Isolation and primary culture of rat cerebral microvascular endothelial cells for studying drug transport in vitro

1996 ◽  
Vol 36 (1) ◽  
pp. 45-52 ◽  
Author(s):  
Nobuhiro Ichikawa ◽  
Kohji Naora ◽  
Hidenari Hirano ◽  
Michio Hashimoto ◽  
Sumio Masumura ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Primary Culture ◽  
Drug Transport ◽  
Microvascular Endothelial Cells ◽  
Microvascular Endothelial

scholarly journals A tumor-promoting role for soluble TβRIII in glioblastoma

2021 ◽  
Author(s):  
Isabel Burghardt ◽  
Judith Johanna Schroeder ◽  
Tobias Weiss ◽  
Dorothee Gramatzki ◽  
Michael Weller
Keyword(s):  
Endothelial Cells ◽  
Ex Vivo ◽  
Microvascular Endothelial Cells ◽  
Smad2 Phosphorylation ◽  
Cell Gene Expression ◽  
Microvascular Endothelial ◽  
Cell Gene ◽  
Mouse Glioma

Abstract Purpose Members of the transforming growth factor (TGF)-β superfamily play a key role in the regulation of the malignant phenotype of glioblastoma by promoting invasiveness, angiogenesis, immunosuppression, and maintaining stem cell-like properties. Betaglycan, a TGF-β coreceptor also known as TGF-β receptor III (TβRIII), interacts with members of the TGF-β superfamily and acts as membrane-associated or shed molecule. Shed, soluble TβRIII (sTβRIII) is produced upon ectodomain cleavage of the membrane-bound form. Elucidating the role of TβRIII may improve our understanding of TGF-β pathway activity in glioblastoma Methods Protein levels of TβRIII were determined by immunohistochemical analyses and ex vivo single-cell gene expression profiling of glioblastoma tissue respectively. In vitro, TβRIII levels were assessed investigating long-term glioma cell lines (LTCs), cultured human brain-derived microvascular endothelial cells (hCMECs), glioblastoma-derived microvascular endothelial cells, and glioma-initiating cell lines (GICs). The impact of TβRIII on TGF-β signaling was investigated, and results were validated in a xenograft mouse glioma model Results Immunohistochemistry and ex vivo single-cell gene expression profiling of glioblastoma tissue showed that TβRIII was expressed in the tumor tissue, predominantly in the vascular compartment. We confirmed this pattern of TβRIII expression in vitro. Specifically, we detected sTβRIII in glioblastoma-derived microvascular endothelial cells. STβRIII facilitated TGF-β-induced Smad2 phosphorylation in vitro and overexpression of sTβRIII in a xenograft mouse glioma model led to increased levels of Smad2 phosphorylation, increased tumor volume, and decreased survival Conclusions These data shed light on the potential tumor-promoting role of extracellular shed TβRIII which may be released by glioblastoma endothelium with high sTβRIII levels.

Isolation and characterization of human and rat cardiac microvascular endothelial cells

1993 ◽  
Vol 264 (2) ◽  
pp. H639-H652 ◽  
Author(s):  
M. Nishida ◽  
W. W. Carley ◽  
M. E. Gerritsen ◽  
O. Ellingsen ◽  
R. A. Kelly ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Microvascular Endothelial Cells ◽  
Microvascular Endothelium ◽  
Adult Rat ◽  
Isolation And Characterization ◽  
Ventricular Tissue ◽  
Microvascular Endothelial ◽  
Cardiac Microvascular Endothelial Cells

Although reciprocal intercellular signaling may occur between endocardial or microvascular endothelium and cardiac myocytes, suitable in vitro models have not been well characterized. In this report, we describe the isolation and primary culture of cardiac microvascular endothelial cells (CMEC) from both adult rat and human ventricular tissue. Differential uptake of fluorescently labeled acetylated low-density lipoprotein (Ac-LDL) indicated that primary isolates of rat CMEC were quite homogeneous, unlike primary isolates of human ventricular tissue, which required cell sorting based on Ac-LDL uptake to create endothelial cell-enriched primary cultures. The endothelial phenotype of both primary isolates and postsort subcultured CMEC and their microvascular origin were determined by characteristic histochemical staining for a number of endothelial cell-specific markers, by the absence of cells with fibroblast or pericyte-specific cell surface antigens, and by rapid tube formation on purified basement membrane preparations. Importantly, [3H]-thymidine uptake was increased 2.3-fold in subconfluent rat microvascular endothelial cells 3 days after coculture with adult rat ventricular myocytes because of release of an endothelial cell mitogen(s) into the extracellular matrix, resulting in a 68% increase in cell number compared with CMEC in monoculture. Thus biologically relevant cell-to-cell interactions can be modeled with this in vitro system.

Hydroxysafflor yellow A promotes angiogenesis in rat brain microvascular endothelial cells injured by oxygen-glucose deprivation/reoxygenation(OGD/R) through SIRT1-HIF-1α-VEGFA signaling pathway

2021 ◽  
Vol 18 ◽  
Author(s):  
Juxuan Ruan ◽  
Lei Wang ◽  
Jiheng Dai ◽  
Jing Li ◽  
Ning Wang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Growth Factor ◽  
Signaling Pathway ◽  
Ischemia Reperfusion ◽  
Tube Formation ◽  
Microvascular Endothelial Cells ◽  
Brain Microvascular Endothelial Cells ◽  
Hydroxysafflor Yellow A ◽  
Microvascular Endothelial

Objective: Angiogenesis led by brain microvascular endothelial cells (BMECs) contributes to the remission of brain injury after brain ischemia reperfusion. In this study, we investigated the effects of hydroxysafflor yellow A(HSYA) on angiogenesis of BMECs injured by OGD/R via SIRT1-HIF-1α-VEGFA signaling pathway. Methods: The OGD/R model of BMECs was established in vitro by OGD for 2h and reoxygenation for 24h. At first, the concentrations of vascular endothelial growth factor (VEGF), Angiopoietin (ang) and platelet-derived growth factor (PDGF) in supernatant were detected by ELISA, and the proteins expression of VEGFA, Ang-2 and PDGFB in BMECs were tested by western blot; the proliferation, adhesion, migration (scratch healing and transwell) and tube formation experiment of BMECs; the expression of CD31 and CD34 were tested by immunofluorescence staining. The levels of sirtuin1(SIRT1), hypoxia-inducible factor-1α (HIF-1α), VEGFA mRNA and protein were tested. Results: HSYA up-regulated the levels of VEGF, Ang and PDGF in the supernatant of BMECs under OGD/R, and the protein expression of VEGFA, Ang-2 and PDGFB were increased; HSYA could significantly alleviate the decrease of cell proliferation, adhesion, migration and tube formation ability of BMECs during OGD/R; HSYA enhanced the fluorescence intensity of CD31 and CD34 of BMECs during OGD/R; HSYA remarkably up-regulated the expression of SIRT1, HIF-1α, VEGFA mRNA and protein after OGD/R, and these increase decreased after SIRT1 was inhibited. Conclusion: SIRT1-HIF-1α-VEGFA signaling pathway is involved in HSYA improves angiogenesis of BMECs injured by OGD/R.

N6-isopentenyladenosine a new potential anti-angiogenic compound that targets human microvascular endothelial cells in vitro

2018 ◽  
Vol 37 (10) ◽  
pp. 533-545
Author(s):  
Sara Castiglioni ◽  
Valentina Romeo ◽  
Silvana Casati ◽  
Roberta Ottria ◽  
Cristiana Perrotta ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Microvascular Endothelial Cells ◽  
Microvascular Endothelial

7-Ketocholesterol activates caspases-3/7, -8, and -12 in human microvascular endothelial cells in vitro

2008 ◽  
Vol 75 (3) ◽  
pp. 343-350 ◽  
Author(s):  
Saurabh Luthra ◽  
Joyce Dong ◽  
Ana L. Gramajo ◽  
Marilyn Chwa ◽  
Dae W. Kim ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Microvascular Endothelial Cells ◽  
Microvascular Endothelial

Abstract 474: Effects of Cigarette Smoke on CD146 Expression and Function in Pulmonary Endothelial Cells

2012 ◽  
Vol 32 (suppl_1) ◽  
Author(s):  
Adelheid Kratzer ◽  
Jonas Salys ◽  
Benjy Gonzalez ◽  
Hong Wei Chu ◽  
Martin Zamora ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cigarette Smoke ◽  
Immunofluorescent Staining ◽  
Microvascular Endothelial Cells ◽  
Pulmonary Microvascular Endothelial Cells ◽  
Pulmonary Endothelial Cells ◽  
Membrane Bound ◽  
Microvascular Endothelial ◽  
Cd146 Expression

Background and Objectives: Cell adhesion molecule CD146 is a transmembrane glycoprotein constitutively expressed in all types of endothelial cells (EC). It exists in two forms: a membrane-anchored form (CD146) and a soluble, extracellular and cleaved form (sCD146). The plasma concentration of sCD146 is modulated in inflammatory diseases that involve endothelial alterations. We investigated the role of endothelial CD146 in cigarette smoke-induced emphysema in vivo and in pulmonary endothelial cells (EC) in vitro . Methods: Sprague Dawley rats exposed to cigarette smoke for 2 months developed significant emphysematous changes (measured by mean linear intercept). Levels of sCD146 were subsequently measured in the circulation as well as in the bronchoalveolar lavage fluid (BALf) via ELISA. In vitro studies were carried out in rat pulmonary microvascular endothelial cells using CSE. Results: CD146 is highly expressed in rat pulmonary microvascular endothelial cells (RPMVEC) and to a much lower extent, in pulmonary macrovascular endothelial cells (RPAEC). Treatment of RPMVEC with cigarette smoke extract (CSE) in vitro resulted in decreased expression of membrane-bound CD146 as well as a reduced gene expression and increased sCD146 levels in the culture medium after 12 hours. Moreover, CSE-induced downregulation of CD146 expression resulted in increased vascular permeability of RPMVEC, as measured by EVANs Blue assay and migration of CFSE-labeled rat alveolar macrophage. Immunofluorescent staining revealed that CSE treatment resulted in translocation of membrane-bound CD146 into the nucleus. Subsequent western blot analysis showed changes in ERK and AKT activation and signaling. Similar results were found upon siRNA silencing of CD146, implicating a role for CD146 in tissue inflammation and integrity. Circulating levels of sCD146 were also elevated in plasma and BALf of patients with COPD and correlated, in part, with the presence of anti-endothelial autoantibodies. Additionally, we found decreased expression of membrane-bound CD146 in lung tissues of COPD patients. Conclusions: Our data suggest that CD146 plays an important role in pulmonary vascular EC function. Moreover, levels of circulating soluble CD146 can be a predictor of vascular endothelial cell injury.

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