Benzene Exposure and Lung Cancer Risk: A Systematic Review and Meta-Analysis of Human Studies.

Lung cancer is a leading cause of death with nearly 1.8 million deaths estimated worldwide in 2020. Although benzene is classified as a human carcinogen (Group 1) on the basis of its association with acute myeloid/non-lymphocytic leukaemia, there is still limited evidence that it may influence lung cancer risk. This study examined the potential link between benzene exposure and risk of lung cancer using a systematic review of epidemiological studies and meta-analysis. We searched through PubMed, Web of Science and Scopus databases up to 10 February 2023 to identify all articles on the association between benzene exposure and lung cancer (incidence or prevalence) and/or mortality. We extracted the risk estimates of the highest and the lowest reported categories of benzene exposure and conducted a meta-analysis using a random-effects model. Heterogeneity and publication bias were analysed using an I2 test and funnel plots asymmetry, respectively. Twenty-one studies were included in the final analysis, with a total of 10,750 lung cancer cases and 2899 lung cancer deaths. Overall, risk estimates of lung cancer prevalence and mortality in association with benzene exposure were 1.20 (n = 14; 95% CI 1.05-1.37) and 1.15 (n = 13; 95% CI 1.02-1.30), respectively. In all cases, heterogeneity was quite large, while no significant publication bias was observed. When only studies that adjusted for smoking habit were selected, the risk for lung cancer increased by up to 34% (n = 9; 95% CI 1.10-1.64). Our data, which show a strong association between benzene exposure and lung cancer risk, may have important public health implications. However, further studies are needed to identify the lung cancer risk associated with benzene exposure considering different smoking conditions.

Adherence to Data-Driven Dietary Patterns and Lung Cancer Risk: A Systematic Review and Dose-Response Meta-Analysis.

The effect of dietary patterns on lung cancer risk is currently debated. In this study, we evaluated the association between different "a posteriori" dietary patterns and lung cancer risk. The search was carried out (February 2023) through Scopus, Web of Science, and PubMed databases. Meta-analysis was performed by a random-effects model using risk values (RR and OR) extracted from the 12 selected studies. Two main dietary patterns were identified and named "Western/meat" and "Healthy/prudent". The highest adherence to the "Western/meat" dietary pattern significantly increased the lung cancer risk (OR = 1.39; 95% CI: 1.17-1.65; p = 0.0002) while the highest adherence to the "Healthy/prudent" pattern reduced it (OR = 0.65; 95% CI: 0.51-0.83; p = 0.001). A linear trend between both dietary patterns and lung cancer risk was observed. However, a statistically significant inverse dose-response trend was found only for the "Healthy/prudent" dietary pattern (regression coefficient = -0.0031, p = 0.003). Subgroup analyses showed that the "Western/meat" pattern significantly increased the lung cancer risk in former (n = 4) (OR = 1.93, 95% CI: 1.11-3.36) and current smokers (n = 7) (OR = 1.35, 95% CI: 1.06-1.71). Similarly, the "Healthy/prudent" pattern exerts a protective effect on former (n = 4) (OR = 0.61, 95% CI: 0.44-0.85) and current smokers (n = 8) (OR = 0.64, 95% CI: 0.46-0.88). For both dietary patterns, no significant effect was observed on never-smokers.

Hysterectomy and thyroid cancer risk: A systematic review and meta-analysis.

Background: Incidence rates of thyroid cancer have increased. Recent studies findings suggest that women who underwent a hysterectomy have an elevated relative risk of thyroid cancer. The aim of our meta-analysis is to summarize the evidence about the association between hysterectomy and thyroid cancer risk. Methods: PubMed, Web of Science, and Scopus database were searched for studies published up to 5 September 2023. The PRISMA statement was followed. Heterogeneity was explored with Q statistic and the I2 statistic. Publication bias was assessed with Begg's and Egger's tests. Results: Sixteen studies met the criteria. The pooled analysis showed a significantly 64% increment of thyroid cancer risk in association with any hysterectomy (OR 1.64, 95% CI 1.48-1.81; I2 = 28.68%, p = 0.156). Hysterectomy without oophorectomy was a stronger predictor of risk than hysterectomy with oophorectomy. The pooled analysis of data regarding hysterectomy without oophorectomy showed a statistically significant increment of thyroid cancer risk by 59%. Hysterectomy with oophorectomy was associated with an increase of thyroid cancer risk of 39% (OR 1.39, 95% CI 1.16-1.67; I2 = 42.10%, p = 0.049). Significant publication bias was not detected. Conclusions: Our findings help with decision making around these surgeries.

Epigenetic Modifications Induced by Olive Oil and Its Phenolic Compounds: A Systematic Review.

Many studies demonstrated that olive oil (especially extra virgin olive oil: EVOO) phenolic compounds are bioactive molecules with anti-cancer, anti-inflammatory, anti-aging and neuroprotective activities. These effects have been recently attributed to the ability of these compounds to induce epigenetics modifications such as miRNAs expression, DNA methylation and histone modifications. In this study, we systematically review and discuss, following the PRISMA statements, the epigenetic modifications induced by EVOO and its phenols in different experimental systems. At the end of literature search through "PubMed", "Web of Science" and "Scopus", 43 studies were selected.Among them, 22 studies reported data on miRNAs, 15 on DNA methylation and 13 on histone modification. Most of the "epigenomic" changes observed in response to olive oil phenols' exposure were mechanistically associated with the cancer preventive and anti-inflammatory effects. In many cases, the epigenetics effects regarding the DNA methylation were demonstrated for olive oil but without any indication regarding the presence or not of phenols. Overall, the findings of the present systematic review may have important implications for understanding the epigenetic mechanisms behind the health effects of olive oil. However, generally no direct evidence was provided for the causal relationships between epigenetics modification and EVOO health related effects. Further studies are necessary to demonstrate the real physiological consequences of the epigenetics modification induced by EVOO and its phenolic compounds.

Hydroxytyrosol Exerts Anti-Inflammatory and Anti-Oxidant Activities in a Mouse Model of Systemic Inflammation.

Hydroxytyrosol (3,4-dihydroxyphenil-ethanol, HT), the major phenol derived from olive oil consumption, has shown different anti-inflammatory and anti-oxidant activities in vitro which may explain the chronic-degenerative diseases preventive properties of olive oil. The aim of this study was to examine the ability of HT reduce inflammatory markers, Cyclooxygenase-2 (COX2) and Tumour Necrosis Factor alfa (TNF-α and oxidative stress in vivo on a mouse model of systemic inflammation. Balb/c mice were pre-treated with HT (40 and 80 mg/Kg b.w.) and then stimulated by intraperitoneal injection of lipopolysaccharide (LPS). Blood was collected to measure COX2 gene expression by qPCR and TNF-α level by ELISA kit in plasma. In addition, the total anti-oxidant power of plasma and the DNA damage were measured by FRAP test and COMET assay, respectively. LPS increased the COX2 expression, the TNF-α production and the DNA damage. HT administration prevented all LPS-induced effects and improved the anti-oxidant power of plasma. HT demonstrated in vivo anti-inflammatory and anti-oxidant abilities. The results may explain the health effects of olive oil in Mediterranean diet. HT represents an interesting molecule for the development of new nutraceuticals and functional food useful in chronic diseases prevention.

Genotoxicity of heterocyclic amines (HCAs) on freshly isolated human peripheral blood mononuclear cells (PBMC) and prevention by phenolic extracts derived from olive, olive oil and olive leaves.

In this study we investigated the genotoxic potential of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, (PhIP); 2-amino-3-methyl-3H-imidazo[4,5-f]quinoline, (IQ); 2-amino-3,8-dimethyl-imidazo[4,5-f]quinoxaline, (MeIQx) and 2-amino-3,4,8-trimethyl-3H-imidazo[4,5-f]quinoxaline (DiMeIQx) on human freshly isolated peripheral blood mononuclear cells (PBMC) by the comet assay. The preventive ability of three different phenolic extracts derived from olive (O-PE), virgin olive oil (OO-PE) and olive leaf (OL-PE) on PhIP induced DNA damage was also investigated. PhIP and IQ induced a significant DNA damage at the lowest concentration tested (100 µM), while the genotoxic effect of MeIQx and DiMeIQx become apparent only in the presence of DNA repair inhibitors Cytosine b-D-arabinofuranoside and Hydroxyurea (AraC/HU). The inclusion of metabolic activation (S9-mix) in the culture medium increased the genotoxicity of all HCAs tested. All three phenolic extracts showed an evident DNA damage preventive activity in a very low concentration range (0.1-1.0 µM of phenols) which could be easily reached in human tissues "in vivo" under a regular intake of virgin olive oil. These data further support the observation that consumption of olive and virgin olive oil may prevent the initiation step of carcinogenesis. The leaf waste could be an economic and simple source of phenolic compounds to be used as food additives or supplements.

Overview of the Biological Activities of a Methanol Extract from Wild Red Belt Conk, Fomitopsis pinicola (Agaricomycetes), Fruiting Bodies from Central Italy.

Fomitopsis pinicola (Sw.) P. Karst. (Fomitopsidaceae) is a medicinal mushroom with a variety of healthy properties. In this study we tested the radical scavenging activity and antimicrobial and anticancer potential of methanol extracts of F. pinicola from central Italy. Molecular identification confirmed that the samples were F. pinicola; a Basic Local Alignment Search Tool search showed a close match (99% sequence identity) with European isolates of this species. The free radical scavenging capacities, measured by DPPH assay, showed that the extract activity was 3.5% that of Trolox. The MTT test, evaluated after 72 hours of treatment with increasing doses of extract (5-500 µg · mL-1), considerably inhibited proliferation in a dose-dependent manner in 2 human tumor cell lines. This reduction was coupled with a relevant induction of apoptosis in the human leukemia THP-1 cell line after 24 hours of treatment, but a relevant toxic effect occurred in the human colon adenocarcinoma HT29 cell line. The genotoxic potential of the methanol extracts was studied by single-cell gel electrophoresis of normal human leukocytes exposed to 20 µg extract at 37°C for 30 minutes; no DNA damage was observed. The F. pinicola methanol extract was found to have varying degrees of antifungal effects against the pathogenic fungi tested (minimum inhibitory concentration from 23.63 to 66.81 µg · mL-1). The results show that the tested F. pinicola extract has strong antimicrobial and chemo-preventive activities, but is a poor antioxidant.

Nutrigenomics of extra-virgin olive oil: A review.

Nutrigenomics data on the functional components of olive oil are still sparse, but rapidly increasing. Olive oil is the main source of fat and health-promoting component of the Mediterranean diet. Positive effects have been observed on genes involved in the pathobiology of most prevalent age- and lifestyle-related human conditions, such as cancer, cardiovascular disease and neurodegeneration. Other effects on health-promoting genes have been identified for bioactive components of olives and olive leafs. Omics technologies are offering unique opportunities to identify nutritional and health biomarkers associated with these gene responses, the use of which in personalized and even predictive protocols of investigation, is a main breakthrough in modern medicine and nutrition. Gene regulation properties of the functional components of olive oil, such as oleic acid, biophenols and vitamin E, point to a role for these molecules as natural homeostatic and even hormetic factors with applications as prevention agents in conditions of premature and pathologic aging. Therapeutic applications can be foreseen in conditions of chronic inflammation, and particularly in cancer, which will be discussed in detail in this review paper as major clinical target of nutritional interventions with olive oil and its functional components. © 2016 BioFactors, 43(1):17-41, 2017.

Oleuropein Prevents Azoxymethane-Induced Colon Crypt Dysplasia and Leukocytes DNA Damage in A/J Mice.

Previous studies have shown that the precursor of olive oil secoiridoids, Oleuropein (OL) has several in vitro chemopreventive properties. OL inhibits proliferation and induces apoptosis in breast, thyroid, prostate, and colorectal cancer (CRC) cells. Much less is known about the effects of OL on animal models of carcinogenesis. In this study, we investigated the ability of OL to prevent the azoxymethane (AOM)-induced colon cancer upset and DNA damage in mice. Animals, fed with a basal diet either enriched or not with OL (125 mg/kg), were injected with AOM (10 mg/kg, once a week for 6 weeks) and sacrificed after either 7 weeks for histological analysis of colon crypt dysplasia and evaluation of DNA damage in leukocytes or 17 weeks for counting the macroscopically observable colon tumors. An OL-enriched diet prevented the AOM-induced preneoplastic lesions in different colon segments, reducing the severity of crypt dysplasia and DNA damage in peripheral leukocytes. In addition, OL significantly reduced the AOM-induced tumor incidence from 57% to 14% (P < .05, chi-square test) in the medial colon segment. This study shows that OL is able to prevent CRC and DNA damage in mice treated with the carcinogen AOM. These results stimulate further human cancer prevention studies with OL-enriched food supplements that are actually available on the market.

Apple intake and cancer risk: a systematic review and meta-analysis of observational studies.

OBJECTIVE: Conflicting results on the association between fruit consumption and cancer risk have been reported. Little is known about the cancer preventive effects of different fruit types. The present meta-analysis investigates whether an association exists between apple intake and cancer risk. DESIGN: Relevant observational studies were identified by literature search (PubMed, Web of Science and Embase). A random-effect model was used to estimate the cancer risk in different anatomical sites. Between-study heterogeneity and publication bias were assessed using adequate statistical tests. RESULTS: Twenty case-control (three on lung, five on colorectal, five on breast, two on oesophageal, three on oral cavity, two on prostate and one each on pancreas, bladder, larynx, ovary, kidney and brain cancer) and twenty-one cohort (seven on lung, two on colorectal, three on breast and one each on oesophageal, pancreas, bladder, kidney, endometrial, head-neck, urothelial and stomach cancer) studies met the inclusion criteria. Comparing the highest v. lowest level of apple consumption, the reduction of lung cancer risk was statistically highly significant in both case-control (OR=0·75; 95% CI 0·63, 0·88; P=0·001, I 2=0 %) and cohort studies (relative risk=0·89; 95% CI 0·84, 0·94; P<0·001, I 2=53 %). Instead, in the case of colorectal (OR=0·66; 95% CI 0·54, 0·81; P<0·001, I 2=55%), breast (OR=0·79; 95% CI 0·73, 0·87; P<0·001, I 2=1 %) and overall digestive tract (OR=0·50; 95% CI 0·36, 0·69; P<0·001, I 2=90 %) cancers a significant preventive effect of apples was found only in case-control studies while prospective studies indicated no effect. No evidence of publication bias could be detected for colorectal, oral cavity, oesophageal and breast cancer. However, some confounding effects may be present and related to the consumption of other fruit which have not been considered as adjusting factors. CONCLUSIONS: The present meta-analysis indicates that consumption of apples is associated with a reduced risk of cancer in different anatomical sites.

In vitro chemo-preventive activities of hydroxytyrosol: the main phenolic compound present in extra-virgin olive oil.

The co-incubation in the culture medium with hydroxytyrosol [3,4-dihydroxyphenyl ethanol (3,4-DHPEA)], the main phenolic compound present in extra-virgin olive oil, and H2O2 reduces the oxidative DNA damage in peripheral blood mononuclear cells (PBMC). In this study we investigate, by the comet assay, the ability of 3,4-DHPEA to inhibit the H2O2 induced DNA damage when pre-incubated with PBMC and then removed before the exposure of cells to H2O2. Low doses of 3,4-DHPEA (10-100 µM) pre-incubated for 30 min with PBMC reduced the DNA damage induced by the treatment with H2O2 200 µM for 5 min at 4 °C. Prolonging the exposure time up to 6 h completely prevented the DNA damage. Furthermore we extensively analysed, by the MTT assay, the anti-proliferative activities of 3,4-DHPEA on breast (MDA and MCF-7), prostate (LNCap and PC3) and colon (SW480 and HCT116) cancer cell lines and correlated these effects with the H2O2 accumulation. The concentration of H2O2 in the culture medium was measured by the ferrous ion oxidation-xylenol orange method. The proliferation of all the cell lines was inhibited but at different levels: the prostate cancer cells were more resistant to the growth inhibition with respect to breast and colon cancer cells. The ability of the different cell lines to remove H2O2 from the culture medium was inversely correlated with their sensitivity to the anti-proliferative effect of 3,4-DHPEA. Therefore, 3,4-DHPEA may act as a chemopreventive agent acting on both initiation and promotion/progression phases of carcinogenesis.

Preventive activity of olive oil phenolic compounds on alkene epoxides induced oxidative DNA damage on human peripheral blood mononuclear cells.

The aim of this study was to investigate the ability of epoxides of styrene (styrene-7,8-oxide; SO) and 1,3-butadiene (3,4-epoxy-1-butene; 1,2:3,4:-diepoxybutane) to cause oxidative stress and oxidative DNA damage on human peripheral blood mononuclear cells (PBMCs) and whether a complex mixture of olive oil phenols (OOPE) could prevent these effects. The DNA damage was measured by the single-cell gel electrophoresis (SCGE; comet assay). We found that the DNA damage induced by alkene epoxides could be prevented by N-acetyl-cysteine (10 mM) and catalase (100 U/ml). Alkene epoxides caused a significant (P < 0.05) increase of both peroxide concentration in extra- and intracellular environment and formamidopyrimidine DNA glycosylase (FPG)- and Endonuclease III (ENDO III)-sensitive sites in PBMCs, demonstrating the presence of oxidized bases. OOPE (1 µg of total phenols/ml) was able to prevent the alkene epoxide induced DNA damage both after 2 and 24 h of incubation. In addition, OOPE completely inhibited the SO-induced intracellular peroxide accumulation in PBMCs and prevented the oxidative DNA damage induced by SO, as evidenced by the disappearance of both FPG- and ENDO III-sensitive sites. This is the first study demonstrating the ability of OOPE to prevent the DNA damage induced by alkene epoxides providing additional information about the chemopreventive properties of olive oil.

Effect of olive oil phenols on the production of inflammatory mediators in freshly isolated human monocytes.

Recent in vitro and in vivo studies suggest that the anti-inflammatory properties of extra virgin olive oil may be involved in the prevention of chronic degenerative diseases. In this study, the ability of olive oil phenols to influence the release of superoxide anions (O2-), prostaglandin E2 (PGE2) and tumor necrosis factor α (TNFα) and the expression of cyclooxygenase2 (COX2) in human monocytes, freshly isolated from healthy donors, was investigated. O2- were measured by superoxide dismutase-inhibitable cytochrome c reduction and PGE2 and TNFα production were determined in culture medium with appropriate enzyme immunoassay kits. COX2 mRNA and protein were evaluated by quantitative reverse transcription-polymerase chain reaction and Western immunoblotting, respectively. Treatment of monocytes for 24 h with 100 µM of hydroxytyrosol (3,4-DHPEA), tyrosol (p-HPEA) and their secoiridoid derivatives (3,4-DHPEA and p-HPEA linked to the dialdehydic form of elenolic acid: 3,4-DHPEA-EDA and p-HPEA-EDA, respectively) significantly (P<.05) inhibited the production of O2(-) as follows: 3,4-DHPEA (40%,), p-HPEA (9%), 3,4-DHPEA-EDA (25%) and p-HPEA-EDA (36%). Hydroxytyrosol also considerably reduced the expression of COX2 at both the mRNA and protein level (P<.05) and caused a clear dose-dependent reduction of PGE2 released into the culture medium (45% and 71% at 50 and 100 µM, respectively, P<.05). The COX2 mRNA was also efficiently inhibited by the secoiridoids. Moreover, it was shown that hydroxytyrosol increased the monocytes TNFα production. In addition to other chemopreventive properties, these results suggest that the health effects of olive oil phenols may be related to their ability to modulate the production of pro-inflammatory molecules, a property common to non-steroidal anti-inflammatory drugs.

Genotoxicity of alkene epoxides in human peripheral blood mononuclear cells and HL60 leukaemia cells evaluated with the comet assay.

Volatile organic compounds (VOCs) exert their carcinogenic activity through the production of epoxide metabolites. Because of their high reactivity some epoxides are also produced in the chemical industry for the synthesis of other compounds. Therefore, human exposure to VOCs epoxides does occur and may be an important human health concern. In this study, the in vitro genotoxic potential of epoxides originating from 1,3-butadiene (3,4-epoxy-1-butene: EB; 1,2:3,4-diepoxybutane: DEB), isoprene (3,4-epoxy-2-methyl-1-butene: IO), styrene (styrene-7,8-oxide: SO), propylene (propylene oxide: PO) and 1-butene (1,2-epoxy-butane: BO) in human peripheral blood mononuclear cells (PBMCs) and promyelocytic leukaemia cells (HL60) was measured with the comet assay (single-cell gel electrophoresis, SCGE). The effect of inclusion of foetal calf serum (FCS, 5%) in the cell-culture medium and different durations of exposure (2h, 24h) were also investigated. All epoxides tested produced DNA damage in a concentration range that did not reduce cell viability. HL60 cells were more resistant than PBMCs to the DNA damage induced by the different epoxides. With the exception of IO, the treatment for 24h resulted in an increase of DNA damage. FCS slightly protected PBMCs from the genotoxic effects induced by IO and BO, whilst no such effect was noted for the other compounds. Overall, the dose-dependent effects that were seen allowed us to define a genotoxicity scale for the different epoxides as follows: SO>EB>DEB>IO>PO>BO, which is in partial agreement with the International Agency for Research on Cancer (IARC) classification of the carcinogenic hazards.

Anti-proliferative and pro-apoptotic activities of hydroxytyrosol on different tumour cells: the role of extracellular production of hydrogen peroxide.

PURPOSE: Several recently published data suggest that the anti-proliferative and pro-apoptotic properties of hydroxytyrosol [3,4-dihydroxyphenyl ethanol (3,4-DHPEA)] on HL60 cells may be mediated by the accumulation of hydrogen peroxide (H2O2) in the culture medium. The aim of this study was to clarify the role played by H2O2 in the chemopreventive activities of 3,4-DHPEA on breast (MDA and MCF-7), prostate (LNCap and PC3) and colon (SW480 and HCT116) cancer cell lines and to investigate the effects of cell culture medium components and the possible mechanisms at the basis of the H2O2-producing properties of 3,4-DHPEA. METHODS: The proliferation was measured by the MTT assay and the apoptosis by both fluorescence microscopy and flow cytometry. The concentration of H2O2 in the culture medium was measured by the ferrous ion oxidation-xylenol orange method. RESULTS: It was found that the H2O2-inducing ability of 3,4-DHPEA is completely prevented by pyruvate and that the exposure of cells to conditions not supporting the H2O2 accumulation (addition of either catalase or pyruvate to the culture medium) inhibited the anti-proliferative effect of 3,4-DHPEA. Accordingly, the sensitivity of the different cell lines to the anti-proliferative effect of 3,4-DHPEA was inversely correlated with their ability to remove H2O2 from the culture medium. With regard to the mechanism by which 3,4-DHPEA causes the H2O2 accumulation, it was found that superoxide dismutase increased the H2O2 production while tyrosinase, slightly acidic pH (6,8) and absence of oxygen (O2) completely prevented this activity. In addition, different transition metal-chelating compounds did not modify the H2O2-producing activity of 3,4-DHPEA. CONCLUSIONS: The pro-oxidant activity of 3,4-DHPEA deeply influences its 'in vitro' chemopreventive activities. The main initiation step in the H2O2-producing activity is the auto-oxidation of 3,4-DHPEA by O2 with the formation of the semiquinone, superoxide ions (O2(-)) and 2H(+).

Influence of cultivar and concentration of selected phenolic constituents on the in vitro chemiopreventive potential of olive oil extracts.

One of the main olive oil phenolic compounds, hydroxytyrosol (3,4-DHPEA), exerts in vitro chemopreventive activities (antiproliferative and pro-apoptotic) on tumor cells through the accumulation of H(2)O(2) in the culture medium. However, the phenol composition of virgin olive oil is complex, and 3,4-DHPEA is present at low concentrations when compared to other secoiridoids. In this study, the in vitro chemopreventive activities of complex virgin olive oil phenolic extracts (VOO-PE, derived from the four Italian cultivars Nocellara del Belice, Coratina, Ogliarola, and Taggiasca) were compared to each other and related to the amount of the single phenolic constituents. A great chemopreventive potential among the different VOO-PE was found following this order: Ogliarola > Coratina > Nocellara > Taggiasca. The antiproliferative and pro-apoptotic activities of VOO-PE were positively correlated to the secoiridoid content and negatively correlated to the concentration of both phenyl alcohols and lignans. All extracts induced H(2)O(2) accumulation in the culture medium, but this phenomenon was not responsible for their pro-apoptotic activity. When tested in a complex mixture, the olive oil phenols exerted a more potent chemopreventive effect compared to the isolated compounds, and this effect could be due either to a synergistic action of components or to any other unidentified extract constituent.

Genotoxic effect of bile acids on human normal and tumour colon cells and protection by dietary antioxidants and butyrate.

BACKGROUND: Colorectal cancer is the second cause of death for tumour worldwide. Among the risk factors for this disease the dietary habits seem to have a pivotal role. An elevated intake of fats causes a high release in the gut lumen of bile acids that are positively correlated with colorectal cancer, since they act as detergents and proliferation promoters. Recently, it was evidenced that bile acids can also be able to induce DNA damage. AIM OF THE STUDY: In this study the genotoxicity of deoxycholic acid (DCA) and chenodeoxycholic acid CDCA) has been evaluated in human normal colonocytes derived from 60 colon biopsies and in tumour cells. The involvement of reactive oxygen species (ROS) and the oxidative DNA damage was assessed. In addition, the protective effect exerted by both two well-known antioxidants commonly present in the diet, beta-carotene and alpha-tocopherol, and butyrate which is known to be involved in the regulation of several cellular functions, has also been tested. METHODS: The DNA damage was evaluated by the "comet assay" or single cell gel electrophoresis (SCGE) both in its conventional use and by the Endonuclease III modified method, which allow to detect the presence of oxidized pyrimidines. RESULTS: Bile acids (CDA and CDCA) resulted genotoxic on both normal and tumour human colon cells. The inclusion of the endonuclease III digestion step in the comet assay demonstrated that bile acids induced an oxidative DNA damage. In addition, treatment of colonocytes with bile acids in the presence of the antioxidants (beta-carotene, alpha-tocopherol) and Na-butyrate caused a reduction of DNA damage. CONCLUSION: Our results suggest that bile acids may be involved in the tumour initiation by inducing a DNA oxidative damage, and so add further evidences to the preventive properties of antioxidants present in the Mediterranean diet.

Oxidative DNA damage is prevented by extracts of olive oil, hydroxytyrosol, and other olive phenolic compounds in human blood mononuclear cells and HL60 cells.

Our aim in this study was to provide further support to the hypothesis that phenolic compounds may play an important role in the anticarcinogenic properties of olive oil. We measured the effect of olive oil phenols on hydrogen peroxide (H(2)O(2))-induced DNA damage in human peripheral blood mononuclear cells (PBMC) and promyelocytic leukemia cells (HL60) using single-cell gel electrophoresis (comet assay). Hydroxytyrosol [3,4-dyhydroxyphenyl-ethanol (3,4-DHPEA)] and a complex mixture of phenols extracted from both virgin olive oil (OO-PE) and olive mill wastewater (WW-PE) reduced the DNA damage at concentrations as low as 1 micromol/L when coincubated in the medium with H(2)O(2) (40 micromol/L). At 10 micromol/L 3,4-DHPEA, the protection was 93% in HL60 and 89% in PBMC. A similar protective activity was also shown by the dialdehydic form of elenoic acid linked to hydroxytyrosol (3,4-DHPEA-EDA) on both kinds of cells. Other purified compounds such as isomer of oleuropein aglycon (3,4-DHPEA-EA), oleuropein, tyrosol, [p-hydroxyphenyl-ethanol (p-HPEA)] the dialdehydic form of elenoic acid linked to tyrosol, caffeic acid, and verbascoside also protected the cells against H(2)O(2)-induced DNA damage although with a lower efficacy (range of protection, 25-75%). On the other hand, when tested in a model system in which the oxidative stress was induced by phorbole 12-myristate 13-acetate-activated monocytes, p-HPEA was more effective than 3,4-DHPEA in preventing the oxidative DNA damage. Overall, these results suggest that OO-PE and WW-PE may efficiently prevent the initiation step of carcinogenesis in vivo, because the concentrations effective against the oxidative DNA damage could be easily reached with normal intake of olive oil.

Inhibition of cell cycle progression by hydroxytyrosol is associated with upregulation of cyclin-dependent protein kinase inhibitors p21(WAF1/Cip1) and p27(Kip1) and with induction of differentiation in HL60 cells.

Recent evidence indicates that the cancer preventive activity of olive oil can be mediated by the presence of minor components, such as antioxidant phenolic compounds. However, their mechanisms of action remain largely unknown. In this study, we investigated the in vitro effects of one of the main olive oil phenols, hydroxytyrosol [3,4-dihydroxyphenylethanol (3,4-DHPEA)], on proliferation, cell cycle progression, apoptosis, and differentiation of HL60 human promyelocytic leukemia cells. 3,4-DHPEA showed a potent inhibitory activity on DNA synthesis, as evidenced by a 92% reduction of [3H]-thymidine incorporation at 100 micromol/L, and an induced apoptosis, as evidenced by the release of cytosolic nucleosomes and flow cytometry. This phenol, 3,4-DHPEA, was also able to inhibit the progression of the cell cycle in synchronized HL60 cells, which accumulated in the G0/G1 phase of the cell cycle after 25 h of treatment. Furthermore, 3,4-DHPEA induced differentiation on HL60 cells with a maximum effect (22% of cells) at 100 micromol/L after 72 h of treatment. Among the different proteins involved in the regulation of the cell cycle, 3,4-DHPEA reduced the level of cyclin-dependent kinase (CDK) 6 and increased that of cyclin D3. With regard to the CDK inhibitors, p15 was not altered by 3,4-DHPEA treatment, whereas the expression of p21(WAF1/Cip1) and p27(Kip1) was increased at both protein and mRNA levels. To our knowledge, these results provide the first evidence that 3,4-DHPEA may effect the expression of genes involved in the regulation of tumor cell proliferation and differentiation.

DNA-damaging ability of isoprene and isoprene mono-epoxide (EPOX I) in human cells evaluated with the comet assay.

Isoprene is produced in combustion processes and is widely used as an industrial chemical. It is a natural product emitted by plants and endogenously produced by humans and other mammals. Therefore, exposure to isoprene from both endogenous and exogenous sources is unavoidable and occurs during the entire human life. Based on evaluations of the International Agency for Research on Cancer (IARC), isoprene has been classified in Group 2B (possibly carcinogenic to humans). In the present work, we have demonstrated, by use of the single-cell gel electrophoresis assay (SCGE or comet assay), that isoprene is able to induce DNA damage in peripheral blood mononuclear cells (PBMCs) in the presence of metabolic activation. In addition, treatment of cells with the main isoprene mono-epoxide (EPOX I) induced time- and dose- dependent DNA damage in both PBMCs and human leukaemia cells (HL60). The metabolic activation system, represented by rat liver post-mitochondrial fractions (S9), was obtained from rats that had been treated - or not - with inducing agents such as phenobarbital and ethanol. The inclusion of S9 fractions (4mg protein/mL) from non-induced or phenobarbital-induced rats resulted in a statistically significant enhancement of isoprene genotoxicity. A different pattern was obtained by the addition of ethanol-induced S9, which appeared highly genotoxic by itself even in the absence of isoprene. Reducing the concentration of ethanol-induced S9 to 0.25mg protein/mL resulted in a considerable enhancement of isoprene genotoxicity. In the absence of clear epidemiological evidence of the carcinogenicity of isoprene in humans, the results of this study seem to be particularly important since they add new findings to support the classification of this chemical as possibly carcinogenic to humans.