ATP-binding cassette transporters in immortalised human brain microvascular endothelial cells in normal and hypoxic conditions
1 Department of Neurology, University of Regensburg, Bezirksklinikum Regensburg, Regensburg, Germany
2 Department of Psychiatry, University of Regensburg, Bezirksklinikum Regensburg, Regensburg, Germany
3 Institute of Clinical Chemistry and Laboratory Medicine, Regensburg University Medical Center, Regensburg, Germany
4 Departement de biologie cellulaire (CNRS UMR 8104), Institut Cochin, Université Paris Descartes, Paris, France and INSERM U567, Departement de biologie cellulaire, Institut Cochin, Paris, France
Experimental & Translational Stroke Medicine 2012, 4:9 doi:10.1186/2040-7378-4-9Published: 3 May 2012
Rapid reperfusion following ischemia is the most effective therapy in stroke therapy. However, the success may be compromised by ischemia & reperfusion (I/R) injury and at the human blood–brain barrier (BBB), therefore the effects on transendothelial transport are of special interest. Current studies suggest the ATP-binding cassette (ABC) transporters to be regulated upon ischemic stroke in a way that impedes the effects of drug therapy. The immortalised human brain microvascular endothelial cell line hCMEC/D3 provides most of the unique properties of the BBB with respect to transport and might be a reliable in vitro model to study transendothelial transport after I/R.
We exposed hCMEC/D3 cells to 24 hours of hypoxia alone and to hypoxia followed by 60 min of reoxygenisation as an in vitro model for I/R. Western blot showed mild upregulation of hypoxia inducible factor (HIF-1α) after hypoxia alone and RNA lysates were analysed with a well-established real-time RT-PCR-based TaqMan low-density array detecting 47 of 48 known human ABC transporters.
No significant increases of ABC mRNA expression levels were detected neither in hypoxic nor in I/R samples. However, slight decrease of ABCC1 in hypoxic and I/R samples and of ABCA10 and ABCD3 in I/R samples was observed.
Our data suggests that hCMEC/D3 cell line and – at the moment – in vitro models in general are a poor basis for stroke research but may be enhanced by co-culturing more cells of the neurovascular unit inducing an overall ischemic response at the BBB.