Establishing a method to isolate rat brain capillary endothelial cells by magnetic cell sorting and dominant mRNA expression of multidrug resistance-associated protein 1 and 4 in highly purified rat brain capillary endothelial cells

Pharm Res. 2007 Apr;24(4):688-94. doi: 10.1007/s11095-006-9188-x. Epub 2007 Feb 22.


Purpose: To establish a method for isolating highly purified brain capillary endothelial cells (BCECs) from rat brain by using magnetic cell sorting, and clarify the expression levels of multidrug resistance-associated protein (Mrp) subtypes in these highly purified BCECs.

Methods: The cells were prepared from the capillary enriched-fraction by enzyme digestion, and reacted with anti-PECAM-1 antibody. The cell sorting was performed by autoMACS. The mRNA levels were measured by quantitative real-time PCR analysis.

Results: From five rats, 2.3 x 10(6) cells were isolated in the PECAM-1(+) fraction and the percentage of labeled cells in this was 85.9%. PECAM-1, claudin-5 and Tie-2 mRNA were concentrated in the PECAM-1(+) fraction compared with rat brain. The contamination by neurons and astrocytes was markedly less than in the brain capillary fraction prepared by the glass bead column method. Mrp1 and 4 were predominantly expressed in the PECAM-1(+) fraction at similar levels to Mdr1a. The mRNA levels of Mrp5 and 3 were 10.6 and 7.60% of that of Mrp1, respectively.

Conclusions: This new purification method provides BCECs with less contamination by neural cells. In the isolated BCECs, Mrp1 and 4 are predominantly expressed, suggesting that they play an important role at the rat blood-brain barrier.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • ATP Binding Cassette Transporter, Subfamily B / analysis
  • ATP Binding Cassette Transporter, Subfamily B / genetics
  • ATP-Binding Cassette Transporters / analysis
  • ATP-Binding Cassette Transporters / genetics
  • Animals
  • Astrocytes / chemistry
  • Brain / blood supply*
  • Capillaries / chemistry
  • Capillaries / cytology
  • Capillaries / immunology
  • Claudin-5
  • Endothelial Cells / chemistry*
  • Endothelial Cells / immunology
  • Flow Cytometry
  • Gene Expression*
  • Immunomagnetic Separation / methods*
  • Male
  • Membrane Proteins / analysis
  • Membrane Proteins / genetics
  • Multidrug Resistance-Associated Proteins / analysis*
  • Multidrug Resistance-Associated Proteins / genetics
  • Neurons / chemistry
  • Platelet Endothelial Cell Adhesion Molecule-1 / analysis*
  • Platelet Endothelial Cell Adhesion Molecule-1 / genetics
  • Polymerase Chain Reaction
  • RNA, Messenger / analysis*
  • Rats
  • Rats, Wistar
  • Receptor, TIE-2 / analysis
  • Receptor, TIE-2 / genetics


  • ATP Binding Cassette Transporter, Subfamily B
  • ATP-Binding Cassette Transporters
  • Abcc4 protein, rat
  • Abcc5 protein, rat
  • Claudin-5
  • Cldn5 protein, rat
  • Membrane Proteins
  • Multidrug Resistance-Associated Proteins
  • Platelet Endothelial Cell Adhesion Molecule-1
  • RNA, Messenger
  • multidrug resistance protein 3
  • Receptor, TIE-2
  • multidrug resistance-associated protein 1