Joint effect of particulate matter and cigarette smoke on women's sex hormones.

BACKGROUND: Although relationships between exposure to air pollution and reproductive health are broadly studied, mechanisms behind these phenomena are still unknown. The aim of the study was to assess whether exposure to particulate matter (PM10) and tobacco smoking have an impact on menstrual profiles of 17ß-estradiol (E2) and progesterone (P) and the E2/P ratio. METHODS: Levels of sex hormones were measured daily in saliva during the entire menstrual cycle among 132 healthy, urban women. Exposure to smoking (active or passive) was assessed by questionnaire, whilst exposure to PM10 with municipal monitoring data. RESULTS: During the early luteal phase, profiles of E2 were elevated among women with higher versus lower exposure to PM10 (p = 0.02, post-hoc tests). Among those who were exposed versus unexposed to tobacco smoking, the levels of mean E2 measured during the entire cycle were higher (p = 0.02). The difference in mean E2 levels between the group of joint exposure (i.e. to high PM10 and passive or active smoking) versus the reference group (low PM10, no smoking) was statistically significant at p = 0.03 (18.4 vs. 12.4 pmol/l, respectively). The E2/P ratios were higher among women with higher versus lower exposure to PM10 and this difference was seen only in the early luteal phase (p = 0.01, exploratory post-hoc tests). CONCLUSIONS: We found that PM10 and tobacco smoking affect ovarian hormones independently and do not interact with each other. Both exposures appear to have estrogenic effects even though women's susceptibility to these effects differs across the menstrual cycle. We propose that the hormonal mechanisms are involved in observed relationships between air pollution and smoking with women's reproductive health.

Apolipoprotein E (ApoE) polymorphism is related to differences in potential fertility in women: a case of antagonistic pleiotropy?

The alleles that are detrimental to health, especially in older age, are thought to persist in populations because they also confer some benefits for individuals (through antagonistic pleiotropy). The ApoE4 allele at the ApoE locus, encoding apolipoprotein E (ApoE), significantly increases risk of poor health, and yet it is present in many populations at relatively high frequencies. Why has it not been replaced by natural selection with the health-beneficial ApoE3 allele? ApoE is a major supplier of cholesterol precursor for the production of ovarian oestrogen and progesterone, thus ApoE has been suggested as the potential candidate gene that may cause variation in reproductive performance. Our results support this hypothesis showing that in 117 regularly menstruating women those with genotypes with at least one ApoE4 allele had significantly higher levels of mean luteal progesterone (144.21 pmol l(-1)) than women with genotypes without ApoE4 (120.49 pmol l(-1)), which indicates higher potential fertility. The hormonal profiles were based on daily data for entire menstrual cycles. We suggest that the finding of higher progesterone in women with ApoE4 allele could provide first strong evidence for an evolutionary mechanism of maintaining the ancestral and health-worsening ApoE4 allele in human populations.

Is the natural PCDD/PCDF mixture toxic for human placental JEG-3 cell line? The action of the toxicants on hormonal profile, CYP1A1 activity, DNA damage and cell apoptosis.

The present study was conducted to define the action of a mixture obtained by the extraction and purification of real fly ash, on specific toxicity endpoints, such as hormonal secretion, CYP1A1 expression, DNA damage and cell apoptosis. JEG-3 cell line was exposed in vitro to different doses of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or Polychlorinated dibenzo-p-dioxin/Polychlorinated dibenzo-P-furan (PCDD/PCDF) mixture. Both TCDD and the mixture decreased hCG secretion, while inhibition of progesterone levels was noted only under the influence of TCDD. The changes in hormone production were not due to the action on cell viability. There were time-dependent differences in CYP1A1 expression in cells exposed to TCDD and PCDD/PCDF mixture. Both TCDD and PCDD/PCDF mixture did not induce the DNA damage, as evaluated by the comet assay. Significantly lower DNA migration from the head of comet into the comet tail was noted after the removal of reagents. The highest efficiency of this process was noted 4 h after the TCDD and 24 h after the PCDD/PCDF mixture removal. These results suggest that the DNA adducts and/or DNA-DNA cross-links were formed. Neither TCDD nor PCDD/PCDF mixture had any effect on cell apoptosis assessed by caspase-3 activity and Hoechst 33258. Taken together, these findings clearly indicate a weaker action of the mixture when compared with TCDD. However, in both cases, their action was not due to the induction of the DNA damage and subsequent cell apoptosis but due to a direct influence of these toxicants on placental hormone production.

Lifespan of etoposide-treated human neutrophils is affected by antioxidant ability of quercetin.

Neutropenia is the primary dose-limiting effect of etoposide toxicity resulting in a decreased efficiency of cancer treatment. Hence, the protection of neutrophils has important clinical implications. We investigated whether quercetin, due to its antioxidant properties, is able to modulate the damaging activity of etoposide. DNA damage, evaluated by the comet assay, and apoptosis, determined by FACScan flow cytometry using Annexin/PI, increased with etoposide doses. The intracellular level of reactive oxygen species (ROS) was enhanced in resting neutrophils incubated with etoposide at concentrations up to 25 microM; above this concentration etoposide revealed antioxidant properties. Only in latex-activated neutrophils, i.e. with latex-stimulated respiratory burst was the ROS production inhibited, as assessed by the luminol amplified chemiluminescence. The characteristic electron spin resonance (ESR) signal of etoposide phenoxyl radical, which occurs in the presence of myeloperoxidase, H2O2 and etoposide, was quenched by quercetin in a dose-dependent manner (0.1-0.5 microM). Quercetin also inhibited DNA damage induced by etoposide and enhanced the inhibitory action of etoposide on the ROS formation in neutrophils. However, quercetin (1 microM) lowered early and late apoptosis/necrosis only when apoptosis was induced by 25 microM etoposide; at higher etoposide concentration apoptosis was enhanced. Summing up, antioxidant adjuvant therapy using quercetin can be beneficial in prolonging neutrophils' lifespan in peripheral blood only when etoposide plasma concentration is low.

Prolonged quercetin administration diminishes the etoposide-induced DNA damage in bone marrow cells of rats.

The DNA damage in bone marrow cells induced by etoposide (E) injected intraperitoneally to rats (100 mg/kg b.w.) decreased to the control level when quercetin (Q) was administered subcutaneously for 10 consecutive days (40 mg/kg b.w.per day) before E was injected. The antioxidant power (FRAP assay) increased significantly after Q or E compared with control rats but did not change when Q preceded the E injection. The superoxide dismutase activity significantly increased in Q+E-treated rats compared with quercetin given alone. The study provides evidence that Q protects bone marrow cells against long-lived E-induced DNA damage and alters the redox balance in lung tissue.

CYP17 genotypes differ in salivary 17-beta estradiol levels: a study based on hormonal profiles from entire menstrual cycles.

Variation in the levels of sex-steroid hormones results from differences in developmental conditions, adult lifestyle, and genetic polymorphism. Genes involved in sex-steroid biosynthesis have been implicated to influence levels of hormones in premenopausal women, but the results were inconclusive. We tested variation among women in levels of salivary estradiol (E(2)) corresponding to CYP17 genotypes. CYP17 encodes cytochrome P450c17alpha, which mediates two enzymes important in E(2) synthesis. In contrast to the earlier studies that relied on one or a few samples for assessing the E(2) levels of an individual woman, our study is based on daily collected saliva samples for one entire menstrual cycle. Sixty Polish women, ages 24 to 36 years, with regular menstrual cycles and no reported fertility problems participated in the study. Women with A2/A2 genotype had 54% higher mean E(2) levels than women with A1/A1 genotype (P = 0.0001) and 37% higher than women with A1/A2 genotype (P = 0.0008). Heterozygous A1/A2 women had 13 % higher E(2) levels than homozygous A1/A1 women (but this difference was significant only in a nonparametric test). Levels of E(2) during the day with highest E(2) (day -1) were 72% higher in A2/A2 compared with A1/A1 (P = 0.01) and 52 % higher compared with A1/A2 (P = 0.03). Our results suggest that CYP17 genotype may serve as a biomarker of endocrine function in women of reproductive age. (Cancer Epidemiol Biomarkers Prev 2006;15(11):2131-5).

Induction of cytochromes P450, caspase-3 and DNA damage by PCB3 and its hydroxylated metabolites in porcine ovary.

Polychlorinated biphenyl (PCBs) levels of tens and hundreds of pg/ml for individual congeners are measured in human follicular fluid. PCB3 (4-chlorobiphenyl), caused a significant increase in estradiol secretion in porcine granulose-theca cell co-cultures and its two metabolites, 4-OH-PCB3 and 3,4-diOH-PCB3, were even more potent than PCB3 itself [Ptak, A., Ludewig, G., Lehmler, H.J., Wojtowicz, A.K., Robertson, L.W., Gregoraszczuk, E.L. 2005. Comparison of the actions of 4-chlorobiphenyl and its hydroxylated metabolites on estradiol secretion by ovarian follicles in primary cells in culture. Reprod. Toxicol. 20, 57-64]. The question is whether these follicle cells are potentially able to metabolize PCB3 to hydroxylated and genotoxic or cytotoxic intermediates. We report here that granulose-theca co-cultures express xenobiotic-metabolizing cytochrome P450 activities, with CYP1A1>CYP2B>>CYP1A2. A significant increase in CYP1A1 and 2B, but not CYP1A2, activity was seen in cells that were exposed to 6 ng/ml PCB3 or 20 nM 17-beta-estradiol. An increase in caspase-3 activity, indicative for apoptosis, was only observed in PCB3-exposed cells after 24 h exposure. Genotoxicity, determined with the Comet assay, was initially reduced after 24 h exposure to PCB3 and both metabolites compared to untreated controls, followed by a significant transient increase in Comets at the 4 and 24 h time point with PCB3 and 4-OH-PCB3. 3,4-diOH-PCB3 induced a significant increase only after 72 h of recovery. We hypothesize that these biphasic damage kinetics may be due to cross-links caused by adduct formation. These results show for the first time that granulose-theca cells in co-culture express CYP1A1, 2B and 1A2 activities and that PCBs at concentrations that are reached in the environment induce genotoxicity in granulosa cells.

A vegetable to meat consumption ratio as a relevant factor determining cancer preventive diet. The Mediterranean versus other European countries.

The observed growth of cancer incidence in certain regions has been usually linked to frequent consumption of 'unhealthy' food. Such food often contains genotoxic substances as heterocyclic aromatic amines (HAAs) and/or polycyclic aromatic hydrocarbones (PAHs), occurring during food preparation, which induce DNA damage in cells. These substances are mainly formed during frying or grilling of meat and they can be removed from the body in a two-stage metabolic process of detoxification (phase 1 and phase 2). If they are not excreted, they form DNA adducts. The effectiveness of detoxification depends on the activity of enzymes encoded by polymorphic genes. A diet containing plenty of fruits and vegetables, due to the presence of biologically active polyphenols, can modulate activity of detoxifying enzymes. Such a diet can decrease the extent of DNA adducts, breaks and oxidative damage, supporting the body's enzymatic system in sufficient removal of DNA damage. The antioxidant vitamins' content in such a diet also enhances the DNA protection by increasing the scavenging of radical oxidative species that occurs during metabolic reactions. The lack of balance between the amount of 'unhealthy' and 'healthy' food leads to the accumulation of unrepaired damage, initiating DNA instability and inducing cancer development. Such damage is often used as a biomarker of cancer risk in epidemiological studies. Moreover, in in vitro studies, the amount of the DNA damage is used as indicator of the protective ability of vitamins, plant extracts and/or individual flavonoids. The incidences of certain dietaryrelated cancers in European Mediterranean countries is lower than in Central and Northern European countries; there is simultaneously variation in the habitual diet in these regions. This suggests that some features of routine nutrition in the Mediterranean countries may be responsible for this preventing effect. However, inconsistency in the epidemiological data, associating the meat and fruit and vegetable intake with cancer risk, suggests that another strategy for evaluation dietary influence on cancer risk should be undertaken. This article argues that it is not the consumption of a single food product or an individual component of diet, but rather a proper ratio of vegetable to meat consumption that is responsible for cancer prevention. This hypothesis is tested comparing the association between certain dietaryrelated cancer incidences (colon & rectum, breast and prostate cancer), registered in 2002, with the ratio between consumption of these two groups of food products in the Mediterranean region and in Central and Northern European region over the last three decades. The results clearly showed that both the ratio between vegetables and meat consumption as well as the ratio between the amount of energy from vegetables and from animal products can be used successfully to evaluate the dietary pattern related to cancer risk.

High tea consumption diminishes salivary 17beta-estradiol concentration in Polish women.

We hypothesized that among reproductive-age women consuming large quantities of tea, the production of estradiol would be suppressed. It has been shown that catechins and theaflavines, the major constituents of tea, inhibit aromatase, an enzyme which catalyses the conversion of androgens to oestrogens. Our study included Polish women living in urban (n 61) and rural (n 48) areas. Women collected daily saliva samples for one complete menstrual cycle and filled out dietary questionnaires. Saliva samples were analysed by RIA for concentration of 17beta-estradiol (E2). Women with high (above the median) average daily consumption of black tea had reduced levels of salivary E2 in comparison with women who drank less black tea (below the median). This effect was observed within the whole study group, as well as separately within urban (P=0.0006) and rural (P=0.013) groups. High intake of the sum of subclasses of tea catechins and epigallocatechin gallate, assessed using the United States Department of Agriculture database (http://www.nal.usda.gov), was also associated with lower concentrations of E2 within all women (P=0.01 and P=0.0001, respectively) and within the urban group (P=0.0001 and P=0.004, respectively). Similar relationships were observed between the sum of subclasses of theaflavines and thearubigines and E2 levels for the whole group (P=0.002) and for urban women (P=0.02). Women with high consumption of tea had lower levels of E2 concentration throughout the entire menstrual cycle. These results may have implications for reducing hormone-related cancer risk by a relatively easy dietary intervention.

The level of endogenous DNA damage in lymphocytes isolated from blood is associated with the fluctuation of 17beta-estradiol concentration in the follicular phase of healthy young women.

The aim of this study was to evaluate whether the differences in plasma 17beta-estradiol concentration in early and late follicular phases of the menstrual cycle can affect the level of endogenous DNA damage in lymphocytes assessed by comet assay, and whether the extent of this damage in the follicular phase is associated with the genotype of catechol-O-methyltransferase (COMT). The level of DNA damage was positively correlated with 17beta-estradiol concentration only in the late follicular phase. Subjects with the COMT L/L homozygous mutated variant revealed more DNA damage as compared to individuals with the COMT wild-type and heterozygous (H/L+HH) genotype.

Uracil misincorporation into DNA of leukocytes of young women with positive folate balance depends on plasma vitamin B12 concentrations and methylenetetrahydrofolate reductase polymorphisms. A pilot study.

Changes in the folate and vitamin B12 status in the body influence the extent of uracil misincorporation (UrMis) into DNA, which is one of the biomarkers of genomic stability and, thus, portends a risk of cancer. In our study, the level of UrMis into DNA was evaluated by the comet assay (using the specific DNA repair enzyme, uracil DNA glycosylase) in leukocytes from blood donated by healthy young women with positive folate balance achieved by 4 weeks of folic acid supplementation (400 microg/day). The nutritional status was evaluated on the basis of nine food diaries recorded by the subjects during two winter months. The data were computerized, and the intake of nutrients and micronutrients was estimated using the DIETA 2 program (Food and Nutrition Institute, Warsaw, Poland) linked to recently updated Polish food tables. The plasma folate and vitamin B12 concentration, as well as methylenetetrahydrofolate reductase (MTHFR) polymorphisms, were evaluated to determine their influence on the level of UrMis into DNA. The mean value of B12 intake for all subjects reached 100% of the Polish recommended dietary allowances (RDA), whereas the mean value of folate intake, before folate supplementation, was 50%, suggesting moderate deficiency. Folic acid supplementation brought the folate intake way above the RDA, and plasma folate concentration in each individual was above the deficient range (mean value 14.67 ng/ml). The UrMis did not correlate with the plasma folate concentration, but the level of UrMis was significantly lower in subjects with plasma vitamin B12 concentration above 400 pg/ml (P=.02) only after folic acid supplementation. The concentration of folate in plasma correlated (P