共查询到20条相似文献,搜索用时 31 毫秒
1.
Zisch AH 《Current opinion in biotechnology》2004,15(5):424-429
Adult bone marrow and peripheral blood contain small subsets of mononuclear cells that can be differentiated into endothelial-like cells in vitro. Experimental and clinical transplantation of such cell isolates--often referred to as endothelial stem/progenitor cells--into ischaemic or infarcted areas shows their incorporation into sites of new vessel growth along with improvement of regional blood flow. Emerging evidence suggests that these beneficial effects on vascular growth can be attributed to the paracrine activation of resident endothelial cells, rather than their integration into new endothelium. Autologous endothelial progenitor cells can also substitute for native vessel-derived endothelial cells in tissue-engineered vascular autografts. 相似文献
2.
Using cultured human umbilical cord vein endothelial cells and human blood neutrophils, the interaction between neutrophils and endothelial cells, in vitro, was studied. The aim of the study was to examine whether a respiratory burst stimulation by neutrophils would be observed by neutrophil/endothelial cell interaction and whether the respiratory burst stimulation of neutrophils by endothelial cells could be enhanced by lipopolysaccharide stimulation of neutrophils. The second aim was whether such an effect, or secretion of elastase, could cause an endothelial cell damage in vitro. Chemiluminescence as an indicator of oxygen-derived metabolites produced by neutrophils, elastase release by neutrophils, and endothelial cell damage, based on111 In-oxine release from labelled endothelial cells, were measured simultaneously. The present investigation demonstrates that neutrophils can be directly stimulated by endothelial cells. A further amplification of this process following lipopolysaccharide priming up to 10 ng/ml blood could be demonstrated. A slight endothelial cell damage occurs following neutrophil stimulation, although elastase secretion does not increase during interaction between neutrophils and endothelial cells. These results raise the possibility that oxygen-derived metabolites rather than elastase contribute to an endothelial cell damage which might occur in conditions such as endotoxin-induced adult respiratory distress syndrome. 相似文献
3.
An in vitro experimental system was developed to study the interaction between endothelial cells and blood as an early event in coagulation. A designed vascular vessel model tube is composed of a monolayer of bovine aorta endothelial cells (BAECs) cultured on an inner surface of a glass tube by means of a rotatory cultivation method. The change of fluidity during coagulation of blood in the tube was measured by a rheological technique. The rate of coagulation of blood in contact with endothelial cells was affected by cell culture conditions such as cell age, passage number of BAECs and substrate beneath endothelial cells. Fibrinolytic activity of the cells was examined by the rheological method. The present experimental system would be useful in examining the mechanism of blood coagulation based on the interaction between blood and endothelial cells as well as in evaluating endothelial cell functions. 相似文献
4.
J Martin-Ramirez M Hofman M van den Biggelaar RP Hebbel J Voorberg 《Nature protocols》2012,7(9):1709-1715
Blood outgrowth endothelial cells (BOECs) are important tools when investigating diagnostic and therapeutic approaches for vascular disease. In this protocol, mononuclear cells are isolated from peripheral blood and plated on type I collagen at ~135,000 cells per cm(2) in endothelial cell differentiation medium. On average, 0.34 colonies of endothelial cells per milliliter of blood can be obtained. Colonies of endothelial cells become visible after 14-28 d. Upon confluence, these rapidly expanding colonies can be passaged and have been shown to propagate up to 10(18)-fold. Isolated BOECs are phenotypically similar to vascular endothelial cells, as revealed by their cobblestone morphology, the presence of endothelial cell-specific Weibel-Palade bodies and the expression of endothelial cell markers such as VE-cadherin. The protocol presented here also provides a particularly useful tool for the ex vivo assessment of endothelial cell function from patients with different vascular abnormalities. 相似文献
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6.
One of the cardinal processes of inflammation is the infiltration of immune cells from the lumen of the blood vessel to the surrounding tissue. This occurs when endothelial cells, which line blood vessels, become adhesive to circulating immune cells such as monocytes. In vitro measurement of this adhesiveness has until now been done by quantifying the total number of monocytes that adhere to an endothelial layer either as a direct count or by indirect measurement of the fluorescence of adherent monocytes. While such measurements do indicate the average adhesiveness of the endothelial cell population, they are confounded by a number of factors, such as cell number, and do not reveal the proportion of endothelial cells that are actually adhesive. Here we describe and demonstrate a method which allows the enumeration of adhesive cells within a tested population of endothelial monolayer. Endothelial cells are grown on glass coverslips and following desired treatment are challenged with monocytes (that may be fluorescently labeled). After incubation, a rinsing procedure, involving multiple rounds of immersion and draining, the cells are fixed. Adhesive endothelial cells, which are surrounded by monocytes are readily identified and enumerated, giving an adhesion index that reveals the actual proportion of endothelial cells within the population that are adhesive. 相似文献
7.
Koizumi K Tsutsumi Y Kamada H Yoshioka Y Watanabe M Yamamoto Y Okamoto T Mukai Y Nakagawa S Tani Y Mayumi T 《Biochemical and biophysical research communications》2003,306(1):219-224
In this study, we attempted to assess the incorporable potential of vascular endothelial cells derived from adult organ blood vessels into tumor blood vessels. Two kinds of adult organ-derived vascular endothelial cells, human aorta endothelial cells (HAEC) and umbilical vein endothelial cells (HUVEC), were administered into murine tumors inoculated to SCID mice. Many human blood vessel networks were visualized in the murine tumors. These cells in solid tumor not only survived and proliferated, but also incorporated into tumor endothelium. These results suggest that adult organ-derived vascular endothelial cells possess the potential to form the neovascular network in various tissues such as vascular endothelial progenitor-like cells in vivo. We propose that these cells can be regarded as a congenic (autologous) vector for vascular regeneration cell therapy and tumor vascular targeting gene therapy. 相似文献
8.
A microcarrier culture system was established for a large-scale production of functional human endothelial cells. It has been difficult to cultivate human endothelial cells in large quantities for the reasons that specific growth factor and extracellular matrix are required for the survival and proliferation of the cells and the life span of the primary cells are limited. A lot of studies have reported that the shear stress gives significant influences on the structure, growth rate and biological functions of endothelial cells. We aimed to develop a convenient microcarrier culture system for human endothelial cells which can reproduce the flow effects experienced in vivo or in vitro. In 200 mL volume culture, human umbilical vein endothelial cells (HUVEC) could be serially sub-cultivated by optimizing the culture conditions such as shear strength, growth factor, beads and seeding cell concentration, serum concentration, and passage timing. The growth rate was enhanced depending on the shear strength and the life span of the cells was elongated until over 43PDL which is much longer than those of monolayer cultures. The cells maintained the diploidy of over 80% without obvious abnormal changes in the chromosomes. The serially sub-cultured microcarrier cells maintained various endothelial cell functions such as the syntheses of von Willebrand factor (vWf), prostacyclin and other biological substances, the expression of CD31, and the VEGF(165) dependent growth characteristic. The synthesis of biological products was affected by shear strength. In the case of prostacyclin, a different synthesis response was observed between steady flow and transiently reduced shear strength. The synthesis of endothelin-1 (ET-1) was down-regulated by increase of shear strength different from those of other products. The culture system was scaled up until 2 L volume under the optimum DO control. The cells synthesized IL-6 in response to shear strength. These results indicate that the established microcarrier system might be able to contribute to the supply of functional human endothelial cells for various medical applications such as the reconstruction of injured blood vessels caused by atherosclerosis or restenosis of coronary arteries after angioplasty, and the construction of an anti-coagulable artificial blood vessel or an artificial skin with good transplant-ability. 相似文献
9.
Endothelial cells in the intact adult are, apart from those in the female reproductive organs, believed to be quiescent. Systematic examination of endothelial cell proliferation in male reproductive organs has not been performed and was therefore the aim of the present study. Intact adult rats were either pulse labeled or long-term labeled with bromodeoxyuridine to label proliferating cells. The roles of Leydig cells and testosterone were examined after castration or treatment with the Leydig cell toxin ethane dimethane sulfonate (EDS) and testosterone substitution. After perfusion fixation, all blood vessels remained open and were easily identified. In all male reproductive organs studied, particularly in the testis and epididymis, endothelial cell proliferation was considerably higher than in other tissues such as the liver, brain, and muscle. Proliferating endothelial cells were observed in all types of blood vessels in male reproductive organs, but other characteristics of new blood vessel formation were not seen. High endothelial cell proliferation may reflect a continuous high turnover of endothelial cells rather than classical angiogenesis. In the epididymis, the ventral and dorsolateral prostate lobes, and the seminal vesicles, endothelial cell proliferation decreased after testosterone withdrawal and increased following testosterone treatment. In the testis, endothelial cell proliferation was decreased after Leydig cell depletion but remained low after testosterone substitution. High, hormonally regulated endothelial cell proliferation is not unique to the female but is also seen in the male reproductive organs. 相似文献
10.
Leah N. Cueni 《Experimental cell research》2009,315(10):1715-1187
Podoplanin is a small, mucin-like membrane glycoprotein highly expressed by lymphatic but not by blood vascular endothelial cells. Although it was shown to be indispensable for the correct formation and function of the lymphatic vasculature, its precise molecular function has remained unknown. In the present study, we identified the mammalian lectin galectin-8 as a novel, glycosylation-dependent interaction partner of podoplanin. Galectin-8 is a tandem-repeat type galectin, which interacts with cell surface glycoproteins, including certain integrins, as well as with extracellular matrix molecules such as fibronectin. Here we show that, similar to podoplanin, galectin-8 is more highly expressed by lymphatic than by blood vascular endothelial cells, and that it promotes lymphatic endothelial cell adhesion as well as haptotactic migration when immobilized onto a surface, while inhibiting the formation of tube-like structures by lymphatic endothelial cells in a collagen matrix when incorporated into the matrix. Importantly, functions of blood vascular endothelial cells, which lack podoplanin expression, are not affected by galectin-8. These data suggest a role for galectin-8 and podoplanin in supporting the connection of the lymphatic endothelium to the surrounding extracellular matrix, most likely in cooperation with other glycoproteins on the surface of lymphatic endothelial cells. 相似文献
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The purpose of these experiments was to evaluate the expression of endothelial markers, such as Tie2 and VEGFR2 in endothelial cells derived from blood mononuclear endothelial progenitor cells. Bovine mononuclear cells were isolated using separation by centrifugation and were grown in endothelial specific media supplemented with growth factors. Isolation of the whole cell population of mononuclear cells (MNC) from bovine peripheral blood gave rise to progenitor-like cells (CD45−) that, although morphologically similar, have different phenotypes revealed by expression of endothelial specific markers Tie2 and VEGFR2. Plating of MNCs on collagen and fibronectin gave rise to more colonies than non-coated dishes. Occasional colonies from MNC isolations had a mural cell phenotype, negative for Tie2 and VEGFR2 but positive for smooth muscle actin and PDGFRβ. Although cells expressing high levels of VEGFR2 and low levels of Tie2, and vice versa were both able to form cords on Matrigel, cells with higher expression of Tie2 migrate faster in a scratch assay than ones with lower expression of Tie2. When these different clones of cells were introduced in mice through tail vein injections, they retained an ability to home to angiogenesis occurring in a subcutaneous Matrigel plug, regardless of their Tie2/VEGFR2 receptor expression patterns, but cells with high VEGFR2/low Tie2 were more likely to be CD31 positive. Therefore, we suggest that active sites of angiogenesis (such as wounds, tumors, etc.) can attract a variety of endothelial cell precursors that may differentially express Tie2 and VEGFR2 receptors, and thus affect our interpretation of EPCs as biomarkers or therapies for vascular disease. 相似文献
13.
Embryonic data and ultrastructural analyses suggest that the primitive endothelium signals undifferentiated mesenchymal cells to migrate to the forming blood vessel and subsequently regulates mural cell growth and behavior. Upon maturation of the blood vessel, chemotactic and mitogenic signals are apparently diminished and differentiated smooth muscle cells normally remain quiescent. This homeostasis is seemingly upset in conditions which lead to pathologies characterized by smooth muscle cell hyperplasia such as atherosclerosis. By culturing endothelial cells at different densities, we attempted to re-create the various stages of vascular development. Whereas media conditioned by sparse endothelial cells stimulate smooth muscle cells, media conditioned by dense endothelial cell cultures are inhibitory. Culture of sparse smooth muscle cells in media conditioned for 3 days by postconfluent endothelial cell cultures leads to dose-dependent and reversible smooth muscle cell inhibition. Furthermore, in the presence of the endothelial cell-derived inhibitor, smooth muscle cells are rendered refractory to mitogens such as fibroblast growth factor and platelet-derived growth factor. The inhibitory activity is not attributable to the well-characterized inhibitors of smooth muscle cell growth, transforming growth factor type-β, prostaglandin I2, or heparan sulfate proteoglycan. Partial characterization of the inhibitory conditioned media suggests that the active molecule is smaller than 1,000 da, and stable to boiling as well as proteinase K and heparinase digestion. These findings support the concept that there is intercellular communication between endothelial cells and smooth muscle cells and provide evidence for a novel endothelial cell-derived smooth muscle cell growth inhibitor. 相似文献
14.
Peter I. Lelkes Esther Ramos Victor V. Nikolaychik Dawn M. Wankowski Brian R. Unsworth Thomas J. Goodwin 《In vitro cellular & developmental biology. Animal》1997,33(5):344-351
Summary The aim of this study was to test the versatility of a new basal cell culture medium, GTSF-2. In addition to traditional growth-factors,
GTSF-2 contains a blend of three sugars (glucose, galactose, and fructose) at their physiological levels. For these studies,
we isolated normal endothelial cells from human, bovine, and rat large blood vessels and microvessels. In addition, GTSF-2
was also tested as a replacement for high-glucose-containing medium for PC12 pheochromocytoma cells and for other, transformed
cell lines. The cell growth characteristics were assessed with a novel cell viability and proliferation assay, which is based
on the bioreduction of the fluorescent dye, Alamar Blue. After appropriate calibration, the Alamar Blue assay was found to
be equivalent to established cell proliferation assays. Alamar Blue offers the advantage that cell proliferation can be measured
in the same wells over an extended period of time. For some of the cell types (e.g., endothelial cells isolated from the bovine
aorta, the rat adrenal medulla, or the transformed cells), proliferation in unmodified GTSF-2 was equivalent to that in the
original culture media. For others cell types (e.g., human umbilical vein endothelial cells and PC12 cells), GTSF-2 proved
to be a superior growth medium, when supplemented with simple additives, such as endothelial cell growth supplement (bFGF)
or horse serum. Our results suggest that GTSF-2 is a versatile basal medium that will be useful for studying organ-specific
differentiation in heterotypic coculture studies. 相似文献
15.
Vascular endothelial cells are located at the innermost layer of the blood vessel wall and are always exposed to three different mechanical forces: shear stress due to blood flow, hydrostatic pressure due to blood pressure and cyclic stretch due to vessel deformation. It is well known that endothelial cells respond to these mechanical forces and change their shapes, cytoskeletal structures and functions. In this review, we would like to mainly focus on the effects of shear stress and hydrostatic pressure on endothelial cell morphology. After applying fluid shear stress, cultured endothelial cells show marked elongation and orientation in the flow direction. In addition, thick stress fibers of actin filaments appear and align along the cell long axis. Thus, endothelial cell morphology is closely related to the cytoskeletal structure. Further, the dynamic course of the morphological changes is shown and the related events such as changes in mechanical stiffness and functions are also summarized. When endothelial cells were exposed to hydrostatic pressure, they exhibited a marked elongation and orientation in a random direction, together with development of centrally located, thick stress fibers. Pressured endothelial cells also exhibited a multilayered structure with less expression of VE-cadherin unlike under control conditions. Simultaneous loading of hydrostatic pressure and shear stress inhibited endothelial cell multilayering and induced elongation and orientation of endothelial cells with well-developed VE-cadherin in a monolayer, which suggests that for a better understanding of vascular endothelial cell responses one has to take into consideration the combination of the different mechanical forces such as exist under in vivo mechanical conditions. 相似文献
16.
Sickle red blood cells display an abnormal propensity to adhere to cultured bovine aortic endothelial cells when compared to normal red blood cells. The adherence was potentiated three-fold by endothelial cell derived conditioned medium, enriched in multimers of von Willebrand factor. Such adherence was ablated by 80% by either the synthetic peptide (RGDS) or antibody to GPIIb/IIIa, indicating the presence of RGD peptide recognition domain/receptor in either endothelial cells or sickle cells or both. The adherence was also inhibited by 70% by phosphatidylserine, but not by other phospholipids, indicating the presence of putative receptors for this phospholipid in endothelial cells. The labeling of cultured bovine aortic endothelial cells with monoclonal antibodies revealed the localization of MAB D2 to regions of cell-cell contact. The antigen on endothelial cells which cross-reacts with this antibody has a Mr of 130,000. The addition of such an antibody during the plating of endothelial cells disrupted monolayer formation. It appears that a 130-kDa polypeptide antigen in endothelial cells which is recognized by MAB D2, may be a cell-cell adhesion molecule. 相似文献
17.
Activation of endothelial cells via antibody-enhanced dengue virus infection of peripheral blood monocytes. 总被引:12,自引:6,他引:6
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Although endothelial cells have been speculated to be a target in the pathogenesis of dengue hemorrhagic fever (DHF), there has been little evidence linking dengue virus infection to any alteration in endothelial cell function. In this study, we show that human umbilical vein endothelial cells become activated when exposed to culture fluids from dengue virus-infected peripheral blood monocytes. Maximum activation was achieved with culture fluids from monocytes in which virus infection was enhanced by the addition of dengue virus-immune serum, thus correlating with epidemiological evidence that prior immunity to dengue virus is a major risk factor for DHF. Activation was strongest for endothelial cell expression of VCAM-1 and ICAM-1. In contrast, activation of endothelial cell E-selectin expression appeared to be more transient, as indicated by its detection at 3 h, but not at 16 h, of treatment. Treatment of monocyte culture fluids with anti-tumor necrosis factor alpha (TNF-alpha) antibody largely abolished the activation effect (as measured by endothelial cell expression of ICAM-1), whereas treatment with IL-1beta receptor antagonist had a much smaller inhibitory effect on activation. Endothelial cells inoculated directly with dengue virus or with virus-antibody combinations were poorly infectable (compared to Vero cells or peripheral blood monocytes), and virus-inoculated endothelial cells showed no increased expression of VCAM-1, ICAM-1, or E-selectin. Taken together, the results strongly indicate that dengue virus can modulate endothelial cell function by an indirect route, in which a key intermediary is TNF-alpha released from virus-infected monocytes. 相似文献
18.
微血管内皮细胞层是一层半选择通透性屏障,可以调节血液中的液体、溶质和血浆蛋白进入组织间隙。在炎症刺激作用下,可通过旁细胞途径和跨细胞途径引起内皮通透性上升。旁细胞通路主要由内皮细胞间的紧密连接、黏附连接和细胞与外基质的黏着斑组成。炎症介质,如脂多糖和肿瘤坏死因子α可激活多种蛋白激酶。活化的蛋白激酶主要包括Rho相关的卷曲蛋白激酶、肌球蛋白轻链激酶、蛋白激酶C、酪氨酸激酶和丝裂原活化蛋白激酶等,参与引发内皮屏障生化和结构改变,旁细胞通路开放,导致通透性上升。该文对上述蛋白激酶在微血管通透性中作用机制的研究进展进行综述。 相似文献
19.
Angiogenesis is the process by which new blood vessels arise from the pre-existing vasculature. Human endothelial cells are known to be involved in three key cellular processes during angiogenesis: increased cell proliferation, degradation of the extracellular matrix during cell migration, and the survival of apoptosis. The above processes depend upon the presence of growth factors, such as vascular endothelial growth factor isoform 165 (VEGF(165)) that is released from the extracellular matrix as it is being degraded or secreted from activated endothelial cells. Thus, the goal of the current study is to develop a system with a backbone of polyethylene glycol (PEG) and grafted angiogenic signals to compare the initial angiogenic response of human umbilical vein endothelial cells (hUVEC) or human microvascular endothelial cells (hMEC). Adhesion ligands (PEG-RGDS) for cell attachment and PEG-modified VEGF(165) (PEG-VEGF(165)) are grafted into the hydrogels to encourage the angiogenic response. Our data suggest that our biomimetic system is equally effective in stimulating proliferation, migration, and survival of apoptosis in hMEC as compared to the response to hUVEC. 相似文献
20.
Morphogenesis of the developing vascular network requires coordinated regulation of an extensive array of endothelial cell behaviors. Precisely regulated signaling molecules such as vascular endothelial growth factor (VEGF) direct some of these endothelial behaviors. Newly forming blood vessels also become subjected to novel biomechanical forces upon initiation of cardiac contractions. We report here the identification of a recessive mouse mutation termed shrunken-head (shru) that disrupts function of the Titin gene. Titin was found to be required for the initiation of proper heart contractions as well as for maintaining the correct overall shape and orientation of individual cardiomyocytes. Cardiac dysfunction in shrunken-head mutant embryos provided an opportunity to study the effects of lack of blood circulation on the morphogenesis of endothelial cells. Without blood flow, differentiating endothelial cells display defects in their shapes and patterns of cell-cell contact. These endothelial cells, without exposure to blood circulation, have an abnormal distribution within vasculogenic vessels. Further effects of absent blood flow include abnormal spatial regulation of angiogenesis and elevated VEGF signaling. The shrunken-head mutation has provided an in vivo model to precisely define the roles of circulation on cellular and network aspects of vascular morphogenesis. 相似文献