Transport in Hepatocytes

In the liver drugs are predominantly taken up by the hepatocytes, e.g. by carrier-mediated uptake, metabolized in the hepatocyte and excreted either via the bile canaliculus into the bile or back into the bloodstream, e.g. by carrier-mediated excretion. 

These results also indicate that human hepatocytes are an appropriate model in which to study inter-species differences and the mechanisms of hepatic transport in man. 

In contrast to isolated hepatocytes, liver slices retain the cellular architecture of the Uver without prior digestion with collagenase. This makes a systematic comparison of the data relating to transport of free drugs as well as drug targeting moieties from isolated hepatocytes and liver slices, an attractive model for studying the potential and limitations of the liver slice model in this area of research. 

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A more recent example of this technique has been the study on human absorption characteristics of fexofenadine [109], Fexofenadine has been shown to be a substrate for P-gp in the in vitro cell lines its disposition is altered in knockout mice lacking the gene for MDRla, and co-administration of P-gp inhibitors (e.g. ketoconazole and verapamil) was shown to increase the oral bioavailability of fexofenadine [110-113], Hence, it is suggested that the pharmacokinetics of fexofenadine appears to be determined by P-gp activity. In the human model, the intestinal permeability estimated on the basis of disappearance kinetics from the jejunal segment is low, and the fraction absorbed is estimated to be 2% [114], Co-administration of verapamil/ketoconazole did not affect the intestinal permeability estimates however, an increased extent of absorption (determined by de-convolution) was demonstrated. The increased absorption of fexofenadine was not directly related to inhibition of P-gp-mediated efflux at the apical membrane of intestinal cells as intestinal Peff was unchanged. Furthermore, the effect cannot be explained by inhibition of intestinal based metabolism, as fexofenadine is not metabolised to any major extent. It was suggested that this may reflect modulation of efflux transporters in hepatocyte cells, thereby reducing hepatobiliary extraction of fexofenadine. 

Figure 9.9 Transport systems participating in drug transport in hepatocyte membranes.

Glucose-6-phosphate transporter in hepatocyte microsomal membrane Phosphate transporter in hepatocyte microsomal membrane... 

ATP1 Transports bile acid, bromosulfophtalein, and conjugate and unconjugated steroid hormones, etc. The main basolateral transporter in hepatocyte, called also moat [50]... 

Because of the high and K , of GLUT-2 transporters in hepatocytes, their cytoplasmic glucose is in quasi-equilibrium with that in the blood. Therefore as tissues consume glucose from the blood, they effectively withdraw newly formed glucose from the liver. 

Keusch and Donta reported in 1975 that the enterotoxin killed HeLa cells. This opened the way for the study on the action of the enterotoxin at cellular level. In 1978, Giger and Pariza reported disturbance of amino acid transport in hepatocyte by the enterotoxin. Matsuda and Sugimoto (1979) and McClane and McDonel (1979) reported that morphological alterations induced by the enterotoxin on HeLa and Vero cells looked like those of epithelial cells in sections of ileal loops treated with the enterotoxin. 

In vitro cytotoxicity assays using isolated cells have been applied intermittently to cyanobacterial toxicity testing over several years." Cells investigated for suitability in cyanobacterial toxin assays include primary liver cells (hepatocytes) isolated from rodents and fish, established permanent mammalian cell lines, including hepatocytes, fibroblasts and cancerous cells, and erythrocytes. Earlier work suggested that extracts from toxic cyanobacteria disrupted cells of established lines and erythrocytes," but studies with purified microcystins revealed no alterations in structure or ion transport in fibroblasts or erythrocytes,... 

ABC Transporters. Figure 2 ABC-transporter expression in hepatocytes and enterocytes (modified according to www.iwaki-kk.co.jp/bio/specialedition/se02.htm). 

Bai CL, Stacey NH. 1993. Mechanism of trichloroethylene-induced elevation of individual serum bile acids. II. In vitro and in vivo interference by trichloroethylene with bile acid transport in isolated rat hepatocytes. Toxicol Appl Pharmacol 121 296-302. 

Figure 9.7 Iron transport by hepatocytes. Known proteins involved in iron transport across the plasma membrane of hapatocytes are represented. LMW = low molecular weight Trf = transferrin Trf-R = transferrin receptor HFE = hamochromatosis gene product 132m = 62-microglobulin 02-= superoxide OH- = hydroxyl radical FR = ferritin receptor SFT = stimulator of iron transport.

TfR is low in these patients, PIT is normal and most iron is stored in hepatocytes. In patients with hypoplastic anaemias and with transfusion iron overload, the BM cannot utilize iron, resulting in low TfR expression and decreased iron absorption. Quantitative analysis of all iron fluxes, which can be deduced from Figure 9.1, can assist in understanding the clinical expression of mutations of proteins involved in iron transport. 

Sandker, G. W., Weert, B., Olinga, P., Wolters, H., Slooff, M. J., Meijer, D. K., Groothuis, G. M., Characterization of transport in isolated human hepatocytes. A study with the bile add taurocholic add, the uncharged ouabain and the organic... 

Breast Cancer Resistance Protein (BCRP, also known as MXR or ABCP), first cloned from mitoxantrone and anthracycline-resistant breast and colon cancer cells [188, 189] is a half-transporter efflux pump believed to function as a homo-or hetero-dimer. Following its identification, BCRP-mediated drug resistance was observed for topoisomerase inhibitors including camptothecins [190, 191] and in-dolocarbazoles [192]. In normal tissues, BCRP was detected in placental syncytio-trophoblasts, hepatocyte canalicular membrane, apical intestinal epithelia and vascular endothelial cells [193]. These findings support the important role BCRP plays in modulating topotecan bioavailability, fetal exposure and hepatic elimination [194]. Considering that the substrates and tissue distributions for BCRP overlap somewhat with MDR1 and MRPs [195], additional studies will be required to define the relative contribution of each of these transporters in the overall and tis-... 

Several kinetic parameters can be measured on different experimental systems to account for the interaction of a compound with CYPs. For example when studying the metabolic stability of a compound, it could be measured in a recombinant CYP system, in human liver microsomes, in hepatocytes and so on. Each system increases in biological complexity. Although in the recombinant CYP system only the cytochrome under consideration is studied, in the case of the human liver microsomes, there is a pool of enzyme present that includes several CYPs, and finally in the hepatocyte cell system, metabolizing enzymes play an important role in the metabolic compound stability. In addition, transport systems are also present that could involve recirculation or other transport phenomena. The more complex the experimental system, the more difficult it is to extract information on the protein/ligand interaction, albeit it is closer to the in vivo real situation and therefore to the mechanism that is actually working in the body. 

Figure 15.2 Transport proteins involved in the intestinal absorption and the renal and hepatic excretion of drugs. In the intestine, drugs are taken up from the luminal side into enterocytes before the subsequent elimination into blood. In hepatocytes, drugs are taken up from the blood over the basolateral membrane and excreted over the canalicular membrane into bile. In the renal epithelium, drugs undergo secretion (drugs are taken up from the blood and excreted into the urine) or reabsorption (drugs are taken up from the urine and are excreted back into blood). Uptake transporters belonging to the SLC transporter superfamily are shown in red and export pumps...

GLUT 2, a low-affinity transporter, is in hepatocytes. After a meal, portal blood from the intestine is rich in glucose. GLUT 2 captures the excess glucose primarily for storage. When the glucose concentration drops below the for the transporter, much of the remainder leaves the hver and enters the peripheral circulation. 

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