Development of a direct contact astrocyte‐human cerebral microvessel endothelial cells blood–brain barrier coculture model

• Published in: Journal of pharmacy and pharmacology 2017-12, Vol.69 (12), p.1684-1696
• Main Authors: Kulczar, Chris, Lubin, Kelsey E., Lefebvre, Sylvia, Miller, Donald W., Knipp, Gregory T.
• Format: Article
• Language: English
• Subjects: Astrocytes; Astrocytes - cytology; Blood-brain barrier; Blood-Brain Barrier - cytology; Blood-Brain Barrier - metabolism; blood–brain barrier coculture; Cell culture; Cerebrovascular Circulation - physiology; Chemical compounds; Coculture Techniques; Endothelial cells; Endothelial Cells - cytology; Endothelium, Vascular - cytology; human astrocytes; human cerebral microvessel endothelial cells; Humans; Membrane permeability; Microvessels - cytology; Monoculture; Neurotoxicity; paracellular permeability; Pericytes; Permeability; Research Paper
• ISSN: 0022-3573; 2042-7158
• DOI: 10.1111/jphp.12803

Objectives In conventional in‐vitro blood–brain barrier (BBB) models, primary and immortalized brain microvessel endothelial cell (BMEC) lines are often cultured in a monolayer or indirect coculture or triculture configurations with astrocytes or pericytes, for screening permeation of therapeutic or potentially neurotoxic compounds. In each of these cases, the physiological relevancy associated with the direct contact between the BMECs, pericytes and astrocytes that form the BBB and resulting synergistic interactions are lost. We look to overcome this limitation with a direct contact coculture model. Methods We established and optimized a direct interaction coculture system where primary human astrocytes are cultured on the apical surface of a Transwell® filter support and then human cerebral microvessel endothelial cells (hCMEC/D3) seeded directly on the astrocyte lawn. Key findings The studies suggest the direct coculture model may provide a more restrictive and physiologically relevant model through a significant reduction in paracellular transport of model compounds in comparison with monoculture and indirect coculture. In comparison with existing methods, the indirect coculture and monoculture models utilized may limit cell–cell signaling between human astrocytes and BMECs that are possible with direct configurations. Conclusions Paracellular permeability reductions with the direct coculture system may enhance therapeutic agent and potential neurotoxicant screening for BBB permeability better than the currently available monoculture and indirect coculture in‐vitro models.