Selective gene silencing of rat ATP-binding cassette G2 transporter in an in vitro blood-brain barrier model by short interfering RNA

Satoko Hori, Sumio Ohtsuki, Masashi Ichinowatari, Takanori Yokota, Takashi Kanda, Tetsuya Terasaki

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

The aim of the present study was to specifically silence the rat ATP-binding cassette transporter G2 (rABCG2) gene in brain capillary endothelial cells by transfection of short interfering RNA (siRNA). Four different siRNAs designed to target rABCG2 were each transfected into HEK293 cells with myc-tagged rABCG2 cDNA. Quantitative real-time PCR and western blot analyses revealed that three of the siRNAs were able to reduce exogenous rABCG2 mRNA and protein levels in HEK293 cells. Moreover, rABCG2-mediated mitoxantrone efflux transport was suppressed by the introduction of these three siRNAs into HEK293 cells. In contrast, the other siRNA and non-specific control siRNA did not significantly affect the mRNA expression, the protein level or the transport activity. Endogenous rABCG2 mRNA and protein expression in a conditionally immortalized rat brain capillary endothelial cell line (TR-BBB13) was suppressed by the most potent siRNA among the four siRNAs tested. Furthermore, this siRNA did not affect the mRNA levels of other ABC transporters, such as ABCB1, ABCC1 and ABCG1, and the protein level of ABCB1 in TR-BBB13 cells, suggesting that it can selectively silence rABCG2 at the blood-brain barrier. This should be a useful and novel strategy for clarifying the contribution of rABCG2 to brain-to-blood transport of substrate drugs and endogenous compounds across the blood-brain barrier.

Original languageEnglish
Pages (from-to)63-71
Number of pages9
JournalJournal of Neurochemistry
Volume93
Issue number1
DOIs
Publication statusPublished - 2005 Apr 1
Externally publishedYes

Fingerprint

ATP-Binding Cassette Transporters
Gene Silencing
Blood-Brain Barrier
Small Interfering RNA
Rats
Genes
Adenosine Triphosphate
HEK293 Cells
Brain
Messenger RNA
Endothelial cells
Proteins
Endothelial Cells
In Vitro Techniques
Mitoxantrone
Transfection
Real-Time Polymerase Chain Reaction
Blood
Complementary DNA
Western Blotting

Keywords

  • 17β-estradiol
  • ABC transporter
  • ATP-binding cassette transporter G2
  • Blood-brain barrier
  • In vitro blood-brain barrier model
  • Short interfering RNA

ASJC Scopus subject areas

  • Biochemistry
  • Cellular and Molecular Neuroscience

Cite this

Selective gene silencing of rat ATP-binding cassette G2 transporter in an in vitro blood-brain barrier model by short interfering RNA. / Hori, Satoko; Ohtsuki, Sumio; Ichinowatari, Masashi; Yokota, Takanori; Kanda, Takashi; Terasaki, Tetsuya.

In: Journal of Neurochemistry, Vol. 93, No. 1, 01.04.2005, p. 63-71.

Research output: Contribution to journalArticle

Hori, Satoko ; Ohtsuki, Sumio ; Ichinowatari, Masashi ; Yokota, Takanori ; Kanda, Takashi ; Terasaki, Tetsuya. / Selective gene silencing of rat ATP-binding cassette G2 transporter in an in vitro blood-brain barrier model by short interfering RNA. In: Journal of Neurochemistry. 2005 ; Vol. 93, No. 1. pp. 63-71.
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