Characterization of Cytochrome P450 Protein Expression along the Entire Length of the Intestine of Male and Female Rats

Abstract

ABSTRACT:Intestinal cytochrome P450 (P450) proteins play an important role in the biotransformation of drugs and may significantly limit their oral absorption and bioavailability. Therefore, we have investigated the amount of P450 proteins via Western blot analysis along the entire intestine of male and female rats. Despite of the use of an inbred rat strain, controlled housing conditions for the animals, and a timed sample preparation, high interindividual differences in the expression of all P450 proteins was observed. CYP3A (135-243 fmol/mg of protein) and CYP2B1 (107-645 fmol/mg of protein) were the most abundant P450 isoforms in the duodenum and jejunum of rat intestine but were present in neither the ileum nor the colon. Compared with CYP2B1 and CYP3A, CYP2D1 (25-71 fmol/mg of protein) and CYP2C6 (3-10 fmol/mg of protein) were only expressed in minor amounts. CYP2C11 could not be identified in the entire rat intestine. In conclusion, this is the first systematic evaluation and quantification of the expression of P450 proteins along the entire length of the intestine in both male and female rats. These data will provide a basis for a better understanding of the extent of intestinal metabolism along the gastrointestinal tract.Absorption through the gut is a key step in the oral delivery of drugs. The main interface between gut lumen and the bloodstream is an epithelial cell layer consisting of polarized enterocytes controlling the passage of exogenous substances into the portal circulation. Enterocytes have a variety of structural features, including tight junctions reducing paracellular permeability, numerous drug transporters, and a set of metabolic enzymes that may all affect the entry of drugs into the body. The extent to which a drug is absorbed also depends on the intrinsic properties of the compound such as solubility, permeability, efflux or uptake transport properties, and susceptibility to metabolic degradation (Martinez and Amidon, 2002;Benet et al., 2004).Although the liver is known as the major site of first-pass extraction, recent studies have indicated that the small intestine also contributes significantly to the first-pass metabolism of many drugs, e.g., cyclosporine (Wu et al., 1995), nifedipine (Iwao et al., 2002), midazolam (Paine et al., 1996, and diltiazem (Iwao et al., 2004). It is known that several uptake and efflux transporter and P450 isoforms are expressed in the human and rat intestine (van de Kerkhof et al., 2007). If a drug is a substrate of efflux transporters, it may enter and exit the enterocytes several times, and, with each cycle, small quantities may be metabolized by cytochrome P450 (P450) enzymes localized in the endoplasmic reticulum. The P450 enzymes belong to a superfamily of heme proteins that show a broad substrate specificity, with substrates ranging in size from ethylene (M r 28) to cyclosporine (M r 1201) (Isin and Guengerich, 2007). Although the liver is regarded as the main organ of drug metabolism, P450 proteins are also expressed in other tissues, e.g., kidn...

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