Proliferation of pulmonary endothelial cells: Time-lapse cinematography of growth to confluence and restitution of monolayer after wounding

Ryan, Una S.; Absher, Marlene; Olazabal, Bertha M.; Brown, Lynn M.; Ryan, James W.

doi:10.1016/0040-8166(82)90054-4

Cited by 20 publications

(7 citation statements)

References 17 publications

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“…The wounded monolayer model has been used to study the mechanisms and kinetics of endothelial regeneration, and it has been shown that wounding initiates both cell migration and division (Sholley et al, 1977;Selden and Schwartz, 1979;Ryan et al, 1982). Cell-cycle dependent variations in PA activity and uPA mRNA levels have been reported (Rohrlich and Rifldn, 1977;Loskutoff and Paul, 1978;Aggeler et al, 1982;Grimaldi et al, 1986;Scott et al, 1987).…”

Section: Discussionmentioning

confidence: 99%

“…Wounding has been demonstrated to induce both cell migration and cell division (Sholley et al, 1977;Selden and Schwartz, 1979;Ryan et al, 1982). Wound-edge caseinolysis was observed in cultures treated with mitomycin C (10 ltg/ ml) (Fig.…”

Section: Piasrainogen-dependent Caseinolysismentioning

confidence: 92%

“…In vivo, this occurs during the regeneration of endothelial lesions in large blood vessels (Fishman et al, 1975;Schwartz et al, 1978;Haudenschild and Schwartz, 1979), and during the formation of new capillary blood vessels in the process of angiogenesis (Ausprunk and Folkman, 1977;Folkman, 1985). In vitro, endothelial cell migration can be triggered by mechanically wounding a confluent monolayer of cells (Sholley et al, 1977;Selden and Schwartz, 1979;Goflieb and Spector, 1981;Ryan et al, 1982;Madri and Stenn, 1982).…”

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confidence: 99%

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Urokinase-type plasminogen activator is induced in migrating capillary endothelial cells.

Pepper

Vassalli

Montesano

et al. 1987

The Journal of Cell Biology

259

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Abstract. Cellular migration is an essential component of invasive biological processes, many of which have been correlated with an increase in plasminogen activator production. Endothelial cell migration occurs in vivo during repair of vascular lesions and angiogenesis, and can be induced in vitro by wounding a confluent monolayer of cells. By combining the wounded monolayer model with a substrate overlay technique, we show that cells migrating from the edges of an experimental wound display an increase in urokinase-type plasminogen activator (uPA) activity, and that this activity reverts to background levels upon cessation of movement, when the wound has closed. Our results demonstrate a direct temporal relationship between endothelial cell migration and uPA activity, and suggest that induction of uPA activity is a component of the migratory process.T HE vascular endothelium consists of a highly ordered monolayer of cells which provides a structural and functional barrier between circulating blood and the surrounding tissues. In response to a variety of stimuli, normally quiescent endothelial cells can be induced to migrate. In vivo, this occurs during the regeneration of endothelial lesions in large blood vessels (Fishman et al., 1975;Schwartz et al., 1978;Haudenschild and Schwartz, 1979), and during the formation of new capillary blood vessels in the process of angiogenesis (Ausprunk and Folkman, 1977;Folkman, 1985). In vitro, endothelial cell migration can be triggered by mechanically wounding a confluent monolayer of cells (Sholley et al., 1977;Selden and Schwartz, 1979; Goflieb and Spector, 1981;Ryan et al., 1982; Madri and Stenn, 1982).Proteases, and in particular plasminogen activators (PAs),1 have been correlated with cell migration in a variety of invasive biological processes (Reich, 1978; Mullins and RJahrlich, 1983;Saksela, 1985;Dan~ et al., 1985; Goldfarb and Liotta, 1986). Using the wounded monolayer model together with a substrate overlay technique, which has the advantage of allowing direct visualization of proteolysis around migrating cells, we demonstrate here that capillary endothelial cells migrating from the edges of an experimental wound display an increase in urokinase-type PA (uPA) activity. Materials and Methods Materials4[~-phorbol 12-myristate 13-acetate (PMA), g-aminocaproic acid, cycloheximide, mitomycin C, and amiloride were purchased from Sigma Chemical 1. Abbreviations used in this paper: BME, bovine microvascular endothelial cells; PA, plasminogen activator; tPA, tissue-type plasminogen activator; uPA, urokinase-type plasminogen activator. BSA (Sigma Chemical Co.) was acid treated as described (Loskutoff, 1978), to remove labile protease inhibitors. Plasminogen was purified from human plasma by lysine-Sepharose (Pharmacia Fine Chemicals, Uppsala, Sweden) affinity chromatography (Deutsch and Mertz, 1970). Endothelial Cell CultureCloned microvascular endothelial cells from bovine adrenal cortex (BME cells) (Furie et al., 1984), a generous gift from Drs. M. B. Furie and S. C. S...

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Section: Discussionmentioning

confidence: 99%

Section: Piasrainogen-dependent Caseinolysismentioning

confidence: 92%

mentioning

confidence: 99%

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Urokinase-type plasminogen activator is induced in migrating capillary endothelial cells.

Pepper

Vassalli

Montesano

et al. 1987

The Journal of Cell Biology

259

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“…Cell migration assay was performed as described previously (Ryan et al, 1982). Briefly, cells were seeded on 24‐well plates.…”

Section: Methodsmentioning

confidence: 99%

Discovery of a benzoxazine derivative promoting angiogenesis in vitro and in vivo

Dong

Cheng

Zhao

et al. 2010

Journal Cellular Physiology

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Angiogenesis is a multi-step process that refers to the growth of new vessels from pre-existing ones. Endothelial proliferation, migration, and tube formation constitute a critical step in angiogenesis. Recently, we demonstrated that a novel benzoxazine derivative, 6-amino-2,3-dihydro-3-hydroxymethyl-1,4-benzoxazine (ABO) could improve the proliferation of human umbilical vein endothelial cells (HUVECs) without basic fibroblast growth factor (bFGF) and serum. In this study, we further tested its effect on endothelial angiogenesis with Matrigel assay, migration assay, and in vivo chick chorioallantoic membrane (CAM) assay. Our results showed that ABO effectively facilitated cell migration and promoted capillary-like tube formation in vitro and in vivo. To elucidate the underlying mechanisms, we examined intracellular reactive oxygen species (ROS) level/nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and superoxide dismutase (SOD) activities, nitric oxide (NO) level/endothelial nitric oxide synthase (eNOS) activity, and mitochondrial membrane potential (MMP). Our data indicated that ABO depressed ROS with inhibition of NADPH oxidase instead of SOD activity, stimulated NO production and eNOS activation, and restored MMP in HUVECs. Our findings suggest that ABO is a promising tool for exploring the mechanisms of angiogenesis and may have a therapeutic potential in ischemic pathologies.

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“…TLC has been used to compare proliferation and migration activity of pulmonary artery endothelial ceils and fibroblasts (5) and to compare growth of endothelial cultures during establishment of a confluent monolayer from low-density seeding to growth of a culture reconstituted after mechanical "wounding" (12). Detailed methodology for preparation of endothelial cell cultures for TLC and for acquisition and analysis of data derived from filmed sequences are presented.…”

Section: Time-lapse Cinematography (Tlc) Is a Valuablementioning

confidence: 99%

Cinematographic analysis of endothelial cell proliferation in culture

Absher

1986

Journal of Tissue Culture Methods

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SUMMARY: A method is described for the application of time-lapse cinematographic techniques (TLC) to analysis of the behavior of individual cells and clones in endothelial cultures. Using TLC, detailed and precise information on cell cycle time, proliferative state, and migration activity can be obtained on several hundred individual cells in a given filmed sequence of proliferating endothelial cultures.

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Proliferation of pulmonary endothelial cells: Time-lapse cinematography of growth to confluence and restitution of monolayer after wounding

Cited by 20 publications

References 17 publications

Urokinase-type plasminogen activator is induced in migrating capillary endothelial cells.

Urokinase-type plasminogen activator is induced in migrating capillary endothelial cells.

Discovery of a benzoxazine derivative promoting angiogenesis in vitro and in vivo

Cinematographic analysis of endothelial cell proliferation in culture

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