Abstract
Important reactions of drug metabolism, including UGT mediated glucuronidation and steroidsulfatase mediated hydrolysis of sulfates, take place in the endoplasmic reticulum (ER) of hepatocytes. Consequently, UGT generated glucuronides, like estradiol-17β-glucuronide, have to be translocated back into the cytoplasm to reach their site of excretion. Also steroidsulfatase substrates, including estrone-3-sulfate, have to cross the ER membrane to reach their site of hydrolysis. Based on their physicochemical properties such compounds are not favored for passive diffusion and therefore likely necessitate transport system(s) to cross the ER membrane in either direction.The current study aims to investigate the transport of taurocholate, estradiol-17β-glucuronide, and estrone-3-sulfate in smooth (SER) and rough (RER) endoplasmic reticulum membrane vesicles isolated from Wistar and TR− rat liver.Time-dependent and bidirectional transport was demonstrated for taurocholate, showing higher uptake rates in SER than RER vesicles. For estradiol-17β-glucuronide a fast time-dependent efflux with similar efficiencies from SER and RER but no clear protein-mediated uptake was shown, indicating an asymmetric transport system for this substrate. Estrone-3-sulfate uptake was time-dependent and higher in SER than in RER vesicles. Inhibition of steroidsulfatase mediated estrone-3-sulfate hydrolysis decreased estrone-3-sulfate uptake but had no effect on taurocholate or estradiol-17β-glucuronide transport.Based on inhibition studies and transport characteristics, three different transport mechanisms are suggested to be involved in the transport of taurocholate, estrone-3-sulfate and estradiol-17β-glucuronide across the ER membrane.
Highlights
Biotransformation and elimination of endogenous and exogenous compounds is an important function of the liver
The current study aims to investigate the transport of taurocholate, estradiol-17βglucuronide, and estrone-3-sulfate in smooth (SER) and rough (RER) endoplasmic reticulum membrane vesicles isolated from Wistar and TR- rat liver
The transport systems localized to the plasma membrane are complemented by intracellular enzymes catalyzing phase 1 and phase 2 reactions, converting poorly water-soluble compounds into more water-soluble metabolites to enable their excretion into the bile or the sinusoidal blood plasma
Summary
Biotransformation and elimination of endogenous and exogenous compounds is an important function of the liver. A corresponding animal model are the so-called TR- rats [7] Both CYPs [8] as well as UGTs [9] are anchored in the membrane of the endoplasmic reticulum (ER). Substrates of STS, like estrone-3-sulfate (E3S) or dehydroepiandrosteronesulfate (DHEAS), have to cross the ER membrane to reach their site of hydrolysis Due to their physicochemical properties, these compounds. Based on the knowledge that STS and UGTs have their catalytic site within the ER lumen, the present study investigated and characterized the transport activity for the UGT model product estradiol-17β-glucuronide (E17βG) as well as the STS model substrate estrone-3-sulfate (E3S) in SER and RER vesicles isolated from rat liver. Transport of sulfated and glucuronidated model compounds across SER and RER membranes were functionally compared. Taurocholate transport into RER and SER was investigated to test for potential common transport pathways with phase 2 products
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