DNA alterations in rat organs after chronic exposure to cigarette smoke and/or ethanol ingestion.

In spite of the epidemiological evidence supporting a synergism between alcohol consumption and cigarette smoking in the pathogenesis of cancers of the aerodigestive tract, there is a paucity of experimental studies evaluating the effects of these agents under well-controlled conditions and exploring the mechanisms involved. We exposed groups of female BD6 rats, aged 8 months, to ethanol (5% in drinking water for 8 consecutive months) and/or whole-body to mainstream cigarette smoke (1 h/day, 5 days/week for 8 months). DNA was purified from different organs and analyzed for the presence of DNA-protein crosslinks and 32P-postlabeled DNA adducts after butanol enrichment. No significant increase of DNA-protein crosslinks, compared to untreated controls, was induced by any treatment in liver, lung, or heart. 'Spontaneous' nucleotidic modifications were detected by 32P-postlabeling in organs of untreated rats, with the highest levels occurring in the heart. Ingestion of ethanol did not affect DNA adduct levels in any of the organs examined: esophagus, liver, lung, and heart. Exposure to cigarette smoke induced formation of DNA adducts in the lung and heart, but not in the esophagus or liver. The combined ingestion of ethanol resulted in a significant formation of smoke-related DNA adducts in the esophagus and in their further, dramatic increase in the heart. It thus appears that ethanol consumption increases the bioavailability of DNA binding smoke components in the upper digestive tract and favors their systemic distribution. The mechanisms responsible for the interaction between ethanol and smoke and for the selective localization of DNA alterations in different organs are discussed. Formation of DNA adducts in the organs examined may be relevant in the pathogenesis of lung and esophageal cancers as well as in the pathogenesis of other types of chronic degenerative diseases, such as chronic obstructive pulmonary diseases and cardiomyopathies.

Genotoxic changes in the pulmonary alveolar macrophages of mice, rats and hamsters treated with tobacco smoke.

To determine whether tobacco smoke (TS) is genotoxic for lung tissue macrophages (pulmonary alveolar macrophages, PAM) as a general result of its inhalatory action BD6 rats, Syrian golden hamsters and BDF1 (C57BlxDBA2) mice were subjected to wholebody exposure for 90 or 60 min daily (600 cm3 mainstream smoke in 16-1 glass chamber, 9 or 6 exposures of 15 min each, respectively), for different periods ranging up to 30 days. A significant enhancement of the frequency of polynucleated macrophages (BiN PAM) was observed in all animal species after more than 10-days of repeated exposure to TS. The increased level of BiN PAM (the number of bi- (+) poly-nucleated PAM) correlates with the duration of exposure to TS: on day 20 after the start of inhalation, more than 25/1000 of mice PAM were polynucleated, while on day 30 this applied to approximately 50/1000. Furthermore, a highly significant increase in the level of micronucleated PAM (MN PAM) was also established after 10 days TS treatment of mice and persisted to the end of these examinations. TS was effective in enhancing the micronucleated and polynucleated PAM levels in hamsters irrespective of their sex, as it was in male BD6 rats aged 2 or 11 months. It appears that TS induces a more pronounced elevation of polynucleated PAM frequency in rats than in hamsters and mice. These data suggest that inhaled TS is genotoxic in alveolar macrophages in all exposed species of laboratory animals. An attempt was made to trace the possible clastogenic effect of a single i.p. administration of cyclophosphamide (CP, 15 mg/kg) in mice simultaneously in bone marrow and in PAM. A definite clastogenic effect in bone marrow 24 h and 48 h after CP injection and a total absence of changes in PAM from the lungs during the 15-day period after clastogen exposure were established. These data may support the hypothesis of local production of PAM in the lung from their proliferative precursor. The results provide evidence that PAM in laboratory animals are a sensitive and useful target for assessing harmful effects associated with environmental chemical factors that can be inhaled, including TS.

Modulation of diethylnitrosamine carcinogenesis in rat liver and oesophagus.

A series of 16 experiments, using a total of 2,000 BD6 rats, was designed in order to assess the ability of 8 individual agents or their combinations to modulate the liver and oesophageal carcinogenesis induced by multiple doses of diethylnitrosamine (DEN). Of the antioxidants tested, sodium selenite, ascorbic acid, and butylated hydroxytoluene generally exhibited protective effects on both types of tumors. In contrast, retinoic acid behaved as a promoter of DEN hepatocarcinogenesis, but this effect could be eliminated by its combination with either selenite or butylated hydroxytoluene. Caffeine and theophylline, when individually assayed, were devoid of significant protective effects, and the latter methylxanthine stimulated oesophageal tumorigenesis when administered after exposure to the carcinogen. Caffeine tended to decrease the multiplicity of liver tumors and potentiated the inhibitory effect of selenite in the liver. Irrespective of combination with caffeine, treatment with phenobarbital before each DEN injection tended to reduce the multiplicity of both liver and oesophageal tumors. On the other hand, the metabolic inhibitor diethyldithiocarbamate, given after each DEN injection, dramatically enhanced the incidence and multiplicity of oesophageal tumors. Thus, on the whole, modulation of DEN carcinogenesis varied depending on test agents, their combinations, dosages, treatment schedules, and target organ.

Modulation of the mutagenic activity of cigarette smoke, cigarette smoke condensate and benzo[a]pyrene in vitro and in vivo.

A series of naturally occurring compounds were tested for the ability to modulate the mutagenicity induced by cigarette smoke (CS), cigarette smoke condensate (CSC) and benzo[a]pyrene (BP) in the Salmonella/microsome mutagenicity assay and the micronucleus test in mouse bone marrow. Sodium selenite, retinol acetate and alpha-tocopherol significantly decreased the mutagenic activity of CS in Salmonella typhimurium TA98. Ascorbic acid, reduced glutathione (GSH), cysteine, caffeine, theophylline, cobalt chloride, folic acid, adenine, adenosine, guanosine, cytidine and cytosine were conversely devoid of any significant effect. Sodium selenite slightly decreased the mutagenic activity of CSC in the same bacterial strain, while caffeine was ineffective and ascorbic acid potentiated its mutagenicity. Ascorbic acid inhibited the mutagenic activity of BP in S. typhimurium TA98, but not in TA100. Retinol acetate diminished the number of BP-induced his+ revertants in TA98 but only at the highest concentrations used, whereas alpha-tocopherol, GSH, cysteine, sodium selenite and caffeine had no effect. Selenite and GSH, which were ineffective when applied individually, inhibited in a dose-dependent manner the BP-induced mutagenesis in S. typhimurium TA98 when simultaneously added to the top agar. All other combinations tested, including selenite plus either GSH, cysteine or caffeine towards CS or CSC, or selenite plus cysteine, or selenite plus retinol acetate and alpha-tocopherol towards BP, failed to produce interactive effects. Sodium selenite and caffeine, given either alone or in combination in drinking water, did not influence the clastogenesis induced in mouse bone marrow by a single treatment with CS or BP. Ascorbic acid was also ineffective towards CS clastogenicity but significantly decreased the number of micronucleated polychromatic erythrocytes induced by BP.(ABSTRACT TRUNCATED AT 250 WORDS)

Coclastogenicity of ethanol with cigarette smoke in rat erythroblasts and anticlastogenicity in alveolar macrophages.

A series of experiments was carried out to assess cytotoxic and cytogenetic effects in bone marrow polychromatic erythrocytes (PCE) and pulmonary alveolar macrophages (PAM) resulting from individual or combined exposure of male BD6 rats to ethanol, cigarette smoke and Aroclor 1254. Addition of 5% ethanol to drinking water did not affect the micronucleus frequency but consistently enhanced the proportion of polynucleated PAM. Moreover, the higher concentration used (10%) was cytotoxic in the bone marrow. Whole-body exposure to cigarette smoke elevated the micronucleus frequency in both PCE (4.0-4.4-fold) and PAM (2.0-3.6-fold) and enhanced the frequency of polynucleated PAM. After 3 weeks of combined exposure, ethanol produced contrasting effects in smoke-exposed rats, i.e. an increase of micronuclei in PCE and a decrease in PAM. An i.p. injection of Aroclor 1254 was per se devoid of any influence on the monitored parameters but tended to attenuate the cytotoxic and cytogenetic changes produced by cigarette smoke or ethanol in both types of cell.

Diminished genotoxicity of mitomycin C and farmorubicin included in polybutylcyanoacrylate nanoparticles.

The genotoxic effects of mitomycin C (MMC) and farmorubicin (FR) in a free form and included in polybutylcyanoacrylate nanoparticles (PBCN) were studied employing the Salmonella/microsome mutagenicity assay and the micronucleus test in mouse bone marrow as well as in mouse fetal liver. The data obtained clearly indicated that MMC (0.25-2.00 micrograms/plate) was a strong mutagen in S. typhimurium TA102, while the same concentrations of this compound in PBCN were ineffective in inducing his+ revertant mutations in bacterial cells. A similar total suppression of mutagenic activity of FR (1.0-20.0 micrograms/plate) was registered in S. typhimurium TA98 when the drug was included in PBCN. Furthermore, the incorporation of MMC (2.0 or 4.0 mg/kg, i.p.) into PBCN strongly diminished or even abolished its clastogenic activity in the bone marrow of virgin and pregnant mice as well as in mouse fetal liver, respectively. In addition, a lack of genotoxic effect of PBCN only was also established. The toxic activity of MMC in mouse bone marrow was significantly reduced or completely abolished after its inclusion in PBCN. A conclusion might be drawn that the genotoxic activity of some antitumor drugs might be markedly diminished or even abolished after their incorporation in PBCN.

Effect of metabolic inhibitors, methylxanthines, antioxidants, alkali metals, and corn oil on 1,2-dimethylhydrazine carcinogenicity in rats.

The effect of the oral administration of 10 compounds on 1,2-dimethylhydrazine (DMH) carcinogenesis was investigated in 180 male Wistar rats and 510 male BD6 rats. DMH, administered s.c. once per week for 20 consecutive weeks (20 mg/kg body wt/dose), produced intestinal (mainly colon) tumors of various histological type in 100% of both rat strains and, in addition, caused Zymbal gland carcinomas in 79.7% of Wistar rats. Pretreatment with disulfiram (DSF, 500 mg/kg), a known inhibitor of DMH metabolism, totally prevented intestinal and Zymbal gland tumors in Wistar rats. When DSF treatment started after the first DMH injection, the protective effect was not total, the incidence and multiplicity of both types of tumors being comparable to those observed following a single injection of the carcinogen alone. This confirms the involvement of DSF in the initiation stage only of DMH carcinogenesis. A complete prevention of intestinal tumors in BD6 rats was also produced not only by the DSF metabolite carbon disulfide (250 mg/kg) but also by the hepatotoxic agent carbon tetrachloride (1.5 ml/kg), which suggests that the block of DMH metabolism in rat liver is not an exclusive property of thiono-sulfur compounds. Butylated hydroxytoluene (BHT) decreased the multiplicity of intestinal tumors, but not to a significant extent. BHT and the aforementioned metabolic inhibitors were administered by gavage in corn oil, which per se did not significantly decrease intestinal or Zymbal gland tumors. All remaining modulators were administered with drinking water. Two additional antioxidants triggered opposite effects on the multiplicity of intestinal tumors. In fact, sodium selenite (10 mg/l) significantly decreased the number of tumors, whereas ascorbic acid (10 g/l), irrespective of its combination with CaCl2, produced a marked enhancement. The alkali metal salts CaCl2 and KCl (both at 5 g/l) as well as the methylxanthines caffeine and theophylline (both at 600 mg/l) were devoid of significant effects. Neither treatment with DMH alone nor its association with test modulators was accompanied by significant changes in body weight gain or survival of animals. On the whole, depending on the mechanisms involved, the comparative study of test compounds led to a broad array of effects on DMH carcinogenesis, ranging from complete inhibition to significant enhancement. The resulting picture can be visualized at a glance in Figure 1 of this article.

Clastogenic activity of urethane in mice.

Single intraperitoneal (i.p.) treatment of male and female BDF1 (C57B1 x DBA2) mice with urethane (0.5 or 1.0 g/kg) caused a significant increase in micronucleated polychromatic erythrocytes (MNPCE) in bone marrow after 24 h. The clastogenic effect observed was dose-, sex- and age-dependent, the male and younger (6-8 weeks old) animals being more susceptible than the female and older (6 months of age) mice. 3-week oral treatment of female Balb/c mice with urethane (3 g/l added to the drinking water) caused an up to 4-fold increase in the number of micronucleated normochromatic erythrocytes (MNNCE) in mouse peripheral blood. In a month after the carcinogen treatment was stopped, the number of MNNCE dropped to the control values. In addition, a single i.p. treatment of pregnant BDF1 mice on day 17 of gestation with urethane (1.0 g/kg) caused a 514.3% (p less than 0.001) elevation of MNPCE in mouse fetal liver after 24 h as well as a 154.4% (p less than 0.05) increase in MNPCE frequency in the fetal peripheral blood. At this time point, the clastogenic response in mouse fetal liver erythroblasts was less pronounced than that detected in the maternal bone marrow cells. Urethane is a strong clastogen in mice when administered either intraperitoneally or orally and the micronucleus test applied to adult and fetal erythroblasts is a convenient method of choice for studying the acute and subchronic clastogenicity of this carcinogen, its transplacental effects as well as the influence of modifying factors on these processes.

Potentiation by caffeine of the frequencies of micronuclei induced by mitomycin C and cyclophosphamide in young mice.

Employing the micronucleus test in mouse bone marrow and in fetal mouse liver, the possible clastogenicity of caffeine as well as its influence on MMC- and CP-induced micronucleus levels were studied. The treatment of male and female C57Bl or BDF1 (C57Bl x DBA2) mice with caffeine (1 or 3 x 50 mg/kg and 100 mg/kg, s.c.) had no clastogenic effect in mouse bone marrow or in the fetal livers and maternal bone marrow when pregnant mice were injected with caffeine on day 16-17 of gestation. MMC (2.0 mg/kg, i.p.) increased up to 10-30-fold the number of MNPCEs in bone marrow compared to a 3-7 fold elevation of MNPCEs in fetal liver. A similar effect was also established in pregnant mice treated with CP (30 mg/kg, i.p.). No significant sex differences in spontaneous and MMC- or CP-induced MNPCEs levels were established in C57Bl and BDF1 mice. However, a significantly higher spontaneous rate of MNPCEs as well as a better-expressed responsiveness to the clastogenic activity of MMC and CP were established in C57Bl compared to BDF1 mice. The pregnancy had no effect on MMC- or CP-induced clastogenicity although a tendency to a decreased sensitivity to the damaging activity of MMC seemed to be detected in pregnant C57Bl mice compared to virgin female animals. The combined treatment of mice with caffeine (3 x 100 mg/kg) and MMC or CP caused an up to 45-49% potentiation of clastogenesis in the bone marrow of male, female and pregnant female C57Bl and BDF1 mice but not in fetal mouse livers.

Modulation of genotoxic activity of tobacco smoke.

Tobacco smoke (TS) caused a three- to nine-fold increase in the frequency of his+ revertants in Salmonella typhimurium TA98 but not in TA97a, TA100 or TA102. Activation by a post-mitochondrial fraction obtained from the liver of rats pretreated with Aroclor-1254 or methylcholanthrene was required; fractions from phenobarbital-pretreated or untreated rats had no effect. Vitamins A and E, but not ascorbic acid, inhibited the TS-induced mutagenesis by up to 63%, whereas glutathione and cysteine increased it slightly. Na2SeO3, but neither CoCl2 nor caffeine, inhibited the mutagenic effect of TS by 46-56%. In Chinese hamster ovary cells, both Na2SeO3 and caffeine strongly potentiated the number of chromosomal aberrations induced by TS, while theophilline slightly reduced its clastogenic effect. Treatment of mice with TS for 60 min/day increased the frequency of micronuclei in polychromatic erythrocytes in bone marrow and in fetal liver and the number of NCE micronuclei in peripheral blood by four to five fold. Simultaneous treatment of mice with TS and Na2SeO3 reduced the clastogenic effect of TS. Ascorbic acid had no effect on clastogenicity but reduced toxicity as measured by body weight loss. Both Na2SeO3 and ascorbic acid suppressed the induction of TS-induced hyperplastic and metaplastic changes in bronchial mucosa but had no effect on the number of urethane-induced lung adenomas. Vitamins A and E and ascorbic acid may have a protective effect against the toxic and genotoxic activities of TS.

[The testing for carcinogenicity of a polyethylene terephthalate vascular prosthesis]. / Testirovane na kantserogennost na polietilentereftalova sudova proteza.

A long-term for possible carcinogenicity was conducted by subcutaneous implantation of 1 cm 2 pieces of polyethylenetherophthalate vascular grafts (PET-VG) to 150 BDF1 mice and 120 Syrian golden hamsters of either sex. A false operation was carried out in the same number of animals of control groups. Implantation was performed in nine-week-old animals. The observation period was 73 weeks for mice and 82 weeks for hamsters. A total of 161 tumors (81 in the experimental group) with 15 localizations were observed in mice and 43 tumors (20 in the experimental group) with 13 localizations--in hamsters. No pathological or statistical evidence of induction of tumors by PET-VG was found. It is concluded that under the conditions of this study PET-VG is not carcinogenic for BDF1 mice and Syrian golden hamsters of either sex.

Tobacco smoke-induced clastogenicity in mouse fetuses and in newborn mice.

The clastogenic activity of tobacco smoke (TS) was established in mouse fetuses employing the micronucleus test. A 1-h exposure of pregnant mice BDF1 (C57Bl x DBA2) to TS (600 cm3 TS in a 14-l glass chamber, 4 exposures of 15 min each with 1 min intervals during which a total air change was made) on day 16/17 caused a 2-3-fold increase in the number of micronucleated polychromatic erythrocytes (MN PCEs) in fetal liver as well as in the liver of newborn mice (1-5 h after birth). A similar, although slightly greater micronucleus response occurred in fetuses obtained from pregnant mice treated repeatedly with TS (60 min/day in total) starting on day 11 of gestation. The in vivo clastogenic activity of TS was also established by evaluating the MN PCE level in the peripheral blood of newborn mice (1-5 h after birth and during the first several days of life) treated transplacentally with TS during the last third of pregnancy. The young animals (1-3 weeks old) were more sensitive to the clastogenic activity of TS as compared to their 6-month-old mothers but no data were obtained showing a possible passing of the TS-contained clastogens from 'smoking' lactating mothers to their suckling offspring. The combined application of micronucleus test proved to be very convenient and useful when studying the different aspects of TS-induced genotoxicity.

The mutagenic and clastogenic activity of tobacco smoke.

Employing the Salmonella/microsome mutagenicity assay it was established that the mutagenic effect of tobacco smoke (TS) (240 cm3 in a 16-l glass chamber, at 1 min or 5 min exposure time) in S. typhimurium TA98 depended on the type of S9 mix used. Addition of S9 mix obtained from the liver of 3-methylcholanthrene- or Aroclor-1254-pretreated rats but not from the liver of phenobarbital-pretreated or untreated rats was required to demonstrate the mutagenic activity of TS. One might suggest that polycyclic aromatic hydrocarbons were involved in TS-induced mutagenesis in S. typhimurium TA98. In addition, treatment of BDF1 mice with TS (600 cm3 TS in a 14-l glass chamber, 2-6 exposures of 30 min each with a 1-min interval between them during which a total change of the air was made) caused an up to 3.5-fold increase of the number of micronucleated polychromatic erythrocytes (PCE) in mouse bone marrow detected 24 h after the TS exposure. Furthermore, a stable 2-5-fold elevation of the number of micronucleated normochromatic erythrocytes (NCE) was detected in the peripheral blood of mice treated daily (2 x 30 min) with TS, starting 48 h after the first TS exposure. The application of the micronucleus test in mouse peripheral blood, a more convenient and useful approach for detecting the chronic clastogenic activity of TS, allowed us to establish the cumulative genotoxic effect of TS in mice.

Investigation of the mutagenic activity of tobacco smoke.

The genotoxic effect of whole tobacco smoke was studied employing the Salmonella/microsome mutagenicity assay, the micronucleus test in mouse bone marrow and UDS in peripheral human lymphocytes. It was established that tobacco smoke (120-480 cm3 in a 16-1 glass chamber, at 1-10 min exposure time) induced a 3-9-fold increase of spontaneous his+ reversion mutation rate in S. typhimurium TA98, but not in strains TA97a, TA100 and TA102. Addition of S9 mix obtained from the liver of Aroclor 1254-treated rats was necessary to reveal the mutagenic activity of tobacco smoke. Treatment of BDF1 mice placed in a 14-1 glass chamber with tobacco smoke (600 cm3 smoke, 2 exposures of 30 min each, with a 1-min interval between them) caused a 2-fold dose-dependent elevation of the number of micronucleated PCE in bone marrow. No cumulative effect was detected when mice were treated with tobacco smoke during 2-28 consecutive days. The effect observed 24 h after tobacco-smoke exposure was abolished 48 h later. Tobacco smoke (180 or 360 cm3) passed through the culture medium (with or without S9 mix) of human peripheral lymphocytes (the cells were then incubated for 60 min at 37 degrees C) did not increase the spontaneous rate of UDS. Both the Salmonella/microsome mutagenicity assay employing S. typhimurium TA98 strain and the micronucleus test in mouse bone marrow might be useful in studying tobacco smoke-induced mutagenesis.

The effect of antioxidants on MNNG-induced stomach carcinogenesis in rats.

The effect of vitamins A, C and E, butylated hydroxytoluene (BHT) and glutathione (GSH) on gastric carcinogenesis induced by N-methyl-N-nitro-N-nitrosoguanidine (MNNG) was investigated. Male and female BD-VI rats 2-3 months old received a single oral application of MNNG dissolved in corn oil. The male rats were divided into four groups: Group-I: MNNG 250 mg/kg by intubation; Group-II: MNNG + vitamin C daily in the drinking water (400 mg/l); Group-III: MNNG + vitamin C (400 mg/l) + 100 g of milk broth (for each of 10 rats) containing vitamin A (40,000 IU), vitamin E (0.5 g) and BHT (0.1 g) three times a week. The treatment with antioxidants started 7 days before the MNNG administration and continued until the end of experiment. Group-IV rats received MNNG + oxyferriscorbone, i.p. as a single dose of 1.0 mg/kg, daily during the week before and the week after MNNG exposure and than 3 times a week till the end of the experiment. Female rats were divided into two groups: Group-I: MNNG 333 mg/kg by intubation; Group-II: MNNG + GSH orally at a dose of 100 mg/rat 1 h before and 5, 24, 48, and 72 h after MNNG intubation. The incidence of gastric tumors after 15 months of treatment was as follows: male rats, 82.4% in Group-I, 40.0% in Group-II, 40.7% in Group-III, and 50.0% in Group-IV; female rats; 72.7% in Group-I, and 36.0% in Group-II.

[Isoenzymes of melanosomal tyrosinase isolated from transplanted hamster melanomas]. / Izoénzimy melanosomal'noi tirozinazy, vydelennoi iz transplantiruemoi melanomy khomiaka.

The isoenzyme profiles of tyrosinase isolated from melanosomal and cytoplasmic fractions of hamster melanoma were investigated. The liberation of the enzyme from the melanosomal membranes was achieved by their treatment with Triton X-100 or alpha-chymotrypsin. The isoenzyme spectrum of tyrosinase liberated by use of Triton X-100 from melanosomes differing in the degree of melanization is identical to that of the cytoplasmic enzyme and gives three peaks on a densitogram. The liberation of melanosomal tyrosinase by alpha-chymotrypsin results in the appearance of an additional isoenzyme due to proteolytic degradation. The role of tyrosinase isoenzymes in the synthesis of melanin is discussed.

Aminoacids--precursors of melanin synthesis in hamster melanoma.

The role of L-tyrosine, L-phenylalanine, L-dihydroxyphenylalanine (dopa) and L-tryptophan in melanin biosynthesis in melanotic hamster melanoma IC-Sofia was investigated with the aid of 14C-aminoacids. Tyrosine and phenylalanine were found to be the main melanin precursors: about 64.5% of total melanin labeling was due to tyrosine incorporation and 27.4% to phenylalanine incorporation. Negligible proportions of melanin radioactivity (5.7% and 2.4%, respectively) resulted from dopa and tryptophan utilization in melanin synthesis. The involvement of each of the aminoacids under investigation in melanin synthesis in vivo is discussed.

Effect of theophyllin and isoproterenol on cAMP level in melanotic and amelanotic hamster melanomas.

The level of cAMP was investigated in the following three types of hamster malignant melanomas: amelanotic, depigmented and melanotic. The amelanotic and depigmented tumors were undifferentiated, with high proliferative activity, and lacked tyrosinase. The melanotic one was slow growing, highly differentiated, and expressed tyrosinase activity. Among the melanomas investigated, the higher concentration of cAMP was found in undifferentiated tumors. The single treatment of the tumor-bearing animals with theophyllin or isoproterenol did not change the cAMP level in melanotic tumors, but significantly enhanced the cAMP content in amelanotic and especially depigmented melanomas. Multiple theophyllin treatment of tumor-bearing animals caused elevation of cAMP content in all tumors, but this effect was accompanied by enhancement of tyrosinase activity only in melanotic melanoma.

The influence of selenium and caffeine on chemical carcinogenesis in rats, mutagenesis in bacteria, and unscheduled DNA synthesis in human lymphocytes.

The influence of sodium selenite (Na2SeO3) and caffeine on chemical carcinogenesis induced in rats by diethylnitrosamine (DEN), N-nitrosomorpholine (NM), andN-methyl-N-nitro-N-nitrosoguanidine (MNNG) was investigated. A dose-dependent inhibitory effect of Na2SeO3 (l-10 ppm) on hepatocarcinogenesis induced by DEN was demonstrated. Na2SeO3 also increased the latency period for stomach tumor formation in rats treated with MNNG. Combined treatment of rats with Na2SeO3 plus vitamin C added to the diet resulted in a slight inhibition of NM-induced liver carcinogenesis. Supplementation of diet with Na2SeO3 plus butylated hydroxytoluene, vitamin C, and vitamin E did not reveal any additive inhibitory effect compared to the inhibitory effect of Na2SeO3 given alone. Caffeine (600 rag/L) reduced the number of liver tumors induced in rats by DEN. Preliminary experiments have indicated that combined treatment of rats with selenium and caffeine could result in more effective inhibition of DEN-induced liver carcinogenesis.Further experiments are being conducted to study the influence of selenium and caffeine on mutagenic activity of 1-methyl-l-nitrosourea (MNU) inSalmonella typhimurium TA 1535. The pretreatment of bacteria cells with Na2SeO3 (3-10 p.g/mL) increased the mutagenic response of bacteria to MNU. A synergistic stimulation of mutagenic activity of MNU was observed in bacteria pretreated simultaneously with Na2SeO3 and caffeine.In addition the influence of Na2SeO3 on UDS induced by DEN in human lymphocytes was investigated. The trace element inhibited the UDS up to 82%.The possible role of potentiation by NazSeO3 of the cell killing effect of DEN in inhibition of liver carcinogenesis was discussed.