Assessment of HBEC-5i endothelial cell line cultivated in astrocyte conditioned medium as a human blood-brain barrier model for ABC drug transport studies

Puech, Clémentine; Hodin, Sophie; Forest, Valérie; Hé, Zhiguo; Mismetti, Patrick; Delavenne, Xavier; Perek, Nathalie

doi:10.1016/j.ijpharm.2018.09.040

Cited by 41 publications

(41 citation statements)

References 40 publications

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“…Astrocytes are the most abundant cell type within the CNS, and they provide structural support, promote the formation of the BBB, and release beneficial factors that maintain brain cell development and homeostasis of the extracellular environment [24]. The function of brain endothelial monolayer cells, ATP-binding cassette transporters, and TJP expression for example, is enhanced when the cells are co-cultured with astrocytes or ACM for a period of time [5, 25]. In the present study, ACM did not alter TJP expression or the integrity of bEnd.3 cell monolayer, potentially due to the relatively short culture period, which was consistent with the results from previous studies [11, 25].…”

Section: Discussionmentioning

confidence: 99%

The glucagon-like peptide-1 receptor agonist reduces inflammation and blood-brain barrier breakdown in an astrocyte-dependent manner in experimental stroke

Shan

Tan

Lin

et al. 2019

J Neuroinflammation

118

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BackgroundPreserving the integrity of the blood-brain barrier (BBB) is beneficial to avoid further brain damage after acute ischemic stroke (AIS). Astrocytes, an important component of the BBB, promote BBB breakdown in subjects with AIS by secreting inflammatory factors. The glucagon-like peptide-1 receptor (GLP-1R) agonist exendin-4 (Ex-4) protects the BBB and reduces brain inflammation from cerebral ischemia, and GLP-1R is expressed on astrocytes. However, the effect of Ex-4 on astrocytes in subjects with AIS remains unclear.MethodsIn the present study, we investigated the effect of Ex-4 on astrocytes cultured under oxygen-glucose deprivation (OGD) plus reoxygenation conditions and determined whether the effect influences bEnd.3 cells. We used various methods, including permeability assays, western blotting, immunofluorescence staining, and gelatin zymography, in vitro and in vivo.ResultsEx-4 reduced OGD-induced astrocyte-derived vascular endothelial growth factor (VEGF-A), matrix metalloproteinase-9 (MMP-9), chemokine monocyte chemoattractant protein-1 (MCP-1), and chemokine C-X-C motif ligand 1 (CXCL-1). The reduction in astrocyte-derived VEGF-A and MMP-9 was related to the increased expression of tight junction proteins (TJPs) in bEnd.3 cells. Ex-4 improved neurologic deficit scores, reduced the infarct area, and ameliorated BBB breakdown as well as decreased astrocyte-derived VEGF-A, MMP-9, CXCL-1, and MCP-1 levels in ischemic brain tissues from rats subjected to middle cerebral artery occlusion. Ex-4 reduced the activation of the JAK2/STAT3 signaling pathway in astrocytes following OGD.ConclusionBased on these findings, ischemia-induced inflammation and BBB breakdown can be improved by Ex-4 through an astrocyte-dependent manner.

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Section: Discussionmentioning

confidence: 99%

The glucagon-like peptide-1 receptor agonist reduces inflammation and blood-brain barrier breakdown in an astrocyte-dependent manner in experimental stroke

Shan

Tan

Lin

et al. 2019

J Neuroinflammation

118

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“…The model was composed of HBEC-5i endothelial cells (from ATCC-Manassas, VA, USA) cultivated with conditioned medium from human astrocytes(HA from ScienCell Research Laboratories, Carlsbad, CA, USA) as previously described (Puech et al, 2018b).Cells were cultured in DMEM-F12 HAM, supplemented with 10% FBS and 1% antibiotic antimycotic solution. Cells were maintained in a humidified incubator at 5% CO2 and 37°C.…”

Section: Hbec-5i Bbb Culturementioning

confidence: 99%

Direct oral anticoagulants are associated with limited damage of endothelial cells of the blood-brain barrier mediated by the thrombin/PAR-1 pathway

et al. 2019

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Anticoagulant therapy presents iatrogenic effects such as intracerebral hemorrhage (ICH). The latest anticoagulants on the market, direct oral anticoagulants (DOACs) such as apixaban, dabigatran and rivaroxaban, are reported to cause less ICH than other anticoagulants. Next to the ICH area, the thrombin is accumulated and the blood-brain barrier (BBB) is opened. The effects of thrombin on the BBB are largely mediated by the protease activated receptor (PAR) family, especially the PAR-1 isoform. Our hypothesis is that DOACs may limit the effects of thrombin on endothelial cells (of the BBB) alteration by a mechanism probably involving PAR-1 activation. To test this hypothesis in vitro, we used HBEC-5i human brain endothelial cells as a human BBB model. The effects of thrombin under warfarin, heparin, rivaroxaban, apixaban, and dabigatran treatment on endothelial cells were then investigated by measuring of permeability and junction proteins: ZO-1 and VE-cadherin expressions and PAR-1 cleavage. Depending on the anticoagulant used, we observed three profiles of response of the endothelial cells after thrombin exposure: i) dabigatran treatment allowed maintaining the tightness of the endothelial monolayer; ii) other DOACs limited thrombin-induced alteration of the endothelial monolayer; and iii) pretreatment with warfarin and heparin did not protect from thrombininduced BBB breakdown. Pretreatment with DOACs clearly limited the impact of thrombin on PAR-1 cleavage in our model, contrary to other anticoagulants, associated with ZO-1 and VEcadherin expressions. In conclusion, DOACs seem to limit the alteration of the monolayer of endothelial cells of the BBB mediated by the thrombin/PAR-1 pathway.

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“…This study was conducted on an in vitro model of culture of cerebral endothelial cells in conditioned media. This model could be improved with astrocytes and pericytes; nevertheless, it has been shown earlier that it is a good pharmacological model 15 . Our work showed that our model may screen patient serum and may be an interesting tool to evaluate risk and may be, one day, the reversal effects in sleep apnea patients.…”

Section: Discussionmentioning

confidence: 99%

“…The BBB model was composed of HBEC-5i endothelial cells and was cultivated with human astrocyte conditioned medium on insert as previously described 15 . All cells were cultured in DMEM-F12 HAM and used before passage number 20, which is the time where these cells may begin to lose their BBB properties.…”

Section: Methodsmentioning

confidence: 99%

“…We have developed in our laboratory a relevant human BBB model composed of an endothelial cell line (HBEC-5i) 15 , which is grown with a conditioned astrocyte culture medium without adjuvant. It has been shown that astrocytic cells have the ability to induce and maintain close junctions between endothelial cells 16,17 .…”

Section: Sera Of Elderly Obstructive Sleep Apnea Patients Alter Bloodmentioning

confidence: 99%

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Sera of elderly obstructive sleep apnea patients alter blood–brain barrier integrity in vitro: a pilot study

Voirin

Celle

Perek

et al. 2020

Sci Rep

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Obstructive sleep apnea syndrome (OSAS) is characterized by repeated episodes of hypoxia during the night. The severity of the disorder can be evaluated using an apnea–hypopnea index (AHI). The physiological consequences are mainly cardiovascular and neuronal dysfunctions. One hypothesis to explain such associated neurological disorders is disruption of the blood–brain barrier (BBB), which protects the brain from endovascular cytotoxic compounds. We selected two subgroups of volunteers from the PROOF cohort study (France), a group of patients suffering newly diagnosed severe OSAS (AHI > 30/h) and a group showing no sleep apnea (AHI < 5/h). We exposed a human in vitro BBB model of endothelial cells (HBEC-5i) with sera of patients with and without OSAS. After exposure, we measured the apparent BBB permeability as well as tight junction and ABC transporter expression using whole cell ELISA. We showed that after incubation with sera from OSAS patients, there was a loss of integrity in the human in vitro BBB model; this was reflected by an increase in permeability (43%; p < 0.001) and correlated with a 50% and 40% decrease in tight junction protein expression of ZO-1 and claudin-5, respectively. At the same time, we observed an upregulation in Pgp protein expression (52%) and functionality, and a downregulation in BCRP expression (52%). Our results demonstrated that severe BBB disorder after exposure to sera from OSAS patients was reflected by an opening of the BBB.

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Assessment of HBEC-5i endothelial cell line cultivated in astrocyte conditioned medium as a human blood-brain barrier model for ABC drug transport studies

Cited by 41 publications

References 40 publications

The glucagon-like peptide-1 receptor agonist reduces inflammation and blood-brain barrier breakdown in an astrocyte-dependent manner in experimental stroke

The glucagon-like peptide-1 receptor agonist reduces inflammation and blood-brain barrier breakdown in an astrocyte-dependent manner in experimental stroke

Direct oral anticoagulants are associated with limited damage of endothelial cells of the blood-brain barrier mediated by the thrombin/PAR-1 pathway

Sera of elderly obstructive sleep apnea patients alter blood–brain barrier integrity in vitro: a pilot study

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