Decreased expression of Bcl-x protein during hepatocarcinogenesis induced exogenously and endogenously in rats.

Dysregulations of apoptosis have been widely recognized as important events in multi-stage carcinogenesis. Bcl-x, a member of the Bcl-2 family, is known to act as a regulator of apoptosis. The present study was conducted to assess the role of altered Bcl-x protein expression in exogenous and endogenous hepatocarcinogenesis in rats. In the short-term exogenous models, male Fischer 344 rats, 6 weeks old, were given a single intraperitoneal injection of diethylnitrosamine (DEN) at a dose of 200 mg / kg body weight, partially hepatectomized at the end of week 3, administered phenobarbital at a concentration of 0.05% from the end of week 2 for 6 weeks, and sacrificed. In the livers, glutathione S-transferase (GST-P)-positive, putative preneoplastic lesions were induced, and Bcl-x protein expression was decreased in 24.7% of such lesions. The incidence of GST-P-positive lesions with decreased Bcl-x increased depending on the size of the lesions; 18.9%, 32.4% and 86.5% in the lesions smaller than 0.03, between 0.03 and 0.3, and larger than 0.3 mm(2), respectively. In GST-P-positive lesions larger than 0.3 mm(2), both apoptosis induction and cell proliferation activity were enhanced when Bcl-x protein expression was decreased. In the long-term exogenous models, rats were given 10 mg / kg of DEN, partially hepatectomized 4 h after treatment, administered 0.5 mg / kg of colchicine at the end of days 1 and 3, subjected to a selection procedure, and sacrificed at the end of week 45. Hepatocellular carcinomas were induced with the decreased Bcl-x protein expression. In the endogenous model, rats were fed a choline-deficient, L-amino acid-defined diet for 16 or 80 weeks and sacrificed. Bcl-x protein expression was decreased both in GST-P-positive lesions and hepatocellular carcinoma. These results suggest that this decrease of Bcl-x protein might serve as an indicator of the advanced form of preneoplastic lesions, and that this decrease could also be associated with a potential to progress into carcinoma in both exogenous and endogenous hepatocarcinogenesis of rats.     

Inhibitory effects of lemon grass (Cymbopogon citratus,Stapf) extract on the early phase of hepatocarcinogenesis after initiation with diethylnitrosamine in male Fischer 344 rats

Effects of lemon grass extract (LGE) on hepatocarcinogenesis were examined in male Fischer 344 rats, administered diethylnitrosamine (DEN) at three weekly intraperitoneal doses of 100 mg/kg body weight and partially hepatectomized at the end of week 5. LGE was given at dietary concentrations of 0, 0.2, 0.6 or 1.8% from the end of week 4 for 10 weeks. All rats were sacrificed at the end of week 14. LGE reduced the number of putatively preneoplastic, glutathione S-transferase placental form-positive lesions and the level of oxidative hepatocyte nuclear DNA injury, as assessed in terms of 8-hydroxydeoxyguanosine production. In contrast, LGE did not affect the size of the preneoplastic lesions, hepatocyte proliferative activity, activities of phase II enzymes or hepatocyte extra-nuclear oxidative injury. These results suggest inhibitory effects of LGE on the early phase hepatocarcinogenesis in rats after initiation with DEN.     

突变型p53蛋白在二乙基亚硝胺诱发大鼠肝癌前病变中的表达

目的　探讨 p53基因与肝细胞癌变的关系。方法　用免疫组化方法 ,观察二乙基亚硝胺( DEN)诱发大鼠肝癌前病变组织及对照组肝组织中突变型 p53( mp53)蛋白、胎盘型谷胱甘肽 S转移酶 ( GST- P)的表达及其与细胞增殖状态的关系。结果　实验组第 5周时仅在 2 / 5例癌前病变肝组织中检测到 mp53蛋白的表达 ,此蛋白仅存在于部分较大的肝细胞增生结节 ( HN)或 GST- P阳性灶内 ;mp53蛋白阳性的 HN或 GST- P阳性灶 ,其细胞的溴化脱氧尿嘧啶标记指数 ( Brd U- LI)较阴性者高( P<0 .0 5)。结论　 p53基因的突变可发生于肝细胞癌变的早期阶段 ,与肝细胞的异常增殖和癌变有关 ,并有可能成为新的肝癌前肿瘤基因标记物。     

Inhibition by Green Tea Extract of Diethylnitrosamine–initiated but Not Cholinedeficient, L–Amino Acid–defined Diet–associated Development of Putative Preneo–plastic, Glutathione S–Transferase Placental Form–positive Lesions in Rat Liver

The effects of green tea extract (GTE) on exogenous and endogenous models of rat liver carcinogenesis using diethylnitrosamine (DEN) and a choline–deficient, L–amino acid–defined (CDAA) diet were studied. For the exogenous carcinogenesis study, male Fischer 344 rats, 6 weeks old, were given a single intraperitoneal dose of 200 mg/Kg body weight of DEN, partially hepatectomized at week 3, and administered GTE at doses of 0, 0.01 and 0.1% in the drinking water from week 2 for 10 weeks. For the endogenous carcinogenesis study, rate were fed the CDAA diet and simultaneously given GTE for 12 weeks. All rats were killed at the end of week 12. After DEN–initiation, the apparent numbers of glutathione S–transferase placental form–positive foci, assayed as putative preneoplastic lesions, were decreased by the administration of GTE, though their sizes were not altered. In contrast, GTE did not significantly reduce the numbers of the lesions induced by the CDAA diet or affect their sizes. While the levels of 8–hydroxyguanine, a parameter of oxidative DNA damage, were reduced by the GTE administration in both experimental models, GTE did not protect against the CDAA–diet–associated liver tissue damage in terms of either histology or plasma marker enzyme levels. We conclude that, while GTE may be a possible chemopreventive agent for nitrosamine–initiated hepato–carcinogenesis in the absence of chronic hepatocyte damage, it does not significantly inhibit lesion development in hepatocarcinogenesis associated with the CDAA diet, a cirrhosis–associated model.     

Inhibition by N-(4-hydroxyphenyl)retinamide and all-trans-retinoic acid of exogenous and endogenous development of putative preneoplastic, glutathione S-transferase placental form-positive lesions in the livers of rats

The effects of N-(4-hydroxyphenyl)retinamide (4-HPR) and all-trans- retinoic acid (tRA) on the exogenous and endogenous models of rat liver carcinogenesis respectively using diethylnitrosamine (DEN) and a choline-deficient, L-amino acid-defined (CDAA) diet were studied. For the exogenous study, male Fischer 344 rats, 6 weeks old, were given a single i.p. dose of 200 mg/kg body wt of DEN, partially hepatectomized at week 3, administered 4-HPR at doses of 0, 0.04, 0.08 and 0.16% or tRA at 0, 0.004, 0.008 and 0.015% in diet from week 2 for 6 weeks, and killed at the end of week 8. For the endogenous study, rats were fed the CDAA diet containing 4-HPR or tRA for 12 weeks and killed at the end of week 12. 4-HPR decreased the numbers and sizes of the glutathione S-transferase placental form-positive foci, assayed as putative preneoplastic lesions, the levels of 8-hydroxyguanine (8-OHG), a parameter of oxidative DNA damage, and the bromodeoxyuridine labeling indices (BrdU L.I.) by all three doses in the DEN-initiated case and, more prominently, in the CDAA diet-associated case. In contrast, while tRA failed to exert inhibitory effects apparently on foci development, 8-OHG formation or BrdU labeling in the DEN-initiated case, it reduced the numbers and sizes of the foci, the 8-OHG levels and the BrdU L.I. by all three doses in the CDAA diet-associated case. Furthermore, both 4-HPR and tRA inhibited the CDAA diet-associated induction of hepatocyte necrosis and connective tissue increase but not intrahepatocellular fat accumulation. These results indicate that 4-HPR exerts chemopreventive effects against the exogenous and endogenous rat liver carcinogenesis, while tRA can inhibit only the latter.      

Cell proliferation and tumour promotion by ethinyl estradiol in rat hepatocarcinogenesis

A two-stage model of hepatocarcinogenesis is used to study the effect of exposure time to ethinyl estradiol (EE) on promotion of preneoplastic lesions in rat liver induced by diethylnitrosamine (DEN). Young male and female Sprague-Dawley rats initiated by a single dose of DEN (100 mg/kg) were subjected to different times of EE administration incorporated into the diet at 10 p.p.m. (0.5 mg/kg x day). Animals were killed 1 year after initiation. Whereas macroscopic tumours were rarely seen in animals with short exposure (3 or 4 months) or in only-initiated controls, all the animals under a long period of administration (8 months) showed macroscopic tumours. Morphometric studies on glutathione-S-transferase (GST) positive preneoplastic lesions revealed an increase in the mean size of foci and nodules corresponding to 8 months of treatment, whereas no changes were observed between animals with short exposure and only-initiated controls. No differences were seen in the incidence of these lesions between any of the protocols. In addition to an acute hyperplastic effect on non-initiated liver described earlier, our preliminary results suggest cytotoxicity and an enhancement of the liver cell turnover after several months of continuous EE administration. These results taken together suggest that promotion of hepatocarcinogenesis by EE largely depends on the time of exposure to the compound and that chronic effects on the liver cell turnover may play an important role in its ability to promote hepatocarcinogenesis.     

饮水微囊藻毒素在大鼠肝癌发生期间对细胞增殖与凋亡的影响

目的: 研究经饮水摄入微囊藻毒素(MC)在大鼠肝癌发生期间对细胞增殖与凋亡的影响,进而探讨微囊藻毒素促肝癌机制.方法:50只Wistar大鼠随机分为4组,A组(空白对照):B组(DEN启动对照);C组(DEN处理,饮含MC藻水):D组(DEN处理,腹腔注射MCLR纯毒素)、应用LSAB免疫组化技术检测大鼠肝癌前病变病灶中PCNA、Bcl-2和Bax蛋白的表达.结果:各处理组PCNA平均积分光密度值(AIOD)均较A组0.0013明显升高,分别为B组0.0033,C组0.0036和D组0.0088,其中D组与B组比较有明显升高(P<0.05).饮水摄入或腹腔注射微囊藻毒素均可使Bcl-2表达上调,Bax表达下调.结论:促进调控细胞增殖与凋亡的基因平衡失调.使细胞失控性生长,可能是微囊藻毒素促肝癌作用的分子机制之一.     

Nonalcoholic steatohepatitis induced by a high‐fat diet promotes diethylnitrosamine‐initiated early hepatocarcinogenesis in rats

It has been suggested that patients with nonalcoholic steatohepatitis (NASH) may have high risk for liver cancer. However, it is unknown whether high‐fat diet (HFD) induced NASH promotes hepatocarcinogenesis. In this study, Sprague‐Dawley rats were injected with a low dose of hepatic carcinogen diethylnitrosamine (DEN) and then fed either Lieber‐DeCarli control diet (CD) or HFD for 6 weeks. Liver histology and the hepatic placental form of glutathione S‐transferase (P‐GST) positive foci were examined. Expression levels of proliferating cell nuclear antigen (PCNA), cyclinD1, phosphorylated mitogen‐activated protein kinase (MAPK) including extracellular signal‐regulated kinase (ERK) and p38, as well as tumor necrosis factor‐alpha (TNF‐α), and nuclear factor‐kappaB (NF‐κB) were measured in the liver. Induction of lipid peroxidation end products (malondialdehyde plus 4‐hydroxynonenal) in liver and apoptotic hepatocytes were also assessed. Results showed that HFD‐fed rats developed significantly higher incidence and multiplicity of P‐GST positive foci along with more fat accumulation, infiltration of inflammatory cells and higher lipid peroxidation in the liver, when compared with rats fed the CD. This high prevalence of hepatic lesions in the liver was accompanied by greater PCNA expression and cyclinD1 protein concentration but little change in hepatocyte apoptosis. HFD feeding elevated hepatic phosphorylated ERK but reduced phosphorylated p38 when compared with the CD feeding. In addition, a significantly higher expression of TNF‐α mRNA and nuclear NF‐κB p65 protein were observed in HFD group than in CD group. These data clearly demonstrate that NASH induced by HFD promoted DEN‐initiated early hepatocarcinogenesis, which was associated with elevated TNF‐α/NF‐κB signaling and MAPK related hepatocyte proliferation. © 2008 Wiley‐Liss, Inc.     

Cleistocalyx nervosum Extract Ameliorates Chemical-Induced Oxidative Stress in Early Stages of Rat Hepatocarcinogenesis

Purpose: To study the effect of Cleistocalyx nervosum extract (CE) on diethylnitrosamine (DEN) andphenobarbital (PB) induced oxidative stress in early stages of rat hepatocarcinogenesis. Materials and Methods:Male Wistar rats were divided into 4 groups, with Group 1 as a negative control and Group 2 was a positivecontrol receiving DEN injections once a week and PB in drinking water for 6 weeks. Two weeks before DENinitiation and PB treatment, Groups 3 and 4, were fed with 500 and 1000 mg/kg of CEs, respectively, for 8weeks. Results: A number of GST-P-positive foci, preneoplastic lesions, in the liver were markedly increased incarcinogen administered rats, but was comparatively decreased in rats treated with 1000 mg/kg of CE. The CEreduced malondialdehyde in serum and in the livers of rats treated with DEN and PB. Moreover, CE significantlyincreased the activities of glutathione peroxidase and catalase in rat liver. Conclusions: CE appeared to exertits chemopreventive effects by modulating antioxidant status during DEN and PB induced early stages ofhepatocarcinogenesis in rats.     

Dietary Administration of Inositol and/or Inositol-6-phosphate Prevents Chemically-induced Rat Hepatocarcinogenesis

Chemoprevention is considered a rational strategy for dietary approaches to prevention of cancer. Multiple lines ‍of evidence suggest that many of our dietary principles are able to intervene in the multistage carcinogenesis process ‍and phytic acid (inositol hexaphosphate, IP6), a phytochemical present in a variety of plant species, has been shown ‍to prevent various cancers, including those of the mammary gland, colon and liver. However, the mechanism of ‍chemoprevention by IP6 has not been fully elucidated. In the present study, we examined the effects of inositol and/ ‍or IP6 supplementation on rat hepatocarcinogenesis initiated by diethylnitrosamine (DEN) and promoted by partial ‍hepatectomy (PH). Supplementation with either inositol or IP6, or their combination, starting one week prior to ‍administration of DEN, resulted in a significant decrease in both the area and the number of placental glutathione Stransferase ‍positive (GST-P+) foci, a preneoplastic marker for DEN-initiated hepatocarcinogenesis. The administration ‍of inositol and/or IP6 in drinking water caused marked enhancement in the glutathione S-transferase (GST) activity. ‍In addition, the production of thiobarbituric acid reactive substances and the catalase activity were significantly ‍reduced in rats supplemented with inositol and /or IP6. Based on these findings, it is likely that the chemopreventive ‍effects of inositol and/or IP6 on rat hepatocarcinogenesis initiated by DEN and promoted by PH are associated with ‍induction of GST activity and suppression of lipid peroxidation.     

Anti-carcinogenic action of phenobarbital given simultaneously with diethylnitrosamine in the rat

The present work has been planned in order to elucidate the effect of phenobarbital (PB: 15 mg per rat of ingested dose) on carcinogenesis when it is administered simultaneously with diethylnitrosamine (DEN: 10 mg/kg/day). Wistar rats (180 g) were treated by DEN alone or by DEN + PB during 2, 4 and 6 weeks according to our schedule for hepatocarcinogenesis. After the end of the treatment, the number and the size of induced PAS positive preneoplastic foci was significantly reduced when PB was given simultaneously with DEN for 4 and 6 weeks. The mitotic inhibition and the production of micronuclei normally observed after partial hepatectomy in DEN treated rats were also significantly decreased in DEN + PB treated rats. When the treatment last only 2 weeks, the presence of PB did not change significantly the last parameters. In DEN + PB treated rats, the survival was prolonged and the tumor incidence decreased as compared with the results obtained by DEN alone. It is concluded that PB, which promotes carcinogenesis when administered after the DEN treatment, reduces the carcinogen effect when given simultaneously with DEN. This ‘anti-carcinogen’ effect acts on the initiation as well as on the promotion of the precancerous lesions. Biochemical investigations are in progress to obtain more information about this ‘paradoxical’ PB effect.     

Presence of a no-observed effect level for enhancing effects of development of the alpha-isomer of benzene hexachloride (alpha-BHC) on diethylnitrosamine-initiated hepatic foci in rats

The dose dependence of the promoting effects of the alpha-isomer of benzene hexachloride (alpha-BHC) on hepatocarcinogenesis was investigated in a medium-term rat liver bioassay (Ito test). A total of 195 F344 male rats, 6 weeks old, were given a single intraperitoneal injection of diethylnitrosamine (DEN) at the start of the experiment and subjected to two-thirds partial hepatectomy at week 3. Two weeks after the administration of DEN, alpha-BHC were fed to rats at doses of 0, 0.01, 0.1, 0.5, 1, 2, 4, 7.5, 15, 30, 60, 125 and 500 ppm in diet for 6 weeks. All surviving animals were killed at week 8, and their livers were examined immunohistochemically for detection of glutathione S-transferase placental form (GST-P)-positive foci, surrogate preneoplastic lesions. Quantitative values for numbers and areas were dose-dependently increased in rats given alpha-BHC at 0.5-500 ppm. However, those for groups treated with 0.01 and 0.1 ppm were decreased, albeit not significantly in comparison to the controls. Cytochrome P450 3A2 (CYP3A2) protein levels and activities showed a good correlation to the number and area of GST-P-positive foci. These results support evidence of hormesis and indicate a no-observed effect level for alpha-BHC promoting potentials may exist regarding rat liver carcinogenesis, which correlates with expression of CYP3A2 in the liver.     

Enhancement by neurotensin of hepatocarcinogenesis by N-nitrosomorpholine in Sprague-Dawley rats

The effect of neurotensin on hepatocarcinogenesis by N-nitrosomorpholine (NNM) was investigated in Sprague-Dawley rats. Rats were given drinking water containing NNM for 8 weeks and alternate-day injections of neurotensin from the beginning until the end of the experiment. Preneoplastic and neoplastic lesions stained for placental type glutathione-S-transferase (GST-P) were examined histochemically. Administration of neurotensin significantly increased the percentage area and the number of GST-P-positive lesions, and the labeling indices of pre-neoplastic lesions and adjacent liver. These findings indicate that neurotensin enhances hepatocarcinogenesis and that this effect may be related to its effect in increasing cell proliferation in preneoplastic lesions.     

Inhibition of Early-phase Exogenous and Endogenous Liver Carcinogenesis in Transgenic Rats Harboring a Rat Glutathione S-Transferase Placental Form Gene

Hepatocarcinogenesis initiated with N -nitrosodiethylamine (DEN) and that initiated by feeding of a choline-deficient, l -amino acid-defined (CDAA) diet were compared in transgenic male Wistar rats harboring a rat glutathione S -transferase placental form ( GST-P ) gene (GST-P-Tg rats) and non-transgenic (N-Tg) rats. Eight-week-old GST-P-Tg and N-Tg rats were administered DEN intraperitoneally at 100 mg/kg body weight, subjected to a selection procedure with 2-acetylaminofluorene and CCl₄, and killed at the end of weeks 5 and 12. Other groups were fed the CDAA diet for 12 weeks and killed. Five weeks after the DEN treatment, numbers and sizes of γ-glutamyltransferase (GGT)- or GST-P-positive lesions and 8-hydroxyguanine (8-OHG) levels in the livers were significantly less in GST-P-Tg rats than in N-Tg rats. The lesion numbers were unchanged between the ends of weeks 5 and 12 in GST-P-Tg rats, but decreased in N-Tg rats. The lesion sizes were increased in GST-P-Tg rats, but unchanged in N-Tg rats. While the proliferating cell nuclear antigen labeling indices (PCNA L.I.) in and surrounding the lesions were decreased, more prominently in GST-P-Tg rats than in N-Tg rats, the 8-OHG levels were also decreased but similarly in both cases. After 12 weeks on the CDAA diet, the lesion incidences, numbers and sizes, 8-OHG levels, PCNA L.I. in and surrounding the lesions, and liver injury were significantly less in GST-P-Tg rats than in N-Tg rats. These results indicate that insertion of a rat GST-P transgene alters the early phase of exogenous and endogenous rat hepatocarcinogenesis, presumably due to enhanced detoxification by GST-P expressed both transiently during the initiation and chronically in the altered hepatocyte populations.     

Chemopreventive effects of Paullinia cupana Mart var. sorbilis, the guaraná, on mouse hepatocarcinogenesis

Guaraná (Paullinia cupana) is originally from Amazon, Brazil. Its effects on mouse hepatocarcinogenesis have been investigated in this study. Mice were treated with N-nitrosodiethylamine (DEN), received three different doses of P. cupana added to commercial food, and euthanized after 25 weeks. Gross lesions were quantified, and preneoplastic lesions (PNL) were histologically measured. Cellular proliferation was evaluated by immunobloting for the proliferating cell nuclear antigen (PCNA). The incidence and multiplicity of macroscopic lesions were reduced. The PNL number and PCNA expression were reduced in the highest P. cupana dose. According to these results, guaraná presented inhibitory effects on DEN hepatocarcinogenesis in mice.     

INHIBITORY EFFECT OF BOSCHNIAKIA ROSSICA ON DEN-INDUCED PRECANCEROUS HEPATIC FOCI AND ITS ANTIOXIDATIVE ACTIVITIES IN RATS

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Resistance of Copenhagen rats to chemical induction of glutathione S- transferase 7-7-positive liver foci

Copenhagen (Cop) rats are completely resistant to the chemical induction of mammary adenocarcinomas, but their susceptibility to hepatocarcinogenesis is virtually unknown. Rat liver is a well- characterized and easily manipulated tissue in which to study carcinogenesis. Therefore, if Cop rats are resistant to hepatocarcinogenesis, studies into resistance mechanisms may be feasible. Male Cop and F344 rats, 7-8 weeks old, were initiated using either N-nitrosodiethylamine (DEN) (200 mg/kg, i.p.) or a two-thirds partial hepatectomy (PH) followed by N-methyl-N-nitrosourea (MNU) (60 mg/kg, i.p.). The rats were then promoted using a modified resistant hepatocyte (RH) protocol (a combination of four doses of 2- acetylaminofluorene (2-AAF) and a single dose of CCl4 that provides a selective mitotic stimulus for initiated cells). Six weeks after initiation the rats were killed and liver sections were stained for glutathione S-transferase 7-7 (GST 7-7), a marker for putative preneoplastic hepatocytes. Cop rats were found to be highly resistant, having a approximately 9- and approximately 27-fold smaller percentage of liver area occupied by GST 7-7-positive foci than susceptible F344 rats following initiation by DEN and MNU respectively. Furthermore, gross liver nodules did not form in any of the Cop rats, whereas all F344 rat livers contained nodules. Hepatic necrosis caused by DEN during initiation, and CCl4 during promotion is necessary to stimulate compensatory hepatocyte division. We demonstrated that these agents do indeed increase serum transaminase levels and produce histologic evidence of necrosis in Cop rats. In order for liver foci to grow rapidly in the RH protocol, the surrounding normal hepatocytes must be mito-inhibited by 2-AAF. We found that the degree of mito-inhibition of normal hepatocytes by 2-AAF is the same in Cop and F344 rats. These results show that the Cop rat is highly resistant to the chemical induction of putative preneoplastic liver foci and nodules.      

Effects of arctiin on PhIP-induced mammary, colon and pancreatic carcinogenesis in female Sprague-Dawley rats and MeIQx-induced hepatocarcinogenesis in male F344 rats

Chemopreventive effects of arctiin, a lignan isolated from Arctium lappa (burdock) seeds, on the initiation or post initiation period of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) induced mammary carcinogenesis in female rats and on 2-amino-3, 8-dimethylimidazo[4,5-f]quinoxaline (MeIQx)-associated hepatocarcinogenesis in male rats were examined. In experiment 1, female Sprague-Dawley (SD) rats were given intragastric doses of 100 mg/kg body wt of PhIP once a week for 8 weeks as initiation. Groups of 20 rats each were treated with 0.2 or 0.02% arctiin during or after PhIP initiation. Control rats were fed 0.2 or 0.02% arctiin, or basal diet alone during the experimental period. Animals were killed at the end of week 48. Although the incidence of mammary carcinomas did not significantly differ among the PhIP-treated groups, multiplicity was significantly decreased in rats given 0.2 (0.7+/-0.7, P<0.05) or 0.02% (1.0+/-1.1, P<0.05) arctiin after PhIP initiation as compared with the PhIP alone controls (2.1+/-2.5). The average number of colon aberrant crypt foci was also significantly decreased in these two groups. Pancreas acidophilic foci were induced in PhIP treated animals with slight decrease in the multiplicity with arctiin during the initiation phase. For liver carcinogenesis, groups of 15 male F344 rats were given a single intraperitoneal injection of diethylnitrosamine (DEN) and starting 2 weeks later, they were administered 0.03% MeIQx in the diet, MeIQx together with 0.5% arctiin, 0.1% arctiin or basal diet for 6 weeks. They were subjected to two-third partial hepatectomy 3 weeks after DEN initiation and killed at the end of week 8 for glutathione S-transferase placental form (GST-P) immunohistochemistry. The numbers and areas of preneoplastic GST-P positive foci were elevated by the treatment with MeIQx, and further increased by the simultaneous treatment with arctiin. These results indicate that arctiin has a protective effect on PhIP-induced carcinogenesis particularly in the mammary gland in the promotion period. On the other hand, it may have a weak co-carcinogenic influence on MeIQx-induced hepatocarcinogenesis. In addition, the results suggested that PhIP is a weak pancreatic carcinogen in female SD rats, targeting acinar cells.     

Suppression of diethylnitrosamine and 2-acetylaminofluorene-induced hepatocarcinogenesis in rats by tocotrienol-rich fraction isolated from rice bran oil.

The anticancer efficacy of tocotrienol-rich fraction (TRF) was evaluated during diethylnitrosamine (DEN)/2-acetylaminofluorene (AAF)-induced hepatocarcinogenesis in male Sprague-Dawley rats. TRF treatment was carried out for 6 months, and was started 2 weeks before initiation phase of hepatocarcinogenesis. Morphological examination of the livers from DEN/AAF rats showed numerous off-white patches and few small nodules, which were significantly reduced by TRF treatment. Cytotoxic damage by DEN/AAF was estimated by alkaline phosphatase (ALP) release into the plasma from the cell membranes. DEN/AAF caused a twofold increase in the activity of ALP in plasma as compared with normal control rats, and this increase was prevented significantly by TRF treatment. We observed an increase of 79% in liver ALP activity in DEN/AAF rats, which was further increased by another 48% after the administration of TRF. Hepatic activity of glutathione S-transferase (GST) was also increased (3.5-fold) during the induction of hepatic carcinogenesis. Lipid peroxidation and low-density lipoprotein (LDL) oxidation increased threefold following initiation by DEN/AAF as compared with normal control rats. However, TRF treatment to DEN/AAF-treated rats substantially decreased (62-66%) the above parameters and thus limited the action of DEN/AAF. We conclude that long-term intake of TRF could reduce cancer risk by preventing hepatic lipid peroxidation and protein oxidation damage due to its antioxidant actions.