Identification of N2-(deoxyguanosin-8-yl)-2-amino-3,8-dimethylimidazo[4,5-f)quinoxaline 3',5'-diphosphate, a major DNA adduct, detected by nuclease P1 modification of the 32P-postlabeling method, in the liver of rats fed MeIQx

The carcinogenic heterocyclic amine 2-amino-3,8-dimethyl-imidazo[4,5-f]quinoxaline(MeIQx) is widely distributed in cooked foods. The nucleaseP1 method increased the sensitivity of the standard ³²P-postlabelinganalysis about 1000-fold for detection of MeIQx-DNA adducts.The recovery of MeIQx-DNA adducts by the nuclease P1 methodwas determined to be about 50% using liver DNA of a rat treatedwith [¹⁴C]MeIQx intragastrically. By the nuclease P1 methodfive adducts were detected in the liver DNA of rats fed MeIQxand two of them, including the most abundant one, were identifiedas MeIQx-deoxyguanosine adducts by comparison with the adductsformed in in vitro reactions of N-acetoxy-2-amino-3,8-dimethylimidazo[4,5-f)quinoxalinewith the four 2'-deoxyribonucleotides. The most abundant adductin vivo was identified as N²-(deoxyguanosin-8-yl)-MeIQx 3',5'-diphosphate(3',5'-pdGp-C8-MeIQx). MeIQx-DNA adduct levels in human tissuescould be determined by the nuclease P1 modification of the ³²P-postlabelingmethod in combination with HPLC, and thus provide informationon the roles of MeIQx in human carcinogenesis.     

Impact of commercial cigarette smoke condensate on brain tissue co-cultured with astrocytes and blood–brain barrier endothelial cells

The purpose of the current study was to investigate the effect of two commercial cigarette smoke condensates (CCSC) on oxidative stress and cell cytotoxicity in human brain (T98G) or astrocytes (U-373 MG) in the presence of human brain microvascular endothelial cells (HBMEC). Cell viability of mono-culture of T98G or U-373 MG was markedly decreased in a concentration-dependent manner, and T98G was more susceptible than U-373 MG to CCSC exposure. Cytotoxicity was less prominent when T98G was co-cultured with HBMEC than when T98G was co-cultured with U-373 MG. Significant reduction in trans-epithelial electric resistance (TEER), a biomarker of cellular integrity was noted in HBMEC co-cultured with T98G (HBMEC-T98G co-culture) and U-373 MG co-cultured with T98G (U-373 MG-T98G co-culture) after 24 or 48 hr CCSC exposure, respectively. TEER value of U-373 MG co-cultured with T98G (79–84%) was higher than HBMEC co-cultured with T98G (62–63%) within 120-hr incubation with CCSC. Reactive oxygen species (ROS) generated by CCSC in mono-culture of T98G and U-373 MG reached highest levels at 4 and 16 mg/ml, respectively. ROS production by T98G fell when co-cultured with HBMEC or U-373MG. These findings suggest that adverse consequences of CCSC treatment on brain cells may be protected by blood–brain barrier or astrocytes, but with chronic exposure toxicity may be worsened due to destruction of cellular integrity.     

Adduct Formation at C-8 of Guanine on in vitro Reaction f the Ultimate Form of 2-Amino-l-methyl-6-phenylimidazo[4,5-b]pyridine with 2'-Deoxyguanosine and Its Phosphate Esters

We examined the reactivity of the N -hydroxyamino derivative of a carcinogenic heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5- b ]pyridine (PhIP), after its O -acetylation with four 2'-deoxyribonucleoside 3'-monophosphates. ³²P-Postlabeling analysis demonstrated that the levels of adducts with 2'-deoxyguanosine 3'-moiiophosphate were much higher than those with the other three nucleotides. ¹H-NMR, mass spectral and UV absorption spectral analyses of the major adducts formed by N -acetoxy-PhIP with 2'-deoxyguanosine and with its phosphate esters indicated that PhIP bound at the C-8 position of guanine, as previously demonstrated with other heterocyclic amines.     

13-Week oral toxicity study of oil derived from squid (Todarodes pacificus) in Sprague-Dawley rats

Recommendations to increase the consumption of the long-chain omega-3 fatty acids are challenged by the global problem of declining fish stocks. Non-traditional and more sustainable sources of the long-chain omega-3 fatty acids are needed. Squid (Todarodes pacificus) represents a uniquely sustainable source of these fatty acids. A 13-week oral toxicity study was conducted in male and female Sprague-Dawley rats administered either 0, 250, 500, or 1000 μl/kg body weight (bw)/day of a refined squid oil. All of the rats survived through to the end of the study. All of the rats grew normally and had normal clinical and ophthalmic observations. No signs of toxicity were evident from clinical chemistry, hematology, and urinalysis data measured. No abnormal findings attributable to exposure to purified squid oil were observed following the necropsy of male and female rats and the histopathological examination of the organs. The no-observed-adverse-effect level for refined squid oil was determined to be 1000 μl/kg bw/day, the highest dose tested.