Isolation and expansion of human and mouse brain microvascular endothelial cells.
Brain microvascular endothelial cells (BMVECs) have an important role in the constitution of the blood-brain barrier (BBB). The BBB is involved in the disease processes of a number of neurological disorders in which its permeability increases. Isolation of BMVECs could elucidate the mechanism involved in these processes. This protocol describes how to isolate and expand human and mouse BMVECs. The procedure covers brain-tissue dissociation, digestion and cell selection. Cells are selected on the basis of time-responsive differential adhesiveness to a collagen type I-precoated surface. The protocol also describes immunophenotypic characterization, cord formation and functional assays to confirm that these cells in endothelial proliferation medium (EndoPM) have an endothelial origin. The entire technique requires ∼7 h of active time. Endothelial cell clusters are readily visible after 48 h, and expansion of BMVECs occurs over the course of ∼60 d.
Laboratory of Cellular Neurobiology, Istituto Di Ricovero e Cura a Carattere Scientifico Foundation Neurological Institute Carlo Besta, Milan, Italy. stefania.navone@unimi.it
Stefania E
SE
Navone
Giovanni
G
Marfia
Gloria
G
Invernici
Silvia
S
Cristini
Sara
S
Nava
Sergio
S
Balbi
Simone
S
Sangiorgi
Emilio
E
Ciusani
Alessandra
A
Bosutti
Giulio
G
Alessandri
Mark
M
Slevin
Eugenio A
EA
Parati
D000818:Animals; D018929:Cell Culture Techniques; D002470:Cell Survival; D018920:Coculture Techniques; D042783:Endothelial Cells / Q000166:cytology*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D051379:Mice; D055806:Microvessels / Q000166:cytology*; D018919:Neovascularization, Physiologic