Inflammation, chromosomal instability, and cancer: the schistosomiasis model.

Evidence is accumulating in support of a role for reactive oxygen species in the etiology of cancer. Inflammatory cells, such as neutrophils, macrophages, and eosinophils, are an important endogenous source of oxygen radicals. Stimulation of these cells by tumor promoters or by foreign bodies (parasites, bacteria, etc.) causes the release of reactive oxygen species. Laboratory studies have shown that genetic damage and neoplastic transformation are induced in vitro in cells cocultured with activated inflammatory cells. We have recently begun to study the role of inflammatory reactions in inducing genetic damage in a human population. This paper describes our initial studies of Egyptian patients infected with Schistosoma haematobium. This infection induces chronic inflammation and irritation in the urinary bladder and is associated with increased cancer at this site. We describe a recently completed population study that shows that infected individuals have elevated levels of genetic damage in their bladders, as measured by the exfoliated cell micronucleus test. Treatment that kills the parasite also reduces the micronucleus frequencies. We also explore the hypothesis that altered sensitivity of clones of cells in these patients to reactive oxygen species could be a force that drives the development of neoplasia by facilitating clonal expansion. Evidence is presented for the possible involvement of loci on chromosome 11 in controlling the level of chromosomal breakage caused by oxidative damage. We have shown that bladder carcinoma cells are sensitive to micronucleus induction by promoter-activated neutrophils and that they can be protected from this damage by insertion of a normal chromosome 11. Further work is in progress to define the source of chromosomal breakage in schistosomiasis patients and to begin to develop an understanding of the host factors protecting bladder cells in these individuals from genetic damage.

Localized induction of micronuclei in the oral mucosa of xeroderma pigmentosum patients.

Xeroderma pigmentosum (XP) patients are predisposed not only to skin cancers but also to tumors on the tip of the tongue. Although this enhanced risk has been attributed to a defect in the repair of DNA damage induced by ultraviolet rays from sunlight there is a lack of data showing that DNA damage is occurring in vivo at these sites. In order to determine whether a relationship exists between exposure to ultraviolet light and the level of chromosomal breakage occurring in epithelial tissue in XP patients, the exfoliated cell micronucleus test was applied to different sites in the oral cavities of four XP patients: the right and left buccal mucosa, the dorsal tip of the tongue and the palate. Six Egyptian controls were sampled concurrently. Micronucleus (MN) frequencies were higher in XP patients than in controls for all sites except the palate, where technical difficulties were encountered. In addition, an unequal distribution of the frequency of micronucleated cells was found in the different sample sites of the oral cavity in the XP patients, with the greatest elevation in frequencies among cells collected from the dorsal tip of the tongue. In contrast, the frequency of micronucleated cells did not vary significantly in samples from different sites obtained from the controls. These data suggest that the complex interplay of host and environmental factors can affect MN frequencies when this endpoint is used to quantify in vivo genotoxic damage in a tissue.

A multiplex PCR procedure for polymorphic analysis of GSTM1 and GSTT1 genes in population studies.

A deletion polymorphism in glutathione S-transferase theta (GSTT1) gene was recently discovered in humans. Similar to the GSTM1 gene, GSTT1 is also recognized as a risk modifier in exposed populations. To evaluate the role of genetic polymorphism in health effects, the combined genetic polymorphism of different genes should be taken into consideration. In the present study, we have developed a multiplex PCR approach for simultaneous replication of both genes for molecular analysis. The multiplex PCR protocol was validated using donor DNA with different polymorphic combinations for both genes from two different ethnic populations (North Americans and Egyptians). The prevalence of the GSTM1 null genotype was 51% among North Americans and 44% among Egyptians. The prevalence of the GSTT1 null genotype was 15% among North Americans and 14.7% among Egyptians. Combined polymorphism analysis of both genes revealed that 6.3% of North Americans harbor the deleted genotype of both genes compared to 8.8% of the Egyptians. The data indicate that there is no major difference in allelic distribution of both genes between the ethnic populations. The multiplex PCR assay used in this study has the advantage of reducing the time, effort and cost required to carry out such analysis. It will also significantly enhance the ability to use genetic screening techniques as a potential tool for early detection of health outcomes in exposed populations.

The CYP2D6 extensive metabolizer genotype is associated with increased risk for bladder cancer.

Inheritance of certain polymorphic metabolizing genes is associated with the development of a number of environmental cancers and may also influence the clinicopathological tumor outcome. We have investigated the association between the inheritance of the polymorphic cytochrome P-450 2D6 (CYP2D6) gene and the development of transitional and squamous cell carcinomas (TCC and SCC) of the bladder in 37 Egyptian cancer patients and 27 matched controls. Genotypic analysis using the polymerase chain reaction (PCR) and the restriction fragment length polymorphism (RFLP) assays revealed that the CYP2D6 extensive metabolizer genotype (CYP2D6*1A) is over represented in bladder cancer patients compared to controls (79 versus 44%, respectively) and is significantly associated with increased risk for bladder cancer (odds ratio (OR) = 4.5, 95% confidence limit (CL) = 1.3-15.7, P = 0.006). Our results also indicate that individuals who have inherited this genotype are more likely to develop TCC (OR = 5.9, 95% CL = 1.4-27.9, P = 0.006) rather than SCC (OR = 3.1, 95% CL = 0.7-15.9; P = 0.09). When the relative risk associated with this genotype was estimated among subjects who were smokers or schistosoma infected, the same tendency towards the development of TCC was observed. These data suggest that the predisposing CYP2D6 gene may not only increase the risk for bladder cancer among Egyptians, but may also influence the clinicopathological tumor outcome.

Cytogenetic monitoring of human populations at risk in Egypt: role of cytogenetic data in cancer risk assessment.

Somatic mutation plays a critical role in carcinogenesis. Numerous environmental agents can increase the probability that somatic mutation will occur. The use of genotoxicity testing is essential for assessing potential human toxicity so that hazards can be prevented. Cytogenetic monitoring of human populations exposed to chemicals has proved to be a useful tool for detecting the chemical mutagenic effects. Cytogenetic analyses of human chromosomes in peripheral lymphocytes allows direct detection of mutation in somatic cells. Different methods can be used for chromosomal analysis (conventional chromosomal analysis, sister chromatid exchange, micronucleus frequency detection). Micronucleus frequency can be detected either in peripheral blood lymphocytes or in exfoliated cells. Different examples of human population studies are presented. Several problems that are found in biomonitoring studies are discussed. These studies should help us learn about individual exposure assessment and biologically relevant doses, leading to quantitative assessment of human cancer risks.

Biomarkers of human exposure to pesticides.

For centuries, several hundred pesticides have been used to control insects. These pesticides differ greatly in their mode of action, uptake by the body, metabolism, elimination from the body, and toxicity to humans. Potential exposure from the environment can be estimated by environmental monitoring. Actual exposure (uptake) is measured by the biological monitoring of human tissues and body fluids. Biomarkers are used to detect the effects of pesticides before adverse clinical health effects occur. Pesticides and their metabolites are measured in biological samples, serum, fat, urine, blood, or breast milk by the usual analytical techniques. Biochemical responses to environmental chemicals provide a measure of toxic effect. A widely used biochemical biomarker, cholinesterase depression, measures exposure to organophosphorus insecticides. Techniques that measure DNA damage (e.g., detection of DNA adducts) provide a powerful tool in measuring environmental effects. Adducts to hemoglobin have been detected with several pesticides. Determination of chromosomal aberration rates in cultured lymphocytes is an established method of monitoring populations occupationally or environmentally exposed to known or suspected mutagenic-carcinogenic agents. There are several studies on the cytogenetic effects of work with pesticide formulations. The majority of these studies report increases in the frequency of chromosomal aberrations and/or sister chromatid exchanges among the exposed workers. Biomarkers will have a major impact on the study of environmental risk factors. The basic aim of scientists exploring these issues is to determine the nature and consequences of genetic change or variation, with the ultimate purpose of predicting or preventing disease.

Monitoring of human populations at risk by different cytogenetic end points.

Humans are exposed to a large number of environmental genotoxic agents. These can increase the probability that somatic mutation will occur. The use of genotoxicity testing is essential for assessment of potential human toxicity so that hazards can be prevented. Cytogenetic monitoring of human populations exposed to chemicals has proved to be a useful tool for detecting the chemical mutagenic effects. Cytogenetic analysis of human chromosomes in peripheral lymphocytes allows direct detection of mutation in somatic cells. Cytogenetic monitoring of a group of traffic policemen from Cairo, Egypt, was an example of a human population study. The induction of chromosomal damage was studied in a group of 28 traffic policemen with exposure of over 10 years and a control group of 15 policemen trainers. Blood lead level was significantly higher in the traffic policemen (30 +/- 8.7) unit compared to the control group (18.2 +/- 1.2) unit. The percentage of chromosomal aberrations (7.7 +/- 3.1), as well as the mean sister chromatid exchanges (7.5 +/- 3.4), were significantly higher among the traffic policemen than in the control group. The percentage of chromosomal aberrations was 2.8 +/- 2.1 and the mean sister chromatid exchanges was 4.8 +/- 2.9 in the control group. On the other hand, the increase in chromosome damage among the traffic policemen was enhanced further by smoking. Several problems that are found in biomonitoring studies are discussed.

Chemical interaction: enhancement and inhibition of clastogenicity.

Most environmental exposures involve concurrent or sequential exposure to multiple chemicals in air, water, and food. Interactive effects in carcinogenesis have been described for certain combinations of agents. They are described in terms of enhancement or inhibition of carcinogenesis. Enhancement effects have been documented for cigarette smoking in combination with exposure to asbestos, radon, alcohol, or other exposures. A variety of inhibitors of carcinogenesis have also been described. They are classified into agents preventing formation of carcinogens; blocking agents; and suppressing agents. Assessment of risk from exposure to multiple agents can be derived either from epidemiological studies in relation to actual exposure or from laboratory studies after controlled exposure to different agents. Prediction of how toxic components of mixtures will interact should be based on an understanding of the mechanisms of such interactions. Compounds may interact chemically, yielding new toxic components or causing a change in the biological availability of the existing components or metabolites. In humans, great individual variability in response is to be expected because of genetic heterogeneity or acquired host susceptibility factors. Interaction is thus a key component in the risk assessment process. In this paper, the definition of interaction and the theoretical basis for different types of interaction in cancer causation are reviewed. Epidemiological and experimental studies showing interactive effects of two chemical carcinogens are also presented.

Hepatitis C in a community in Upper Egypt: I. Cross-sectional survey.

The prevalence of antibody to hepatitis C virus (anti-HCV) was determined in a cross-sectional survey in a village in Upper Egypt. Exposure and demographic characteristics were obtained through a questionnaire. Antibody to hepatitis C virus was assessed using a second generation enzyme immunoassay, and the presence of HCV RNA was tested using a reverse transcriptase-polymerase chain reaction. Collection of blood samples was targeted at those > or = 5 years old, and obtained from 62.8%. This report describes the community, the HCV infection characteristics of the subjects, and evaluates some factors associated with presence of anti-HCV. Of the 6,031 participants, 522 (8.7%) were anti-HCV positive. Prevalence was higher among males than females (11.3% versus 6.5%; P < 0.001). It was greater among those > 30 years of age than among those < or = 30 years of age (20.0% versus 3.6%; P < 0.001). Those who were less educated, farmed, provided health care, and were currently married had a significantly higher anti-HCV prevalence than those who were not; however, these associations were not significant after adjusting for age. Although active infections with Schistosoma haematobium were not associated with anti-HCV, a history of past infection was (age-adjusted risk ratio [RR] = 2.1, 95% confidence interval [CI] = 1.8, 2.4); 134 persons who had a history of receiving parenteral anti-schistosomal therapy had a higher age-adjusted RR (3.0; 95% CI = 2.5, 3.7) for anti-HCV than those who did not. Hepatitis C virus RNA was detected in 62.8% of the anti-HCV positive subjects, without significant variation by age, gender, education, or marital status. The prevalence of anti-HCV in Upper Egypt is high, albeit lower than in Lower Egypt, with continuing but limited transmission indicated by the lower prevalence in residents < or = 30 years old.

The burden of trachoma in the rural Nile Delta of Egypt: a survey of Menofiya governorate.

BACKGROUND: Evidence of widespread distribution of trachoma in Egypt had not been clarified as previous surveys were limited to individual communities which may not have been representative of the general population. The Nile Delta of Egypt presents a unique environment for trachoma to persist. Economic improvements in the past decade have affected even the poorest rural environments; availability of electricity is now found in many rural communities. Availability of water in Nile Delta has always been good but poor hygienic conditions have been the primary factor in trachoma transmission. A survey of trachoma was undertaken in Menofiya governorate to determine if Egypt should be identified as trachoma endemic and targeted for trachoma control efforts. METHODS: A multistage random cluster study design was used with the target population defined as adults aged 50 and over and children aged 2-6 years from throughout the governorate. Among preschool children only trachoma was graded while among adults presenting visual acuity and cause of vision loss or blindness were also recorded. Adults were interviewed regarding past trichiasis surgery; those currently with trichiasis or a history of trichiasis surgery were also interviewed regarding outcome of surgery. RESULTS: A total of 3272 children aged 2-6 and 3322 adults age 50+ were enumerated. Among the children 81.3% were examined and among the adults 73.0% were examined. Active trachoma (follicles (TF) and/or intense inflammation (TI)) was found among 36.5% (95% confidence interval (CI) 34.7-38.3%) of the children. TI was 1.89 (95% CI 1.22-2.94) times more common in rural children compared to urban children. The prevalence of trichiasis (TT) in adults was 6.5%; women had an age adjusted odds of trichiasis of 1.68 (95% CI 1.18-2.39) compared to men. Trichiasis was 2.11 times (95% CI 1.33-3.37) more common in rural Menofiya compared to urban Menofiya. TT accounts for blindness (presenting vision <3/60) in 8% of patients and accounts for 13.2% of visual impairment. Overall, trichiasis surgical coverage was 34.4%, slightly higher among men than women. The outcome of trichiasis surgery was poor in 44.4% of cases. CONCLUSION: Trachoma is a serious public health problem in Menofiya governorate and a significant contributor to vision loss. These findings would suggest that continued poor hygienic conditions in rural Egypt have limited the reduction of active trachoma even in the face of significant improvements in socioeconomic status. Furthermore, the high proportion of trichiasis surgery cases with a poor outcome would indicate a need to reassess current surgical practices in Egypt and improve training and monitoring.

Field studies of aflatoxin exposure, metabolism and induction of genetic alterations in relation to HBV infection and hepatocellular carcinoma in The Gambia and Thailand.

The relative contribution of aflatoxins (AF) and hepatitis B virus (HBV) to the aetiology of liver cancer remains to be determined, as does the mechanism of any interaction between these two factors. Methods to measure individual exposure to AF permit the assessment of this possible interaction in field studies. The measurement of AF covalently bound to albumin in peripheral blood has been particularly useful in this respect. In east and west African countries the majority (75-100%) of individuals has been found positive (> 5 pg AFB1-lysine eq./mg albumin) for the AF-albumin adduct with levels ranging up to 720 pg/mg. Levels of adduct to date have been age- and sex-independent, although marked seasonal variations were seen in The Gambia. Exposure also occurs in utero, with the AF-adduct being found in umbilical cord blood. In a study in The Gambia involving 323 children (age 3-8 years) the AF-albumin adduct levels were examined with respect to HBV infection and ethnic group. Over 95% of all sera contained detectable adduct but children positive for HBV surface antigen (HBsAG) had significantly higher adduct levels than children with markers of past infection or who had never been infected (mean (log) AF-albumin adduct levels 4.41 +/- 0.95, 4.04 +/- 0.99, and 4.05 +/- 1.03 respectively, p = 0.04). In addition, there were highly significant differences in adduct levels between the three major ethnic groups (Wollof 4.41 +/- 0.69: Fula 4.05 +/- 1.1; Mandinka 3.7 +/- 1.14). Wollof children were also more likely to be HBsAg positive than the other two groups. These data suggest that ethnic group and HBV infection can influence AF metabolism and this is being examined in this population with respect to genetic polymorphisms in cytochrome P450 and glutathione-S-transferase enzymes. In addition, these biomarkers are being compared to the nature and frequency of mutations in somatic and tumour cells.

Praziquantel (antischistosomal drug): is it clastogenic, co-clastogenic or anticlastogenic?

Schistosoma haematobium infection is the most common health problem in Egypt. It is strongly associated with the development of urinary bladder carcinoma. The actual cause for the development of cancer is still under investigations, it can be due to mechanical irritation from schistosomiasis ova, the infection itself or the drugs which are used to treat the patients. Praziquantel (PQ) is a commonly used drug to treat schistosomiasis patients. In mice, an in vivo cytogenetic study showed that PQ is not clastogenic in mice. The frequency of micronuclei in all the study groups were insignificantly different from the control group (p > 0.05). However, it enhanced the clastogenicity of benzene at a very high dose. Results from combined exposure with benzene and PQ showed a significant PQ dose-dependent increase in micronucleus frequency (p < 0.05). A metabolite study revealed that PQ enhanced the metabolism of benzene to form muconaldehyde which may be responsible for the enhancement effect. In schistosomiasis patients, two cytogenetic studies were carried out before and after treatment with PQ. In one of these studies, peripheral blood lymphocytes were examined from schistosomiasis patients to detect chromosomal aberrations (CAs) and micronuclei (MN) before and after treatment with PQ. There was no significant increase in CAs in patients compared with the controls (p > 0.05). The frequency of MN was significantly higher in the infected persons (0.59 +/- 0.44) than the control individuals (0.23 +/- 0.23) (p < 0.05). After treatment, there was no significant change in both parameters. The other study was conducted to determine whether infection with this parasite resulted in an increase of chromosomal breakage, micronuclei, in exfoliated urothelial cells. Micronucleus frequencies were significantly higher in the infected group (mean frequency, 0.84 +/- 0.69%) than among controls (mean frequency, 0.12 +/- 0.21%, p < 0.001). Micronucleus frequencies were higher in infected individuals who smoked compared with those who were non-smokers, although this effect was not significant (p > 0.05). The mean micronucleus frequencies were reduced significantly in the group of patients who were followed up (before treatment, 0.80 +/- 0.70%, after treatment, 0.19 +/- 0.23%, p < 0.001), thus supporting a direct involvement of the infection in increased chromosomal breakage in the urothelium and provide proof of the role of PQ in decreasing the risk of cancer development. At this stage, we still need to study the cytogenetic effect of human exposure to environmental agents such as pesticides, smoking, etc., together with treatment with PQ.

Assessment of cytogenetic changes in human populations at risk in Egypt.

Humans are exposed to numerous environmental agents that can increase the probability of mutagenicity and carcinogenicity. Most of environmental exposures involve concurrent or sequential exposure to several agents in air, water, and food. Interactive effects in carcinogenesis have been described for a certain number of combinations of agents. They are described in terms of enhancement or inhibition of carcinogenesis. Risk assessment of exposure to environmental agents can start either from laboratory studies after exposure to different agents or from epidemiological studies in relation to actual exposure. The use of genotoxicity testing is essential for assessment of potential human toxicity so that hazards can be prevented. Cytogenetic monitoring of human populations exposed to environmental agents has proved to be a useful tool for detecting their mutagenic effects. Cytogenetic analysis of human chromosomes in peripheral lymphocytes allows direct detection of mutation in somatic cells. Various methods can be used for chromosomal analysis (conventional chromosomal analysis, sister chromatid exchange, micronucleus frequency detection). Micronucleus frequency can be detected either in peripheral blood lymphocytes or in exfoliated cells. Different examples of human population studies are presented in this review. Several problems which are found in biomonitoring studies are discussed.

Cytogenetic effects in a group of traffic policemen in Cairo.

The aim of this study was to evaluate the cytogenetic effects in humans exposed to automobile exhaust. The induction of chromosome damage was studied in an exposed group of 28 traffic policemen with exposure of over 10 years and a control group of 15 policemen trainers from the Faculty of Police. The percentage of chromosomal aberrations as well as the mean sister-chromatid exchanges were significantly higher among the traffic policemen than in the control group. The cause for this elevated chromosome damage is most likely due to their exposure to pollutants from automobile exhaust, however, the increase is not correlated with the blood lead level or the duration of employment. On the other hand, the increase in chromosome damage among the traffic policemen is enhanced further by smoking.

Chromosomal aberrations and micronuclei in reinforced plastics workers exposed to styrene.

The aim of this study was to investigate the cytogenetic changes induced in humans exposed to styrene in a reinforced plastics plant. Blood and urine samples were collected from 18 styrene exposed workers and 18 age and sex matched control subjects from the administrative department of the same factory. Chromosome aberrations (CAs) and micronuclei (MN) (cytokinesis block method) were analyzed in blood lymphocytes. All of the subjects included in the study were male non-smokers. The duration of employment ranged from 10 to 22 years (14.3 +/- 4.4). In order to monitor exposure to styrene, urinary mandelic acid (MA) levels were measured using a standard colorimetric method. The level of thioethers in the urine was also determined colorimetrically. The mean level of mandelic acid was significantly higher in the exposed workers (328.44 +/- 266.21 mg/g creatinine) compared with that of the controls (50.09 +/- 16.84 mg/g creatinine) (p < 0.05). The level of urinary thioethers was found to be higher among the exposed workers. The number of cells with chromosomal aberrations was significantly higher in the workers (6.06 +/- 4.41) compared with the controls (3.44 +/- 2.28) (p < 0.05). There was no significant increase in the frequency of micronuclei in the exposed workers compared to controls. Our results support earlier findings on increased rates of chromosomal aberrations in reinforced plastics workers.

Reduction in chromosomal damage in schistosomiasis patients after treatment with praziquantel.

Schistosoma haematobium infection is the most common health problem in Egypt. It is strongly associated with the development of urinary bladder carcinoma. The aim of this study was to determine whether infection with this parasite resulted in an increase of chromosomal breakage in the exfoliated urothelial cells. Urine samples were collected from 211 Egyptian male farmers and schoolchildren with Schistosoma haematobium ova in their urine. 66 samples were suitable for scoring. Accordingly, 66 non-infected males attending the same village clinic were included in the study as a control group. The urine was centrifuged and the urothelial cells present in the sediment were assayed for the presence of micronuclei, a quantitative indicator of chromosomal breakage. Micronucleus frequencies were significantly higher in the infected group (mean frequency, 0.84 +/- 0.69%) than among controls (mean frequency, 0.12 +/- 0.21%, p < 0.001). There was a trend towards an increase in micronucleus frequencies in infected individuals who smoked (17 of the 66 subjects) compared with those who were non-smokers, although this effect was not significant (p > 0.05). Bladder infection was associated with a 6.1-fold increase in micronucleus frequencies among non-smokers. In this study, patients were treated with praziquantel, an anti-schistosomal agent. Out of the 66 studied patients, only 37 gave scoreable samples after treatment. The mean micronucleus frequencies were reduced significantly (before treatment, 0.80 +/- 0.70%, after treatment, 0.19 +/- 0.23%, p < 0.001), thus supporting a direct involvement of the infection in increased chromosomal breakage in the urothelium.

Micronuclei, chromosomal aberrations and aflatoxin-albumin adducts in experimental animals after exposure to aflatoxin B1.

Rats and mice differ markedly in sensitivity to aflatoxin B1 (AFB1) hepatocarcinogenicity, the former being sensitive and the latter resistant. Animals were treated with single doses of different concentrations of AFB1, between 0.01 and 1.0 microgram AFB1/g body weight. The frequency of chromosomal aberrations and micronuclei in the bone marrow was measured and compared to the level of AFB1 bound covalently to albumin in the peripheral blood. Both chromosomal aberrations and micronuclei were significantly increased in treated rats compared to the control group at doses above 0.1 microgram/g. In contrast, in mice, a slight increase in chromosome aberrations was seen in the highest dose group (1.0 microgram/g) but no increase in micronuclei was observed at any of the doses. The level of chromosomal aberrations was about 10 times higher in rats than in mice at the highest dose of AFB1. AFB1-albumin (AF-alb) adducts did not show a strong dose-response increase after treatment in mice, whereas in rats the levels increased linearly with dose of AFB1 and there were strong correlations at the individual rat level with both chromosomal aberrations (r = 0.92; p < 0.0001) and micronucleus frequency (r = 0.86; p < 0.0001). These data suggest that the AF-alb may reflect the level of genetic alteration resulting from the initial binding of this carcinogen to cellular DNA. Therefore, this adduct used as a biomarker in studies of human exposure to aflatoxin may provide information not only on exposure but also on the risk of genetic alterations consequent to that exposure.

Enhancement of benzene clastogenicity by praziquantel in mice.

Praziquantel (PQ) is a commonly used drug to treat patients with schistosomiasis. Previous studies using cells in vitro have shown that PQ can enhance the mutagenic activities of known mutagens. We have conducted a cytogenetic - urine metabolite study to determine the in vivo clastogenic and co-clastogenic potential of PQ with a ubiquitous environmental contaminant, benzene (BZ). 16 groups of adult male ICR mice (5 animals per group) were used. They were negative control, solvent controls (cremophore E1 3%, olive oil and combined), positive control (BZ 440 mg/kg b.w.) and 11 exposed groups. To test for clastogenicity of PQ, mice were treated orally with 100, 400, 800 and 1200 mg/kg b.w. PQ and sacrificed 30 h later for determination of micronuclei (MN) frequency in bone-marrow polychromatic erythrocytes (PCE). None of these PQ does induced an increase of MN frequency. On the other hand, BZ induced, as expected, a high frequency of MN (46.4 +/- 6.34/1000 PCE). The enhancement effect of PQ was tested in 7 groups of mice using 3 different protocols. Mice were treated with 440 mg/kg b.w. BZ and 1 h later with 0, 100, 200, 400, 800 and 1200 mg/kg b.w. PZ. In another group, 800 mg/kg PQ was administered at 3 h after BZ exposure. In the last group, PQ (800 mg/kg) was administered at 1 h prior to BZ exposure. Results from the first combined exposure group showed a significant PQ dose-dependent increase in the frequency of MN in PCE (p less than 0.05). The increase with the two high doses of praziquantel is significantly higher (p less than 0.05) than the MN frequencies in the benzene control and the expected value based on the additive effects of the two agents. Studies with other combined treatment groups showed that the induction of MN was highest when PQ was administered at 1 h before BZ exposure. Moreover, the presence of BZ metabolites (muconic acid, phenol, catechol and hydroquinone) in urine was studied in 6 of the combined treatment groups. This metabolite study revealed that PQ enhanced the metabolism of BZ towards the pathway to form muconaldehyde which is converted to muconic acid in urine. In conclusion, our study showed that PQ is not a clastogen but can enhance the clastogenic activity of BZ in vivo by shifting the metabolic pathways of BZ towards formation of muconaldehyde which may be responsible for the enhancement effect.

Involvement of inflammatory reactions and elevated cell proliferation in the development of bladder cancer in schistosomiasis patients.

Schistosoma haematobium infection is strongly associated with urinary bladder cancer. Although numerous explanations have been proposed for this association, the nature of this relationship remains unresolved. This paper explores the hypothesis that inflammation and elevated cell proliferation play a major role in the development of bladder cancer in infected patients, possibly by increasing the level of genetic instability in the urothelium. The paper details in vivo and in vitro studies being done in our laboratories to test this hypothesis. These studies include population studies in which chromosomal breakage in the bladder of infected individuals is assayed using the micronucleus (MN) test on exfoliated urothelial cells. The approach also includes parallel studies in Vancouver with patients with long-term catheter drainage, a population with many similarities to schistosomiasis patients. In the in vitro studies we are co-incubating bladder cells with activated neutrophils or experimental conditions simulating inflammation. These studies show that inflammatory cells when activated can induce micronuclei in bladder cells and that this response is associated with loci on chromosome 11, a chromosome commonly altered during bladder carcinogenesis. A final approach being used is to assay chromosomal change (MN frequencies and numerical chromosome alterations) and level of proliferation (expression of proliferating cell nuclear antigen) in archival biopsies from schistosomiasis patients. Preliminary results show that a dysregulation of cell proliferation is occurring during cystitis in these patients. The extent to which this alteration affects the level of chromosomal breakage is yet to be determined.

The mutagenicity of Gramoxone (paraquat) on different eukaryotic systems.

The possible mutagenicity of the herbicide Gramoxone was evaluated using five different living systems: Allium cepa, Vicia faba, yeast, Drosophila melanogaster and human lymphocytes. The results indicate that Gramoxone has mutagenic activity at the cytological level in Allium cepa, Vicia faba and human lymphocytes. All doses were effective in inducing chromosomal abnormalities and a clear dose-response relationship was observed in the various cytological tests. Analysis of chromosomal abnormalities revealed that this herbicide displays clastogenic and turbagenic activities. At the gene mutation level Gramoxone induced gene conversion at the trp-5 locus and reversion at the ilv locus in Saccharomyces cerevisiae. In Drosophila melanogaster, Gramoxone proved to be mutagenic to germ cells and induced a high frequency of sex-linked recessive lethals (SLRL). At the protein level, Gramoxone had detectable mutagenic effects on the genetic background of two enzymes, Adh and Est-6. Gramoxone should be considered a mutagenic herbicide.