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Keywords:

  • aortic endothelial cells;
  • astrocytes;
  • blood-brain barrier;
  • murine brain microvascular endothelial cell;
  • coculture

Abstract

Construction of in vitro models of the blood-brain barrier (BBB) using primary brain microvascular endothelial cells (BMEC) is time intensive and not high throughput, in part due to a lack of culture purity, low yields, and cellular dedifferentiation after the first passage. This problem has created interest in the substitution of BMEC with immortalized brain endothelial cells (EC), or peripheral EC such as bovine aortic EC (BAEC). Many BBB models have focused on further inducing the brain and peripheral ECs by incorporating astrocyte back-to-back or nonback-to-back cocultures. However, previous studies demonstrating induction effects of astrocytes on BAEC in back-to-back cocultures failed to recognize the extensive barrier properties of astrocytes alone, which can have a significant effect on interpreting the results. This manuscript reports the establishment of back-to-back and nonback-to-back cocultures between astrocytes and BAEC or BMEC (as a control) with primary focus on the properties of astrocytes alone and with a linear contrast statistical methodology to interpret the results. Transendothelial electrical resistance and permeability studies revealed that astrocytes can significantly increase the barrier tightening of BMEC by 167%, while having no effect on BAEC. Immunocytochemical studies also revealed the reorganization of BMEC occludin junctions in the presence of astrocytes, while indicating the absence of this junctional protein in BAEC. In contrast to a previous report, here the linear contrast statistical analysis revealed that observed decreases in permeability of BAEC in back-to-back cocultures is due to the addition of astrocytes' properties in series and not due to induction. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011