Modulation of N-nitrosomethylbenzylamine metabolism by black raspberries in F344 rats

2484 Previosly our laboratory has demonstrated that dietary freeze-dried black raspberries (BRB) inhibit N-nitrosomethylbenzylamine (NMBA)-induced tumorigenesis in the F344 rat esophagus. The present study was undertaken to determine the mechanistic basis of the anti-initiating effects of BRB, and to identify the active inhibitory component(s). Rats were administered AIN-76A diet containing either 5% or 5μmole phenylethylisothiocyanate (PEITC), a known inhibitor of cytochrome P450s (CYP450s) for three weeks. During the third week of dietary treatments, rats were administered three s.c. doses of NMBA (0.5 mg/kg b.w.) or the vehicle and sacrificed 24 h after the last dose. The livers were snap frozen in liquid nitrogen and the esophagus aseptically removed and explant cultures prepared for metabolic studies. Our preliminary results showed that addition of 5%BRB or PEITC to the diet inhibited NMBA metabolism in the esophageal explant by 25% and 80% respectively compared to the control rats. To identify the active inhibitory component(s) of BRB that influence NMBA metabolism, the explant cultures from control rats were further treated in vitro with either an ethanol extract of BRB or with individual components of BRB; ellagic acid and anthocyanidins (cyanidin glycoside and cyanidin rutinoside) known to be present in BRB. Results show that [3H]-NMBA metabolism compared to untreated explants was inhibited maximally in explants treated with cyanidin rutinoside (42%) followed by ellagic acid (25%), cyanidin glycoside (21%) and ethanol extract of BRB (11%). Gene expression analysis was also performed using Agilent oligonucleotide microarrays to examine the effects of BRB treatment on Phase I and II genes in the rat esophagus. Preliminary data was obtained using a single sample and respective microarray from each of: untreated, NMBA, 5% BRB, and 5% BRB+NMBA treatments. Using a minimum 2-fold change cut off, many genes were identified as differentially expressed between the treatments, including several phase I and phase II metabolizing enzymes. Currently, we are using DNA microarray. Currently similar studies are being conducted in liver cytosol and microsomes to examine the effects of BRB treatment, both in vivo and in vitro on Phase I and II enzyme activities in the rat liver. Supported by the NIH grant CA 745152.