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1.
Electron-microscopic and immunocytochemical analyses of Weibel-Palade bodies in the human umbilical vein during pregnancy 总被引:3,自引:0,他引:3
Summary The present study was done to elucidate the biological significance of the Weibel-Palade body of human umbilical vein endothelial cells. Quantitative determinations of these endothelial-specific granules throughout pregnancy revealed that their numbers and size per cell profile were maintained at low levels from 12 to 19 weeks of gestation; then both rapidly increased from 33 weeks to full term. This increase coincided with the development of the rough endoplasmic reticulum and an increase in the number of endothelial cell pinocytotic vesicles. Light-microscopic peroxidase anti-peroxidase and electron-microscopic protein A-gold techniques provided evidence that factor VIII-related antigen was localized in the Weibel-Palade bodies. Furthermore, in vitro treatment of incubated umbilical vein tissue with compound 48/80, a histamine releaser, induced degranulation of Weibel-Palade bodies from the endothelium. The present study indicates that Weibel-Palade bodies are storage sites of both histamine and factor VIII-related antigen and have an important role in the obliteration of this vessel. 相似文献
2.
Tissue culture of human and canine thoracic duct endothelium 总被引:7,自引:0,他引:7
Gnepp Douglas R. Chandler Wayne 《In vitro cellular & developmental biology. Plant》1985,21(4):200-206
Summary Endothelial cells from the canine or human thoracic duct were harvested using 0.2% collagenase digestion and grown in Media
199, supplemented with fetal bovine serum. The canine endothelial cells grew to confluence (4.4 to 12×104 cells/cm2) in 6 to 10 d; doubling times ranged from 1.5 to 2.8 d. There was a minimum critical density for cell growth between 500
and 10 000 cells/cm2. The canine endothelial cells have been maintained in culture for periods up to 11 mo. The human thoracic duct endothelial
cells are more difficult to grow and maintain. Endothelial cells were isolated from 5 out of 35 human thoracic ducts and grew
for periods of up to 2 wk before degenerating. Both human and canine endothelial cells were Factor VIII positive. It has thus
been demonstrated that it is possible to grow canine and, less easily, human thoracic duct endothelium in tissue culture. 相似文献
3.
M. P. Carson I. Saenz de Tejada I. Goldstein C. C. Haudenschild 《In vitro cellular & developmental biology. Plant》1989,25(3):248-254
Summary A method for culturing endothelial cells (HCC-EC) from surgical specimens of human corpus cavernosum has been developed. The
approach involves selective endothelial outgrowth from explants and may be generally applicable to tissue whose endothelium
is not amenable to isolation by routine mechanical or enzymatic methods. The tissue is minced into pieces which are placed
onto gelatin-or fibronectin-coated tissue culture plastic, and grown in medium suitable for microvascular endothelial cell
growth (Carson and Haudenschild, In Vitro 22:344–354, 1986). By Days 5 to 7 EC colonies are found. Within a day or two after
the appearance of the EC colonies, a non-EC cell type appears and, if undisturbed, quickly overgrows the EC. An exploitable
temporal separation between the emergence of EC and non-EC is obtained when both conditioned medium (from bovine aortic endothelium)
and retinal extract are present during the outgrowth period. Explants are removed by pipetting at the first sign of the emergence
of the non-EC cell type. Once isolated, HCC-EC do not require conditioned medium but do require either retinal extract or
acidic fibroblast growth factor for survival and growth. Approximately 60% of the first passage cultures are at least 80%
EC as judged by DiI-Ac-LDL labeling. One corpus (0.3×0.3×0.5 cm) usually produces 120 cm2 of primary culture within 2 wk. These EC form contact-inhibited monolayers and stain positively for Factor VIII. They have
a doubling time at 6th passage of 48 h and a plateau density of 5 to 7×104 cells/cm2. The availability of such cultures should facilitate the study of endothelium-mediated responses which play an important
role in the erectile function of human penile corpus cavernosum.
Supported by NIH R01 HL23567-09, DK-39080-01, DK40025-01, DK40487-01. 相似文献
4.
Lymphatic endothelial and smooth-muscle cells in tissue culture 总被引:9,自引:0,他引:9
Summary Endothelial and smooth-muscle cells from bovine mesenteric lymphatic vessels have been collected and cultured in vitro. The
endothelial cells grew as a monolayer exhibiting a “cobblestone” appearance with individual cells tending to be more flattened
at confluence than their blood vascular counterparts. Approximately 30% of these cells expressed Factor VIII antigen compared
with bovine mesenteric artery or human umbilical-vein endothelium in which the majority of cells were positive. The lymphatic
smooth-muscle cells exhibited focal areas of multilayering and were Factor VIII negative. The availability of lymphatic endothelial
and smooth-muscle cells in culture will provide a new tool for the investigation of the biological properties of the lymphatic
vessels and their role in homeostasis.
Supported by the Medical Research Council of Canada, Grant MA-7925 相似文献
5.
Isolation of adult canine venous endothelium for tissue culture 总被引:6,自引:0,他引:6
John W. Ford William E. Burkel Raymond H. Kahn 《In vitro cellular & developmental biology. Plant》1981,17(1):44-50
Summary In order to provide autologous adult endothelial cells for the production of cell-lined artificial vascular prostheses, we
have developed a method for harvesting large numbers of cells with minimal contamination by other cellular types. In this
technique, the vein to be stripped is isolated, removed, and everted over a stainless steel rod. After washing, the vein is
incubated in trypsin-EDTA solution followed by collagenase and the endothelial cells flushed off with a stream of culture
medium. With care and appropriate timing, the endothelium can be selectively removed leaving the underlying basal lamina intact.
This research was supported by National Heart, Lung and Blood Institute Contract NIH-NO1-HV-2054 and Grant HL23345. 相似文献
6.
Summary Lymphatic endothelial cells grown long term in culture form lymphatic capillarylike tubes. Examination by light and transmission
electron microscopy showed that these structures were closed loops composed of one to several cells connected by intercellular
junction to form a luminal space. This first demonstration of lymphangiogenesis in confluent monolayer cultures of lymphatic
endothelial cells (a) showed that collagen type I accelerated lymphatic capillary tube formation, whereas fibronectin and
matrigel had no effect; b) provided a model to study lymphatic endothelial cell function and differentiation; and c) offered
a possibility to distinguish differences between the process of lymphangiogenesis and angiogenesis by testing various factors
and conditions that effect endothelial cell behavior. 相似文献
7.
Summary A long-term culture of a spontaneously transformed endothelial cell line derived from the choroid-retina of a rhesus macaque
fetus is reported. It has been carried in vitro by serial propagation more than 548 passages in 17 yr. Cells were identified
as being of endothelial origin by cellular morphology, growth pattern, ultrastructure, immunocytochemistry (immunofluorescence
and immunoperoxidase), and immunodiffusion. The transformants are characterized by (a) an infinite life span, (b) a changed
expression of Factor VIII-related antigen, and (c) chromosomal aberrations. Throughout long-term serial passages and after
repeated freeze-storage, thawing, and reculture the cells retain the specific organelles, Weibel-Palade bodies, and most of
the other characteristic morphologic features. For this long-term cultured endothelial cell line, Weibel-Palade bodies seem
to be a more reliable marker than Factor VIII-related antigen.
This is publication No. 1475 of the Oregon Regional Primate Research Center. Research was supported by grant RR-00163 from
Animal Resources Branch, Division of Research Resources, National Institutes of Health; by NIH grant EY02086, basic research
support grant from the Oregon Regional Primate Research Center, and the Collins Medical Trust. 相似文献
8.
LPS-induced IL-6, IL-8, VCAM-1, and ICAM-1 expression in human lymphatic endothelium. 总被引:2,自引:0,他引:2
Yoshihiko Sawa Takeshi Ueki Minoru Hata Kana Iwasawa Eichi Tsuruga Hiroshi Kojima Hiroyuki Ishikawa Shigemitsu Yoshida 《The journal of histochemistry and cytochemistry》2008,56(2):97-109
We have previously reported the TLR4 expression in human intestinal lymphatic vessels. In the study here, microarray analysis showed the expression of the TLR4, MD-2, CD14, MyD88, TIRAP, TRAM, IRAK1, and TRAF6 genes in cultured human neonatal dermal lymphatic microvascular endothelial cells (LEC). The microarray analysis also showed that LEC expressed genes of IL-6, IL-8, VCAM-1, and ICAM-1, and the real-time quantitative PCR analysis showed that mRNA production was increased by lipopolysaccharide (LPS). The LPS-induced IL-6, IL-8, VCAM-1, and ICAM-1 production in LEC was suppressed by the introduction of TLR4-specific small interfering RNA, and also by anti-TLR4, nobiletin, and CAPE pretreatment. These findings suggest that LEC has TLR4-mediated LPS recognition mechanisms that involve at least activation of NF-kappaB, resulting in increased expression of IL-6, IL-8, VCAM-1, and ICAM-1. Both the LPS effect on the gene expression and also the suppression by nobiletin and CAPE pretreatment on the protein production were larger in IL-6 and in VCAM-1 than in IL-8 and in ICAM-1 in LEC. The signal transduction of NF-kappaB and AP-1-dependent pathway may be more critical for the expression of IL-6 and VCAM-1 than that of IL-8 and ICAM-1 in LEC. 相似文献
9.
10.
Substance P and gastrin releasing peptide in bovine mesenteric lymphatic vessels: chemical characterization and action 总被引:1,自引:0,他引:1
Alcoholic extracts of bovine mesenteric lymphatic vessels were assayed for the presence of SP, GRP, VIP, PHI, GIP and NT using specific radioimmunoassays. SP and GRP immunoreactivities were detected at concentrations of 190 +/- 20 and 1,000 +/- 130 pg.g-1, respectively. No significant levels of immunoreactivity were detected for any of the other peptides. SP and GRP immunoreactivities coeluted with their synthetic counterparts from both Sephadex G-50 and reversed phase HPLC columns. Synthetic SP (10(-9)-10(-7) M) and the naturally occurring analogue of GRP, bombesin (10(-9)-10(-7) M), increased spontaneous contraction rate in isolated vessel segments. This excitatory effect was not blocked by the alpha-adrenoceptor antagonist phentolamine (3 x 10(-6) M). 相似文献
11.
12.
Glenn K. Sherer Timonthy P. Fitzharris W. Page Faulk E. Carwile LeRoy 《In vitro cellular & developmental biology. Plant》1980,16(8):675-684
Summary A procedure is described for the isolation and cultivation of microvascular endothelium from human skin. Neonatal foreskins
are pooled, washed, minced, and dissociated by a mixture of collagenase and dispase. Microvascular endothelium, liberated
in the form of intact capillary fragments, is incompletely separated from fibroblasts and epidermal cells by sieving through
nylon mesh, followed by velocity sedimentation on 5% bovine serum albumin. The endothelium-enriched fraction has been maintained
in primary culture for up to 3 weeks. The resulting epithelioid colonies have been characterized morphologically by both light
and transmission electron microscopy and manifest all of the structural features that distinguish other, large-vessel endothelia
in culture. In addition, immunohistochemical studies using an indirect fluorescent antibody technique demonstrate that these
cells contain the endothelium-specific product, Factor VIII antigen.
This work was supported by National Institutes of Health Grants AM18904 and AM20571, the RGK Foundation, the Charlotte and
Sidney Lifschultz Foundation, the Juvenile Diabetes Foundation, and the South Carolina Geenral Medical Faculty Research Appropriation. 相似文献
13.
Lawrence E. DeBault Larry E. Kahn Stephen P. Frommes Pasquale A. Cancilla 《In vitro cellular & developmental biology. Plant》1979,15(7):473-487
Summary Microvessels isolated from mouse forebrain were used as the source material for the derivation of cerebral vascular endothelium
and smooth-muscle cells in culture. The microvessels were isolated by a mechanical dispersion and filtration technique, and
were maintained in vitro as organoid cultures. A microvessel classification system was developed and proved to be useful as
a tool in monitoring culture progress and in predicting the type(s) of microvessel(s) that would give rise to migrating and/or
proliferating cells. The isolated cerebral microvessels were heterogeneous in diameter, size of individual vascular isolate,
and proliferative potential. The isolated microvessels ranged in diameter from 4 μm to 25 μm and in size from a single microvascular
segment to a large multibranched plexus with mural cells. The initial viability, determined by erythrosin B exclusion, was
approximately 50% on a per cell basis. All microvessel classes had proliferative potential although the rate and extent of
proliferation were both microvessel class- and density-dependent. The smaller microvessels gave rise to endothelial cells,
whereas the large microvessels gave rise to endothelial and smooth-muscle cells. The viability and progress of a microvessel
toward derived cell proliferation seemed to be directly proportional to the number of mural cells present.
This work was supported in part by an Arteriosclerosis Specialized center of Research grant from the National Heart, Lung
and Blood Institute, National Institutes of Health (HL-14230) and Grant 584-127703 from the Veterans Administration. 相似文献
14.
Microvascular endothelium and pericytes: High yield,low passage cultures 总被引:17,自引:0,他引:17
Mary Pat Carson Christian C. Haudenschild 《In vitro cellular & developmental biology. Plant》1986,22(6):344-354
Summary Cultured microvascular endothelial cells (MEC) have become a valuable model for studies of microvascular physiology and pathology.
Most current techniques involve manual removal of undesirable cell types or cloning, require one to several months, and yield
high population doubling level cultures derived from a relatively small sample of the original population. We have devised
a technique to more rapidly produce larger numbers of MEC. This method provided primary cultures consisting predominantly
of MEC within 1 wk. The technique involves selective aspiration of gray matter from the bovine cerebral cortex followed by
homogenization, sieving, enzymatic dissociation, and then dense plating (104 to 105 vessel fragments/cm2) onto gelatin- or fibronectin-coated plastic. Typical yields were 0.1 to 0.5 × 106 fragments/g of aspirated gray matter. The optimal culture medium for these cells was 15% equine plasma derived serum, 20%
conditioned medium, 2% retinal extract, 60% fresh medium, and 500 μg/ml heparin. Cells attached within 24 h, well-spread colonies
were present within 1 to 2 d, and cultures approached confluence within 2 to 3 d. Alkaline phosphatase staining confirmed
the microvascular origin of the material plated. Morphology, Factor VIII-related antigen staining and 1,1′-dioctacecyl-3,3,3′3,-tetramethyl-indocarbocyanine
perchlorate acetylated low density lipoprotein uptake suggested that MEC predominated. Cultures could be passaged and additionally
purified by sequential exposure to pancreatin and trypsin-EDTA. Pancreatin selectively removed MEC colonies leaving a relatively
homogeneous pericyte population. The relative ease with which such cultures can be produced should facilitate the in vitro
study of brain microvascular function and may also provide insights useful for growing MEC from other vascular beds.
This work was supported by grants from the Hubert H. Humphrey Cancer Research Center, Boston University School of Medicine
(American Cancer Soc. IN97G) and from the National Institutes of Health (HL26895 and HL07224), Bethesda, MD. 相似文献
15.
Selection and characterization of bovine aortic endothelial cells 总被引:17,自引:0,他引:17
Stephen M. Schwartz 《In vitro cellular & developmental biology. Plant》1978,14(12):966-980
Summary This paper reports techniques for isolation, selection and long-term passage of bovine aortic endothelium (BAE). A [3H]thymidine-selection technique was developed to limit overgrowth of cultures by contaminating smooth-muscle cells. The resulting
cultures could be passaged for a replicative life span of 35 to 40 doublings and maintained a stable, normal karyotype throughout
this period. Despite the fact that these cultures reached a stable monolayer with density-inhibited growth state, postconfluent
cells showed focal areas of a second growth pattern called “sprouting.” This was seen only when cultures were maintained at
high densities for periods of 1 to 2 weeks. Ultrastructural analysis, as well as immunofluorescence studies with markers for
endothelial cells (factor VIII) and smooth-muscle cells (actin), indicates that this phenomenon is not due to overgrowth of
a residual population of smooth-muscle cells, but may represent a second growth pattern of the endothelial cells themselves.
This research was supported by NIH Grant HL 18645. This work was done during the tenure of an Established Investigationship
of the American Heart Association. 相似文献
16.
G. Azzali 《Cell and tissue research》1990,262(1):191-193
Summary The passage of cells across the lymphatic endothelium of rat lacteals in both normal and non-pathological experimental conditions (fasting, lymphatic stasis) was studied by means of serial thin sections and three-dimensional models. Two different pathways of transendothelial migration were observed: (1) macrophages enter the lymphatic lumen via the cytoplasm of endothelial cells, without involvement of intercellular junctions, whereas (2) lymphocytes migrate through intraendothelial channels, dynamic structures organized by the lymphatic endothelium under physiological conditions. 相似文献
17.
18.
Johanna Plendl Christine Neumüller Fred Sinowatz 《Biology of the cell / under the auspices of the European Cell Biology Organization》1996,87(3):179-188
Summary— The purpose of the present study was to investigate potential modulations of endothelial cells of the bovine corpus luteum (CL) during pregnancy. Luteal endothelia of pregnant and non-pregnant cows were isolated and purity of cultures was verified by flow cytometric quantification of three independent endothelial markers (von Willebrand factor, angiotensin converting enzyme, Bandeiraea simplicifolia agglutinin I ligands). Different cellular parameters including light and electron microscopical investigation of morphology and growth characteristics as well as quantification of cellular lectin binding sites were compared. Extensive heterogeneity between luteal endothelial cells in pregnant and non-pregnant animals could be demonstrated, reflected in functional attributes like angiogenic activity, ultrastructural characteristics and the quantitative expression of cellular carbohydrates. Two different morphological types of cells (‘cobblestone growth pattern’ and ‘arcuate growth pattern’) were isolated from the CL of pregnancy as well as from the cyclic CL. Spontaneous angiogenic activities, including cellular migration in band-like structures and formation of ring-like structures, were observed in endothelial cells isolated from the CL of pregnant cows exclusively. This strongly suggests that microvascular luteal endothelium of pregnant animals, in contrast to the one of non-pregnant animals, is able to produce quantitatively and/or qualitatively specific angiogenesis factor(s). Heterogeneity between luteal endothelial cells in the pregnant and non-pregnant animal could also be demonstrated by quantification of lectin (Bandeiraea simplicifolia agglutinin I, concanavalin A, Dolichos biflorus agglutinin, Ulex europaeus agglutinin I, wheat germ agglutinin) binding sites: quantitative expression of specific endothelial cell surface carbohydrates could be correlated to be status of pregnancy, thus emphasizing the actual need of quantification of lectin binding. 相似文献
19.
Ingeburg E. Goetz Jean Warren Carmen Estrada Eugene Roberts Diana N. Krause 《In vitro cellular & developmental biology. Plant》1985,21(3):172-180
Summary Cerebrovascular endothelial cells from adult bovine brain were carried successfully in long-term, serial culture. Endothelial
cells were obtained from the middle and anterior cerebral arteries and from capillaries isolated from grey matter of the cerebral
cortex or caudate nucleus. Capillary cells were found to grow best in RPMI 1640 with 20% fetal bovine serum. They did not
require tumor-conditioned medium or matrix-coated surfaces, although fibronectin was used to enhance the initial plating efficiency
of the primary cultures. The same conditions were used to support satisfactory growth of arterial endothelial cells; however
they did not grow as rapidly as the capillary cells. Retention of endothelial-specific characteristics were shown for capillary-derived
cells carried up to Passage 28, arterial-derived cells up to Passage 11, and after frozen storage of both types of cultured
cells. Cultures of both arterial and capillary cells stained positively for Factor VIII antigen, exhibited a nonthrombogenic
surface, and produced prostacyclin in response to arachidonic acid. Arterial endothelial cells produced more prostacyclin
than capillary endothelium. The capillary cells had a unique tendency to assume a ringlike morphology after subculture and
sometimes formed capillarylike networks of cell cords in dense cultures. When cultured in a three-dimensional plasma clot,
capillary and arterial endothelial cells, but none of the other cell types studied, organized into tubelike structures reminiscent
of capillary formation in vivo. The availability of long-term cultures of cerebrovascular endothelial cells provides an opportunity
to compare properties of arterial and capillary endothelium from the same tissue and to investigate such processes as angiogenesis
and blood-brain barrier induction.
This work was supported in part by Public Health Service grant NS-18586 from the National Institute of Neurological and Communicative
Disorders and Stroke and by a grant from The Council for Tobacco Research—U.S.A., Inc. C. E. was supported by the Universidad
Autonoma of Madrid and the Ministerio de Universidades e Investigacion of Spain. 相似文献