Cultured cells of the blood–brain barrier from apolipoprotein B-100 transgenic mice: effects of oxidized low-density lipoprotein treatment

Lénárt, Nikolett; Walter, Fruzsina R.; Bocsik, Alexandra; Sántha, Petra; Tóth, Melinda E.; Harazin, András; Tóth, Anna Zsófia; Vízler, Csaba; Török, Zsolt; Pilbat, Ana Maria; Vı́gh, László; Puskás, László G.; Sántha, Miklós; Deli, Mária A.

doi:10.1186/s12987-015-0013-y

Cited by 28 publications

(33 citation statements)

References 56 publications

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“…Chloromethyl-dichloro-dihydro-fluorescein diacetate (DCFDA, Molecular Probes, Life Technologies, USA) was used for ROS detection, and 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate (DAF-FM, Molecular Probes, Life Technologies, USA) for NO detection as fluorescent probes, as described in our previous studies [39,40]. Brain endothelial cells were cultured in 96-well plates with black walls and transparent plastic bottoms (Corning, NY, USA).…”

Section: Measurement Of Reactive Oxygen Species and Nitric Oxide Prodmentioning

confidence: 99%

“…Cells treated with kainate and protective drugs for 24 h were fixed with 1% paraformaldehyde for 20 min at 4 °C, permeabilized with 0.2% Triton-X100 for 10 min at 4 °C, then blocked with 3% BSA-PBS for 1 h at room temperature. Samples were incubated with rabbit anti-claudin-5 polyclonal (SAB4502981, 1:800 dilution in 3% BSA-PBS, antibody registry ID: AB_10753223; Sigma-Aldrich, USA) or rabbit anti-ZO-1 polyclonal (61-7300, 1:400 dilution in 3% BSA-PBS, antibody registry ID: AB_2533147; Invitrogen, USA) primary antibodies overnight at 4 °C, then with anti-rabbit secondary antibody conjugated with Cy3 (C2306, 1:400 dilution in PBS, antibody registry ID: AB_258792; Sigma-Aldrich, USA) and H33342 nucleus stain (1 µg/ml) for 1 h at room temperature [40]. Cells were washed with PBS between the incubations.…”

Section: Immunohistochemistrymentioning

confidence: 99%

“…M: 1 Kb Plus DNA ladder used as a reference inducer of ROS in the assay (Fig. 4), as in our previous study [29], while sodium nitroprusside was given to increase NO production [40].…”

Section: Effect Of Kainate On Reactive Oxygen Species and Nitric Oxidmentioning

confidence: 99%

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Simvastatin, edaravone and dexamethasone protect against kainate-induced brain endothelial cell damage

Barna

Walter

Harazin

et al. 2020

Fluids Barriers CNS

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Background: Excitotoxicity is a central pathological pathway in many neurological diseases with blood-brain barrier (BBB) dysfunction. Kainate, an exogenous excitotoxin, induces epilepsy and BBB damage in animal models, but the direct effect of kainate on brain endothelial cells has not been studied in detail. Our aim was to examine the direct effects of kainate on cultured cells of the BBB and to test three anti-inflammatory and antioxidant drugs used in clinical practice, simvastatin, edaravone and dexamethasone, to protect against kainate-induced changes.Methods: Primary rat brain endothelial cell, pericyte and astroglia cultures were used to study cell viability by impedance measurement. BBB permeability was measured on a model made from the co-culture of the three cell types. The production of nitrogen monoxide and reactive oxygen species was followed by fluorescent probes. The mRNA expression of kainate receptors and nitric oxide synthases were studied by PCR.Results: Kainate damaged brain endothelial cells and made the immunostaining of junctional proteins claudin-5 and zonula occludens-1 discontinuous at the cell border indicating the opening of the barrier. The permeability of the BBB model for marker molecules fluorescein and albumin and the production of nitric oxide in brain endothelial cells were increased by kainate. Simvastatin, edaravone and dexamethasone protected against the reduced cell viability, increased permeability and the morphological changes in cellular junctions caused by kainate. Dexamethasone attenuated the elevated nitric oxide production and decreased the inducible nitric oxide synthase (NOS2/iNOS) mRNA expression increased by kainate treatment. Conclusion:Kainate directly damaged cultured brain endothelial cells. Simvastatin, edaravone and dexamethasone protected the BBB model against kainate-induced changes. Our results confirmed the potential clinical usefulness of these drugs to attenuate BBB damage. Publisher's NoteSpringer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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Section: Measurement Of Reactive Oxygen Species and Nitric Oxide Prodmentioning

confidence: 99%

Section: Immunohistochemistrymentioning

confidence: 99%

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Simvastatin, edaravone and dexamethasone protect against kainate-induced brain endothelial cell damage

Barna

Walter

Harazin

et al. 2020

Fluids Barriers CNS

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“…decreased membrane fluidity (17,18), we hypothesized that alterations in cholesterol composition or lipid packaging in lipid rafts in response to OxLDL or atheroprone flow may alter integrin α5 distribution and/or activity. Furthermore, given that activation of NF-κB in turn activates Nod-like receptor protein 3 (NLRP3) inflammasome (19), integrin α5 would be upstream of the atheroprone flow-induced NLRP3 inflammasome (20).…”

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confidence: 99%

Activation of integrin α5 mediated by flow requires its translocation to membrane lipid rafts in vascular endothelial cells

Sun

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Local flow patterns determine the uneven distribution of atherosclerotic lesions. Membrane lipid rafts and integrins are crucial for shear stress-regulated endothelial function. In this study, we investigate the role of lipid rafts and integrin α5 in regulating the inflammatory response in endothelial cells (ECs) under atheroprone versus atheroprotective flow. Lipid raft proteins were isolated from ECs exposed to oscillatory shear stress (OS) or pulsatile shear stress, and then analyzed by quantitative proteomics. Among 396 proteins redistributed in lipid rafts, integrin α5 was the most significantly elevated in lipid rafts under OS. In addition, OS increased the level of activated integrin α5 in lipid rafts through the regulation of membrane cholesterol and fluidity. Disruption of F-actin-based cytoskeleton and knockdown of caveolin-1 prevented the OS-induced integrin α5 translocation and activation. In vivo, integrin α5 activation and EC dysfunction were observed in the atheroprone areas of low-density lipoprotein receptor-deficient (Ldlr −/− ) mice, and knockdown of integrin α5 markedly attenuated EC dysfunction in partially ligated carotid arteries. Consistent with these findings, mice with haploinsufficency of integrin α5 exhibited a reduction of atherosclerotic lesions in the regions under atheroprone flow. The present study has revealed an integrin-and membrane lipid raft-dependent mechanotransduction mechanism by which atheroprone flow causes endothelial dysfunction.integrin | lipid rafts | shear stress | proteomics | endothelial dysfunction S hear stress imposed on vascular endothelial cells (ECs) influences vascular phenotype and function. Atherosclerosis preferentially develops at branches and curvatures in the arterial tree where flow is disturbed. In contrast, pulsatile shear stress (PS) in the straight parts of the arteries is atheroprotective (1). At the cellular and molecular levels, disturbed flow pattern increases, while PS inhibits, the inflammatory response in ECs, including the expression of intercellular adhesion molecule 1 (ICAM-1), vascular adhesion molecule 1 (VCAM-1), and interleukin 1β (IL-1β) (2). We found that integrins in ECs are shear stress-sensitive (3). Many subsequent studies demonstrated that integrin activation is essential for transmitting mechanical stimuli to intracellular biochemical pathways (4-6). Additionally, mounting evidence indicates that lipid raft microdomains, membrane receptors, cytoskeletal proteins, and extracellular matrices are linked to integrin activation in the context of mechanotransduction (7-11). However, the key mechanism underlying the differential effects of atheroprone versus atheroprotective flows in activating integrins that in turn induces inflammatory response in ECs remains to be elucidated.Lipid rafts are membrane microdomains that are enriched in cholesterol, sphingolipids, and a variety of signaling molecules, which function as cellular signaling platforms. These microdomains are more ordered and tightly packed than the surrounding membran...

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“…In addition, dyslipidemia may alter BBB function in AD [42], contributing to the transportation of inflammatory lipids from the blood [6]. Further, studies have found ApoB-100 protein in the membrane microdomains of primary brain endothelial cells [43, 44]. Using the LDL-receptor binding domain of apoB could be an effective way to deliver recombinant proteins across the BBB, and such an approach can be used to treat a neurodegenerative disease [45, 46].…”

Section: Discussionmentioning

confidence: 99%

Ratio of apoB/LDL: a potential clinical index for vascular cognitive impairment

Qian

Tan

2016

BMC Neurol

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BackgroundVascular cognitive impairment (VCI), compared to vascular dementia (VD), has a broader definition and highlights the effect of vascular disease in dementia, and stroke seems play an important role in the development of VCI. However, not all patients with brain infarcts suffer from VCI; unique risk factors appear to cause such progression. This study aimed to find potential risk factors of vascular cognitive impairment among patients with brain infarcts.MethodsThirty-seven dementia patients and 74 brain infarction patients were included; all had infarcts in both basilar ganglia. The frequencies of risk factors, such as age, hypertension, and hyperlipidemia, were compared between the two groups.ResultsThe incident rate of hyperlipidemia in the patients with dementia was 35.14%, which was significantly lower than that in the patients with infarction (59.46%, P = 0.015). In the dementia group, there was a positive correlation between the ratio of apoprotein B (apoB)/low density lipoprotein (LDL) and the Mini Mental State Examination (MMSE) score (R = 0.411, P = 0.011).ConclusionOur study indicated that the ratio of apoB/LDL may be a potential clinical index for vascular cognitive impairment.Electronic supplementary materialThe online version of this article (doi:10.1186/s12883-016-0766-1) contains supplementary material, which is available to authorized users.

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Cultured cells of the blood–brain barrier from apolipoprotein B-100 transgenic mice: effects of oxidized low-density lipoprotein treatment

Cited by 28 publications

References 56 publications

Simvastatin, edaravone and dexamethasone protect against kainate-induced brain endothelial cell damage

Simvastatin, edaravone and dexamethasone protect against kainate-induced brain endothelial cell damage

Activation of integrin α5 mediated by flow requires its translocation to membrane lipid rafts in vascular endothelial cells

Ratio of apoB/LDL: a potential clinical index for vascular cognitive impairment

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