Interaktion zwischen Metabolismus und Transport von Pyren in Caco-2-Zellen als Modell der gastrointestinalen Barriere ; Interaction between metabolism and transport of pyrene in Caco-2 cells representing the gastrointestinal barrier

Polycyclic aromatic hydrocarbons (PAH´s) represent a class of substances formed in all types of incomplete combustion of organic compounds. Some of its members have - due to certain molecular structure chracteristics – procarcinogenic properties in both man and laboratory animals. PAH´s are metabolized within the organism by the almost ubiquitous classes of cytochrome P450 enzymes and epoxide hydrolases (functionalization, phase 1) and sulfotransferases, UDP-glucuronosyltransferases and glutathione-S-transferases (conjugation, phase 2). The enzymatic reactions of phase 1 result in an increased reactivity with cellular macromolecules (including DNA), but are also the prerequisite for phase 2 reactions which increase the molecules´ hydrophilicity decisively, enabling the organism to excrete the PAH´s via the kidney, the bile and (as recently shown for the carcinogenic PAH benzo[a]pyrene) the intestinal mucosa. In the present work, the existence of a biphasic metabolism and a subsequent metabolite transport via the intestinal epithelium was examined and demonstrated for the noncarcinogenic PAH pyrene, including the identification of the participating enzymes, their products and the proteins involved in the transport. The human intestinal cell line Caco-2 was chosen as a model for the intestinal epithelium by reason of its close similarity (in morphology and cellular biochemistry) with small bowel enterocytes. The ability of different human sulfotransferases to metabolize pyrene and pyrene-1-hydroxide was examined in the hamster cell line V79, the role of the transport protein cMOAT/MRP2 in the metabolite transport was investigated using the canine cell line MDCK II. The results can be summarized as follows: 1. In Caco-2 cells, pyrene is metabolized to pyrene-1-hydroxide (phase 1). 2. The hydroxilation of pyrene is carried out mainly by the cytochromes P450 1A1, P450 1A2 and/or P450 1B1. This reaction is inducible by several CYP1A1 inductors and can be inhibited by α-naphtoflavone, an inhibitor of CYP1A1, CYP1A2 and ...