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“…In contrast, in almost all reports on SV40 T antigen-immortalized endothelial cells, the growth characteristics of primary cells were changed or only some of the characteristic endothelial cell markers remained (Ades et al, 1992;Fickling et al, 1992;Gimbrone and Fareed, 1976;Hohenwarter et al, 1994;Reznikoff and DeMars, 1981;Wautier et al, 1999). The detrimental effects of immortalization on the expression of endothelial characteristics were also observed in a commonly used cell line of endothelial origin EA.hy926 (a fusion product between HUVEC and lung carcinoma) (Edgell et al, 1983) and in HUVEC immortalized with human papilloma virus-16 (HPV-16) oncoproteins E6/E7 (Burger et al, 1998;Fontijn et al, 1995).…”

Section: Krump-konvalinkova Et Almentioning

confidence: 94%

Generation of Human Pulmonary Microvascular Endothelial Cell Lines

Krump‐Konvalinkova

Bittinger

Unger

et al. 2001

Laboratory Investigation

143

102

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SUMMARY:The limited lifespan of human microvascular endothelial cells in cell culture represents a major obstacle for the study of microvascular pathobiology. To date, no endothelial cell line is available that demonstrates all of the fundamental characteristics of microvascular endothelial cells. We have generated endothelial cell lines from human pulmonary microvascular endothelial cells (HPMEC) isolated from adult donors. HPMEC were cotransfected with a plasmid encoding the catalytic component of telomerase (hTERT) and a plasmid encoding the simian virus 40 (SV40) large T antigen. Cells transfected with either plasmid alone had an extended lifespan, but the cultures eventually entered crisis after several months of proliferation. Only those cells that were transfected with both plasmids acquired the capacity to grow in vitro without demonstrating major crisis, and these cells have been in culture for 24 months. HPMEC isolated from two different donors were used, generating two populations of immortalized cells, HPMEC-ST1 and HPMEC-ST2. Single cell-derived clones of the immortalized cells HPMEC-ST1 exhibited growth characteristics that were similar to those of the parental HPMEC. One selected clone, HPMEC-ST1.6R, displayed all major constitutively expressed and inducible endothelial phenotypic markers, including platelet endothelial cell adhesion molecule (PECAM-1, CD31), von Willebrand factor (vWF), and the adhesion molecules, intercellular adhesion molecule (ICAM-1), vascular adhesion molecule (VCAM-1), and E-selectin. In addition, an angiogenic response was demonstrated by sprout formation on a biological extracellular matrix (Matrigel). The HPMEC-ST1.6R cells did not form tumors in nude mice. The microvascular endothelial cell line, HPMEC-ST1.6R, will be a valuable tool for the study of microvascular endothelial physiology and pathology including gene expression, angiogenesis, and tumorigenesis. (Lab Invest 2001, 81:1717-1727.

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Section: Krump-konvalinkova Et Almentioning

confidence: 94%

Generation of Human Pulmonary Microvascular Endothelial Cell Lines

Krump‐Konvalinkova

Bittinger

Unger

et al. 2001

Laboratory Investigation

143

102

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“…Enforced expression of SV40 T antigens (SV40Tag) or the human papilloma virus E6 and E7 oncogenes, which inactivate pRB and TP53 functions, extends the replicative life span of human endothelial cells (20,21). However, telomeres in viral antigentransformed cells continue to shorten, resulting in extensive chromosomal instability and eventual genetic catastrophe, which terminates the culture in a growth crisis (4).…”

Section: Introductionmentioning

confidence: 99%

Telomere-Driven Karyotypic Complexity Concurs with p16INK4a Inactivation in TP53-Competent Immortal Endothelial Cells

Wen¹,

Wu²,

Baksh³

et al. 2006

Cancer Research

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Critically short telomeres promote chromosomal fusions, which in TP53-defective cells initiate the formation of cytogenetic aberrations that are typical of human cancer cells. Expression of the enzyme telomerase stabilizes normal and aberrant chromosomes by maintaining telomere length. However, previous investigations, including our own, have shown that overexpression of telomerase reverse transcriptase (hTERT) does not prevent net telomere shortening in human endothelial cells. In the present study, two mass cultures of hTERT-transduced bone marrow endothelial cells (BMhTERT) and 26 clones were employed to further investigate the immortalization process and consequences of telomere shortening. Eighty-five percent (22 of 26) of the clones and both mass cultures were immortalized. However, cytogenetic analyses revealed recurring cytogenetic aberrations in the mass cultures and 12 representative clones. Several of the recurring aberrations, including +5p, +11, À13, +19, and +20, and nonreciprocal translocations involving 17p and 2p were previously implicated in human carcinogenesis. One mass culture and a subset of clones (5 of 12) had complex karyotypes, characterized by cytogenetic heterogeneity and at least five chromosomal abnormalities. p16 INK4a was silenced exclusively in the five clones and mass culture with complex karyotypes, whereas the p53/p21 cip1 pathway was defective in only one clone. Telomere dysfunction was implicated in the evolution of complex karyotypes by the presence of anaphase bridges, telomere associations, and dicentric chromosomes. These results show that complex karyotypes can evolve in TP53-competent cells and provide evidence that p16 INK4a functions as a gatekeeper to prevent telomere-driven cytogenetic evolution. These investigations provide new insight to the role of p16 INK4a as a tumor suppressor. (Cancer Res 2006; 66(22): 10691-700)

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“…Establishment of a continuous cell line of endothelial cells would facilitate studies on the physiologic and pathologic factors that induce endothelial mitosis. In addition, transformation of the endothelial cells may offer a clue to a better understanding of tumors of endothelial origin, such as Kaposi's sarcoma (4, 11).Transformation of human endothelial cells with SV40 was first accomplished by Gimbrone and Fareed, in 1976, by transfection with intact circular DNA or linear fragments containing the entire early-gene region, but not by infection with SV40 virions. However, the transformed endothelial cells were negative for Factor VIIIrelated antigen (FVIII-RAG) (6).…”

mentioning

confidence: 99%

“…Transformation of human endothelial cells with SV40 was first accomplished by Gimbrone and Fareed, in 1976, by transfection with intact circular DNA or linear fragments containing the entire early-gene region, but not by infection with SV40 virions. However, the transformed endothelial cells were negative for Factor VIIIrelated antigen (FVIII-RAG) (6).…”

mentioning

confidence: 99%

“Transformation” of Human Endothelial Cells by SV40 Virions

Ide

Minamishima²,

Eizuru³

et al. 1988

Microbiology and Immunology

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Human endothelial cells derived from the umbilical vein were transformed with SV40 virions. A cell line subcultured for over 60 serial passages was characterized in comparison with its untransformed counterpart which was culturable for less than five passages.The SV40-transformed human endothelial cells, designated SV-HUVEC, were positive not only for tumor (T) antigen specific to the SV40-transformed cell, but also for two markers of endothelial cells, Factor VIII-related antigen and a receptor for Ulex europaeus agglutinin I. By transformation the growth potential of the human endothelial cells was increased and their serum requirement was decreased. The SV40-transformed endothelial cells were, however, unable to form colonies in soft agar or to form tumors in athymic nude mice, although a small nodule was produced at the site of inoculation. Subcultivation of these cells up to the 62nd passage eventually resulted in crisis and loss of further cell division. Thus, the human endothelial cells were transformed by SV40 while retaining certain normal functions but without showing tumorigenicity.Endothelial cells cover all the vascular walls in a one-wall-thick layer and form a continuous lining between the circulating blood and the surrounding tissues. Endothelial cells also play an important role in wound healing (19), the reendothelialization of vessel-wall injury (23), and tumor growth (1, 3). Establishment of a continuous cell line of endothelial cells would facilitate studies on the physiologic and pathologic factors that induce endothelial mitosis. In addition, transformation of the endothelial cells may offer a clue to a better understanding of tumors of endothelial origin, such as Kaposi's sarcoma (4, 11).Transformation of human endothelial cells with SV40 was first accomplished by Gimbrone and Fareed, in 1976, by transfection with intact circular DNA or linear fragments containing the entire early-gene region, but not by infection with SV40 virions. However, the transformed endothelial cells were negative for Factor VIIIrelated antigen (FVIII-RAG) (6).Recently, transformation of endothelial cells by infection with SV40 virions was achieved and a cell line positive for FVIII-RAG was subcultured up to the 62nd passage in our laboratory. The characteristics of the SV40-transformed endothelial cells, designated SV-HUVEC, are described in this paper.

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Transformation of cultured human vascular endothelium by SV40 DNA

Cited by 105 publications

References 35 publications

Generation of Human Pulmonary Microvascular Endothelial Cell Lines

Generation of Human Pulmonary Microvascular Endothelial Cell Lines

Telomere-Driven Karyotypic Complexity Concurs with p16INK4a Inactivation in TP53-Competent Immortal Endothelial Cells

“Transformation” of Human Endothelial Cells by SV40 Virions

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