Establishment of Madin-Darby Canine Kidney Cell Line with High P-Glycoprotein Expression

LIU Yo;YU Chun-n;ZENG Su

Chinese Pharmaceutical Journal ›› 2009, Vol. 44 ›› Issue (21) : 1608-1613.

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PDF(3361 KB)
Chinese Pharmaceutical Journal ›› 2009, Vol. 44 ›› Issue (21) : 1608-1613.
Article

Establishment of Madin-Darby Canine Kidney Cell Line with High P-Glycoprotein Expression

  • LIU Yao , YU Chun-na , ZENG Su*
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Abstract

OBJECTIVE To establish Madin-Darby canine kidney (MDCK) cell line with stable expression of human P-glycoprotein (P-gp) and scree an effective model to study drug transportation. METHODS The plasmid pcDNA3.1(+)/MDR1 was transfected into MDCK cell line using LipofectamineTM 2000 transfection reagent. Several stable transfected clones were obtained after selection with G418. Rhodamine123 (Rho123), the fluorescence probes substrate, was used to study the profiles of efflux and uptake to test P-gp activities, which was performed with MDCK and MDCK-MDR1 (Madin-Darby canine kidney/multidrug resistance 1)cells in a one-chamber system with the presence or absence of verapamil as inhibitor. The expression of P-gp messenger ribonucleic acid (mRNA) and protein was detected by using reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. This study measured the bidirectional transport of Rho123 across MDCK and MDCK-MDR1 monolayers with the absence or presence of four well-known P-gp inhibitors (verapamil, cyclosporina, ketoconazole, quinine). RESULTS The RT-PCR and Western blot result indicated high homo sapiens P-gp expression in transfected cells comparing with controls. The Rho123 efflux ratios of MDCK to MDCK-MDR1 were 5.94 and 15.45, respectively. Several known P-gp inhibitors could significantly reduced the polarized efflux. CONCLUSION The developed MDCK-MDR1 cell line is an effective model to study drug transportation in body.

Key words

Madin-Darby canine kidney cell / multidrug resistance 1 / P-glycoprotein / transport

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LIU Yo;YU Chun-n;ZENG Su . Establishment of Madin-Darby Canine Kidney Cell Line with High P-Glycoprotein Expression[J]. Chinese Pharmaceutical Journal, 2009, 44(21): 1608-1613

References


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( 收稿日期 : 2009-01-04 )


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