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Evaluation of 99mTechnetium-Mebrofenin and 99mTechnetium-Sestamibi as Specific Probes for Hepatic Transport Protein Function in Rat and Human Hepatocytes

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ABSTRACT

Purpose

This study characterized 99mTc-Mebrofenin (MEB) and 99mTc-Sestamibi (MIBI) hepatic transport and preferential efflux routes (canalicular vs. basolateral) in rat and human sandwich-cultured hepatocytes (SCH).

Methods

99mTc-MEB and 99mTc-MIBI disposition was determined in suspended hepatocytes and in SCH in the presence and absence of inhibitors and genetic knockdown of breast cancer resistance protein (Bcrp).

Results

The general organic anion transporting polypeptide (Oatp/OATP) inhibitor rifamycin SV reduced initial 99mTc-MEB uptake in rat and human suspended hepatocytes. Initial 99mTc-MIBI uptake in suspended rat hepatocytes was not Na+-dependent or influenced by inhibitors. Multidrug resistance-associated protein (Mrp2/MRP2) inhibitors decreased 99mTc-MEB canalicular efflux in rat and human SCH. 99mTc-MEB efflux in human SCH was predominantly canalicular (45.8 ± 8.6%) and ∼3-fold greater than in rat SCH. 99mTc-MIBI canalicular efflux was similar in human and rat SCH; basolateral efflux was 37% greater in human than rat SCH. 99mTc-MIBI cellular accumulation, biliary excretion index and in vitro biliary clearance in rat SCH were unaffected by Bcrp knockdown.

Conclusion

99mTc-MEB hepatic uptake is predominantly Oatp-mediated with biliary excretion by Mrp2. 99mTc-MIBI appears to passively diffuse into hepatocytes; biliary excretion is mediated by P-gp. The SCH model is useful to investigate factors that may alter the route and/or extent of hepatic basolateral and canalicular efflux of substrates.

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Abbreviations

99mTc-MEB:

99mTechnetium-mebrofenin

99mTc-MIBI:

99mTechnetium-sestamibi

Bcrp:

breast cancer resistance protein

BEI:

biliary excretion index

Clbiliary :

in vitro biliary clearance

DMEM:

Dulbecco’s modified Eagle’s medium

E217G:

estradiol-17-β-D-glucuronide

FBS:

fetal bovine serum

GF120918:

N-(4-[2-(1,2,3,4-tetrahydro-6,7-dimethoxy-2-isoquinolinyl)ethyl]-phenyl)-9,10-dihydro-5-methoxy-9-oxo-4-acridine carboxamide

HBSS:

Hanks’ balanced salt solution

Mrp:

multidrug resistance-associated protein

NTCP:

sodium taurocholate cotransporter.

OAT:

organic anion transporter

OCT:

organic cation transporter

PAH:

para-aminohippuric acid

P-gp:

multidrug resistance protein P-glycoprotein

RNAi:

RNA interference

SCH:

sandwich-cultured hepatocytes

siBcrp:

RNAi short hairpin knockdown of rat Bcrp

siNT:

non-target siRNA

TEA:

tetraethylammonium

WT:

wild-type

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ACKNOWLEDGMENTS

This research was supported by a grant from the National Institutes of Health (R01GM41935). Brandon Swift was supported by an Eli Lilly and Company Foundation predoctoral fellowship. The authors would like to thank Arlene S. Bridges, Ph.D., for her analytical support; Yiwei Rong for her technical expertise in the isolation of rat hepatocytes; and CellzDirect, Life Technologies, for kindly providing freshly isolated suspended and sandwich-cultured human hepatocytes.

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Authors and Affiliations

  1. Division of Pharmacotherapy and Experimental Therapeutics UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, 27599-7569, USA

    Brandon Swift, Wei Yue & Kim L. R. Brouwer

  2. UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, 311 Pharmacy Lane, CB#7569, 3205, Kerr Hall Chapel Hill, North Carolina, 27599-7569, USA

    Kim L. R. Brouwer

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  1. Brandon Swift
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Correspondence to Kim L. R. Brouwer.

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Swift, B., Yue, W. & Brouwer, K.L.R. Evaluation of 99mTechnetium-Mebrofenin and 99mTechnetium-Sestamibi as Specific Probes for Hepatic Transport Protein Function in Rat and Human Hepatocytes. Pharm Res 27, 1987–1998 (2010). https://doi.org/10.1007/s11095-010-0203-x

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