Potent protection against aflatoxin-induced tumorigenesis through induction of Nrf2-regulated pathways by the triterpenoid 1-[2-cyano-3-,12-dioxooleana-1,9(11)-dien-28-oyl]imidazole.

Synthetic triterpenoid analogues of oleanolic acid are potent inducers of the phase 2 response as well as inhibitors of inflammation. We show that the triterpenoid, 1-[2-cyano-3-,12-dioxooleana-1,9(11)-dien-28-oyl]imidazole (CDDO-Im), is a highly potent chemopreventive agent that inhibits aflatoxin-induced tumorigenesis in rat liver. The chemopreventive potency of CDDO-Im was evaluated by measuring inhibition of formation of putative preneoplastic lesions (glutathione S-transferase P positive foci) in the liver of rats exposed to aflatoxin B1. CDDO-Im produces an 85% reduction in the hepatic focal burden of preneoplastic lesions at 1 micromol/kg body weight and a >99% reduction at 100 micromol/kg body weight. CDDO-Im treatment reduces levels of aflatoxin-DNA adducts by approximately 40% to 90% over the range of 1 to 100 micromol/kg body weight. Additionally, changes in mRNA levels of genes involved in aflatoxin metabolism were measured in rat liver following a single dose of CDDO-Im. GSTA2, GSTA5, AFAR, and EPHX1 transcripts are elevated 6 hours following a 1 micromol/kg body weight dose of CDDO-Im. Microarray analysis using wild-type and Nrf2 knockout mice confirms that many phase 2 and antioxidant genes are induced in an Nrf2-dependent manner in mouse liver following treatment with CDDO-Im. Thus, low-micromole doses of CDDO-Im induce cytoprotective genes, inhibit DNA adduct formation, and dramatically block hepatic tumorigenesis. As a point of reference, oltipraz, an established modulator of aflatoxin metabolism in humans, is 100-fold weaker than CDDO-Im in this rat antitumorigenesis model. The unparalleled potency of CDDO-Im in vivo highlights the chemopreventive promise of targeting Nrf2 pathways with triterpenoids.

Response of two rodents, Mastomys natalensis and Mystromys albicaudatus, to the pancreatic carcinogen azaserine.

The response of two rodents to azaserine carcinogenicity for the pancreas was evaluated. Mystromys albicaudatus was not responsive; however, Mastomys natalensis developed large numbers of atypical acinar cell nodules and several adenomas in a 6-month study. Mastomys is the most responsive of several animals in which azaserine has been studied as a pancreatic carcinogen.

Species and rat strain variation in pancreatic nodule induction by azaserine.

Subtoxic doses of azaserine induced atypical acinar cell nodules (AACN) in the pancreases of outbred Wistar rats, inbred W/LEW and F344 rats, and outbred Charles River CD-1 albino mice 4-6 months after initiation of treatment in the growing animal. These AACN apparently represented preneoplastic lesions, some of which have the potential to develop into adenomas or adenocarcinomas. Wistar and W/LEW rats were highly responsive to nodule induction; AACN developed in about 90% of the outbred Wistar rats and in all of the W/LEW rats tested. F344 rats were less susceptible and developed about 10% as many AACN as the Wistar rats. Female rats developed approximately half as many AACN as males. The mouse was intermediate in response between the F344 and the two Wistar rats. Syrian golden hamsters and strain 13 guinea pigs were relatively unresponsive. These studies of azaserine-induced AACN provided a basis for selection of carcinogenic azaserine regimens and suggested that the young male W/LEW rat was the most sensitive of the animals studied.

Enhancement of pancreatic carcinogenesis by a dietary unsaturated fat in rats treated with saline or N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine.

The effect of diets high in an unsaturated fat on the enhancement of pancreatic carcinogenesis in saline-treated rats and in rats treated with N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine (HPOP) was examined. Young male LEW rats were treated with a single dose of HPOP (160 mg/kg body wt) or saline, fed diets containing 5 or 20% corn oil, and then autopsied 12 months later. The pancreata of HPOP-treated rats fed the diet with 5% fat contained multiple foci and nodules of atypical acinar cells (AACN), acinar cell adenomas, and localized carcinomas. Rats fed the diet with 20% fat developed a similar spectrum of pancreatic lesions and also developed carcinomas that showed local invasion or metastasis to regional lymph nodes. The incidence and multiplicity of localized carcinomas was significantly higher in the group that was fed the high-fat diet. HPOP also induced neoplasms in the liver, lungs, and kidneys, but none of these had a higher incidence in the group fed the high-fat diet. Among rats that received no carcinogen, the incidence of AACN was high, but the multiplicity of these lesions was low, an average of three per pancreas in groups fed both levels of fat; however, the average area of AACN transections was larger in the high-fat diet group. One acinar cell adenoma and 1 carcinoma developed in the group of 11 rats fed the 20% corn oil diet, whereas no neoplasms developed in the group of 12 rats fed the 5% corn oil diet. Although the incidence of pancreatic neoplasms is not significantly different in these 2 groups, the data are consistent with the hypothesis that initiated foci are promoted to grow and become neoplasms in the pancreas of rats that are fed diets with a high content of unsaturated fat--as was demonstrated in the HPOP-treated rats.

Pathologic and biochemical effects of azaserine in inbred Wistar/Lewis rats and noninbred CD-1 mice.

Inbred W/LEW rats and noninbred CD-1 mice were compared for responsiveness to induction of pancreatic adenocarcinomas by azaserine. At 1 year following the first of 15 weekly ip injections of 10 mg azaserine/kg body weight, the rats had a pancreatic adenoma incidence of 71% (12/17) and a pancreatic adenocarcinoma incidence of 35% (6/17), with 1 invasive adenocarcinoma. The mice showed none of these advanced lesions, although numerous pancreatic atypical acinar cell nodules (AACN) were present. An examination of the number and size of the AACN showed the rats to hve more and larger AACN thatn did the mice. The concentration of [14C]azaserine and/or its metabolites was greater in rat pancreas than in mouse pancreas. Alkaline sucrose gradients were used to compare azaserine-induced pancrewtic and liver DNA damage in W/LEW and F344 rats, the CD-1 mouse, the Syrian golden hamster, and the strain 13 guinea pig. DNA damage was detected 1 hour following azaserine administration in both pancreata and livers of all animals tested and persisted for at least 1 week in the pancreata of all animals except the CD-1 mouse; however, neither the degree nor persistence of DNA damage accurately reflected the differing responsiveness of these species to induction of pancreatic AACN or neoplasms by azaserine.

Modulation of azaserine-induced pancreatic foci by phenolic antioxidants in rats.

Effects of the dietary phenolic antioxidants butylated hydroxyanisole [(BHA) CAS: 25013-16-5; (1,1-dimethylethyl)-4-methoxyphenol] and butylated hydroxytoluene [(BHT) CAS: 128-37-0; 2,6-di-tert-butyl-p-cresol] on pancreatic tumorigenesis were examined. Male LEW inbred rats were given injections of 30 mg azaserine [CAS: 115-02-6; diazoacetate (ester) serine] per kg body weight once a week for 3 weeks and maintained on either a control diet or 0.45% BHA- or 0.45% BHT-supplemented control diet throughout the initiation and post-initiation phases of the experiment. At 4 months post initiation, pancreatic tissue sections were quantitatively examined for the number and size of preneoplastic foci. BHT and BHA treatments reduced the number of acidophilic foci per pancreas by 32 and 48%, respectively, but were without effect on focal size. By contrast, basophilic foci were not subject to modulation by these antioxidants. A constellation of enzyme activities involved in carcinogen inactivation and known to be perturbed by antioxidant treatment was examined in liver and pancreas. The hepatic activities of glucose-6-phosphate dehydrogenase, glutathione reductase, and glutathione-S-transferases were markedly elevated while catalase and superoxide dismutase activities were unchanged. Glutathione peroxidase activity was diminished. In the pancreas, only glutathione peroxidase activity was affected, and it was reduced in both the BHA and BHT treatment groups. Although the pancreas is refractory to the enzyme inductive effects of these antioxidants, morphometric analysis of foci demonstrated chemoprevention by BHA and BHT of azaserine-induced foci. Whether this reduction reflected inhibition of an initiation, postinitiation , or a combination of effects was not known.

Inhibition by retinoids of the growth of azaserine-induced foci in the rat pancreas.

The usefulness of a short-term azaserine [CAS: 115-02-6; diazoacetate serine (ester)]-rat model for the screening of retinoids (known chemopreventive agents) and the effect of two retinoids on the growth of azaserine-induced, presumptive preneoplastic foci of acinar cells were examined. At 14 days of age, male Lewis rats were each given injections of a single dose of 30 mg azaserine/kg body weight. These rats were weaned to test diets to which retinoids were added. At 4 months post initiation, pancreata were examined by quantitative stereologic methods to determine number and mean size of foci. Two phenotypically different populations of foci were observed and characterized as acidophilic or basophilic. Retinylidene dimedone and N-2-hydroxyethylretinamide decreased the number and size of the acidophilic foci but not the basophilic foci. The inhibition of growth of the acidophilic foci correlates well with the known effects of these retinoids in long-term carcinogenicity studies.

Induction of pancreatic carcinomas in rats with N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine: histopathology.

The carcinogenicity of N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine (HPOP) for rat pancreas was evaluated. Two-week-old male LEW rats were given a single ip injection of HPOP, 160 mg/kg body weight; the rats were autopsied 4, 6, or 12 months later. Histologic examination showed that the pancreata contained multiple foci of atypical acinar cells and nodules of atypical acinar cells (AACN), acinar cell adenomas, localized carcinomas, and carcinomas. The incidence of carcinomas was 77%. The carcinomas were composed of poorly differentiated acinar cells and ductlike structures. Pancreatic ducts were unaffected. The prominence of AACN, the histologic type of the neoplasms, and the absence of hyperplastic changes in ductal epithelium suggest that the pancreatic carcinomas were derived from acinar cells. The incidence of liver cell carcinomas and pulmonary adenomas was similar to that of localized pancreatic carcinomas. Neoplasms of other organs were less frequent. HPOP has been shown to induce pancreatic carcinomas in hamsters but has not previously been reported to be a pancreatic carcinogen in rats.

Dietary intervention during the postdosing phase of L-azaserine-induced preneoplastic lesions.

The effects of intervention by diets with high or low levels of dietary fat on the development of preneoplastic pancreatic lesions were examined. Wistar rats were treated ip at 14 days of age with a 30-mg/kg dose of L-azaserine [CAS: 115-02-6; diazoacetate serine (ester)] and weaned onto the test diets. Animals fed 5% corn oil had fewer preneoplastic lesions compared to animals fed 20% corn oil throughout the 4-month posttreatment period. The strong response observed in rats fed 20% corn oil could be markedly reduced by intervention with a 5% corn oil diet halfway through the posttreatment period. Similarly, the low response in animals fed 5% corn oil could be markedly elevated by intervention with a high-fat diet. These results provide evidence for the hypothesis that tumor development may be modified by dietary means.

Effect of dietary intake of fish oil and fish protein on the development of L-azaserine-induced preneoplastic lesions in the rat pancreas.

The effect of dietary intake of fish (menhaden) oil and fish (cod) protein on the development of pancreatic preneoplastic lesions was examined in male Wistar rats. Fourteen-day-old animals were given a single ip injection of 30 mg L-azaserine/kg body weight [CAS: 115-02-6; diazoacetate serine (ester)]. At 21 days of age they were weaned and maintained on dietary treatment for 4 months. Fish protein did not appear to produce a significantly different preneoplastic response when compared to casein as a protein source. However, a 20% menhaden oil diet, rich in omega 3 fatty acids, produced a significant decrease in the development of both the size and number of preneoplastic lesions when compared to a 20% corn oil diet rich in omega 6 fatty acids. This study provides evidence that fish oils, rich in omega 3 fatty acids, may have potential as inhibitory agents in cancer development.

Effect of dietary omega-3 and omega-6 fatty acids on development of azaserine-induced preneoplastic lesions in rat pancreas.

We examined the effect of varying the ratio of dietary omega-3 (omega 3) to omega-6 (omega 6) on the development of pancreatic preneoplastic lesions in male Wistar rats given azaserine at 14 days of age. As the ratio of dietary omega 3 to omega 6 fatty acids increased in a diet totaling 20% by weight of fat, the development of preneoplastic atypical acinar cell nodules (AACNs) at 4 months after dosing with azaserine decreased significantly. In addition, serum levels of prostaglandin thromboxane B2, prostaglandin E2, and 6-keto-prostaglandin F1 alpha decreased significantly. The fatty acid composition of the rbc membrane was also significantly influenced by the ratio of dietary omega 3 to omega 6 fatty acids. In a second experiment, we examined the effect of dietary intervention with a different type of fat (corn oil or menhaden oil) 2 months into the 4-month postdosing period on AACN development at the end of the post-dosing period. Intervention of the omega 6 fatty acid-rich diet with the omega 3 fatty acid-rich diet significantly decreased focal development. The opposite was true when intervention involved substituting the omega 3 fatty acid-rich diet with the omega 6 fatty acid-rich diet.

Species comparison of in vitro metabolism of aflatoxin B1.

The metabolism of [14C]aflatoxin B1 by 9000 X g supernatant fraction of livers of duck, rat, mouse, monkey, and humans was compared by incubating the compound and liver fractions in the presence of cofactors and air for 30 min. The incubation medium was extracted with chloroform, and the soluble metabolites were separated on thinlayer plates and quantified; radioactivity remaining in the aqueous phase was determined in order to quantify metabolism to water-soluble derivatives. Duck, monkey, and human livers were most active in total conversion, each metabolizing approximately 80% of available substrate in 30 min. Rat and mouse livers had lower activites, metabolizing 15 to 20%. Duck liver produced mainly (60%) chloroform-insoluble derivatives, but all other species produced larger quantities of chloroform-soluble than insoluble metabolites. Aflatoxin Q1 was the principle chloroform-soluble metabolite produced by monkey, human, and rat liver, whereas duck liver produced mainly aflatoxicol in that fraction. Aflatoxin P1 was produced by monkey, human, and mouse liver, but not by duck and rat. The chromatographic region containing M1 and B2alpha contained low levels of radioactivity in all species except human. No consistent pattern of metabolism emerged which could be correlated with species differences in response to aflatoxin B1 toxicity or carcinogenicity.

Caloric restriction and intervention in pancreatic carcinogenesis in the rat.

In two experiments, the effects of caloric restriction during the postinitiation phase of pancreatic carcinogenesis were evaluated. Male Lewis rats were given injections of azaserine at 14 days of age and weaned to the postinitiation test protocols at 21 days of age. In the first experiment, the caloric content of the diets was restricted by 10, 15, 20, and 30% of the intakes of the ad libitum-fed rats. A sixth group was fed diet ad libitum for only 5-6 h/day; i.e., they were "meal-fed". The development of putative preneoplastic lesions (henceforth termed foci) was evaluated by quantitative stereological (morphometric) analysis of the pancreas. Caloric restriction during the 4-month postinitiation phase resulted in a significant reduction in focal development beginning at 10% caloric restriction and increasing with more severe restriction. The caloric intake of the meal-fed group closely matched the caloric intake of the 10 or 15% caloric restriction groups and the focal response of the meal-fed rats was similar to the groups restricted in calories by 15 to 20%. In the second experiment, rats were initiated with azaserine and weaned to one of four groups: ad libitum; meal-fed; meal-fed for 2 months and ad libitum thereafter; or ad libitum for 2 months and meal-fed thereafter. Foci were evaluated at 2 and 4 months; neoplasm incidence and multiplicity were determined at 14 months postinitiation. Compared to the ad libitum group, the meal-fed group had significantly fewer foci at all times of evaluation and significantly fewer neoplasms. When rats were meal fed for 2 months and then switched to ad libitum feeding for the remainder of the experiment, the focal outcome at 4 months was similar to the group meal fed for all 4 months; and at 14 months the neoplastic outcome was intermediate between the ad libitum and the meal-fed group. Intervention in the ad libitum feeding regimen at 2 months by meal feeding for the remainder of the experiment resulted in a significant decrease in the focal and neoplastic development, as compared to the group fed ad libitum continuously. These two intervention groups were intermediate in response between the meal-fed and ad libitum-fed groups. These results indicate that the postinitiation phase of pancreatic carcinogenesis can be modulated by relatively simple dietary interventions such as moderate caloric restriction.

In vitro metabolism of aflatoxin B2 by animal and human liver.

The metabolism of aflatoxin B2 by postmitochondrial supernatant fractions of duck, rat, mouse, and human livers was studied in an in vitro system. Duck liver had a much higher level of activity than had tissues from other species. Postmitochondrial supernatant equivalent to 0.2 g whole liver metabolized 40 to 80% of the initial substrate in 30 min, compared to less than 6% for the other species. Among several metabolites formed by duck liver, aflatoxin B1 was produced in amounts equivalent to 2 to 8% of the initial substrate, and metabolites having chromatographic properties postualted for aflatoxicols 1 and 2 and aflatoxins M1 and M2 were also formed in small amounts. In contrast, rat, mouse, and human liver preparations produced no detectable aflatoxin B1 and only small amounts of compounds thought to be aflatoxins Q2 and P2. The greater susceptibility of duck liver to the toxicity of aflatoxin B2 may be attributable to its ability to form aflatoxin B2 may be attributable to its ability to form aflatoxin B1, which could then be activated through further metabolism.

Dietary modulation of azaserine-induced pancreatic carcinogenesis in the rat.

Because diet has been shown to modulate the incidence of a wide variety of chemically induced cancers in experimental animals, various dietary constituents were evaluated for their ability to modulate the incidence of pancreatic exocrine cancer in male Wistar/Lewis rats given injections of the pancreatic carcinogen, azaserine. Ten different diet regimens were fed. The incidence of pancreatic cancers in rats fed a control diet was compared to that in groups fed diets formulated to evaluate the effect of caloric restriction, high protein, low protein, low fat, cyclopropenoid fatty acids, lipotrope deficiency, high unsaturated fat, and high saturated fat. The incidence of pancreatic adenomas and carcinomas was evaluated by light microscopy. The number of pancreatic neoplasms was reduced in carcinogen-treated groups which were underfed the control diet or fed the diet high in protein. Pancreatic carcinogenesis appeared to be enhanced in two groups which were fed diets containing 20% corn oil, i.e., high in unsaturated fat; whereas, the group fed a diet high in saturated fat had the same incidence of neoplasms as did the group fed the control diet. The pancreatic neoplasms from groups in which the incidence was enhanced by diet showed less evidence of acinar cell differentiation and displayed diverse histological types. In the lipotrope-deficient group, there was a significantly increased incidence of hepatocellular carcinoma; however, a low incidence of liver tumors was encountered in all other dietary groups.

Protective effects of voluntary exercise during the postinitiation phase of pancreatic carcinogenesis in the rat.

Studies were undertaken to evaluate the effects of exercise on the development of pancreatic cancer. Exercise is one life-style factor that has received little attention with regard to its role in the etiology of cancer. Male Lewis and female F344 rats were initiated with azaserine during the suckling period and weaned to the experimental protocols. Food and water were available ad libitum. A purified diet of 20% unsaturated fat was fed to both the sedentary and exercise groups. Rats of the exercise group had free access to voluntary exercise wheels. At approximately 2 and 4 months postinitiation, pancreases were evaluated for the number and size of azaserine-induced putative preneoplastic foci by quantitative stereology. Voluntary exercise activity peaked at approximately 2 months postinitiation with a gradual decline in activity there-after. Male Lewis rats averaged 0.95 +/- 0.13 km/day (SE) and female F344 rats averaged 2.73 +/- 0.26 km/day of voluntary wheel running. Compared with the sedentary groups, male Lewis and female F344 rats with access to the running wheels had significantly smaller foci at 4 months postinitiation. Azaserine-induced foci were evaluated in the male Lewis rats at both 2 and 4 months postinitiation. At 4 months postinitiation, the size and growth rate (as measured by [3H]thymidine autoradiography) of foci were less in the rats with access to the exercise wheels. No differences were observed at 2 months postinitiation. Access to voluntary exercise reduced the growth rate of azaserine-induced pancreatic foci. The effect occurred late in the postinitiation phase and was not directly related to the extent of running activity early in the postinitiation phase.

Effects of dietary fats and soybean protein on azaserine-induced pancreatic carcinogenesis and plasma cholecystokinin in the rat.

Both dietary unsaturated fat and raw soybean products are known to enhance pancreatic carcinogenesis when fed during the postinitiation phase. A comparison of these two dietary components was made to evaluate the relative potency of each ingredient for enhancing pancreatic carcinogenesis and to determine if this enhancement was correlated with an increase in plasma cholecystokinin (CCK) levels. Male Wistar rats were initiated with a single dose of azaserine (30 mg/kg body weight) at 14 days of age. The rats were weaned to test diets formulated from purified ingredients. Dietary protein at 20% by weight was either casein or soy protein isolate (heat treated or raw). Corn oil was the unsaturated fat of major interest and it was fed at either 5 or 20% by weight. Pancreases were quantitatively evaluated for carcinogen-induced lesions at 2- and 4-month postinitiation. In a second experiment designed to closely mimic the above experiment, rats were implanted with cannulae which allowed plasma to be repetitively sampled over a 2.5-week period during which the test diets were fed. Plasma was collected both prior to introduction of the test diets and afterwards. Plasma CCK was measured by a specific radioimmunoassay. Both the 20% corn oil diet and the raw soy protein isolate diet enhanced pancreatic carcinogenesis. The effects of the raw soy protein isolate on the growth of the carcinogen-induced lesions were significantly greater than the effects of the 20% corn oil diet. Plasma CCK values were not elevated in the rats fed the 20% corn oil diet, but they were significantly elevated in the rats fed the raw soy protein isolate. Heat-treated soy protein isolate neither enhanced carcinogenesis nor elevated the plasma CCK level. This study demonstrates that certain plant proteins enhance the growth of carcinogen-induced pancreatic foci and that this effect is considerably greater than the enhancement by high levels of dietary unsaturated fat. Furthermore, the enhancement by the raw soy protein isolate may be mediated by CCK; but this does not appear to be the mechanism by which the unsaturated fat, corn oil, enhances pancreatic carcinogenesis.

Inhibition of pancreatic carcinogenesis by retinoids in azaserine-treated rats.

Chemoprevention by retinoids of the progression of carcinomas induced in rats by azaserine was evaluated. Wistar/Lewis rats were given 15 weekly injections of azaserine, 10 mg/kg, while fed a chow diet; after the completion of carcinogen treatment, they were fed a chow diet supplemented with four different retinoids at the level of 0.5 to 2 mmol/kg diet for 1 year. The incidence of neoplasms was determined by autopsy and histological study. The incidence of pancreatic carcinoma among a male positive control group (azaserine treated, but not retinoid treated) was 42%. The incidence of pancreatic carcinoma among male rats treated with retinoids was: N-2-hydroxyethylretinamide, 6%; N-4-propionyloxyphenylretinamide, 17%; and retinylidene dimedone, 12%. The incidence in rats fed these three retinoids was significantly (p less than 0.05) below the control group incidence. Thus, these three retinoids appeared to be effective in inhibiting the progression of pancreatic carcinomas in the azaserine-induced model. A similar trend was demonstrated in females, but statistical significance was shown only in rats fed N-2-hydroxyethylretinamide. A fourth retinoid, N-4-carboxyphenylretinamide, was more toxic and less effective in chemoprevention. Since retinoids were fed after exposure to carcinogen, the effect was exerted during the postinitiation phase of carcinogenesis. The ratio of invasive pancreatic carcinomas to localized carcinomas (carcinoma in situ) was clearly higher among non-retinoid-treated rats than among those treated with retinoids. This is consistent with retarded progression in the retinoid-treated groups.

Promotion by unsaturated fat of azaserine-induced pancreatic carcinogenesis in the rat.

Diet has been shown to modulate the incidence of a wide variety of chemically induced cancers in animals. Various diets fed either during the initiation stage or the postinitiation (promotion) stage of carcinogenesis were evaluated for their ability to modulate the incidence of pancreatic cancer. Male Wistar/Lewis rats were treated with multiple injections of the pancreatic carcinogen, azaserine, during a 6- to 7-week-long initiation phase and were autopsied after a postinitiation phase of 34 or 44 weeks. The following diets were evaluated for their effects on the incidence of pancreatic neoplasms during each stage of carcinogenesis: high saturated fat; two high unsaturated fats (corn oil and safflower oil); low protein; and caloric restricted. A purified control diet was fed during that stage when the test diets were not fed. The incidence of pancreatic adenomas and adenocarcinomas was evaluated by light microscopy. Feeding of the caloric-restricted diet during the initiation phase suppressed the pancreatic neoplasm incidence. None of the ther diets tested had an effect on the incidence of pancreatic cancer during the initiation phase. During the postinitiation phase, both high-unsaturated-fat diets but not the high-saturated-fat diet significantly elevated the pancreatic neoplasm incidence. The low-protein and caloric-restricted diets had no effect on the neoplasm incidence when fed during the postinitiation phase. Thus, diets high in unsaturated fat appear to promote pancreatic carcinogenesis in the azaserine-treated rat while a diet high in saturated fat failed to show a similar degree of enhancement of pancreatic carcinogenesis.

Inhibitory effects of estrogen and castration on the early stage of pancreatic carcinogenesis in Fischer rats treated with azaserine.

Effects of sex steroids on pancreatic carcinogenesis during the early stage were studied in azaserine-treated rats of both sexes. Fischer rats were given weekly i.p. injections of azaserine (30 mg/kg) [CAS:115-02; diazoacetate serine(ester)] at 2 and 3 weeks of age and were divided into six groups. Castration, ovariectomy, and s.c. implantations of either a 0.3-mg or a 1.0-mg 17 beta-estradiol (CAS:50-28.2; estradiol) pellet were performed at 7 weeks of age. The groups were as follows: group 1, intact male; group 2, castrated; group 3, castrated plus 0.3 mg estradiol; group 4, castrated plus 1.0 mg estradiol; group 5, ovariectomized; and group 6, intact female. Rats were killed 4 months after the last injection of azaserine. Azaserine treatment induced atypical acinar cell foci and nodules (AACN) in both sexes. The acidophilic AACN are considered preneoplastic lesions. An apparent sex difference was observed; the number of acidophilic AACN was greater in male rats than in female rats. Castration caused a significant decrease in both the serum testosterone levels and the number of acidophilic AACN, which were comparable to those in ovariectomized female rats. Furthermore, when estradiol treatment was administered to the castrated male rats, a linear decrease in the number of acidophilic AACN and an elevation in the serum estradiol levels were observed and were dose dependent. There were also positive relationships between estradiol treatments and the mean pituitary and pancreas weights. These results showed that estradiol treatment and the drop in testosterone levels caused by castration were highly effective in inhibiting the development and growth of preneoplastic lesions of the pancreas of the rats treated with azaserine. This estradiol effect was dose dependent. The present study, therefore, provides evidence that estrogen may act as an inhibitor and androgen as a promoter in the early stage of pancreatic carcinogenesis in rats.