Yoko Matsuda

Antioxidant Effects of Flavonoids Used as Food Additives (Purple Corn Color, Enzymatically Modified Isoquercitrin, and Isoquercitrin) on Liver Carcinogenesis in a Rat Medium-Term Bioassay

To clarify the effects of purple corn color, enzymatically modified isoquercitrin (EMIQ), and isoquercitrin (IQ), registered as natural food additives in Japan, on liver carcinogenesis in vivo, a medium-term bioassay was employed. A total of 100 male F344 rats were divided into 5 groups; groups 1 to 4 were given a single intraperitoneal injection of diethylnitrosamine (200 mg/kg b.w.) on day 1. From weeks 2 to 8, they were administered basal diet purple corn color, EMIQ, or IQ as containing test chemicals at doses of 1.0% (groups 1 and 5), 0.1% (group 2), 0.01% (group 3), or 0% (group 4) (experiments 1, 4, and 5). All rats were subjected to two-thirds partial hepatectomy at week 3 and were sacrificed at week 8. Purple corn color exerted no significant modifying effects on GST-P positive foci, preneoplastic foci, development in the liver. However, serum of rats treated with purple corn color provided evidence of antioxidant power significantly by potential antioxidant (PAO) test in vivo (experiment 2). And microarray analyses showed purple corn color to induce RNA expression such as P450 (cytochrome) oxidoreductase, phosphatidylinositol 3-kinase, and phospholipase A2 (experiment 3). Higher doses of EMIQ or IQ with strong antioxidant power in vivo by PAO test treated groups were correlated with smaller numbers of GST-P positive foci, with Spearmans rank correlation coefficients of P= 0.002 and P= 0.049, respectively (experiments 4 and 5). Therefore, the tested food additives may be effective as antioxidants in vivo and have chemopreventive potential against liver preneoplastic lesion development.

Equivocal Colonic Carcinogenicity of Aloe arborescens Miller var. natalensis Berger at High‐Dose Level in a Wistar Hannover Rat 2‐y Study

A 2-y carcinogenicity study of Aloe, Aloe arborescens Miller var. natalensis Berger, a food additive, was conducted for assessment of toxicity and carcinogenic potential in the diet at doses of 4% or 0.8% in groups of male and female Wistar Hannover rats. Both sexes receiving 4% showed diarrhea, with loss of body weight gain. The survival rate in the 4% female group was significantly increased compared with control females after 2 y. Hematological and biochemical examination showed increase of RBC, Hb, and Alb in the 4% males. The cause of these increases could conceivably have been dehydration through diarrhea. AST and Na were significantly decreased in the males receiving 4%, and Cl was significantly decreased in both 4% and 0.8% males. A/G was significantly increased in the 4% females, and Cl was significantly decreased (0.8%) in the female group. Histopathologically, both sexes receiving 4% showed severe sinus dilatation of ileocecal lymph nodes, and yellowish pigmentation of ileocecal lymph nodes and renal tubules. Adenomas or adenocarcinomas in the cecum, colon, and rectum were observed in 4% males but not in the 0.8% and control male groups. Similarly, in females, adenomas in the colon were also observed in the 4% but not 0.8% and control groups. In conclusion, Aloe, used as a food additive, exerted equivocal carcinogenic potential at 4% high-dose level on colon in the 2-y carcinogenicity study in rats. Aloe is not carcinogenic at nontoxic-dose levels and that carcinogenic potential in at 4% high-dose level on colon is probably due to irritation of the intestinal tract by diarrhea.

Establishment of a bioassay model for lung cancer chemoprevention initiated with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in female A/J mice

AIMSnIn order to prevent lung cancer development in people at high risk, identification of chemopreventive agents may be important. The present study was conducted to establish a bioassay model for this purpose. In particular, the time course of 4-(methylnitrosamno)-1-(3-pyridyl)-1-butanone (NNK)-induced lung tumor development was examined to determine the most appropriate shortest period to assess effects of test agents, with 8-methoxypsoralen (8-MOP) as a typical example.nnnMETHODSnA total of 124 mice were separated into two groups (Group A: 60 mice, Group B: 64 mice), pretreated with 100ppm 8-MOP (Group A) or basal diet (Group B) for 3 days before receiving single doses of NNK (2mg/0.1ml saline/mouse i.p.) on days 0 and 7. Subgroups of 15 mice of each group were then sacrificed after 8, 10, 12, and 16 weeks.nnnRESULTSnMicroscopically, the earliest time point when significant differences in data for hyperplasia, adenoma and hyperplasia and adenoma could be detected was 12 weeks. A trend was noted for 8-MOP to reduce adenomas to a greater extent than hyperplasia.nnnDISCUSSIONnIn conclusion, the results of this study showed that the double i.p. treatment with NNK and 12 weeks duration are effective for detection of lung cancer chemoprevention in our A/J mouse lung tumorigenesis model.

Potential Inhibitory Effects of D-Allose, a Rare Sugar, on Liver Preneoplastic Lesion Development in F344 Rat Medium-Term Bioassay

D-allose, the C-3 epimer of d-glucose, is a monosaccharide present in minute quantities in nature and a rare sugar. The effects of D-allose on diethyl nitrosamine (DEN)-induced hepatocarcinogenesis were examined in male F344 rats by a rat medium-term bioassay based on the two-step model of hepatocarcinogenesis (experiment 1). In addition, a DNA microarray analysis was employed to clarify possible mechanisms of action of D-allose (experiment 2). The antioxidation potential of D-allose solution itself or of serum in rats treated with D-allose was also examined directly by measuring Cu(+)-reducing antioxidation power (experiment 3). Furthermore, to investigate the effects of D-allose in vivo under conditions of oxidative stress, it was administered with a choline-deficient, L-amino acid-defined diet (CDAA) in the medium-term liver carcinogenesis bioassay (experiment 4). Experiment 1 demonstrated no effects of D-allose on the development of glutathione S-transferase placental form (GST-P) positive foci in the liver. From DNA microarray analysis, several mRNA markers were found to be altered with functions related to apoptosis and cell proliferation (experiment 2), although D-allose itself and serum in vivo exhibited no antioxidation power directly (experiment 3). When D-allose was administered with the CDAA diet, decreases in the area and number of GST-P positive foci were noted with P values of 0.158 for area (%) and 0.061 for number (/cm(2)) (experiment 4). These results suggest the potential inhibitory effect of D-allose on liver carcinogenesis, particularly under oxidative stress conditions.

Overexpression of CYP2A6 in human colorectal tumors

CYP2A6 metabolizes various nitrosamines, such as those in the diet and in tobacco smoke, which have been implicated as risk factors for colorectal tumors. To determine whether changes in expression levels could contribute to their progression, we carried out immunohistochemistry for CYP2A6 in human colon tumors. Colon specimens (n = 53) were diagnosed as adenoma (n = 16), adenocarcinoma (n = 30) or carcinoma in or with adenoma (n = 7). Colon tumor cells showed cytoplasmic granular immunoreactivity for CYP2A6. Adenocarcinomas and adjacent mucosa showed similar highly elevated degrees of CYP2A6 expression, whereas carcinomas in or with adenoma and adenomas showed lesser increases. To further determine whether CYP2A6 mRNA was expressed at the same level as the CYP2A6 protein, we carried out in situ hybridization of CYP2A6 in two cases of adenocarcinoma. In situ hybridization for CYP2A6 revealed mRNA expression in adenocarcinoma cells. The data indicate that CYP2A6 may have important roles in human colorectal tumorigenesis and progression, so that it could be a candidate therapeutic and chemopreventive target for colorectal cancers. (Cancer Sci 2007; 98: 1582–1585)

Promotion potential of madder color in a medium-term multi-organ carcinogenesis bioassay model in F344 rats.

A medium-term multi-organ carcinogenesis bioassay in rats was conducted to assess any possible tumor promoting effects of madder color extracted from the root of madder. Male F344 rats were divided into 5 groups of 20 each. All rats of groups 1 to 4 were given DMD treatment, consisted of multicarcinogens, N-nitrosodiethylamine (DEN), N-methyl-N-nitrosourea (MNU), and N-bis (2-hydroxypropyl) nitrosamine (DHPN), for 4 wk, while group 5 served as untreated control without carcinogens. The animals were then administered a basal diet containing madder color at doses of 5.0% (group 1), 2.5% (group 2 with 0.75% additional dextrin), or 0 (groups 3 with 1.5% additional dextrin, 4 without dextrin and 5) for the following 28 wk (total 32 wk). The total amount of dextrin in groups 1 to 3 diets was adjusted to 1.5% by extra dextrin because madder color powder contained dextrin. Key organs were observed histopathologically and glutathione S-transferase placental form (GST-P) positive foci of the liver were quantified. In the liver, 5.0% and 2.5% treated groups showed statistically significant dose-related increases in both number and area of GST-P positive foci, number: 2.81 +/- 0.90 and 1.96 +/- 0.93 (groups 1 and 2), area: 0.99 +/- 2.49 and 0.37 +/- 0.77, as compared with control, number: 0.87 +/- 0.72, area: 0.06 +/- 0.06 (group 3). In the kidneys, incidences (and numbers) of adenoma treated with 5.0% and 2.5%, 47.4% (0.20 +/- 0.24), and 47.4% (0.13 +/- 0.15) (groups 1 and 2) were significantly increased compared to control, 0% (0) (group 3). In conclusion, madder color demonstrated significant tumor promoting effects in the liver and kidneys in the DMD model.

A 90-day toxicity study of l-asparagine, a food additive, in F344 rats

L-asparagine is an amino acid listed as an existing food additive in Japan. The present 90-day toxicity study in F344/DuCrlCrj rats was conducted for safety assessment and to determine a no observed adverse effect level (NOAEL) of L-asparagine. Groups of 10 males and 10 females were given the material at dose levels of 0%, 1.25%, 2.5% or 5% in diet for 90 days. During the experiment, there were no remarkable changes in general conditions and no deaths occurred in any group. Final body weights of male 5% and 1.25% groups were significantly decreased. There were also significant increases in relative organ weights of the brain, kidney and testis in 5% males. On serological examination, GLU, PL, K and ALT were increased significantly in 5% females, and GLU was increased significantly and CRN was decreased significantly in the female 1.25% group. However, histopathological examination did not reveal any significant variation in development of lesions among the groups. Changes in body and organ weights, as well as other parameters, were concluded to be due to treatment with 5% L-asparagine. The NOAEL was determined to be 2.5% in the diet (males, 1.65 g/kg body weight/day; females, 1.73 g/kg body weight/day).

Enhancing effects of a high fat diet on 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline-induced lung tumorigenesis in female A/J mice

Both heterocyclic amines and a high fat diet are associated with an increased risk of cancer in many organs. Female A/J mice were fed a diet supplemented with 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and a high fat diet to test for the development of lung tumors. In experiment 1, the mice were divided into 6 groups. Groups 1, 2, 3 and 4 were fed a diet supplemented with MeIQx at a concentration of 600?ppm for 0-12 weeks. A high fat diet containing 20% corn oil was given to Groups 1 and 5 for 0-32 weeks, Group?2 for 12-32 weeks and Group 3 for 0-12 weeks. Group 6 was fed a basal diet without supplements. MeIQx-treated groups (Groups?1, 2, 3 and 4) showed a significant increase in macroscopic and microscopic lung nodules compared with the control (Group 6). Areas of adenomas were increased dependent on the duration of exposure to the high fat diet. In experiment 2, Group 1 mice were fed MeIQx and a high fat diet, Group?2 a MeIQx alone diet, Group 3 a high fat alone diet, and Group?4 a basal diet without supplements. CYP1A2 mRNA in the liver was significantly decreased by a high fat diet (Group?3). The MeIQx alone group (Group 2) showed a tendency towards increased CYP1A2 expression, which was partially reduced in the MeIQx + high fat-treated group (Group 1). In the lungs, CYP1A2 mRNA expression was at an extremely low level, with no intergroup differences. In conclusion, MeIQx exerts tumorigenic potential in the lungs, and a high fat diet increases the size of induced lesions. The expression level of CYP1A2 in relation to MeIQx and a high fat diet may be associated with lung carcinogenesis.