Acetoxy-4-methylcoumarins confer differential protection from aflatoxin B(1)-induced micronuclei and apoptosis in
1V.P. Chest Institute, University of Delhi, 110007, Delhi, India. rajhg@yahoo.com
Abstract:
The ability of various acetoxy derivatives of 4-methylcoumarins to inhibit the genotoxic changes due to aflatoxin B(1) (AFB(1)) is reported here. Several 4-methylcoumarins (test compounds), such as 7,8-diacetoxy-4-methylcoumarin (DAMC), monoacetoxy-4-methylcoumarin (MAC), 5-N-acetyl-6-acetoxy-4-methylcoumarin (NAMC) and 7,8-dihydroxy-4-methylcoumarin (DHMC) were separately administered intraperitoneally (i.p.) to male wistar rats followed by AFB(1) administration i.p. or intratracheally (i.t.) (2-8 mg/kg b.wt.) and another dose of the test compound. The animals were sacrificed 26h after AFB(1) administration. From animals receiving AFB(1) i.p., bone marrow (BM) cells were isolated and stained with Mayer's haematoxylin and eosin. Micronuclei (MN) in BM were scored by light microscopy. From animals receiving AFB(1) i.t., bronchoalveolar lavage (BAL) was obtained, lung cells (LG) were isolated and stained with fluorochrome 6-diamidino-2-phenylindole (DAPI) for the analysis of MN, apoptotic bodies (AP) and cell cycle variations. Rats were separately treated with the vehicle DMSO to serve as the proper control. AFB(1) caused significant dose-dependent induction of MN in BM as well as LG. AP were observed in LG of rats receiving AFB(1) and was found to correlate with MN induction. DAMC injection caused significant decrease in AP due to AFB(1) in LG and MN in both BM and LG. The effectiveness of MAC was approximately half that of DAMC, thereby indicating that number of acetoxy groups on the coumarin molecule determine the efficacy. The fact that NAMC had no effect either on MN or AP indicate that neither acetoxy group at C-6 nor the N-acetyl group at C-5 facilitate the transfer of acetyl group to P-450 required for inhibition of AFB(1)-epoxidation. DHMC, the deacetylated product of DAMC had no normalizing effect on the induction of MN and AP. These findings confirm our earlier hypothesis that DAMC-mediated acetylation of microsomal P-450 (catalysing epoxidation of AFB(1)) through the action of microsomal transacetylase is responsible for the protective action of DAMC. The relative number and position of acetoxy groups on the coumarin nucleus determine the specificity to the transacetylase necessary for the chemopreventive action.
