Plasma canalicular

If the unbound drug concentrations in plasma are higher than their K values on the transporters, then transporter function may be significantly affected [106], Following a pharmacokinetic analysis of the effect of probenecid on the hepatobiliary excretion of methotrexate, it has been shown the extent of an in vivo drug-drug interaction can be quantitatively predicted from the kinetic parameters for transport across the sinusoidal and bile canalicular membranes determined in vitro [107]. 

The phosphatidylcholine in bile is synthesised in the endoplasmic reticulum of the hepatocyte and must be transported to the canalicular membrane. One possibility involves the nonspecific phosphatidylcholine transfer protein but a mouse null for this protein did not show reduced phosphatidylcholine secretion into bile and there was no compensatory increase in other phospholipids transfer proteins. However, the plasma membrane would receive a ready supply of phospholipid by insertion of vesicles, and the MDR3 protein translocates this molecule from the inner leafiet to the outer surface where there is contact with bile acids, as suggested by Smit and colleagues. The role of this transporter is shown in Figure 2.2. 

LIVER Use of isolated perfused liver in studies of biological transport processes, 192, 485 measurement of unidirectional calcium ion fluxes in liver, 192, 495 preparation and specific applications of isolated hepatocyte couplets, 192, 501 characterizing mechanisms of hepatic bile acid transport utilizing isolated membrane vesicles, 192, 517 preparation of basolateral (sinusoidal) and canalicular plasma membrane vesicles for the study of hepatic transport processes, 192, 534. 

BCRP is classified in ABCG subfamily other members of this subfamily are involved in sterol transport (269). Unlike P-gp and MRPs, BCRP consists of a single ABC cassette in the amino terminal followed by six putative transmembrane domains however, it forms a homodimer linked by a disulfide bond in the plasma membrane (270,271). Initially, ABCG2 was identified as an mRNA expressed in placenta (272) and as a non-MDRl- and non-MRP-type resistance factor from cell lines selected in the presence of anthracy-clines and mitoxantrone (273). BCRP is expressed widely in the normal tissues (274) and localized on the canalicular membrane of the hepatocytes and apical membranes of epithelial cells (274,275) and brain capillary endothelial cells (276,277). 

Meier PJ, St. Meier-Abt A, Barrett C, et al. Mechanisms of taurocholate transport in canalicular and basolateral rat liver plasma membrane vesicles. Evidence for an electrogenic canalicular organic anion carrier. J Biol Chem 1984 259 10614—10622. 

Rat liver canalicular membrane vesicles (CMV) have been used to examine the mechanisms of uptake of P-gp substrates such as daunomycin, daunorubicin, and vinblastine, whose biliary excretion is extensive (47,137, 408,409). Early work with plasma membrane vesicles, partially purified from MDR human KB carcinoma cells that accumulated [3H]vinblastine in an ATP-dependent manner, definitively showed how P-gp can act to efflux substrates from cancer cells (410). Additionally, these vesicles have been used to study microscopic aspects of P-gp-mediated efflux, such as the relationship of P-gp function to the membrane fluidity (137). 

The preparation of plasma membrane vesicles from liver canalicular membrane is highly enriched with the canalicular (apical) isoform MRP2 (Buchler et al. 1996). Methods for the isolation of hepatocyte canalicular membranes from liver tissue have been described in detail (Bohme et al. 1994 and Boyer and Meier et al. 1990). The percentage of inside-out-oriented vesicles in these preparations amounts to 32%. Alternatively, transfected I ILK and MDCK cells are often used to study ATP-dependent transport into inside-out vesicles (Cui et al. 1999 Leier et al. 2000). 

Wojtal KA, De Vries E, Floekstra D, van Ijzendoorn SC. Efficient trafficking of MDRl/P-glycoprotein to apical canalicular plasma 52. membranes in HepG2 cells requires PKA-RIIalpha anchoring and glucosylceramide. Mol. Biol. Cell 2006 17 3638-3650. 

The surfece-cormected open canalicular system (OCS) provides access to the interior for plasma-bome substances and it serves as a conduit for products secreted during the release reaction (Fig. 3). 

The bile canaliculus is formed as a bile capillary by means of a groove-like canal in the intercellular space, bounded by 2 adjacent liver cells. The bile canaliculi have no walls of their own, but are surrounded by a special zone of the cell membrane (so-calledpericanalicular ectoplasm). Their diameter amounts to 0.5-1.0 pm. They are interconnected and form an extensive polygonal network. The surface area of the bile capillaries is increased by microvilli, which show great functionally determined variability. The canalicular membrane constitutes 10% of the total plasma membrane in the hepatocytes. Similar to the pericanalicular ectoplasm, the hepatocytes contain contractile microfilaments and other components of the cytoskel-eton. These canaliculi are supplied with carrier proteins and enzymes to control bile secretion. (2,34)... 

Ciclosporin can cause cholestasis and cellular necrosis by an inhibitory effect on hepatocyte membrane transport proteins at both sinusoidal and canalicular levels. It induces oxidative stress by accumulation of various free radicals. Ademetionine (5-adenosylmethionine) is a naturally occurring substance that is involved in liver detoxification processes. The efficacy of ademetionine in the treatment and prevention of ciclosporin-induced cholestasis has been studied in 72 men with psoriasis (89). The patients who were given ciclosporin plus ademetionine had low plasma and erythrocyte concentrations of oxidants and high concentrations of antioxidants. The authors concluded that ademetionine may protect the hver against hepatotoxic substances such as ciclosporin. 

Enzymes are found at different locations within cells. AST, ALT, and LD are cytosolic enzymes. As such, they can be released with cell injury, and appear in plasma relatively rapidly. In the case of AST and ALT, there are both mitochondrial and cytosolic isoenzymes in hepatocytes and other cells containing these enzymes. In the case of ALT, the relative amount of mitochondrial isoenzyme is small, and its plasma half-life is extremely short, making it of no diagnostic significance. In the case of AST, the mitochondrial isoenzyme represents a significant fraction of total AST within hepatocytes. In contrast, ALP and GGT are membrane-bound glycoprotein enzymes. The most important location of both enzymes is on the canalicular membrane of hepatocytes. 

Laboratory features of cholestasis vary, depending on whether the process causes complete or partial impairment of biliary drainage. The common feature of aU cholestatic disorders is an increase in plasma activities of canalicular enzymes, such as ALP and GGT. Because this process involves both increased synthesis of enzyme and release of enzyme from its membrane bound forms, there is generally a short lag period between the onset of cholestasis and the increase in plasma activities. In the early stages of an acute mechanical obstruction (especially from gallstones), there may be transient increases in plasma activities of liver cytosolic enzymes, such as AST and ALT. Plasma AST and ALT may exceed 400 lU/L, and in 1% to 2% of cases are more than 2000 lU/L. Even in the presence of continued obstruction, AST and ALT activity gradually decrease, and AST is typically within the reference interval within 8 to 10 days. 

Interestingly, the distribution of BCRP in normal tissues is similar to that of P-glycoprotein. High levels of BCRP expression were detected in the human placenta syncytiotrophoblast plasma membrane, facing the maternal bloodstream, in the canalicular membrane of the liver hepatocytes, the apical membrane of the epithelium in the small and large intestines, in the ducts and lobules of the breast, and in the luminal surface of brain capillaries.260 261 In addition, significant amounts of... 

Biliary excretion of contrast medium is affected by the bile flow (688, 734, 745, 777, 778). Bile is isosmotic with plasma and is produced from the transport of water from the liver cell into the bile canaliculi (canalicular bile flow) and from the excretion and reabsorption of water and electrolytes in the bile ductules (ductular bile flow). Bile flow is increased by taurocholate and dehydrocholate their presence in the canaliculi creates an osmotic gradient that produces the flow of water and solute. There is a positive correlation between the canalicular bile flow stimulated by taurocholate and the amount of iopanoic acid excreted by the liver. Feeding the patient a fatty meal or taurocholate at the time that iopanoic acid is administered can improve the quality of cholecystograms(734). 

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