Use of membrane vesicles isolated from numerous species (including humans) for transport characterization studies has been popular for some time now (Murer and Kinne, 1980; Hillgren et al., 1995; Sinko et al., 1995). Membrane vesicles prepared from intestinal tissues provide the flexibility of examining the interaction of drugs to a specific membrane of interest (e.g., brush-border membrane vs. basolateral membrane of enterocytes). Vesicles offer a unique opportunity to study the properties of drug and nutrient transport at the cellular level (brush-border as well as basolateral side). Membrane vesicle studies allow a complete manipulation of solute environment both inside and outside of the vesicle, thus making it an ideal system for mechanistic absorption studies. Studies with vesicles also facilitate isolation and identification of transporter proteins that are specifically expressed either on the brush-border or the basolateral side of the membranes.
Compared to the other conventional in vitro techniques (Ussing chamber and intestinal perfusions), the vesicles studies are much better suited when availability of sufficient quantities of drug becomes difficult. Drug uptake study in membrane vesicle can be performed with a small amount of drug (mg quantity), and typically multiple experiments can be performed with vesicles prepared from a single laboratory animal. A unique advantage of this method is that vesicles can be cryop-reserved and used for a long duration. Thus the vesicle system offer unique experimental capabilities not possible with more integrated methodologies. However, prepared vesicles are not pure and very often contain other membrane or organelle fragments. It is practically not feasible to isolate 100% pure brush-border membrane vesicles (or basolateral vesicles) without the contamination with the other type of vesicles. During the process of isolation of vesicles often leads to damage of the transporter proteins and enzymes. Since the volume of the vesicles is very small, they typically require a sensitive analytical method.
Varieties of cell monolayer models that mimic in vivo intestinal epithelium in humans have been developed and currently enjoy widespread popularity. Unlike enterocytes, human immortalized (tumor) cells grow rapidly into confluent mono-layers followed by a spontaneous differentiation providing an ideal system for transport studies. Table 5.2 lists a few of these cell models commonly used for permeability assessment in drug discovery and development.
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