CHARACTERIZATION OF THE CYTOCHROME-P450 CYP2J4 - EXPRESSION IN RAT SMALL-INTESTINE AND ROLE IN RETINOIC ACID BIOTRANSFORMATION FROM RETINAL

The sites of expression in the small intestine and the function of CYP 2J4, a recently identified rat cytochrome (P450) isoform found to be p redominantly expressed in the small intestine, were characterized. Imm unoblot analysis with a polyclonal antibody to heterologously expresse d CYP2J4 revealed that expression of CYP2J4 was at the highest level i n the distal duodenum and jejunum and decreased toward the ileum, Vill ous cells expressed higher levels of CYP2J4 than crypt cells. Isoform- specific RNA polymerase chain reaction indicated that a related P450 i soform, CYP2J3, was only a minor form in rat small intestine. Since th e intestinal mucosa is exposed to high levels of dietary nutrients, we hypothesized that CYP2J4 may be active toward diet-derived factors. W e determined that purified, heterologously expressed CYP2J4 is active toward all-trans-and 9-cis-retinal in reconstituted systems, producing the corresponding retinoic acids as the major products. Apparent K-m values for the formation of retinoic acids were 54 and 49 mu M, respec tively, and apparent V-max values were 20 and 21 nmol/min/nmol P450, r espectively. These activities were readily inhibited by a polyclonal a nti-CYP2J4 antibody, Rat enterocyte microsomes were also active with a ll-trans-retinal to produce all-trans-retinoic acid in the presence of NADPH, and the majority of retinoic acid synthesis activity was inhib ited by the polyclonal anti-CYP2J4 antibody. These findings suggest th at CYP2J4 plays a major role in intestinal microsomal metabolism of re tinal to retinoic acid and may be involved in the maintenance of retin oid homeostasis in the small intestine in vivo. (C) 1998 Academic Pres s.