[IMMUNOCYTOCHEMICAL ANALYSIS OF THE DISTURBANCES IN THE STRUCTURE OF SYNAPTONEMAL COMPLEXES IN SPERMATOCYTE NUCLEI IN MICE UNDER EXPOSURE TO ROCKET FUEL COMPONENT].

There was performed an assessment of genotoxic effects of rocket fuel component--unsymmetrical dimethylhydrazine (UDMH, heptyl)--on forming germ cells of male mice. Immunocytochemically there was studied the structure of meiotic nuclei at different times after the intraperitoneal administration of UDMH to male mice. There were revealed following types of disturbances of the structure of synaptonemal complexes (SCs) of meiotic chromosomes: single and multiple fragments of SCs associations of autosomes with a sex bivalent, atypical structure of the SCs with a frequency higher than the reference level. In addition, there were found the premature desinapsis of sex bivalents, the disorder offormation of the genital corpuscle and ring SCs. Established disorders in SCs of spermatocytes, analyzed at 38th day after the 10-days intoxication of animal by the component of rocket fuel, attest to the risk of permanent persistence of chromosomal abnormalities occurring in the pool of stem cells.

Calcium and α-tocopherol suppress cured-meat promotion of chemically induced colon carcinogenesis in rats and reduce associated biomarkers in human volunteers.

BACKGROUND: Processed meat intake has been associated with increased colorectal cancer risk. We have shown that cured meat promotes carcinogen-induced preneoplastic lesions and increases specific biomarkers in the colon of rats. OBJECTIVES: We investigated whether cured meat modulates biomarkers of cancer risk in human volunteers and whether specific agents can suppress cured meat-induced preneoplastic lesions in rats and associated biomarkers in rats and humans. DESIGN: Six additives (calcium carbonate, inulin, rutin, carnosol, α-tocopherol, and trisodium pyrophosphate) were added to cured meat given to groups of rats for 14 d, and fecal biomarkers were measured. On the basis of these results, calcium and tocopherol were kept for the following additional experiments: cured meat, with or without calcium or tocopherol, was given to dimethylhydrazine-initiated rats (47% meat diet for 100 d) and to human volunteers in a crossover study (180 g/d for 4 d). Rat colons were scored for mucin-depleted foci, putative precancer lesions. Biomarkers of nitrosation, lipoperoxidation, and cytotoxicity were measured in the urine and feces of rats and volunteers. RESULTS: Cured meat increased nitroso compounds and lipoperoxidation in human stools (both P < 0.05). Calcium normalized both biomarkers in rats and human feces, whereas tocopherol only decreased nitro compounds in rats and lipoperoxidation in feces of volunteers (all P < 0.05). Last, calcium and tocopherol reduced the number of mucin-depleted foci per colon in rats compared with nonsupplemented cured meat (P = 0.01). CONCLUSION: Data suggest that the addition of calcium carbonate to the diet or α-tocopherol to cured meat may reduce colorectal cancer risk associated with cured-meat intake. This trial was registered at clinicaltrials.gov as NCT00994526.

Ras signaling pathway in the chemopreventive action of different ratios of fish oil and corn oil in experimentally induced colon carcinogenesis.

Dietary factors play a significant role in colon cancer. The essential polyunsaturated fatty acids (PUFAs), n-3 PUFAs, and n-6 PUFAs exert inverse effect on cancer. This study was designed to understand the mechanism of chemopreventive action of different ratios of fish oil (FO) and corn oil (CO) in colon carcinoma. Wistar rats were divided into 3 groups: Group 1 received purified diet whereas Groups 2 and 3 received modified diet with FO:CO (1:1) and FO:CO (2.5:1), respectively. The groups were further subdivided into controls receiving ethylenediamine-tetra acetic-acid and treated groups received dimethylhydrazine-dihydrochloride (DMH)/wk for 4 wk. Animals sacrificed 48 h after last injection constituted initiation phase and that sacrificed after 16 wk constituted post-initiation phase. Differential effect of different ratios of FO and CO was analyzed in isolated colonocytes. In both phases, DMH treatment showed an increase in pan Ras, Raf, MEK1/2, extracellular signal regulated kinase (Erk)1/2, and c-fos levels. Akt levels were increased in post-initiation phase only. Treatment with FO + CO (1:1) + DMH decreased pan Ras, MEK1/2 and Erk1/2 levels in post-initiation phase whereas Raf and c-fos were decreased in both phases. Treatment with FO + CO (2.5:1) + DMH decreased Ras, Raf, MEK1/2, Erk1/2, and c-fos levels in both phases. Akt was decreased in post-initiation phase only. The chemo-preventive action of FO and CO may be mediated by time- and dose-dependent effect.

Suppression of colonic aberrant crypt foci by soy isoflavones is dose-independent in dimethylhydrazine-treated rats.

The potential of soy isoflavones (SIs) to reduce colon cancer has been investigated in animal models. These studies have found that outcomes are variable and depend on SI dose. The present study investigated dose-response effects of SIs on colon carcinogenesis in a chemically induced rat cancer model. Sprague-Dawley male rats were injected with 1,2-dimethylhydrazine (DMH) and were provided experimental diets that contained 0, 10, 50, 150, or 500 mg of SI aglycones/kg of diet for 12 weeks. Plasma concentrations of genistein, daidzein, and equol were determined using time-resolved fluoroimmunoassay. Plasma concentrations of these SIs tended to increase in a dose-dependent manner in DMH-treated rats. The numbers of aberrant crypt foci (ACF) and the expression of cyclooxygenase-2 (COX-2) proteins of colons were significantly decreased in the SI-fed groups compared with the control group; however, suppression was not dose-dependent. Furthermore, there were no significant correlations between plasma SI concentrations and ACF or COX-2 expression. Increased SI intake and increased plasma levels of SIs and metabolites were not associated with tissue levels of lipid peroxidation. We conclude that dietary supplementation of SIs suppresses DMH-induced ACF formation and COX-2 expression in a dose-independent manner.

[Effects of asymmetric dimethyl hydrazine on physical and chemical properties of blood plasma if experimental animals].

Asymmetric dimethyl hydrazine (ADH), a highly toxic propellant component, is known to cause metabolic disturbances in humans and laboratory animals. Focus of this investigation was placed on crystal-forming properties of blood serum, and types of plasma protein destructions in ADH-intoxicated rats. Object of investigation was blood plasma collected from rats exposed to 100 mg/kg of per oral ADH total (5 mg/kg per a day, 5 days a wk, 4 wks.). The crystal-forming ability of plasma was assessed with the teziographic technique. Severity of proteins destruction was determined by the content of carbonyl products of oxidative protein modification in plasma. Chronic exposure to low ADH doses caused oxidative destruction of proteins in blood plasma and, consequently, significant degradation of the plasma ability to form crystals reflected in an altered zonal phase structure and phase bursting on teziograms. Symmetry gave way to pathological structures, e.g. petals and loops. Amorphous areas grew in size, bursting lines thinned out. Concretions became less numerous. These data suggest a close relationship between the impairments in physical and chemical properties of blood plasma of experimental animals by highly toxic asymmetric dimethyl hydrazine.

Effect of defunctionalization on colon carcinogenesis in the rat.

OBJECTIVE: to examine the effect of fecal absence on experimental colon carcinogenesis in both male and female rats. MATERIAL AND METHODS: a total of 138 10-week-old Sprague-Dawley, male and female rats were divided into five groups: A) 20 rats, no treatment; B) 26 rats, colonic defunctionalization; C) 30 rats, 18 weekly doses of dimethylhydrazine (DMH), 21 mg/kg body weight each, from the beginning of the study; D) 20 rats, ethylen-diamine-tetraacetic acid for 18 weeks; and E) 42 rats, same surgical procedure as rats in group B plus DMH injections at the same doses as rats in group C. Animals were sacrificed after 25-27 weeks. Number of tumors, their location, and pathological findings were all compared between groups. RESULTS: no tumors developed in the dimethylhydrazine-free groups. No differences were obtained either in number of tumors or tumors per rat for group C as compared to group E. Fecal absence was associated with smaller-sized tumors (p = 0.007), greater numbers of non-mucinous tumors (p = 0.00009), better differentiation (p = 0.0054), and lesser penetration into the wall (p = 0.015) for group E as compared to group C. In the dimethylhydrazine group, fecal absence altered the number of tumors developing in males as compared to female rats (p = 0.025). Moreover, this fecal absence showed no inhibitory effect on right colonic tumors (p = 0.0065). CONCLUSIONS: fecal absence alters the DMH-carcinogenic pattern in the defunctionalized colon when using an experimental model in both male and female rats.

A proteomics approach to identify changes in protein profiles in pre-cancerous colon.

The development of colon cancer is characterised by alterations in multiple genetic and epigenetic pathways in colon tissue leading ultimately to deregulation of colon epithelial cells. Early detection is an important factor in decreasing colon cancer deaths. Proteomic techniques were used to identify potential early markers in colon tissue exhibiting pre-cancerous activity that may characterise pathological changes in a chemically induced colon cancer rat model. Protein profiles were assessed in soluble and insoluble fractions prepared from distal colon of rats treated with the colonotropic carcinogen, dimethylhydrazine. Alterations in protein profiles were associated with the presence of aberrant crypt foci, hyperplasia and dysplasia, microanatomical changes, and metabolic changes in rat colon. These changes may have a potential role in the identification of pre-pathological features preceding colon tumorigenesis.

Dietary folate and selenium affect dimethylhydrazine-induced aberrant crypt formation, global DNA methylation and one-carbon metabolism in rats.

Several observations suggest a role for DNA methylation in cancer pathogenesis. Although both selenium and folate deficiency have been shown to cause global DNA hypomethylation and increased cancer susceptibility, the nutrients have different effects on one-carbon metabolism. Thus, the purpose of this study was to investigate the interactive effects of dietary selenium and folate. Weanling, Fischer-344 rats (n = 23/diet) were fed diets containing 0 or 2.0 mg selenium (as selenite)/kg and 0 or 2.0 mg folate/kg in a 2 x 2 factorial design. After 3 and 4 wk of a 12-wk experiment, 19 rats/diet were injected intraperitoneally with dimethylhydrazine (DMH, 25 mg/kg) and 4 rats/diet were administered saline. Selenium deficiency decreased (P < 0.05) colonic DNA methylation and the activities of liver DNA methyltransferase and betaine homocysteine methyltransferase and increased plasma glutathione concentrations. Folate deficiency increased (P < 0.05) the number of aberrant crypts per aberrant crypt foci, the concentration of colonic S-adenosylhomocysteine and the activity of liver cystathionine synthase. Selenium and folate interacted (P < 0.0001) to influence one-carbon metabolism and cancer susceptibility such that the number of aberrant crypts and the concentrations of plasma homocysteine and liver S-adenosylhomocysteine were the highest and the concentrations of plasma folate and liver S-adenosylmethionine and the activity of liver methionine synthase were the lowest in rats fed folate-deficient diets and supplemental selenium. These results suggest that selenium deprivation ameliorates some of the effects of folate deficiency, probably by shunting the buildup of homocysteine (as a result of folate deficiency) to glutathione.

Frequency of proliferation, apoptosis, and their ratio during rat colon carcinogenesis and their characteristic pattern in the dimethylhydrazine-induced colon adenoma and carcinoma.

The imbalance between the cell proliferation and cell loss plays a crucial role in the carcinogenesis and tumor progression. However, the direction of these changes is still the matter of discussion. Thus, the aim of this study was to evaluate the proliferative activity, apoptotic activity, and proliferation/apoptosis ratio (P/A) assessed every 6 weeks in the colonic epithelium during 21 weeks of dimethylhydrazine (DMH) treatment in male Wistar rats. Moreover, it is necessary to answer the question whether these analyzed parameters correlate with the grade of differentiation or dysplasia of the induced tumors. It was found that DMH administration enhanced the proliferation in week 12 and 18 when compared with week 6. The proliferation in the control group did not change during the study. Up to week 12 of the experiment, there were no statistically significant differences between proliferative activity in the control and DMH-treated groups. In week 18, the proliferation in DMH-treated group was higher than in the control group. At all time points of the study, the apoptotic activity in the DMH-treated groups was significantly higher than in controls and in both groups, they dropped during the study. In the control group, apoptotic activity decreased in week 18 and was lower in comparison to that in week 6 and 12. In the group treated with DMH, apoptosis dropped at week 12 and was lower than in week 6. The P/A ratio did not change during the study in the control group, but increased in the DMH-treated group. After 21 weeks of DMH administration, 28 cases of colon adenocarcinoma and nine cases of colon adenoma were obtained and classified according to the WHO classification (1989) for human colon tumors. The adenocarcinomas were divided into four groups: well, moderately, poorly differentiated, and signet-ring cell carcinoma. The colon adenomas were divided into three groups: adenoma with mild, moderate, and severe grade of dysplasia. The proliferative activity in signet-ring cell carcinoma was significantly smaller than in well, moderately, and poorly differentiated adenocarcinoma and apoptotic activity was smaller than in well-differentiated adenocarcinoma. A weak (statistically nonsignificant) negative correlation was also observed between the proliferative and apoptotic activity in adenocarcinoma or adenoma and their grade of dedifferentiation or dysplasia, respectively.

Organ-specific genotoxicity of the potent rodent colon carcinogen 1,2-dimethylhydrazine and three hydrazine derivatives: difference between intraperitoneal and oral administration.

We used a modification of the alkaline single cell gel electrophoresis (SCG) (Comet) assay to test the in vivo genotoxicity of four hydrazine derivatives--1,2-dimethylhydrazine (SDMH), 1,1-dimethylhydrazine (UDMH), hydrazine (HZ), and procarbazine (PCZ)--in mouse liver, lung, kidney, brain, and bone marrow, and in the mucosa of stomach, colon, and bladder. Mice were sacrificed 3 and 24 h after intra-peritoneal (i.p.) and oral (p.o.) administration. SDMH at 20 mg/kg i.p. yielded statistically significant DNA damage in all tested organs except for lung. In the gastrointestinal tract, SDMH was genotoxic in the stomach and the colon after i.p. treatment but only in the colon after 20 and 30 mg/kg p.o. treatment. UDMH at 50 mg/kg i.p. yielded DNA damage in the liver and lung at 3 h. PCZ at 200 mg/kg i.p. caused DNA damage in the liver, kidney, lung, brain, and bone marrow. UDMH and PCZ were positive in the stomach and colon p.o. but not by i.p. treatment. HZ at 100 mg/kg yielded DNA damage in the stomach, liver, and lung when given i.p. and in the brain when p.o. Thus, the administration route is important when evaluating organ-specific genotoxicity in multiple organs.

Inhibition of dimethylhydrazine-induced aberrant crypt foci and induction of apoptosis in rat colon following oral administration of the glucosinolate sinigrin.

Glucosinolates are sulphur compounds that occur as glycosides in brassica vegetables. In response to tissue disruption they are degraded by thioglucosidase, releasing a range of highly reactive breakdown products, including the isothiocyanates, which we have previously shown to be selectively cytotoxic to undifferentiated colorectal tumour cells (HT29). In the present study we explored the effect of sinigrin on the intestinal mucosa of rats previously treated with dimethylhydrazine (DMH). In the first experiment, a semisynthetic feed containing sinigrin (400 microg/g diet) was provided 6 h after the second of two injections of DMH. The level of apoptosis was measured by morphological assessment of intact microdissected crypts obtained at 18, 24, 38, 48 and 72 h after injection, and compared with control groups given DMH only, or a sham-injection. Higher numbers of apoptotic nuclei were present in colonic tissue from both groups of DMH-treated rats compared with the controls, and the level was significantly higher in DMH-treated rats fed sinigrin compared with those given DMH only (P < 0.02). In a second experiment, rats were given sinigrin (400 microg/g diet) 22 h after the second of two injections of DMH; the level of apoptosis was measured after 48 h and the numbers of aberrant crypt foci (ACF) were measured after 42 days. The level of apoptosis was significantly higher in DMH-treated rats given sinigrin compared with controls (P < 0.05), and the numbers of ACF were significantly lower in sinigrin-treated rats (P < 0.001). There was no statistically significant induction of apoptosis in animals fed sinigrin alone. Sinigrin administered after DMH suppresses induction of ACF. This may be due to increased apoptotic deletion of damaged stem cells in the crypts of animals fed sinigrin.

Influência do processo de reparaçäo de ferida cutânea na carcinogênese química experimental do cólon no rato / Influence of wound healing on the experimental chemical carcinogenesis of the colon of mouse

O crescimento neoplásico pode ser estimulado por um trauma cirúrgico e pelo processo reparativo subseqüente. A retirada parcial de uma neoplasia, pode favorecer a proliferaçäo de células tumorais remanescentes. Da mesma forma, a remoçäo de um tumor primário, a cirurgia simulada ou a amputaçäo de um membro sadio influenciam positivamente o crescimento de diferentes tipos de neoplasias. No presente estudo, avaliamos a influência do processo reparativo cutâneo sobre as lesöes pré-neoplásicas induzidas pela 1,2 Dimetilhidrazina (DMH) no cólon de ratos Wistar. Foram utilizados cinquenta animais, divididos em seis grupos. Quatro grupos, constituídos por dez animais cada, receberam injeçöes subcutâneas de DMH (20mg/Kg/semana), durante oito semanas. Dois grupos controles, constituídos por cinco animais cada, receberam injeçöes de soluçäo salina por igual período. Na 9ª semana dois grupos de animais, tratados com DMH e dois controles, sofreram intervençäo cirúrgica para retirada de retalho cutâneo do flanco direito, medindo 16 cm². Os animais foram submetidos à eutanásia na 12ª e 20ª semanas do experimento. O número de focos de lesöes pré-neoplásicas, assim como o número e a área das neoplasias bem ou moderadamente diferenciadas, näo foram influenciadas pelo processo reparativo cutâneo, nos momentos estudados. No entanto, o número e o tamanho dos tumores indiferenciados foi maior, na 12ª semana, no grupo submetido a retirada do retalho cutâneo. Estes resultados demonstram que o processo reparativo cutâneo influenciou positivamente o crescimento de neoplasias colônicas indiferenciadas, induzidas pela 1,2 Dimetilhidrazina. O estímulo induzido foi transitório, sendo detectado no período correspondente a fase de reparaçäo da ferida cutânea.

Dose-response relationship for rat liver DNA damage caused by 1,2-dimethylhydrazine.

An experimental approach was taken to the question of dose-response curves for chemical carcinogenesis, using DNA damage as a biomarker. Female rats were give 13 different doses of 1,2-dimethylhydrazine (from 1.4 to 135,000 micrograms/kg) and the subsequent hepatic DNA damage was determined by the alkaline elution technique. DMH doses below 450 micrograms/kg did not significantly damage DNA; all DMH doses of 1000 micrograms/kg or higher damaged rat hepatic DNA (P < 0.05). In this study the x values (dose) ranged over five orders of magnitude and the y values (DNA damage) ranged 30-fold. Ten different regression models (linear, quadratic, cubic, power, and six nonlinear transition models) were compared in their ability to fit the experimental data. With respect to log transformed dose, the six nonlinear transition equations fit the data considerably better than the four power type of equations. A sigmoid model fit to the log transformed dose of 1,2-dimethylhydrazine had an r2 of 0.9979, a degree of freedom adjusted r2 of 0.9969, a F-statistic of 1,457, and a fit standard error of 0.50. With respect to untransformed dose, only three equations (sigmoid, cascade and gaussian cumulative) could creditably fit the DMH data. The experimental results are interpreted with respect to hormesis, use of log transformed dose, sigmoid dose-response models, thresholds of biological response and cancer risk assessment.

Selective induction of micronuclei in the rat/mouse colon and liver by 1,2-dimethylhydrazine: a seven-tissue comparative study.

1,2-Dimethylhydrazine (DMH) was administered to both genders of mice and rats by oral gavage for 3 days. Twenty-four hours later, an assessment of the incidence of micronucleated cells was made in the bone marrow and sections of the gastrointestinal tract. An increase in micronucleated cells was observed in the colon of both genders of both species of rodent. Negative responses were observed in the forestomach, stomach, duodenum, intestine of both species. The bone marrow micronucleus assays were essentially negative, but the absence of a precise definition of the MTD precludes a definitive conclusion from being drawn. These results are consistent with the selective carcinogenicity of DMH to the colon of the rodent GI-tract. DMH is also known to be carcinogenic to rat and mouse liver and, although it is known to induce micronuclei in the hepatocytes of rats, no such data exist for the mouse. Consequently, mice were administered DMH on 13 successive days, followed by 2/3 partial hepatectomy and assessment of micronucleated hepatocytes. A strong positive liver micronucleus assay response was observed. Thus, DMH selectively induces micronuclei in the colon and liver of rats and mice, consistent with its carcinogenicity to these two tissues. No qualitative differences between the genders was observed in any of the assays. These results indicate that the assessment of genetic toxicity in rodents should not rely solely on assays made in bone marrow.

A comparative study of hepatic and colonic metabolic enzymes in inbred mouse lines before and after treatment with the colon carcinogen, 1,2-dimethylhydrazine.

1,2-Dimethylhydrazine (DMH) is an organotropic colon carcinogen that undergoes metabolic activation to DNA-reactive metabolites. Twenty hours after parenteral treatment of AKR/J (colon tumor resistant) and SWR/J (susceptible) mice with DMH.2HCl (70 mg/kg), functional levels of Cyp1a1 and Cyp2e1 were examined by measuring O-deethylation of ethoxyresorufin (EROD) and hydroxylation of p-nitrophenol, respectively. In control animals, SWR/J mice exhibited higher hepatic EROD activity (1.4-fold) when compared with AKR/J mice. In carcinogen-treated animals, EROD activity was decreased 20-30% in both mouse lines. Hepatic p-nitrophenol hydroxylase activity, similar in control animals of both strains, was reduced comparably (45-50% of control) after DMH administration. In liver, a decrease in immunoreactive Cyp2e1 protein paralleled the decline in enzyme activity, whereas in the colon, no significant treatment-related differences were detected in either strain. In liver and colon cytosols, alcohol dehydrogenase activity was not significantly different in either mouse line, both in control and DMH-treated animals. Glutathione levels were elevated (1.7-fold) in livers of AKR/J mice after DMH administration. Total glutathione-S-transferase (GST) activity was significantly increased (1.8-fold) in the colons of SWR/J mice and in the livers (1.4-fold) of AKR/J mice. Furthermore, the GST isoform, GST-Yp, was reduced 40% in the SWR/J colon. These data demonstrate the importance of metabolic capacity as a factor in conferring differential tumor susceptibility in a murine cancer model to the indirect-acting colon carcinogen, DMH.

Lipid peroxidation and activities of antioxidant enzymes in iron deficiency and effect of carcinogen feeding.

Iron deficiency has been implicated in increasing the risk of GI tract cancers in humans. Among various mechanisms of carcinogenesis, oxidative damage to DNA is well known and, hence, the present experimental study was undertaken to investigate lipid peroxidation and activities of different antioxidant enzymes in iron deficiency to explain the higher risk of tumorigenesis. Two groups of male weanling Fischer rats maintained on iron sufficient (C) or iron deficient (D) diets for a period of 32 weeks were subdivided, from 3 weeks onwards, into two subgroups each. The carcinogen, dimethyl hydrazine was fed at a dose of 30 mg/kg/week IG for a period of 9 weeks to groups that were designated as (C+) and (D+). The other two subgroups (C-) and (D-) served as controls. After the experimental period, hepatic assays for lipid peroxidation (MDA production) and activities of various antioxidant enzymes were carried out. The results showed that MDA production was elevated by 50% and activity of superoxide dismutase significantly depressed in carcinogen-fed, iron-deficient group (D+) by 28% compared to deficient (D-) group. There was an increase in hepatic selenium-dependent glutathione peroxidase activity in iron-deficient and iron-deficient, carcinogen-treated groups to the extent of 57 and 59%, respectively, as compared to controls; however, induction of enzyme in response to carcinogen feeding, observed in the control group, was not evident in iron deficiency. Liver catalase was not altered between control and deficient groups. These results suggest that prolonged iron deficiency superimposed with carcinogen ingestion may render the host susceptible to a greater risk of tumorigenesis through oxidative stress.

Lactobacillus- and bifidobacterium-mediated antigenotoxicity in the colon of rats.

Lactic acid bacteria (LAB) are proposed to have several beneficial effects, including the inactivation of carcinogens. We have studied the potential of Lactobacillus acidophilus (from a commercially available yogurt), Lactobacillus gasseri (P79), Lactobacillus confusus (DSM20196), Streptococcus thermophilus (NCIM 50083), Bifidobacterium breve and Bifidobacterium longum (from human infant stool) to prevent the induction of DNA damage by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG, 7.5 mg/kg body wt) in colon cells of the rat. Using the new technique of single cell microgel electrophoresis, all investigated strains were antigenotoxic toward MNNG after a single dose of 10(10) viable cells/kg body wt p.o. eight hours before the carcinogen. One-half and one-tenth of this initial dose resulted in a loss of protective activity. High doses of heat-treated L. acidophilus strains were also not antigenotoxic. One mechanism of the preventive effect could be that bacterial metabolites or components are responsible. Accordingly, selected examples were investigated in vitro in colon cells of the rat. Metabolically active L. acidophilus cells, as well as an acetone extract of the culture, prevented MNNG-induced DNA damage. Different cell fractions from L. acidophilus (cytoplasm, cell wall skeleton, cell wall) were devoid of antigenotoxic activity, whereas the peptidoglycan fraction and whole freeze-dried cells were antigenotoxic. As a second carcinogen, 1,2-dimethylhydrazine (DMH) was used. A dose- and time-response study was first performed to assess the effects of DMH in several segments of the gastrointestinal (GI) tract. Exposure for 16 hours to 15 or 25 mg DMH/kg body wt p.o. induced DNA damage in cells of the distal colon of rats, whereas no cytotoxicity was seen. Pretreatment orally with LAB on four consecutive mornings before DMH gavage (8 hours after the last LAB application) revealed that L. acidophilus, L. confusus, L. gasseri, B. longum, and B. breve inhibited the genotoxic effect of DMH. One of four S. thermophilus and one of three Lactobacillus delbrueckeii ssp. bulgaricus strains were also protective. Heat-treated L. acidophilus did not inhibit DMH-induced genotoxicity. A few aliquots of the colon cells were processed immunohistochemically for the presence of the "proliferation cell nuclear antigen" (PCNA). DMH treatment did not increase PCNA, nor was there any modulation by LAB. The effect of L. acidophilus on foreign compound-metabolizing enzymes (Phase I and Phase II) in liver and colon cells of rats revealed only one parameter to be modulated, namely, a two- to three-fold increase in the levels of NADPH-cytochrome P-450 reductase. The meaning of this finding, in terms of possible chemoprevention by LAB, remains unclear. In conclusion, our studies show that most, but not all, LAB tested could strongly inhibit genotoxicity in the GI tract of the rat and that viable LAB organisms are required for the protective effect in vivo. The comet assay technique is a powerful tool to elucidate such in vivo antigenotoxic activities in tumor target tissues.

Inhibition of 1,2-dimethylhydrazine-induced oxidative DNA damage by green tea extract in rat.

Following subcutaneous injection of 1,2-dimethylhydrazine (DMH), which is carcinogenic to rat colon and liver, to Sprague-Dawley rats, a significant increase of 8-hydroxydeoxyguanosine (8-OHdG) was observed in the DNA of colonic mucosa and liver. The 8-OHdG formation reached the maximal level at about 24 h after the DMII injection. On the other hand, no increase of 8-OHdG was observed in the DNA of the kidney. Drinking green tea extract (GTE) for ten days prior to the DMH injection significantly inhibited the formation of 8-OHdG in the colon. These findings demonstrate that DMH causes oxidative damage to the DNA of its target organ, and that GTE protects colonic mucosa from this oxidative damage.

Effect of Cocus nucifera on beta-glucoronidade and fecal mucinase activity experimental colon cancer

Foram analisados os efeitos da Cocos nucifera (polpa de noz de coco) na atividade de beta-glucoronidase e mucinase fecal de animais, aos quais foi administrada a 1,2-dimetiltrazina, com o objetivo de induzir aparecimento de cancer do colon. Os animais foram divididos em tres grupos (grupo 2 que recebeu apenas coco, grupo 3 que recebeu 1,2-dimetildrazina e grupo 4 que recebeu polpa de coco e 1,2-dimetildrazina. Quando os tres grupos foram comparados com os ratos controle (grupo 1), verificou-se que a atividade tanto da mucinas, quanto da beta-glucoronidase diminuiu no grupo que recebeu polpa de coco, havendo aumentado no grupo que recebeu 1,2-dimetildrazina e nao correndo alteracao significante no grupo 4...

A model for sensitivity determination of anticancer agents against chemically-induced colon cancer in rats.

Chemically-induced colon cancer was used to test the sensitivity of tumors to chemotherapeutic agents. Seventy-one Sprague-Dawley rats received dimethylhydrazine (20mg/kg) s.c. once weekly for 20 weeks to induce colon cancer. Then a barium enema was performed to see the size of colon tumors. The animals were divided into three groups that were subjected to the following treatments: 5-fluorouracil (5 FU); 1-(2-tetrahydrofuryl)-5 FU(FT); and a mixture of FT and uracil (UFT). After 5 weeks of treatment, the barium enema was repeated. "Response" was assessed on the basis of tumor doubling time. Response rates in the 5-FU, FT, and UFT groups were 25%, 33% and 36%, respectively and this reflects the clinical data of these drugs. The present system may be a predictive model for screening anticancer drugs for human colorectal cancer.