Enhancing effects of dimethylnitrosamine on aflatoxin B1 hepatocarcinogenesis in rats

In a study of possible enhancing effects of dimethylnitrosamine (DMN) on aflatoxin B₁ (AFB₁) hepatocarcinogenesis, male Buffalo strain rats were fed diets containing 1 ppm AFB₁, 25 ppm DMN, and a combination of 1 ppm AFB₁ and 25 ppm DMN (AFB₁ + DMN). The diets were replaced by chow pellets after 6 months, and animals were killed 3,6,9 and 12 months after the onset of the experiment. In the untreated control group animals were free of hepatocellular carcinoma but the treated groups fed AFB₁, DMN and AFB₁ plus DMN developed hepatic lesions ranging from multiple cysts, altered cell foci and neoplastic nodules to hepatocellular carcinomas. Hepatocellular carcinomas developed in 79%, 45% and 5% of rats fed AFB₁ plus DMN, AFB₁ and DMN respectively at the end of the experiment. Multiple cysts were also found in all periods in animals fed AFB₁ plus DMN, whereas rats fed AFB₁ and DMN separately developed a few multiple cysts by the end of the experiment. These findings suggest that DMN potentiates the hepatocarcinogenesis induced by AFB₁ in rats.     

黄曲霉毒素B1诱发大鼠肝癌过程中对CYP3A4活性的影响

目的 探讨黄曲霉毒素B1(AFB1)诱发肝癌过程中对CYP3A4活性的影响.方法 采用大鼠肝微粒体混合酶体外代谢体系,利用荧光定量法动态检测AFB1诱发肝癌过程中不同时期CYP3A4酶活性.结果 AFB1组CYP3A4含量从实验开始逐渐升高,至23周达顶峰,然后逐渐降低,到43周又升高,出现双波峰变化,阶段性比较差异有统计学意义(P=0.000);对照组CYP3A4含量在整个实验过程中,除33周和63周出现明显降低外,其他各阶段变化不大;组间比较显示AFB1组在诱癌过程中有抑制CYP3A4的趋势,于63周抑制最明显,但尚未达统计学意义(P=0.638);结论AFB1在诱癌过程中有抑制CYP3A4的趋势,可能是与早期癌变的细胞减少对特定基因毒性物质的活化有关.     

Effect of protein calorie malnutrition and cell replication on aflatoxin B1-induced hepatocarcinogenesis in rats

Malnourished and well-fed neonatal Holtzman rats 10 days of age were exposed to 3 doses of aflatoxin B1 [(AFB1) CAS: 1162-65-8] at intervals of 96 hours to study the combined effect of malnutrition and cell replication in AFB1-induced hepato-carcinogenesis. The neonatal model made use of the fact that cell replication persists in the liver for 3 weeks of postnatal life. Malnutrition during suckling was induced by adopting the techniques of Widdowson and McCance of increasing the litter size to 16. Following AFB1 administration, the malnourished animals were rehabilitated on a high-protein pellet diet given ad libitum. Preneoplastic lesions and neoplastic nodules were identified in the livers of the 2 groups. Alpha fetoprotein (AFP) was detected in the sera by immunoprecipitation. The preneoplastic lesions appeared earlier, and their progression was faster in the malnourished group as compared to the well-fed animals. By 65 weeks following AFB1 exposure, 6 of 17 (35%) animals from the malnourished group showed neoplastic nodules, whereas no such nodules were observed in the animals from the well-fed group. Neoplastic nodules showed a variable pattern of enzyme activities. Under the electron microscope the changes were again more marked in the animals of the malnourished group as compared to those of the well-fed group. In the former group serum AFP was detected as early as 46 weeks, and by 55-65 weeks almost 50% of the animals from the same group showed positivity for serum AFP. None of the animals from the well-fed group showed any positivity for serum AFP throughout the study. This study thus indicates that preneoplastic lesions-neoplastic nodules are enhanced when cell replication and malnutrition coexist during AFB1-induced hepatocarcinogenesis.     

Potency of dietary indole-3-carbinol as a promoter of aflatoxin B1-initiated hepatocarcinogenesis: results from a 9000 animal tumor study

Indole-3-carbinol (I3C), a metabolite of glucobrassicin found in cruciferous vegetables, is documented as acting as a modulator of carcinogenesis and, depending on timing and dose of administration, it may promote hepatocarcinogenesis in some animal models. In this study we demonstrate that, when given post-initiation, dietary I3C promotes aflatoxin B1 (AFB1)-induced hepatocarcinogenesis in the rainbow trout model at levels as low as 500 p.p.m. Trout embryos (approximately 9000) were initiated with 0, 25, 50, 100, 175 or 250 p.p.b. AFB1 by a 30 min immersion. Experimental diets containing 0, 250, 500, 750, 1000 or 1250 p.p.m. I3C were administered starting at 3 months and fish were sampled for liver tumors at 11-13 months. Promotion at the level of tumor incidence was statistically significant for all dietary levels, except 250 p.p.m. Relative potency for promotion markedly increased at dietary levels >750 p.p.m. We propose that more than one mechanism could be involved in promotion and that both estrogenic and Ah receptor-mediated pathways could be active. The estrogenicity of I3C, measured as its ability to induce vitellogenin (an estrogen biomarker in oviparous vertebrates) was evident at the lowest dietary level (250 p.p.m.), whereas CYPIA (a P450 isozyme induced through the Ah receptor pathway) was not induced until dietary levels of 1000 p.p.m. Therefore, at lower dietary levels, promotion by I3C in this model could be explained by estrogenic activities of I3C acid derivatives, as it is known that estrogens promote hepatocarcinogenesis in trout. Much stronger promotion was observed at high dietary I3C levels (1000 and 1250 p.p.m.), at which levels both CYP1A and vitellogenin were induced.     

金蒲抑瘤片在实验诱发大鼠肝癌过程中的作用

目的探讨金蒲抑瘤片对黄曲霉毒素B1(aflatoxin B1,AFB1)致肝癌作用的影响。方法实验动物随机分为高剂量、低剂量和对照组。用AFB1处理各组动物,高、低剂量组大鼠在接受AFB1期间分别喂含量为9.3和2.3g/kg的金蒲抑瘤片混合饲料,对照组喂基础饲料。8周后处死动物,观察各组动物肝组织内γ-谷氨酰转肽酶阳性肝细胞增生(γ-glu-tamyltranspeptidase-positive hyperplastic livercell,γ-GT)灶的数量和大小。结果高、低剂量金蒲抑瘤片均能减少AFB1诱发的γ-GT灶的数量和大小高、低剂量组的数量分别为0.90和3.72个/cm2,均低于对照组6.10个/cm2,抑制率分别为85%和39%,高剂量组与对照组相比差异有统计学意义,t=2.597,P=0.028。高、低剂量组的大小分别为0.24和1.94mm2/个,均低于对照组2.36mm2/个,抑制率分别为90%和17%,但差异无统计学意义,P>0.05。结论金蒲抑瘤片有减少AFB1诱发大鼠肝γ-GT灶的数量和大小的作用,而高剂量金蒲抑瘤片的减少趋势更强。     

乙肝病毒和黄曲霉毒素B1在树鼩肝癌形成中的协同作用

目的:动态观察乙肝病毒(HBV)和黄曲霉毒素B1(AFB1)在树鼩原发性肝细胞癌(HCC)形成过程中的作用.方法:成年树鼩按不同处理分为四组:A-HBV+AFB1组;B-HBV组;C-AFB1组;D-空白对照组.整个实验期间,各组所有动物定期抽血及肝活检.实验于160周结束,处死所有动物.血及肝组织标本进行HBV感染标志及常规病理组织学等检测.结果:第一例HCC于实验99周时出现于A组.A、C组的HCC发生率分别为66.7%和30.0%;HCC的平均出现时间在A、C组分别为120.3±16.6周和153.3±5.8周(P<0.01).B、D组于实验结束时均无一例HCC发生,但动态观察中见B组的肝细胞增生结节不仅发生较早而且较多,其中一例于实验130周死亡时已有直径大至0.5cm的增生结节形成.结论:HBV作为独立的致肝癌因素时作用较弱,但HBV与AFB1有很强的协同致肝癌作用;在HCC的形成过程中可能存在着"病毒-化学协同机制".     

Inhibition of hepatocarcinogenesis by adrenocorticotropin in aflatoxin B1-treated rats.

We examined whether hormones would modify the carcinogenic action of aflatoxin B1 (AFB1). Four groups of inbred Fischer rats received AFB1, 125 mug per animal, weekly per os. In three of the groups, certain hormones were administered simultaneously: One group received 1 U growth hormone (GH) sc weekly, another was given 4 U adrenocorticotropin (ACTH) weekly, and a third received 0.5 U insulin weekly sc. AFB1, ACTH, and insulin were given for 20 weeks; GH was given for only 10 weeks. The control group did not receive hormone adjuvant. In each group, 4 animals were killed at 7, 14, 21, 28, and 35 weeks; the remaining rats were killed at 77 weeks. Their livers were carefully examined and samples prepared for light and electron microscopy. Animals receiving AFB1 and ACTH failed to exhibit hepatocellular carcinoma. On the other hand, malignant lymphoma appeared at 56 weeks in 3 of the 6 surviving males on this regime. AFB1, alone or when given with insulin or GH, caused hepatocellular carcinoma in all animals; in these, lymphoma was not observed. Lymphoma comprised two cell types, each with similar neclear characteristics but differing in their nucleocytoplasmic ratios and in the amount and distribution of cytoplasmic organelles. Alterations leading to hepatocellular carcinoma were examined at various stages of development. "Basophilic hyperplasia" reflected an increase in free ribosomes. "Hyperplastic nodules" were composed of hepatocyte aggregates with characteristics similar to those encountered in the earlier stage. Both the "neoplastic nodules" and hepatocellular carcinomas were formed by cells containing large, "smooth fingerprints" and free ribosomal aggregates. These features supported the concept that AFB1 impairs ribosomal binding to endoplasmic reticulum membranes. The failure of ACTH-treated animals to develop hepatocellular carcinoma was ascribed to the effect of adrenal cortical stimulation upon membrane-polysome binding.     

Mechanism of the resistance to cytotoxicity which precedes aflatoxin B1 hepatocarcinogenesis

In vivo there is evidence for a decreased covalent binding ofaflatoxin B₁ to DNA following low-level feeding with the toxin.It is suggested that this is due to increased detoxificationof aflatoxin B₁ by conjugation with reduced glutathione. Evidenceindicating increased levels of reduced glutathione and glutathioneS-trans-ferase activity are presented.     

Inhibition of ebselen on aflatoxin B(1)-induced hepatocarcinogenesis in Fischer 344 rats

Aflatoxin B₁ (AFB₁), a potent hepatocarcinogen, enhances ROSformation and causes oxidative DNA damage, which may play arole in its carcinogenicity. We have demonstrated recently thatebselen, an organic selenium compound, protects against thecytotoxicity of AFB₁ through its antioxidant capability. Thepresent study was designed to investigate the effect of ebselenon AFB₁-induced hepatocarcinogenesis in an animal model. Fischer344 rats were first treated with either deionized water or ebselen(5 mg/kg, 5 days/week) via gavage for 4 weeks, then given AFB₁(0.4 mg/kg, gavage, once a week) or AFB₁ plus ebselen (5 mg/kg,5 days/week) for another 24 weeks. The results showed that thehepatocarcinogenicity of AFB₁ in rats was significantly reducedby ebselen treatment as indicated by a decrease in: (i) serum     

Reliability of a short-term test for hepatocarcinogenesis induced by aflatoxin B1.

The reliability of a short-term test for hepatocarcinogenesis induced by aflatoxin B1 (AFB1) was tested by comparing the early appearance of gamma-glutamyl transpeptidase (GGT)-positive foci with the occurrence of primary liver cancer at a later stage. All rats received a basic short-term treatment with AFB1 intraperitoneally, during which three experimental groups received Chinese green tea or 2000 or 5000 ppm butylated hydroxyanisole in the diet and a control group received basic diet. Some of the rats in each group were sacrificed at the end of the short-term procedure, and the remainder were observed up to 92 weeks. The livers of all animals were examined for GGT-positive foci or primary liver tumours. The GGT-positive foci were most numerous and largest and the incidence of liver tumours was highest in the control group. These findings suggest that GGT-positive foci are a valuable preneoplastic marker for AFB1-induced hepatocarcinogenesis, that the short-term model is fairly reliable, and that both Chinese green tea and butylated hydroxyanisole inhibit AFB1-induced hepatocarcinogenesis.     

Enhancing effect of ethanol on aflatoxin B1-induced hepatocarcinogenesis in male ACI/N rats

The modifying effect of ethanol (EtOH) on aflatoxin B1 (AFB1)-induced hepatocarcinogenesis was examined in male ACI/N rats by chronic treatment at the post-initiation phase. Rats received an ip injection of AFB1 (1.5 mg/kg) twice a week for 10 weeks (a total of 20 doses). Following a week of acclimation, they were given 10% EtOH as drinking water for 56 weeks. The effect of EtOH on the hepatocarcinogenesis was evaluated in terms of the incidence of altered hepatocellular foci and neoplasms at the end of the experiment. Exposure to AFB1 alone induced a substantial number of altered foci (6.98 iron-excluding foci/cm2) in rats. The number of altered liver cell foci in rats receiving AFB1 followed by EtOH was significantly increased (26.39 iron-excluding foci/cm2). In the rats given EtOH after AFB1, the total area and mean diameter of both iron-excluding foci and altered foci identified in hematoxylin and eosin-stained sections were significantly higher than in the rats exposed to AFB1 alone. The incidence of liver cell tumors of the group given AFB1 and EtOH (3/15, 20%) was higher than that of the group treated with AFB1 alone (0/14, 0%). Treatment with EtOH alone for 56 weeks did not induce either. These results indicate an enhancing effect of EtOH on AFB1-induced hepatocarcinogenesis when it is given in the promotion phase.     

Enhancement of glutathione S-transferase placental-form positive liver cell foci development by microcystin-LR in aflatoxin B1-initiated rats

The objective of this study was to elucidate whether microcystin-LR(MC-LR), a hepatotoxic blue-green algal toxin in drinking water,is carcinogenic or possesses the ability to modulate aflatoxinB₁ (AFB₁)-induced hepatocarcinogenicity. In a medium-term liverbioassay, male Fischer 344 rats were given a single i.p. injectionof diethylnitrosamine (DEN, 200 mg/kg) followed by an i.p. injectionof MC-LR for 6 weeks after 2 weeks of DEN treatment. To studythe synergism between AFB₁ and MC-LR, DEN-treated rats weregiven an i.p. injection of AFB₁ (0.5 mg/kg) dissolved in dimethylsulfoxide (DMSO) followed by MC-LR at 2 weeks after the treatment.In a separate experiment, the rats were first given AFB₁ (0.5mg/kg) and 2 weeks later an i.p. injection of 1 or 10 µg/kgof MC-LR twice a week for 6 weeks. Most rats were subjectedto a two-thirds partial hepatectomy (PH) at week 3 and werekilled under anesthesia at week 8. Liver sections were analyzedfor glutathione S-transferase placental form (GST-P) expression,and subjected to histopathological examination for phenotypicalteration of hepatocellular foci. In rats that did not receiveDEN, MC-LR did not cause a significant increase in the numbersof GST-P-positive foci, whereas AFB₁ induced a slight increasein GST-P-positive foci development. In rats given DEN, MC-LRenhanced the expression of GST-P-positive foci, as did AFB₁but no synergism was observed. Histopathological analysis revealedthat the area of eosinophilic foci, a biomarker for preneoplasticliver lesion, markedly increased because of MC-LR. In rats givenAFB₁ as an initiator, treatment with MC-LR resulted in a synergisticincrease in the development of GST-P-positive foci. These resultssuggest that the hepatocarcinogenicities of MC-LR and AFB₁ canbe predicted in experimental animals with a medium-term bioassay.Furthermore, tumor promoting activity of MC-LR was demonstratedin rats treated with AFB₁.     

黄曲霉毒素B_1诱发大鼠肝癌过程中对CYP3A4活性的影响

目的探讨黄曲霉毒素B1（AFB1）诱发肝癌过程中对CYP3A4活性的影响。方法采用大鼠肝微粒体混合酶体外代谢体系,利用荧光定量法动态检测AFB1诱发肝癌过程中不同时期CYP3A4酶活性。结果 AFB1组CYP3A4含量从实验开始逐渐升高,至23周达顶峰,然后逐渐降低,到43周又升高,出现双波峰变化,阶段性比较差异有统计学意义（P=0.000）;对照组CYP3A4含量在整个实验过程中,除33周和63周出现明显降低外,其他各阶段变化不大;组间比较显示AFB1组在诱癌过程中有抑制CYP3A4的趋势,于63周抑制最明显,但尚未达统计学意义（P=0.638）;结论AFB1在诱癌过程中有抑制CYP3A4的趋势,可能是与早期癌变的细胞减少对特定基因毒性物质的活化有关。     

Effects of Ginkgo biloba extract on expression of biomarkers during aflatoxin B1-induced hepatocarcinogenesis in Wistar rats

Objective

The aim of this study was to study the effect of Ginkgo biloba extract (EGb761) on metabolism of aflatoxin B₁ (AFB₁) in Wistar rats.

Methods

Seventy one Wistar rats were assigned at random to groups A, B and C. Rats in groups A, B were injected with AFB₁ (intraperitoneal, 100–200 μg/kg body weight, 1–3 times/week). Group C was normal control. Rats in group B were fed in food with EGb761, while rats in groups A, C were given normal food. Blood samples were collected and liver biopsies were performed on the 14th, 28th and 42nd week. All the rats were sacrificed on the 64th week. The incidence of hepatocarcinoma was investigated. The hepatic phase I drug-metabolizing enzyme Cytochrome-P450 (CYP450) and phase II metabolizing enzyme glutathione S-transferase (GST) were analyzed with spectrometry. Serum AFB₁-lysine adduct levels were assessed with high performance liquid chromatography (HPLC). The expression of 8-hydroxydeoxyguanosine (8-OHdG) was measured with immunohistochemistry.

Results

The incidence of hepatocellular carcinoma (HCC) in group B was significantly lower than that in group A (26.92% vs 76.00%, P < 0.001). No HCC developed in group C. EGb761 showed no effects on the activities of CYP450 and GST in rat liver tissues. The level of AFB1-lysine adduct reached the peak (4356.01 pg/mg albumin) at the 14th week in group A. EGb761 significantly inhibited the formation of AFB1-lysine adduct in serum by 13.07% at the 14th week (P = 0.033), and 73.63% at the 42nd week (P = 0.002). The expression of 8-OHdG protein in rat liver tissues in group B was significantly lower than that in group A at the 28th, 42nd, and 64th week (P < 0.05).

Conclusion

The main mechanism underlying the effect of EGb761 in blocking hepatocarcinogenesis induced by AFB1 may not be fully attributable to its influence on the activity of liver phase I and phase II metabolizing enzymes. EGb761 inhibits the production of AFB₁-lysine adducts, decreases the expression of 8-OHdG protein, and finally alleviates the DNA oxidative injury, which may be one of the mechanisms for the effects of EGb761 in inhibiting or delaying AFB1-induced hepatocarcinogenesis.     

Metabolic basis for the protective effect of the antioxidant ethoxyquin on aflatoxin B1 hepatocarcinogenesis in the rat

The effect of dietary administration of 0.5% ethoxyquin (EQ) on the in vivo induction of enzymes and effect on aflatoxin B1 (AFB1)-DNA binding in liver and the consequent in vitro metabolism of AFB1 by male Fischer F344 rat liver-derived fractions have been examined. EQ increased microsomal cytochrome P-450s, in particular those isozymes classed as phenobarbital inducible, and the in vitro rate of metabolism of AFB1. The formation of the presumed detoxified metabolites, aflatoxins M1 and Q1, was enhanced to a greater extent than was the formation of the active metabolite, aflatoxin B1-8,9 epoxide (assessed by the level of aflatoxin B1-8,9-dihydrodiol). Prolonged feeding with EQ was accompanied eventually by a reduction in the initially elevated cytochrome P-450 content, but this was not reflected in any significant decrease in the rate of AFB1 metabolism in vitro. EQ increased the glutathione S-transferase activity of the liver cytosol fractions as assessed with the model substrate 1-chloro-2,4-dinitrobenzene. The capacity of these fractions specifically to catalyze the conjugation of AFB1 with glutathione was induced to a far greater extent than was the conjugation of 1-chloro-2,4-dinitrobenzene. gamma-Glutamyl transpeptidase was induced in the periportal areas of the liver lobule. Reduced in vivo binding of [3H]AFB1 to DNA of liver and kidney was found to result from EQ treatment. It is concluded that the reduced hepatocarcinogenesis which results from feeding EQ simultaneously with AFB1 is due to the reduction in DNA-adduct formation which in turn is due at least in part to increased detoxifying metabolism in the microsomal, cytosolic, and plasma membrane compartments of the liver cells.     

A comparative study on aflatoxin B 1 metabolism in mice and rats

In vivo metabolic studies on rats and mice revealed a marked difference in the fluorescent compounds produced after ingestion of aflatoxin B₁. The mouse converted aflatoxin B₁ to three unknown fluorescent compounds, designated x₁, x₂ and x₃ and the known aflatoxin M₁, while the rat was only capable of producing aflatoxin M₁. The results suggested that metabolites x₁, x₂, x₃ and aflatoxin M₁ were not part of a major metabolic pathway, but produced independently. These unknown yellowish-green fluorescent compounds did not seem to be conjugated with sulphate or glucuronic acid.     

银杏叶提取物对黄曲霉毒素B1所致大鼠肝癌相关蛋白表达影响的研究

目的:探讨银杏叶提取物(extract761 from Ginkgo giloba,EGb761)对黄曲霉毒素B1(aflatox B1,AFB1)致大鼠肝癌后相关蛋白表达的影响。方法:71只大鼠随机分为AFB1组(28只),AFB1+EGb761组(29只),空白对照组(14只)。第64周观察大鼠肝癌发生率,应用免疫组化法检测硫酸乙酰肝素类蛋白聚糖(MXR7)、环氧化酶(COX-2)以及细胞周期蛋白依赖性蛋白激酶抑制蛋白(p16)的表达情况。结果:AFB1组肝癌发生率明显高于AFB1+EGb761组(76%vs28%),χ2=10.602,P<0.05,对照组无肿瘤发生。AFB1+EGb761组MXR7蛋白含量明显低于AFB1组〔(1.488±1.178)vs(5.133±2.725)〕,F=18.638,P<0.001;AFB1+EGb761组p16蛋白含量明显高于AFB1组〔(2.456±1.014)vs(1.533±0.856)〕,F=24.091,P=0.03,AFB1组COX-2蛋白含量为3.305±0.566;AFB1+EGb761组为2.704±0.431,F=9.352,P=0.783。结论...     

Effect of butylated hydroxyanisole pretreatment on in vitro hepatic aflatoxin B1-DNA binding and aflatoxin B1-glutathione conjugation in rats

The effect of 3(2)-tert-butyl-4-hydroxyanisole (BHA) pretreatment of rats on both in vitro hepatic aflatoxin B1 (AFB1)-DNA binding and AFB1-glutathione (AFB1-SG) conjugation has been examined. For these studies, young male F344 rats were fed AIN-76 A diet with or without 0.75% BHA for 2 weeks. There were no significant differences either in microsomal cytochrome P-450 content or microsome-mediated exogenous DNA binding to AFB1 with cytochrome P-450 from control or BHA-treated animals. There were large differences in reduced glutathione S-transferase activity with treated cytosols showing 2.5-fold higher activity than the controls. Hepatic reduced glutathione levels were 25% higher in treated than in controls. Kinetics of cytosolic inhibition of microsome-mediated AFB1-DNA binding and formation of AFB1-SG conjugate when examined at two levels of AFB1 (2 and 10 microM) and a 4-fold range of cytosolic concentrations showed that inhibition of AFB1-DNA binding was greater with cytosol from the treated compared to the controls. However, AFB1-SG conjugation was 3- to 4-fold greater in treated than in controls. Inhibition of AFB1-DNA binding by cytosol was reversed in the presence of 1 mM level of various epoxides with concomitant inhibition of AFB1-SG conjugation. In reconstitution studies with 2 microM AFB1, intact nuclei alone from either group did not yield significant amounts of either DNA binding or AFB1-SG conjugation. However, addition of microsomes from either group to these nuclei generated a large amount of AFB1-DNA binding (82-111 pmol) and a smaller amount of AFB1-SG conjugate (9-28 pmol). The presence of cytosols from the control group reduced AFB1-DNA binding to a much lesser extent than the cytosols from the treated group. However, AFB1-SG conjugation was much higher with the cytosol from treated than with the controls. These reconstitution studies with endogenous DNA show more AFB1-DNA binding with the control than with BHA-treated animals and are in agreement with the studies in vivo. It appears that induced levels of cytosolic reduced glutathione S-transferase modulate AFB1-DNA binding and AFB1 hepatocarcinogenesis.