A comparison of primary cultures of rat cerebral microvascular endothelial cells to rat aortic endothelial cells

1991 ◽  
Vol 27 (4) ◽  
pp. 312-326 ◽  
Author(s):  
Ellen L. Gordon ◽  
Per E. Danielsson ◽  
Thien-Son Nguyen ◽  
H. Richard Winn
Keyword(s):  
Endothelial Cells ◽  
Primary Cultures ◽  
Microvascular Endothelial Cells ◽  
Aortic Endothelial Cells ◽  
Microvascular Endothelial ◽  
Aortic Endothelial

Production of Platelet-Activating Factor by Porcine Brain Microvascular Endothelial Cells in Culture

1995 ◽  
Vol 74 (05) ◽  
pp. 1335-1339 ◽  
Author(s):  
Kei Satoh ◽  
Hidemi Yoshida ◽  
Tada-Atsu Imalzumi ◽  
Masayuki Koyama ◽  
Shigeru Takamatsu
Keyword(s):  
Endothelial Cells ◽  
Platelet Activating Factor ◽  
Microvascular Endothelial Cells ◽  
Ionophore A23187 ◽  
Brain Microvascular Endothelial Cells ◽  
Brain Endothelial Cells ◽  
Aortic Endothelial Cells ◽  
Porcine Brain ◽  
Microvascular Endothelial ◽  
Aortic Endothelial

SummaryEndothelial cells produce platelet-activating factor (PAF), which is the key process in the interactions between the vascular wall and blood cells. To examine the production of PAF in brain microvasculature we have cultured brain endothelial cells and performed a comparative study with aortic endothelial cells. Fresh porcine brain was homogenized, and microvascular endothelial cells were separated by enzyme digestion. The cells were cultured in medium containing epidermal growth factor and bovine brain extract. Endothelial cells from the aorta of the same animal were cultured in a similar manner. Production of PAF was assessed by ǀ3Hǀacetate incorporation into phospholipids or by radioimmunoassay. Prostacyclin was measured by radioimmunoassay of 6-ketoprostaglandin F1α. The cells produced 1760 ± 403 and 2892 ± 347 dpm/106 cells (n = 4) of PAF when stimulated with brady- kinin and calcium ionophore A23187, each at 1 μM, respectively. Aortic endothelial cells produced 3911 ± 2006 and 8052 ± 2270 dpm/106 cells (n = 4), respectively, and these values were significantly higher than those in brain endothelial cells (p<0.01, U-test). Prostacyclin production was also higher in aortic cells as compared to brain microvascular endothelial cells. In aortic endothelial cells both Ca ionophore A23187 and bradykinin significantly stimulated PMN adherence whereas in brain microvascular cells only Ca ionophore enhanced the adherence. Brain microvascular endothelial cells produce smaller amount of PAF and prostacyclin as compared to aortic endothelial cells, and this fact may imply that the functional integrity of the brain microvascular endothelium is maintained at a low level.

Agonist-Stimulated Calcium Entry in Primary Cultures of Human Cerebral Microvascular Endothelial Cells

1999 ◽  
Vol 57 (3) ◽  
pp. 211-226 ◽  
Author(s):  
Li Li ◽  
Brian Bressler ◽  
Rukmini Prameya ◽  
Katerina Dorovini-Zis ◽  
C. Van Breemen
Keyword(s):  
Endothelial Cells ◽  
Primary Cultures ◽  
Calcium Entry ◽  
Microvascular Endothelial Cells ◽  
Microvascular Endothelial

scholarly journals L-type voltage-dependent Ca2+ channels in cerebral microvascular endothelial cells and ET-1 biosynthesis

2002 ◽  
Vol 283 (6) ◽  
pp. C1687-C1695 ◽  
Author(s):  
Momoh A. Yakubu ◽  
Charles W. Leffler
Keyword(s):  
Endothelial Cells ◽  
Primary Cultures ◽  
Chelating Agent ◽  
Microvascular Endothelial Cells ◽  
Vasoactive Agents ◽  
Cerebral Microvessels ◽  
Thromboxane Receptor ◽  
High K ◽  
Voltage Dependent ◽  
Microvascular Endothelial

We investigated the role of intracellular calcium concentration ([Ca2+]i) in endothelin-1 (ET-1) production, the effects of potential vasospastic agents on [Ca2+]i, and the presence of L-type voltage-dependent Ca2+ channels in cerebral microvascular endothelial cells. Primary cultures of endothelial cells isolated from piglet cerebral microvessels were used. Confluent cells were exposed to either the thromboxane receptor agonist U-46619 (1 μM), 5-hydroxytryptamine (5-HT; 0.1 mM), or lysophosphatidic acid (LPA; 1 μM) alone or after pretreatment with the Ca2+-chelating agent EDTA (100 mM), the L-type Ca2+ channel blocker verapamil (10 μM), or the antagonist of receptor-operated Ca2+ channel SKF-96365 HCl (10 μM) for 15 min. ET-1 production increased from 1.2 (control) to 8.2 (U-46619), 4.9 (5-HT), or 3.9 (LPA) fmol/μg protein, respectively. Such elevated ET-1 biosynthesis was attenuated by verapamil, EDTA, or SKF-96365 HCl. To investigate the presence of L-type voltage-dependent Ca2+channels in endothelial cells, the [Ca2+]isignal was determined fluorometrically by using fura 2-AM. Superfusion of confluent endothelial cells with U-46619, 5-HT, or LPA significantly increased [Ca2+]i. Pretreatment of endothelial cells with high K+ (60 mM) or nifedipine (4 μM) diminished increases in [Ca2+]i induced by the vasoactive agents. These results indicate that 1) elevated [Ca2+]i signals are involved in ET-1 biosynthesis induced by specific spasmogenic agents, 2) the increases in [Ca2+]i induced by the vasoactive agents tested involve receptor as well as L-type voltage-dependent Ca2+ channels, and 3) primary cultures of cerebral microvascular endothelial cells express L-type voltage-dependent Ca2+ channels.

scholarly journals Clinopodium tomentosum (Kunth) Govaerts Leaf Extract Influences in vitro Cell Proliferation and Angiogenesis on Primary Cultures of Porcine Aortic Endothelial Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-11
Author(s):  
Irvin Tubon ◽  
Chiara Bernardini ◽  
Fabiana Antognoni ◽  
Roberto Mandrioli ◽  
Giulia Potente ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Primary Cultures ◽  
Ethanolic Extract ◽  
Total Phenolic ◽  
Dependent Manner ◽  
Aortic Endothelial Cells ◽  
Phenolic Components ◽  
Porcine Aortic Endothelial Cells ◽  
Aortic Endothelial

Clinopodium tomentosum (Kunth) Govaerts is an endemic species in Ecuador, where it is used as an anti-inflammatory plant to treat respiratory and digestive affections. In this work, effects of a Clinopodium tomentosum ethanolic extract (CTEE), prepared from aerial parts of the plant, were investigated on vascular endothelium functions. In particularly, angiogenesis activity was evaluated, using primary cultures of porcine aortic endothelial cells (pAECs). Cells were cultured for 24 h in the presence of CTEE different concentrations (10, 25, 50, and 100 μg/ml); no viability alterations were found in the 10-50 μg/ml range, while a slight, but significant, proliferative effect was observed at the highest dose. In addition, treatment with CTEE was able to rescue LPS-induced injury in terms of cell viability. The CTEE ability to affect angiogenesis was evaluated by scratch test analysis and by an in vitro capillary-like network assay. Treatment with 25-50 μg/ml of extract caused a significant increase in pAEC’s migration and tube formation capabilities compared to untreated cells, as results from the increased master junctions’ number. On the other hand, CTEE at 100 μg/ml did not induce the same effects. Quantitative PCR data demonstrated that FLK-1 mRNA expression significantly increased at a CTEE dose of 25 μg/ml. The CTEE phytochemical composition was assessed through HPLC-DAD; rosmarinic acid among phenolic acids and hesperidin among flavonoids were found as major phenolic components. Total phenolic content and total flavonoid content assays showed that flavonoids are the most abundant class of polyphenols. The CTEE antioxidant activity was also showed by means of the DPPH and ORAC assays. Results indicate that CTEE possesses an angiogenic capacity in a dose-dependent manner; this represents an initial step in elucidating the mechanism of the therapeutic use of the plant.

Mechanisms of MAdCAM-1 gene expression in human intestinal microvascular endothelial cells

2005 ◽  
Vol 288 (2) ◽  
pp. C272-C281 ◽  
Author(s):  
Hitoshi Ogawa ◽  
David G. Binion ◽  
Jan Heidemann ◽  
Monica Theriot ◽  
Pamela J. Fisher ◽  
...  
Keyword(s):  
Gene Expression ◽  
Endothelial Cells ◽  
Protein Expression ◽  
Molecular Mechanisms ◽  
Primary Cultures ◽  
Microvascular Endothelial Cells ◽  
Cellular Density ◽  
Tnf Α ◽  
Gene And Protein Expression ◽  
Microvascular Endothelial

Mucosal addressin cell adhesion molecule-1 (MAdCAM-1) is a homing receptor preferentially expressed on gut-associated endothelial cells that plays a central role in leukocyte traffic into the mucosal immune compartment. Although the molecular mechanisms underlying endothelial ICAM-1 or E-selectin expression have been intensively investigated, the mechanisms that regulate human MAdCAM-1 expression have not been defined. We report MAdCAM-1 gene and protein expression in primary cultures of human intestinal microvascular endothelial cells (HIMEC) that was not demonstrated in human umbilical vein endothelial cells. Similar to ICAM-1 and E-selectin expression, MAdCAM-1 gene expression in HIMEC was inducible with TNF-α, IL-1β, or LPS activation. However, in striking contrast to ICAM-1 and E-selectin expression, MAdCAM-1 mRNA and protein expression in HIMEC was heavily dependent on culture duration and/or cellular density, suggesting a prominent role for cell-cell interaction among these endothelial cells in the expression of the mucosal addressin. MAdCAM-1 expression was inhibited by both SN-50 (NF-κB inhibitor) and LY-294002 [phosphatidylinositol 3-kinase (PI3-K) inhibitor], whereas ICAM-1 and E-selectin expression was inhibited by SN-50 but not by LY-294002. The Akt phosphorylation by TNF-α or LPS was greater at higher cell density, demonstrating a pattern similar to that of MAdCAM-1 expression. NF-κB activation was not affected by cellular density in HIMEC. MAdCAM-1 expression in human gut endothelial cells is regulated by distinct signaling mechanisms involving both NF-κB and PI3-K/Akt. These data also suggest that PI3-K/Akt is involved in the gut-specific differentiation of HIMEC, which results in expression of the mucosal addressin MAdCAM-1.

scholarly journals Epoxygenase-driven angiogenesis in human lung microvascular endothelial cells

2003 ◽  
Vol 284 (1) ◽  
pp. H215-H224 ◽  
Author(s):  
Meetha Medhora ◽  
John Daniels ◽  
Kavita Mundey ◽  
Beate Fisslthaler ◽  
Rudi Busse ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Primary Cultures ◽  
Tube Formation ◽  
Microvascular Endothelial Cells ◽  
Arachidonic Acid Metabolite ◽  
Epoxyeicosatrienoic Acid ◽  
Adult Rats ◽  
Microvascular Endothelial ◽  
Cloned Cdna

Angiogenesis is one of the most recent physiological functions attributed to products of cytochrome P-450 (CYP450) enymes. To test this at a molecular level in human cells, we used a cloned cDNA for the human endothelial enzyme CYP450 2C9 (CYP2C9) to study growth as well as differentiation of human microvascular endothelial cells from the lung (HMVEC-L). Using adenoviral vectors overexpressing mRNA for CYP2C9, we show that the presence of CYP2C9 doubles thymidine incorporation and stimulates proliferation of primary cultures of endothelial cells compared with Ad5-GFP (control) in 24 h. In addition, there is a significant increase of tube formation in Matrigel after infection of HMVEC-L with Ad5-2C9 than with Ad5-GFP. More interestingly, Ad5-2C9 expressing the antisense product of CYP2C9 (2C9AS) inhibited tube formation compared with both Ad5-GFP as well as the Ad5-2C9 constructs. Finally, we tested the most abundant arachidonic acid metabolite of CYP2C9, 14,15-epoxyeicosatrienoic acid, which induced angiogenesis in vivo when embedded in Matrigel plugs and implanted in adult rats. These data support an important role for CYP2C9 in promoting angiogenesis.

Endotoxin enhances arachidonic acid metabolism by cultured rabbit microvascular endothelial cells

1992 ◽  
Vol 263 (4) ◽  
pp. H1213-H1221 ◽  
Author(s):  
P. M. Renzi ◽  
J. T. Flynn
Keyword(s):  
Endothelial Cells ◽  
Arachidonic Acid ◽  
Time Course ◽  
Lipid A ◽  
Primary Cultures ◽  
Microvascular Endothelial Cells ◽  
Prostaglandin Endoperoxide ◽  
Eicosanoid Production ◽  
Prostaglandin Endoperoxide Synthase ◽  
Microvascular Endothelial

This study demonstrates that bacterial lipopolysaccharide and lipid A exert a significant effect on eicosanoid formation by primary cultures of microvascular endothelial cells (MECs). Qualitative studies using [14C]-arachidonic acid demonstrated that prostaglandin E2 was the primary eicosanoid formed by MECs after 20 h of treatment with either vehicle or lipopolysaccharide. Significant, dose-dependent productions of PGE2 and prostacyclin, beginning at an endotoxin dose of 0.01 ng/ml, were quantified by radioimmunoassay in supernatants of cells treated for 20 h with lipopolysaccharide or lipid A. This eicosanoid production was inhibited by meclofenamate and cycloheximide and occurred without cellular injury. The time course and kinetics of eicosanoid production in response to endotoxin demonstrate a significant, time-related enhancement. Endotoxin-treated MECs responded to exogenous substrate with augmented PGE2 production, suggesting enhanced prostaglandin endoperoxide synthase activity. These results demonstrate a significant interaction of endotoxin with endothelial cells of microvascular origin that results in an enhanced potential for eicosanoid metabolism. This effect may be mediated in part through induction of prostaglandin endoperoxide synthase.

scholarly journals iNOS expression in human intestinal microvascular endothelial cells inhibits leukocyte adhesion

1998 ◽  
Vol 275 (3) ◽  
pp. G592-G603 ◽  
Author(s):  
David G. Binion ◽  
Sidong Fu ◽  
Kalathur S. Ramanujam ◽  
Yuh Cherng Chai ◽  
Raed A. Dweik ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Intestinal Inflammation ◽  
Leukocyte Adhesion ◽  
Primary Cultures ◽  
Inos Expression ◽  
Microvascular Endothelial Cells ◽  
No Production ◽  
Nos Inhibitors ◽  
Microvascular Endothelial

Increased nitric oxide (NO) production by inducible nitric oxide synthase (iNOS) has been associated with intestinal inflammation, including human inflammatory bowel disease. However, NO can downregulate endothelial activation and leukocyte adhesion, critical steps in the inflammatory response. Using primary cultures of human intestinal microvascular endothelial cells (HIMEC), we determined the role of NO in the regulation of HIMEC activation and interaction with leukocytes. Both nonselective ( N G-monomethyl-l-arginine) and specific ( N-iminoethyl-l-lysine) competitive inhibitors of iNOS significantly increased binding of leukocytes by HIMEC activated with cytokines and lipopolysaccharide. Increased adhesion was reversible with the NOS substratel-arginine and was not observed in human umbilical vein endothelial cells (HUVEC). Activation of HIMEC significantly upregulated HIMEC iNOS expression and NO production. NOS inhibitors did not augment cell adhesion molecule levels in activated HIMEC but did result in sustained increases in intracellular reactive oxygen species. In addition, antioxidant compounds reversed the effect of NOS inhibitors on HIMEC-leukocyte interaction. Taken together, these data suggest that after HIMEC activation, iNOS-derived NO is an endogenous antioxidant, downregulating leukocyte binding and potentially downregulating intestinal inflammation.

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