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21.
Daniel E. Gomez Jacqueline L. Hartzler Robert H. Corbitt Alexander M. Nason Unnur P. Thorgeirsson 《In vitro cellular & developmental biology. Animal》1993,29(6):451-455
Summary We describe a fast and reproducible method that can be used as a final step in obtaining pure populations of liver endothelial
cells. This method employs endothelial cell specific lectin covalently bound to magnetic polystyrene beads (Dynabeads). Evonymus
europaeus agglutinin (EEA)-coated Dynabeads were used to purify monkey liver endothelium from Percoll gradient separated nonparenchymal
cells. EEA-coated beads were also successfully used to purify monkey aortic endothelial cells. The endothelial cells grew
to confluence as a cobblestonelike monolayer, expressed Factor VIII related antigen, and incorporated acetylated-low density
lipoprotein. The magnetic beads seemed not to modify the normal properties of the isolated endothelium, thus facilitating
their use in experimental studies. This immunomagnetic separation technique may be applicable for purification of endothelial
cells from a wide variety of tissue sources. 相似文献
22.
Eleonore Jonas Alexander Dwenger Michael Jonas Michael Nerlich Harald Tscherne 《Luminescence》1993,8(5):253-260
We investigated the effect of prostaglandin E1 on human polymorphonuclear leukocytes, in vivo. Polymorphonuclear leukocytes of a prostaglandin E1 and placebo study group were harvested and their function, as production of oxygen-derived metabolites and adherence to human cultured endothelial cells, was compared. Additionally, data obtained from polymorphonuclear leukocytes of a prostaglandin E1 and placebo group were compared with data obtained from polymorphonuclear leukocytes from 28 blood donors, who served as a control group. Production of oxygen-derived metabolites by polymorphonuclear leukocytes during contact with endothelial cells was measured by chemiluminescence. Chemiluminescence was significantly (p < 0.01) increased in the placebo group in comparison to the control group decreasing to values of control group after 6 d (post-trauma). Chemiluminescence response was not significantly suppressed in patients treated with prostaglandin E1 in comparison to the placebo group. Adherence of polymorphonuclear leukocytes (placebo group) to endothelial cells was significantly increased (p < 0.01) within the first 6 d post-trauma Following day 6, values were in the same range as values for the control group. Adherence was not significantly suppressed in patients treated with prostaglandin E1 in comparison to the placebo group. In conclusion, prostaglandin E1 at a dose of 20 ng/kg bw/min does not influence production of oxygenderived metabolites and adherence in polytraumatized patients in comparison to a placebo group. Additionally, production of oxygen-derived metabolites by polymorphonuclear leukocytes in response to endothelial cells is shown and it is evident that endothelial cells might influence production of oxygen derived metabolites by polymorphonuclear leukocytes. 相似文献
23.
24.
B. L. Coomber 《Journal of cellular biochemistry》1993,52(3):289-296
Monolayers of endothelial cells respond to physical denudation with a characteristic sequence of lamellipodia extrusion, cell migration, and cell proliferation. Basic fibroblast growth factor (bFGF) has been implicated as a necessary component of this process: addition of exogenous bFGF enhances monolayer regeneration both in vitro and in vivo, and monolayer regeneration can be inhibited in vitro by treatment with neutralizing antibodies raised against bFGF. Centrosome reorientation from a random location to one preferentially situated between the nucleus and the denudation edge has been postulated as a mechanism essential for cell polarization and subsequent migration. This present study examined the effects of a polyclonal antibody to bFGF and suramin on monolayer regeneration, actin microfilament staining, and centrosome orientation at the wound edge of partially denuded bovine large vessel endothelial monolayers. Treatment with anti-bFGF or suramin abolished monolayer repair in these cultures. Cells at the denudation edge showed altered actin staining patterns and reduced lamellipodia extrusion, and there was complete inhibition of centrosome reorientation in treated cultures. Monolayer repair and centrosome reorientation could be restored by addition of exogenous bFGF in antibody but not suramin treated cultures. Recent evidence suggests that preferential centrosome location in migrating cells may be a consequence of lamellipodia protrusion and cell spreading, rather than an indication of cell polarization. However, these results indicate that agents which interfere with bFGF availability prevent endothelial monolayer regeneration via mechanisms involving cell spreading and/or centrosome reorientation. 相似文献
25.
Shur-Jen Wang Peter Greer Robert Auerbach 《In vitro cellular & developmental biology. Animal》1996,32(5):292-299
Summary We report on the isolation and propagation of endothelial cells from the mouse embryonic yolk sac, the earliest site of blood
vessel development, and on the advantages of a hypervascular transgenic mouse source of these cells. These transgenic mice
express multiple copies of an activated allele of the humanfps/fes proto-oncogene and display hypervascularity progressing to multifocal hemangiomas. This phenotype suggested a role of thefps/fes proto-oncogene in vasculogenesis and angiogenesis and led us to investigate the growth characteristics of yolk-sac-derived
endothelial cells from transgenicfps/fes embryos. We have established eight independent cell clones from a mixture of transgenic and control yolk sacs from Day 12
embryos. Southern blot hybridization analysis showed all eight clones to be derived from transgenic cells suggesting a growth
advantage of cells carrying the activatedfps/fes gene. A cell line, Clone 166 (C166), established from one of these clones, was more fully characterized. C166 exhibits normal
endothelial characteristics, such as rearrangement into tubelike structures when placed on Matrigel, expression of angiotensin
converting enzyme, retention of cobblestone morphology at confluence, and the presence of cell surface receptors for acetylated
low density lipoprotein. The cells constitutively express murine endothelial cell adhesion molecule VCAM-1 and the vascular
addressin identified by antibody MECA-99. As expected, the cell line expresses high levels of the cytoplasmic protein-tyrosine
kinase encoded by thefps/fes proto-oncogene. The clone we have described as well as other endothelial cell lines that we have established from the mouse
embryonic yolk sac should prove useful for the study of endothelial cell differentiation and for the determination of the
mechanisms underlying the establishment of organ-specific endothelial cell heterogeneity. 相似文献
26.
Effect of anoxia and reoxygenation on antioxidant enzyme activities in immortalized brain endothelial cells 总被引:1,自引:0,他引:1
Olivier Rabin Michèle Piciotti Katy Drieu Jean-Marie Bourre Françoise Roux 《In vitro cellular & developmental biology. Animal》1996,32(4):221-224
Summary The effects of anoxia and reoxygenation on major antioxidant enzyme activities were investigatedin vitro in immortalized rat brain endothelial cells (RBE4 cells). A sublethal anoxic period of 12 h was assessed for RBE4 cells using the neutral red uptake test. Anoxia markedly influenced the specific activity of catalase and superoxide dismutase,
with no major effect on glutathione peroxidase or glutathione reductase. After 24 h postanoxia, the superoxide dismutase activity
modulated by the presence or absence of oxygen returned to control value.
Damage and recovery of RBE4 immortalized rat brain endothelial cells in culture after exposure to free radicals and other oxygen-derived species provides
a usefulin vitro model to study anoxia-reoxygenation trauma at the cellular level. 相似文献
27.
James B. Hoying Carl A. Boswell Stuart K. Williams 《In vitro cellular & developmental biology. Animal》1996,32(7):409-419
Summary During angiogenesis, the microvasculature displays both vessel remodeling and expansion under the control of both cellular
and extracellular influences. We have evaluated the role of angiogenic and angiostatic molecules on angiogenesis in anin vitro model that more appropriately duplicates the cellular and extracellular components of this process. Freshly isolated microvessel
fragments from rat adipose tissue (RFMF) were cultured within three-dimensional collagen I gels. These fragments were characterized
at the time of isolation and were composed of vessel segments observed in the microvasculature of fatin situ (i.e., arterioles, venules, and capillaries). Fragments also exhibited characteristic ablumenally associated cells including
smooth muscle cells and pericytes. Finally, fragments were encased in an extracellular matrix composed of collagen type IV
and collagen type I/III. The elongation of microvascular elements was subsequently evaluated using morphologic and immunocytochemical
techniques. The proliferation, migration, and elongation of cellular elements in microvessel fragments from rat adipose tissue
was dependent on initial fragment density, matrix density, and required serum. Inclusion of endothelial cell growth factors
to microvessel fragments from rat adipose tissue 3-D cultures resulted in the accelerated elongation of tube structures and
the expression of von Willebrand factor in cells constituting these tubes. Molecules with reported angiostatic capacity (e.g.,
transforming growth factor and hydrocortisone) inhibited vessel tube elongation.
In vitro methods have been developed to evaluate numerous mechanisms associated with angiogenesis, including endothelial cell proliferation,
migration, and phenotypic modulation. Microvascular endothelial cell fragments described in this study represent anin vitro population of cells that accurately duplicate thein vivo microcirculatory elements of fat. The proliferation of cells and elongation of microvascular elements subsequently observed
in three-dimensional cultures provides anin vitro model of angiogenesis. Microvascular formation in this system results from pre-existing microvessel fragments unlike tube
formation observed when cultured endothelial cells are placed in three-dimensional gels. This form of tube formation from
cultured endothelium is more characteristic of vasculogenesis. Thus, the formation of microvascular elements from microvessel
fragments provides the opportunity to examine the mechanisms regulating angiogenesis in anin vitro system amenable to precise experimental manipulation. 相似文献
28.
The impact of air perfusion on the endothelial function of the rat mesenteric arterial bed (MAB; perfused with Krebs' bicarbonate plus indomethacin) was compared to that of the NO synthase inhibitor, Nω-nitro-L-arginine methyl ester (L-NAME). Air shifted the dose-response curve for the alpha-adrenoceptor agonist, norepinephrine (NE) to the left (ED50%: 2.9 ± 0.7 to 0.9 ± 0.7 μg, P < 0.05); maximal vasoconstriction did not change. L-NAME produced a similar increase in midrange sensitivity (ED50% 1.4 ± 0.7 μg, P < 0.05) and a 20% increase in maximum (152 ± 6 to 183 ± 7 mmHg, P < 0.05). Electromechanical stimulation with potassium chloride (KCL) was not modified by reserpine. Neither air nor L-NAME modified midrange sensitivity to KCL. L-NAME produced a 17% increase in maximum (91 ± 4 to 107 ± 5 mmHg, P < 0.05); reserpine abolished the latter effect. Air and L-NAME diminished endothelium-dependent vasodilation elicited by carbachol. Air did not modify endothelium-dependent vasodilation elicited by sodium nitroprusside; this response was potentiated by L-NAME. In summary, air and L-NAME produced similar effects on receptor-dependent activation of the endothelial L-arginine nitric oxide (NO) pathway. Potentiation by L-NAME of the maximal electromechanical response suggests the existence of a tone-dependent NO system. Abolition of the latter response by reserpine suggests that this system is of sympathetic origin. 相似文献
29.
30.
Brain Micro vessel 12-Hydroxyeicosatetraenoic Acid Is the (S) Enantiomer and Is Lipoxygenase Derived
Steven A. Moore Michael J. Giordano Hee-Yong Kim Norman Salem Jr. Arthur A. Spector 《Journal of neurochemistry》1991,57(3):922-929
12-Hydroxyeicosatetraenoic acid (12-HETE) production from arachidonic acid by cerebral microvessels isolated from perfused adult murine brain was reduced by the lipoxygenase inhibitors baicalein, esculetin, gossypol, nordihydroguaiaretic acid, and quercetin. Except for quercetin and gossypol, the IC50 did not exceed 10 microM. Each inhibitor, except baicalein, also decreased microvessel prostaglandin production when present in concentrations above their IC50 value for 12-HETE. In contrast, inhibitors of the cytochrome P450 monooxygenase system, clotrimazole, metyrapone, and proadifen (SKF-525A), had little effect on microvessel 12-HETE production. Chiral phase HPLC analysis revealed that only the (S) enantiomer of 12-HETE was formed. The major microvessel metabolite of eicosapentaenoic acid co-eluted with 12-hydroxyeicosapentaenoic acid (12-HEPE) on reverse-phase HPLC and the (S) enantiomer of 12-HEPE on chiral phase HPLC. Furthermore, like 12-HETE, 12-HEPE production was blocked by lipoxygenase inhibitors. These studies demonstrate that brain microvessels produce only the (S) enantiomeric 12-hydroxy derivatives of both arachidonic acid and eicosapentaenoic acid by the action of a lipoxygenase that can be selectively inhibited by baicalein. Since arachidonic acid and eicosapentaenoic acid are available to cerebral blood vessels in certain pathological settings, these 12-hydroxy acid lipoxygenase products may mediate some of the cerebrovascular dysfunction that occurs following stroke, brain trauma, or seizures. 相似文献