Daniela Reali

Antioxidant effects of flavonoid from Croatian Cystus incanus L. rich bee pollen.

Oxidant/antioxidant status, estrogenic/anti-estrogenic activity and gene expression profile were studied in mice fed with Cystus incanus L. (Cistaceae) reach bee pollen from location in Central Croatias Dalmatia coast and offshore islands. Seven phenolic compounds (out of 13 tested) in bee pollen sample were detected by high performance liquid chromatography (HPLC) analysis. Phenolics detected in C. incanus L. bee pollen belong to flavonol (pinocembrin), flavanols (quercetin, kaempferol, galangin, and isorhamnetin), flavones (chrysin) and phenylpropanoids (caffeic acid). Bee pollen as a food supplement (100mg/kgbw mixed with commercial food pellets) compared to control (commercial food pellets) modulated antioxidant enzymes (AOE) in the mice liver, brain and lysate of erythrocytes and reduced hepatic lipid peroxidation (LPO). Bee pollen induced 25% of anti-estrogenic properties while no estrogenic activity was found. Differential gene expression profile analyses after bee pollen enriched diet identify underexpressed gene Hspa9a, Tnfsf6 (liver) and down-regulated gene expression of Casp 1 and Cc121c (brain) which are important in the apoptosis pathway and chemotaxis. These results indicate that used bee pollen possess a noticable source of compounds with health protective potential and antioxidant activity.

Characterization of 'faecal streptococci' as indicators of faecal pollution and distribution in the environment

The recent revision of the taxonomy of ‘faecal streptococci’ prompted us to verify the importance of identifying the species of this group of cocci. During a study carried out to assess the hygienic quality of environmental samples from a variety of sources, we isolated 198 strains named faecal streptococci on the basis of conventional international tests (EVA broth multiple tube test) used for Public Health purposes. The predominant species were Enterococcus faecalis (39%) and Ent. faecium (29%), followed by Ent. durans/hirae, Ent. casseliflavus/gallinarum, Ent. raffinosus, with a different prevalence of the species depending on the source. Eighty‐four per cent of isolates were true faecal species. Only one isolate was identified as belonging to the Streptococcus genus. The authors stress the opportunity to identify the species. This may help to clarify the ecological and epidemiological characteristics of intestinal enterococci and streptococci in the environment, in drinking and recreational waters and their meaning as indicators of faecal pollution. All isolates were tested for their susceptibility to some antimicrobial agents widely used in medical therapy and the pattern was compared with the pattern of isolates from clinical specimens.

Screening of estrogen-like activity of mineral water stored in PET bottles

Bottled mineral water consumption is steadily rising in the World. Italy is the largest natural mineral water consumer in Western Europe, about 200L per capita per annum. Recently, research has concentrated upon emerging toxicological problems such as the presence in drinking water of substances which interfere with the function of the endocrine system; defined as endocrine disruptors (EDs). The aim of this study was to assess the risk of exposure of the population to chemicals with estrogen-like activity through mineral water consumption by monitoring the presence of estrogenic compounds in mineral water bottled in polyethylene terephthalate (PET). A solid phase extraction (SPE) with C18 cartridges was carried out. The estrogenic activity of the extracts was assayed using a yeast assay expressing the human estrogen receptor alpha (hERalpha). This preliminary study shows that more than 90% of the water samples did not exhibit any appreciable estrogenic activity. The highest estrogenic activity detected in water extracts was equivalent to the activity induced by 23.1ng/L of the natural hormone 17beta-estradiol. Some mineral water samples showed toxicity on yeast cells.

Sister-chromatid exchanges in lymphocytes and mutagenicity in urine of nurses handling cytostatic drugs

Sister-chromatid exchanges (SCE) in peripheral blood lymphocytes and mutagenicity of urine (Ames test) were measured in a group of 21 nurses professionally handling antineoplastic drugs and in a group of 21 unexposed controls. No differences in SCE frequencies and in urinary mutagenic activity between exposed and unexposed groups were detected. A clear positive increase in urinary mutagenicity in the TA98 Salmonella strain was observed with increasing number of cigarettes smoked, whereas no evident influence of smoking on SCE was seen. Age, coffee and alcohol consumption did not show any detectable effect in the two tests.

National pathfinder survey on children's oral health in Italy: pattern and severity of caries disease in 4-year-olds.

This paper describes the dental health status of Italian 4-year-olds in 2004/2005 and analyzes the association between caries in preschool children and some background characteristics in children and parents. Caries was recorded according to WHO criteria. 5,538 subjects were examined. Information on the children’s and their parents’ social, behavioral, ethnic and demographic status was obtained through a series of closed questions. Bivariate analysis was performed to investigate the association between caries and background characteristics. The probability of being an extra zero for the dmfs index was estimated via the zero-inflated negative binomial regression model (ZINB). 78.4% (95% CI = 77.2–79.6) of the children were caries-free. The national mean dmfs index was 1.36 (95% CI = 1.15–1.57), ranging from 1.22 (95% CI = 1.03–1.42) in the Italian North-East to 1.73 (95% CI = 0.83–2.63) in the South section. Significant bivariate associations between caries experience and risk factors were observed for parents’ nationality (not Italian vs. Italian: p < 0.001), parents’ educational levels (low vs. high: p < 0.001), preterm birth (yes vs. no: p = 0.011), prolonged breastfeeding (≤13 months vs. >13 months: p = 0.038) and early tooth eruption (<6 months as reference: p = 0.004). Multivariable analysis (ZINB) showed that children with a low caries risk level had a higher probability of being an extra zero; in particular, children from fathers with a high educational level showed a probability of being an extra zero of 0.22. The results suggest a need to plan preventive programs to reduce oral health disparities among Italian preschool children, based on educational intervention with parents and children concerning oral health and caries prevention.

National Pathfinder Survey of 12-Year-Old Children’s Oral Health in Italy

No recent data on the experience of caries among Italian 12-year-olds are available. In 2004, an epidemiological survey called ‘National Pathfinder among Children’s Oral Health in Italy’ was promoted and carried out. This study reports the actual oral health status of Italian 12-year-olds according to gender, residence area and geographical distribution. Clinical examinations were carried out from March 2004 to April 2005, according to WHO criteria, and included dental caries (decay at the dentinal lesion level) and Community Periodontal Index (CPI). 5,342 children (2,670 males, 2,672 females) were examined by 7 ad hoc calibrated raters. Dental caries experience was found in 43.1% (95% CI 41.8–44.4%) of the study population. The mean DMFT score was 1.09 (95% CI 0.98–1.21). Significant differences (p < 0.05) were observed among geographical sections for DT, FT and DMFT. An inverse relationship was observed between mean DMFT and gross national product per capita (p < 0.001). Gingival bleeding was observed in 23.8% of children, while 28.7% had calculus. Significant differences in CPI scores among sections were found throughout the sample in both males and females (p < 0.001). Over the past two decades, mean DMFT fell from over 5 to its present level, halving every decade; consequently, the recorded level of dental caries has become aligned with that in other Western European countries. Nevertheless, differences in DMFT values remain between children from different socioeconomic backgrounds.

Estrogen-like activity of seafood related to environmental chemical contaminants

BackgroundA wide variety of environmental pollutants occur in surface waters, including estuarine and marine waters. Many of these contaminants are recognised as endocrine disrupting chemicals (EDCs) which can adversely affect the male and female reproductive system by binding the estrogen receptor and exhibiting hormone-like activities. In this study the estrogenic activity of extracts of edible marine organisms for human consumption from the Mediterranean Sea was assayed.MethodsMarine organisms were collected in two different areas of the Mediterranean Sea. The estrogenic activity of tissues was assessed using an in vitro yeast reporter gene assay (S. cerevisiae RMY 326 ER-ERE). Concentrations of polychlorinated biphenyls (PCBs) (congeners 28, 52, 101, 118, 138, 153, 180) in fish tissue was also evaluated.ResultsThirty-eight percent of extracts showed a hormone-like activity higher than 10% of the activity elicited by 10 nM 17b-estradiol (E2) used as control.Total PCB concentrations ranged from 0.002 up to 1.785 ng/g wet weight. Chemical analyses detected different levels of contamination among the species collected in the two areas, with the ones collected in the Adriatic Sea showing concentrations significantly higher than those collected in the Tyrrhenian Sea (p < 0.01).ConclusionThe more frequent combination of chemicals in the samples that showed higher estrogenic activity was PCB 28, PCB 101, PCB 153, PCB 180.The content of PCBs and estrogenic activity did not reveal any significant correlation.

Antiestrogenic and antigenotoxic activity of bee pollen from Cystus incanus and Salix alba as evaluated by the yeast estrogen screen and the micronucleus assay in human lymphocytes.

The estrogenic/antiestrogenic activity and the genotoxicity/antigenotoxicity of bee pollen from Salix alba L. and Cystus incanus L. and its derivative extracts in yeast and human cells was investigated. All samples showed a marked inhibitory effect on the activity of the natural estrogen 17 beta-estradiol (higher than 90% for extracts 2) and failed to cause estrogenic activity and chromosome damage. At least one preparation from each species showed a marked antigenotoxic effect against the action of the anticancer drugs mytomicin C, bleomycin, and vincristine. Bee pollens from C. incanus and S. alba were found to be neither genotoxic nor estrogenic as well as effective estrogen inhibitors, and able to reduce the chromosome damage induced by the three cancer drugs used, thus supporting their use as a safe food supplement and future chemoprotective/chemopreventive agents.

Estradiol-antagonistic Activity of Phenolic Compounds from Leguminous Plants

Natural flavonoids are currently receiving much attention because of their estrogenic and antiestrogenic properties. Six isoflavones (isoprunetin, isoprunetin 7‐O‐β‐d‐glucopyranoside, isoprunetin 4′,7‐di‐O‐β‐d‐glucopyranoside, genistein, genistein 7‐O‐β‐d‐glucopyranoside, daidzein), four flavones (luteolin, luteolin 7‐O‐β‐d‐glucopyranoside, luteolin 4′‐O‐β‐d‐glucopyranoside, licoflavone C), isolated from Genista morisii and G. ephedroides (two Leguminosae plants of the Mediterranean area) together with two structurally related pterocarpans, bitucarpin A and erybraedyn C, isolated from Bituminaria bituminosa (Leguminosae), were tested for the antagonist activity by a yeast based estrogen receptor assay (Saccharomyces cerevisiae RMY326 ER‐ERE). Most compounds inhibited the estradiol‐induced transcriptional activity in a concentration dependent manner. In particular, for the flavone luteolin 77% inhibition of the induced β‐galactosidase activity was observed. Interestingly, licoflavone C exhibited a dose‐dependent antagonistic activity at concentrations up to 10−4 m, but stimulated β‐galactosidase expression at higher concentrations resulting in a U‐shaped‐like dose‐response curve. Copyright

Determination of trace endocrine disruptors in ultrapure water for laboratory use by the yeast estrogen screen (YES) and chemical analysis (GC/MS)

High purity water for endocrine disruptors (EDs) analysis in experimental tests is an indispensable requirement for the preparation of reagents and solutions employed in biological laboratories. Commercial ultrapure water may contain traces of organic compounds, which can interfere with in vitro bioassays carried out to detect the potential estrogen-like activity of pure compounds and complex mixtures. This paper shows that solid-phase extracts of different types of ultrapure water (UPW) purchased or produced in situ for laboratory analysis (mQ-UPW) may contain organic molecules able to antagonize the binding of E(2) to the human estrogen receptor alpha in the yeast estrogen screen (YES) assay. GC/MS analysis detected the presence of bis(2-ethylhexyl) phthalate (DEHP) (0.033 ppm+/-0.006) in mQ-UPW extracts. The dose-response curve of DEHP in the YES assay showed a relevant antagonist effect of this phthalate. Agreement between content of DEHP chemically detected in UPW extract and the magnitude of biological effects induced was pointed out. It would be appropriate that chemical analyses were complemented by biological tests to establish concentration limits for chemical contaminants in UPW that do not induce biological effects detectable in vitro. The yeast assay used in this study has previously proved to be a sensitive tool in assessing the presence of agonistic/antagonistic chemicals at the ng/l level in complex mixtures and may be successfully used to identify trace amounts of estrogenic/antiestrogenic chemicals, which can represent critical issues influencing the experimental results in environmental testing laboratories.