Establishment of primary cultures of human brain microvascular endothelial cells to provide an in vitro cellular model of the blood-brain barrier

Michael J. Bernas, Filipa L. Cardoso, Sarah K. Daley, Martin E. Weinand, Alexandre R. Campos, António J.Gonçalves Ferreira, James B. Hoying, Marlys H. Witte, Dora Brites, Yuri Persidsky, Servio H. Ramirez, Maria A. Brito

Research output: Contribution to journalArticlepeer-review

138 Scopus citations

Abstract

We describe a method for generating primary cultures of human brain microvascular endothelial cells (HBMVECs). HBMVECs are derived from microvessels isolated from temporal tissue removed during operative treatment of epilepsy. The tissue is mechanically fragmented and size filtered using polyester meshes. The resulting microvessel fragments are placed onto type I collagen-coated flasks to allow HBMVECs to migrate and proliferate. The overall process takes less than 3 h and does not require specialized equipment or enzymatic processes. HBMVECs are typically cultured for approximately 1 month until confluent. Cultures are highly pure (97% endothelial cells; 3% pericytes), are reproducible, and show characteristic brain endothelial markers (von Willebrand factor, glucose transporter-1) and robust expression of tight and adherens junction proteins as well as caveolin-1 and efflux protein P-glycoprotein. Monolayers of HBMVECs show characteristically high transendothelial electric resistance and have proven useful in multiple functional studies for in vitro modeling of the human blood-brain barrier.

Original languageEnglish (US)
Pages (from-to)1265-1272
Number of pages8
JournalNature Protocols
Volume5
Issue number7
DOIs
StatePublished - Jul 2010

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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