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The presence of ras family and p53 gene mutations in rat forestomach, intestine and liver tumors and soft tissue sarcomas induced by N-methyl-N-nitrosourea (MNU) was examined using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) followed by direct sequencing analysis. In the forestomach squamous cell carcinomas (SCC), Ha-ras and p53 mutations were detected in 2 (40%) and 4 (80%) of 5 cases, respectively. The figures for Ki-ras and p53 gene mutations in adenocarcinomas of the large and small intestines were 3 (18.8%) and 5 (31.3%) of 16 cases. Soft tissue sarcomas in different sites were found to have mutations of Ki-ras in 7 (23.3%) and of p53 in 9 (30%) of 30 cases. One forestomach SCC and 2 soft tissue sarcomas had double p53 mutations in different exons. Single cases of forestomach SCC and intestinal adenocarcinoma had mutations in both Ki-ras and p53 genes. No mutations were found in counterpart benign tumors or hepatocellular adenomas. The p53 mutation spectrum revealed preferential clustering within exon 8 for the forestomach SCCs, and exons 5 and 8 for the intestinal adenocarcinomas, whereas the distribution was evenly spread through exons 5 to 8 in soft tissue sarcomas. All the detected ras or p53 mutations were G:C to A:T transitions. These results indicate firstly that specific Ki-ras, Ha-ras and p53 gene mutations in MNU-induced lesions are related to particular alkylation sites (G:C to A:T transitions) and secondly, although not essential, Ki-ras, Ha-ras or p53 gene mutations may be involved in the progression stage of forestomach, intestine and soft tissue neoplasms induced by MNU.

To assess mechanisms of chemoprevention of hepatocarcinogenesis by trans-beta-carotene (beta-C), DL-alpha-tocopherol (alpha-T), and freeze-dried whole leaves of Kidachi aloe (Aloe), formation of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ)-DNA adducts was measured by 32P-post-labeling analysis, and CYP1A1 and CYP1A2 protein levels were analyzed by ELISA. Group 1 rats were fed diet containing 0.02% beta-C, 1.5% alpha-T or 30% Aloe over an 8-day period, while group 2 was given basal diet alone. On day 7, all animals were subjected to two-thirds partial hepatectomy (PH). Twelve hours after PH, they received a single dose of the carcinogenic food pyrolysate IQ (100 mg/kg) intragastrically, to initiate hepatocarcinogenesis. Rats were killed 6, 12, 24 and 48 h after IQ administration. The levels of adducts, expressed as relative adduct labeling values in rats treated with beta-C, alpha-T and Aloe, were decreased as compared with the control group at hour 24 (36 h after PH), with a significant difference in the case of the beta-C group (46.4% of the control value). Similarly, all showed a tendency for decrease at hour 48. Furthermore, the levels of CYP1A2, known to be responsible for activation of IQ, showed a significant reduction at hour 24. It is concluded that beta-C, and possibly also alpha-T and Aloe, have the potential to reduce IQ-DNA adduct formation, presumably as a result of decreased formation of active metabolites. The results may explain, at least in part, the previously observed inhibitory effects of these compounds on induction of preneoplastic hepatocellular lesions.

In a development trial for an initiation bioassay system, 7 known carcinogens and 1 suspected carcinogen were examined. In experiment 1, group 1 animals were initially subjected to partial hepatectomy (PH) 12 h before administration of diethylnitrosamine, 2-amino-3-methylimidazo[4,5-f]-quinoline (IQ), captafol, alpha-hexachlorocyclohexane or diethylstilbestrol (DES), then 2 weeks later underwent a promotion procedure comprising administration of phenobarbital (0.05% in diet) for 8 weeks and D-galactosamine (300 mg/kg, i.g.) at week 3. Group 2 received the promotion protocol alone as in group 1. Initiating potential was assayed on the basis of significant increase in values of preneoplastic placental form glutathione S-transferase-positive (GST-P+) foci of more than 3 cells in cross section at week 10. Numbers and areas of GST-P+ foci in group 1 given IQ, captafol and DES were significantly increased as compared to group 2, confirming the validity of the protocol as an initiation assay. In Experiment 2, group 1 rats were subjected to PH and 12 h later received a suspected carcinogenic mixture of opium pyrolysate (OP) or carcinogenic pesticide p,p'-dichloro-diphenyltrichloroethane or hexachlorobenzene. Application of a modified promotion procedure comprising cholic acid (0.15%) and carbon tetrachloride (1 ml/kg, i.g.) revealed significant initiation potential for OP. Overall the results indicate that the current protocols may be useful for detection of the initiation potential of carcinogens irrespective of their mutagenicity.

The prevalence of Ki-ras, Ha-ras and N-ras point mutation within exons 1 and 2 was studied in 17 cases of renal cell tumors (8 carcinomas and 9 adenomas) induced by N-ethyl-N-hydroxyethyl-nitrosamine or N-nitrosomorpholine. DNA samples prepared from acetone-fixed, paraffin-embedded tissues were amplified by means of the polymerase chain reaction, and point mutations at codons 12, 13 and 61 were analyzed by direct sequence methods with oligonucleotide primers. No mutations were detected in any of the renal tumors. The results thus indicated that ras family point mutation is not necessary for kidney tumor development in rats, supporting the view that ras mutations may not be generally relevant to neoplastic development in various organs in different species.

Hepatocyte growth factor (HGF), a potent mitogen for adult rat hepatocytes in primary culture, has previously been shown to be primarily expressed in the nonparenchymal cells of the liver. Using polyclonal antisera against human and rat HGFs we studied the tissue distribution of HGF immunohistochemically and found the most intense staining in the pancreas islet cells in both man (autopsy cases) and the rat. Differential localization of 4 pancreas islet hormones, glucagon, insulin, somatostatin and pancreatic polypeptide, revealed HGF to be preferentially expressed within the glucagon-positive cells. The results indicate that HGF is primarily produced or stored in A-cells and may act as a growth factor in a paracrine and an endocrine fashion, like various other hormones.

Carcinogenicity of Agaricus bisporus, the edible mushroom, was studied in rats. Female Charles River Sprague - Dawley rats (CD rats) were given a diet containing a 30% dry powder of A. bisporus for 500 days until the termination of the experiment. A control group was given a basal diet without A. bisporus. The rats in the experimental group had mammary tumor, thymoma, adrenal adenoma and pituitary adenoma. However, there was no significant difference in the incidence of these tumors between the experimental group and control group. No carcinogenic activity of A. bisporus was observed in this experiment.

BACKGROUND. Pancreaticobiliary maljunction (PBM), an anomalous union of the pancreatic duct with the common bile duct, has frequently been shown to be associated with biliary carcinoma. However, the mechanism of carcinogenesis is unknown. METHODS. Mutations of the K-ras oncogene were examined in cancerous and noncancerous biliary tract epithelium of 20 patients with PBM by an extraction of DNA from surgically resected histologic specimens. DNA was analyzed by a polymerase chain reaction single strand conformation polymorphism (PCR-SSCP) method and direct sequencing. RESULTS. An abnormally mobilized DNA band was detected not only in cancerous epitlielium but also in hyperplastic, metaplastic, and inflammatory epithelium of the gallbladder and/or common bile duct in patients with PBM. Among the biliary epithelium of patients with PBM, point mutation of K-ras oncogenes were detected in 4 of 5 (80%) cancerous epithelium, 7 of 12 (58%) hyperplastic and metaplastic epithelium, and 8 of 18 (44%) inflammatory epithelium, whereas no point mutation of the K-ras oncogene was detected in the gallbladder epithelium in 3 control patients without PBM. Direct sequence analysis of the K-ras oncogene revealed the mutation at codon 12 substituting the wild-type glycine (GGT) for aspartic acid (GAT) in all cancerous lesions of patients with PBM. Simultaneous two-point mutations from the wild-type glycine (GGC) to arginine (CGC) at codon 13 associated with the mutation at codon 12 were also found in one case of gallbladder carcinoma and one case of bile duct carcinoma. CONCLUSIONS. K-ras gene mutation is involved in the carcinogenesis of biliary tract epithelium in patients with PBM, and appears to be a high risk factor for carcinogenesis of the biliary tract.

Inhibitory effects of naturally occurring antioxidants on the initiation stage of hepatocarcinogenesis were studied. Group 1 rats were given a diet containing beta-carotene (beta-CT, 0.02%), alpha-tocopherol (alpha-TP, 1.5%), glutathione (GLT, 5%), vanillin (VNL, 1%), quercetin (QCT, 1%) or ellagic acid (ELA, 1%), or 3 doses of diallyl sulfide (DAS, 200 mg/kg, i.g.) over an 8-day period. On day 7, the animals received a single dose of 2-amino-3-methylimidazo[4,5-f] quinoline (IQ, 100 mg/kg, i.g.), 12 h after two-thirds partial hepatectomy for initiation and 2 weeks thereafter, were placed on promotion regimen comprising phenobarbital (0.05% in diet) and a single dose of D-galactosamine (100 mg/kg, i.p.). Groups 2 and 3 were treated as described for Group 1, but without test material or IQ, respectively. Survivors were killed at week 11 and antioxidant influence was assessed by comparing values for preneoplastic glutathione S-transferase placental form-positive (GST-P+) foci between Groups 1 and 2. All lesions larger than 70 microns in diameter consisting of approximately 5 cells in cross section were counted. Numbers of GST-P+ foci/cm2 in Group 1 were: beta-CT, 7.99; alpha-TP, 8.21; GLT, 9.71; DAS, 10.37; VNL, 10.57; QCT, 11.1; ELA, 12.5 (n = 11-15). All, except ELA, showed a significant decrease as compared with the Group 2 value of 14.54 (n = 15). Only beta-CT showed a significant decrease for the area value. This is the first report to show that beta-CT, alpha-TP, GLT, DAS, VNL, QCT exert inhibitory effects on initiation of hepatocarcinogenesis by the food carcinogen IQ, suggesting that these antioxidants might find application as chemopreventive agents. Furthermore, the current protocol proved practical for the assessment of chemopreventive agents within 11 weeks, a relatively short period.

A high intake of salted food is thought to be related to the high incidence of stomach cancer in Japan. In the present study, female F344 rats were divided into four groups. They were fed a nutritionally deficient purified diet (Group 1) and standard purified diet (Group 3) for 113 weeks and the same diets supplemented with salted cuttlefish guts, broiled, salted, dried sardines, pickled radish, and soy sauce (Groups 2 and 4). The incidence of papillomas and ulcers of the forestomach was highest in Group 4, which was given the standard diet supplemented with the salty food materials (p less than 0.05). These results suggest the importance of salted food as a suspicious causal factor in human stomach cancer in Japan.

The sulfated polysaccharides degraded carrageenan and dextran sulfate sodium were studied for genotoxicity using DNA repair tests employing cultured rat hepatocytes or intestinal mucosal cells. No evidence of unscheduled DNA synthesis was obtained. In addition, both alone and in combination with the comutagen norharman, these substances were nonmutagenic in the Salmonella microsome/mutagenicity test. These findings suggest that the carcinogenicity of the sulfated polysaccharides may be due to nongenotoxic mechanisms.

The carcinogenicity of Farfugium japonicum and Senecio cannabifolius was studied in ACI rats. In Group I, rats were given a diet containing 20% Farfugium japonicum. Groups II, III, IV and V were given diets containing 8%, 4%, 1% and 0.2% Senecio cannabifolius until the end of the experiment, respectively. The experiment was terminated after 480 days, except for Group V which was terminated 560 days after the start of feeding. Hemangioendothelial sarcoma of the liver and liver cell adenoma were induced in Groups I, IV and V. All rats in Groups II and III died of hepatotoxicity within a short period.

The effect of vitamin A deficiency on the carcinogenicity of betel quid was studied, using long-term feeding of vitamin-A-deficient and -sufficient diets with and without betel nut and calcium hydroxide in ACI rats. A high incidence of focal epithelial hyperplasia was observed in the upper digestive tract (tongue, buccal oral mucosa, esophagus, and forestomach) of rats in the group given the vitamin-A-deficient diet mixed with betel nut and calcium hydroxide. The vitamin-A-deficient group also showed a high incidence of squamous papilloma in the tongue, buccal mucosa, and forestomach. The incidence of hyperplastic lesions of the tongue and buccal oral mucosa was significantly higher in this group than in the group receiving the vitamin-A-sufficient diet with betel nut and calcium hydroxide. These results suggest that the vitamin-A-deficient condition enhanced the growth of epithelial hyperplasia that was due to the administration of the betel quid ingredients. However, vitamin A did not protect against the development of altered liver cell foci, which were frequently seen with a small number of hepatocellular neoplasms in all groups given the diets containing betel nut and calcium hydroxide (both vitamin-A-deficient and -sufficient groups).

Isolation of the carcinogen in the boiling water extract of bracken fern was conducted by following the active principle with a carcinogenicity bioassay. Fractionation of the bracken extract was carried out using adsorption on resin (Amberlite XAD-2 and TOYOPEARL HW-40 (c] and organic solvent extraction. A diet containing each of the fractions was given to 7 female Charles River Sprague-Dawley rats (CD rats) of 4 weeks old, except for the second fraction. All 7 rats given the last carcinogenic fraction developed mammary and intestinal tumors and 5 rats had urinary bladder tumors. Ptaquiloside (PT) which induced mammary cancer in female CD rats and rho-hydroxystyrene glycosides were isolated from this fraction.

The genotoxicity of cycasin was examined in the standard hepatocyte primary culture (HPC)/DNA repair test and in the test supplemented with β-glucosidase. Generally, no DNA repair was elicited by cycasin in the standard test except for one assay which showed a strong response. With the addition of β-glucosidase to the test medium, cycasin elicited DNA repair with clear dependence on both dose and amount of β-glucosidase. These results indicate that supplementation of the HPC/DNA repair test with the appropriate glucosidase should be useful in detecting potentially genotoxic glucosides and suggests that supplementation with other specific enzymes could compensate for extrahepatic biotransformation processes required prior to final activation by hepatocytes.

The genotoxicity of ptaquiloside (PT), recently isolated from bracken fern and shown to be carcinogenic, was examined by means of the hepatocyte primary culture/DNA-repair test. PT elicited clear unscheduled DNA synthesis with a dose-response effect. The result indicates that PT is a genotoxic carcinogen.

Some sulfated polysaccharides, such as d-CGN, APS, and DSS, have carcinogenicity to the rat colorectum. These materials first induced colitis, secondly squamous metaplasia, and finally tumors at the colorectum. Initially, colitis was located in the columnar epithelium of the rectum and extended proximally thereafter. Squamous metaplasia persisted in almost all experimental rats and progressed irreversibly. The tumors were adenoma, adenocarcinoma, squamous cell papilloma, and squamous cell carcinoma. Macrophages containing these materials were observed in the lamina propria mucosa and submucosa of the colorectum. There were differences in the molecular weight of the substances and their tumor incidences. However, with regard to their carcinogenicity, these sulfated polysaccharides were inferred to be similar to each other in their target organs and process of tumor development. Consequently, these sulfated polysaccharides may be one entity of carcinogenic sulfates.

(1985). Recent advances in research on bracken carcinogen and carcinogenicity of betel nut. Journal of Environmental Science and Health. Part C: Environmental Carcinogenesis Reviews: Vol. 3, No. 2, pp. 145-187.

The carcinogenic activity of petasitenine, a new pyrrolizidine alkaloid isolated from young flower stalk of Petasites japonicus, was studied in ACI rats. All rats that had received a 0.05% solution of petasitenine in drinking water died or were killed in moribund condition 72 days after the start of experiment. They showed necrosis, hemorrhage, and remarkable proliferation of the bile ducts in the liver. In another group that had received a 0.01% solution, 8 of 10 animals surviving beyond 160 days developed tumors in the liver, i.e., hemangioendothelial sarcomas in 5 rats and liver cell adenomas in 5 rats, 2 of which simultaneously developed hemangioendothelial sarcomas. No tumors were observed in the livers of the control animals.

The carcinogenic activity of clivorine, a pyrrolizidine alkaloid isolated from Ligularia dentata, was studied in inbred ACI rats. Twelve animals in the experimental group received a 0.005% solution of clivorine in drinking water for 340 days and survived beyond 440 days after the beginning of the experiment. Of this group, 8 developed tumors in the liver; 2 developed hemangioendothelial sarcomas, and 6 developed neoplastic nodules. The hemangioendothelial sarcoma showed metastasis in the lung of one rat. No tumors were observed in the liver of the control animals.

The carcinogenicity of dextran sulfate sodium was studied in inbred ACI rats. In group 1, 10 male rats were fed a 10% dextran sulfate sodium diet; in group 2, 14 male and 12 female rats were fed a 5% dextran sulfate sodium diet; and in the control group, 9 male and 9 female rats were given the basal diet without dextran sulfate sodium. After the start of the feeding regimen, all rats given the 10% dextran sulfate sodium diet died of severe diarrhea and anemia within 14 days and 15 of 23 surviving rats fed a 5% dextran sulfate sodium diet developed intestinal tumors between 134 and 215 days. These tumors were induced in the colon and cecum and consisted of adenomas, adenocarcinomas, and papillomas.

Fifteen male and 15 female Charles River Sprague-Dawley rats (CD rats) of 6 weeks old were given a diet containing 30% bracken fronds throughout the experimental period of 260 days. Male rats fed bracken diet developed multiple ileal tumors and urinary bladder tumors with incidences of 100% and 60%, respectively. Female rats developed multiple mammary tumors with an incidence of 87% in addition to ileal and urinary bladder tumors. The earliest mammary tumor was detected on day 78, and the last on day 256. The average number of tumors per rat was 5.6. Most mammary tumors were papillary carcinomas and adenocarcinomas. No tumors were found in rats in the control group.

The genotoxicity of a variety of hydrazine derivatives was examined in the DNA-repair test on rat or mouse hepatocytes. Out of 32 hydrazine derivatives, 6 chemicals, i.e., N-acetyl-4-(hydroxymethyl)phenylhydrazine, 1,2-dimethylhydrazine - 2HCl, 1-hydrazinophthalazine - HCl, methylhydrazine-sulfate, p, p′-oxybisbenzene disulfonylhydrazide and phenylhydrazine-HCl, elicited positive DNA repair responses in the test on rat hepatocytes. In the test on mouse hepatocytes, 4 more hydrazine derivatives, i.e., 1,1-dimethylhydrazine, hydrazine hydrate, hydrazine sulfate and 2-methyl-4-chlorophenoxyacetic acid hydrazide-HCl also generated positive responses, in addition to the 6 positive compounds in the rat assay. These results suggest that mouse hepatocytes are more susceptible to the genotoxicity of hydrazine derivatives, and that the species differences in genotoxicity appear to he in agreement with the in vivo carcinogenicity of these agents.

Fifteen female ACI rats initially 5 weeks old were each given a diet containing 0.027-0.08% ptaquiloside [(PT) CAS: 87625-62-5], a carcinogen in bracken, throughout the 210-day experimental period. A control group of 20 female ACI rats was given basal diet without PT. Both ileal and urinary bladder tumors developed in all rats in the experimental group. The ileal tumors were multiple and mostly developed in the distal 10 cm of the ileum. These ileal tumors were identified histologically as epithelial tumors, such as adenomas and adenocarcinomas, and also as nonepithelial malignant tumors, malignant fibrous histiocytomas. The urinary bladder tumors were transitional cell carcinomas, keratinizing squamous cell carcinomas, and sarcomas. Papillomas of the urinary bladder were found in 4 rats in the control group. These results show that, like bracken diet, PT induces tumors in both the ileum and urinary bladder.

The effect of the fecal stream on the induction of intestinal tumors was studied in 3 groups of SD rats. Rats in group 1 were subjected to single-barreled colostomies for the complete exclusion of the fecal stream at the proximal one-third level of the colons and were given consecutive iv injections of methylazoxymethanol acetate. Rats in group 2 were given methylazoxymethanol acetate alone. Rats in group 3 were not treated and served as controls. Tumors were noted in the small and large intestines of almost all rats in both groups 1 and 2. Even animals with single-barreled colostomies frequently developed tumors in the colon distal to the colostomy where the mucosa did not have contact with the fecal stream. These results indicated that carcinogens could probably reach the intestinal mucosa via the vascular system as well as by biliary transport.

Carcinogenicity of quercetin and rutin were examined in inbred ACI strain rats. Rats were given a diet containing 1% or 5% quercetin or 5% rutin for 540 days, or 10% quercetin and 10% rutin for 850 days. Rats in control groups were fed a normal basal diet. Most tumors found in experimental groups were also found in the corresponding control groups. Furthermore, there was no significant difference between the incidence of tumors in the experimental or control groups (P greater than 0.05). Thus, quercetin and rutin tested were not shown to be carcinogenic to ACI rats.

The effect of a lipotrope‐deficient diet on methylazoxymethanol acetate (MAM acetate) intestinal carcinogenesis was studied in 2 groups of ACI rats. For 10 weeks, weanling rats were fed a semisynthetic diet that was either high in fat and marginally deficient in the lipotropes (Group 1), or adequate for rats in all known respects (Group 2). Four weeks after the start of the experimental diets, the animals were given a weekly SC injection of MAM acetate, 25 mg/kg body weight, for a total of 5 times. Following the termination of the experimental diets, the animals were fed a basal diet until the end of the experiment (480 days). The tumor incidence in the jejunum‐ileum and the number of tumors in the colon were significantly higher in Group 1 than in Group 2. These results suggest that a marginally lipotrope‐deficient diet enhances MAM acetate carcinogenesis, probably in its early stage.

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.

The effect of betel nut on chemical carcinogenesis in the upper digestive tract and liver was examined in two different experimental models with ACI rats. The incidences of neoplasms and preneoplastic lesions of the tongue in animals given 5 ppm 4-nitroquinoline-1-oxide (4-NQO; CAS: 56-57-5) in the drinking water for 16 weeks and followed by 20% betel nut in the diet for 40 weeks were significantly higher than those in animals given 4-NQO alone. No enhancing effect from betel nut on the incidences of neoplastic and preneoplastic lesions in the upper digestive tract was found in animals administered 4-NQO for 12 weeks. The number of altered liver cell foci in rats given 200 ppm N-2-fluorenylacetamide (FAA; CAS: 53-96-3) in the diet for 8 weeks and followed by the betel nut diet for 16 weeks was significantly greater than that in animals fed the FAA diet alone. These results indicate enhancing effects of dietary administration of betel nut on oral carcinogenesis by 4-NQO and hepatocarcinogenesis initiated by FAA.

The carcinogenicity of dextran and 3 kinds of dextran sulfate sodium with different molecular weights and almost the same sulfur content were compared in ACI rats. Dextran sulfate sodium of molecular weight 54,000 showed a strong carcinogenic activity when it was given orally as 2.5% diet, whereas dextran sulfate sodium of molecular weight 520,000 and 9500 and dextran showed no significant carcinogenicity, i.e. the peak of carcinogenic activity of dextran sulfate sodium appeared at molecular weight 54,000, and dextran sulfates with larger or smaller molecular weights had no carcinogenic activity.

The carcinogenic activity of 3-amino-1-methyl-5 H -pyrido[4,3- b ]indole (Trp-P-2), a pyrolysis product of tryptophan, was studied in inbred strain ACI rats. Twenty rats received a diet containing 0.01% Trp-P-2 for 870 days. Nineteen rats survived for more than 400 days after the start of feeding, 1 rat developed a hemangioendothelial sarcoma of the liver, and 6 rats developed neoplastic liver nodules. These liver tumors were found only in female rats that died 666–870 days after the start of feeding. No liver tumors were found in control animals.

The carcinogenicity of the pyrrolizidine alkaloids senkirkine and symphytine was studied in male inbred ACI rats. Animals were divided into 3 groups: Group I received ip injections of freshly prepared senkirkine at a dose of 10% of the median lethal dose (LD50) twice weekly for 4 weeks and then once a week for 52 weeks. Group II received ip injections of symphytine at a dose of 10% of the LD50 by the same injection schedule as in group I. The control group was given ip injections of a 0.9% NaCl solution following the same injection schedule as in experimental groups. All group I rats survived for more than 290 days after the start of injections, and 9 of 20 rats developed liver cell adenoma. All group II animals survived for more than 330 days after the start of injections. Of 20 rats, 4 had liver tumors, 3 had hemangioendothelial sarcomas, and 1 had liver cell adenoma. The hemangioendothelial sarcomas showed metastasis in the lungs of 2 rats. The control group had no liver tumors.