Immortalized human cerebral microvascular endothelial cells maintain the properties of primary cells in an in vitro model of immune migration across the blood brain barrier.
The immortalized human cerebral microvascular endothelial cell line HCMEC/D3 presents a less expensive and more logistically feasible alternative to primary human brain microvascular endothelial cells (HBMEC's) for use in constructing in vitro models of the blood brain barrier (BBB). However, the fidelity of the HCMEC/D3 cell line to primary HBMEC's in studies of immune transmigration has yet to be established. Flow cytometric analysis of primary human leukocyte migration across in vitro BBB's generated with either HCMEC/D3 or primary HBMEC's revealed that HCMEC/D3 maintains the immune barrier properties of primary HBMEC's. Leukocyte migration responses and inflammatory cytokine production were statistically indistinguishable between both endothelial cell types, and both cell types responded similarly to astrocyte coculture, stimulation of leukocytes with phorbol myristate acetate (PMA) and ionomycin, and inflammatory cytokine treatment. This report is the first to validate the HCMEC/D3 cell line in a neuroimmunological experimental system via direct comparison to primary HBMEC's, demonstrating remarkable fidelity in terms of barrier resistance, immune migration profiles, and responsiveness to inflammatory cytokines. Moreover, we report novel findings demonstrating that interaction effects between immune cells and resident CNS cells are preserved in HCMEC/D3, suggesting that important characteristics of neuroimmune interactions during CNS inflammation are preserved in systems utilizing this cell line. Together, these findings demonstrate that HCMEC/D3 is a valid and powerful tool for less expensive and higher throughput in vitro investigations of immune migration at the BBB.
Department of Anatomy and Neurobiology, Washington University School of Medicine, Saint Louis, MO 63110, USA.
Brian P
BP
Daniels
Lillian
L
Cruz-Orengo
Tracy Jo
TJ
Pasieka
Pierre-Olivier
PO
Couraud
Ignacio A
IA
Romero
Babette
B
Weksler
John A
JA
Cooper
Tamara L
TL
Doering
Robyn S
RS
Klein
D000704:Analysis of Variance; D019540:Area Under Curve; D001253:Astrocytes / Q000187:drug effects / Q000502:physiology; D001812:Blood-Brain Barrier / Q000187:drug effects / Q000502:physiology*; D061207:Calcium Ionophores / Q000494:pharmacology; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism / Q000494:pharmacology; D015194:Diffusion Chambers, Culture; D042783:Endothelial Cells / Q000502:physiology*; D005434:Flow Cytometry; D006801:Humans; D015759:Ionomycin / Q000494:pharmacology; D007962:Leukocytes / Q000187:drug effects / Q000502:physiology*; D008954:Models, Biological; D010634:Phenobarbital / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors
D061207:Calcium Ionophores; D016207:Cytokines; D015759:Ionomycin; D013755:Tetradecanoylphorbol Acetate; D010634:Phenobarbital