The role of reactive oxygen species and subsequent DNA-damage response in the emergence of resistance towards resveratrol in colon cancer models.

In spite of the novel strategies to treat colon cancer, mortality rates associated with this disease remain consistently high. Tumour recurrence has been linked to the induction of resistance towards chemotherapy that involves cellular events that enable cancer cells to escape cell death. Treatment of colon cancer mainly implicates direct or indirect DNA-damaging agents and increased repair or tolerances towards subsequent lesions contribute to generate resistant populations. Resveratrol (RSV), a potent chemosensitising polyphenol, might share common properties with chemotherapeutic drugs through its indirect DNA-damaging effects reported in vitro. In this study, we investigated how RSV exerts its anticancer effects in models of colon cancer with a particular emphasis on the DNA-damage response (DDR; PIKKs-Chks-p53 signalling cascade) and its cellular consequences. We showed in vitro and in vivo that colon cancer models could progressively escape the repeated pharmacological treatments with RSV. We observed for the first time that this response was correlated with transient activation of the DDR, of apoptosis and senescence. In vitro, a single treatment with RSV induced a DDR correlated with S-phase delay and apoptosis, but prolonged treatments led to transient micronucleations and senescence phenotypes associated with polyploidisation. Ultimately, stable resistant populations towards RSV displaying higher degrees of ploidy and macronucleation as compared to parental cells emerged. We linked these transient effects and resistance emergence to the abilities of these cells to progressively escape RSV-induced DNA damage. Finally, we demonstrated that this DNA damage was triggered by an overproduction of reactive oxygen species (ROS) against which cancer cells could adapt under prolonged exposure to RSV. This study provides a pre-clinical analysis of the long-term effects of RSV and highlights ROS as main agents in RSV's indirect DNA-damaging properties and consequences in terms of anticancer response and potent resistance emergence.

Resveratrol, a phytochemical inducer of multiple cell death pathways: apoptosis, autophagy and mitotic catastrophe.

Cancers are the largest cause of mortality and morbidity in industrialized countries. In the field of the medicinal chemistry of natural products, numerous studies have reported interesting properties of trans-resveratrol as a chemopreventing agent against cancers, inflammation, and viral infection. Tumor growth inhibition has been linked to the ability of resveratrol to arrest cell cycle progression and to trigger cell death. This review focuses on the pathways that mediate resveratrol-induced cell death. Resveratrol impacts on the mitochondrial functions (respiratory chain, oncoproteins, gene expression, etc), in which p53 protein can be involved and its acetylated or phosphorylated forms. This polyphenol also affects death receptor distribution in ceramide-enriched membrane platforms which serve to trap and cluster receptor molecules, and facilitates the formation of a death-inducing signaling complex in the cell. To induce apoptosis, resveratrol also activates the ceramide / sphingomyelin pathway, which promotes ceramide generation and the downstream activation of kinase cascades. Resveratrol can activate alternative pathways to cell death such as those leading to autophagy, senescence or mitotic catastrophe. Furthermore, numerous attempts have been made using resveratrol analogs to improve the molecule's ability to block cell proliferation and induce cell death. Moreover, structural modification of natural phenolics is expected to produce analogs that may be useful tools to study the structure-activity relationships. Lastly, in various cancer types, resveratrol behaves as a chemosensitizer that lowers the threshold of cell death induction by classical anticancer agents and counteracts tumor cell chemoresistance.

Effects of oxysterols on cell viability, inflammatory cytokines, VEGF, and reactive oxygen species production on human retinal cells: cytoprotective effects and prevention of VEGF secretion by resveratrol.

BACKGROUND AND AIMS: Oxysterols are assumed to play important roles in age-related macular degeneration, a major cause of blindness. So we characterized the cytotoxic, oxidative, inflammatory, and angiogenic activities of oxysterols (7ß-hydroxycholesterol (7ß-OH), 7-ketocholesterol (7KC), 25-hydroxycholesterol (25-OH)) in human retinal ARPE-19 cells, and evaluated the protective effects of resveratrol (Rsv: 1 µM), a polyphenol from red wine. METHODS: ARPE-19 cells were treated with 7ß-OH, 7KC, or 25-OH (5-40 µg/mL; 24-48 h) without or with Rsv. Cell viability was determined using trypan blue and the MTT assay. Cell death was characterized by electron microscopy and in situ detection of activated caspases with fluorochrome-labeled inhibitors of caspases. Reactive oxygen species (ROS) production was measured with hydroethidine. ELISA methods and a cytometric bead assay were used to quantify cytokines involved in inflammation (IL-8, IL-1ß, IL-6, IL-10, IL-12p70, TNF-α, MCP-1) and VEGF. RESULTS: 7ß-OH and 7KC triggered a caspase-independent cell death process associated with the presence of multilamellar cytoplasmic structures evocating phospholipidosis, increased ROS production, and IL-8 secretion. 7ß-OH enhanced VEGF secretion. No cytotoxic effects were identified with 25-OH, which highly stimulated ROS production, MCP-1, and VEGF secretion. With oxysterols, no IL-10, TNF-α, and IL-12p70 secretion were detected. 25-OH induced IL-8 secretion through the MEK/ERK½ signaling pathway, and Rsv showed cytoprotective activities and inhibited VEGF secretion. CONCLUSION: 7ß-OH, 7KC, and 25-OH have cytotoxic, oxidative, inflammatory, and/or angiogenic activities on ARPE-19 cells. As Rsv has some protective effects against oxysterol-induced cell death and VEGF secretion it could be valuable in ARMD treatment.

Obstetric interventions for babies born before 28 weeks of gestation in Europe: results of the MOSAIC study.

OBJECTIVE: To describe obstetric intervention for extremely preterm births in ten European regions and assess its impact on mortality and short term morbidity. DESIGN: Prospective observational cohort study. SETTING: Ten regions from nine countries participating in the 'Models of Organising Access to Intensive Care for Very Preterm Babies in Europe' (MOSAIC) project. POPULATION: All births from 22 to 29 weeks of gestation (n = 4146) in 2003, excluding terminations of pregnancy. METHODS: Comparison of three obstetric interventions (antenatal corticosteroids, antenatal transfer and caesarean section for fetal indication) rates at 22-23, 24-25 and 26-27 weeks to that at 28-29 weeks and the association of the level of intervention with pregnancy outcome. MAIN OUTCOME MEASURES: Use of antenatal corticosteroids, antenatal transfer and caesarean section by two-week gestational age groups as well as a composite score of these three interventions. Outcomes included stillbirth, in-hospital mortality and intraventricular haemorrhage (IVH) grades III and IV and/or periventricular leucomalacia (PVL) and bronchopulmonary dysplasia (BPD). RESULTS: There were large differences between regions in interventions for births at 22-23 and 24-25 weeks. Differences were most pronounced at 24-25 weeks; in some regions these babies received the same care as babies of 28-29 weeks, whereas elsewhere levels of intervention were distinctly lower. Before 26 weeks and especially at 24-25 weeks, there was an association between the composite intervention score and mortality. No association was observed at 26-27 weeks. For survivors at 24-25 weeks, the intervention score was associated with higher rates of BPD, but not with IVH or PVL. CONCLUSIONS: There are large differences between European regions in obstetric practices at the lower limit of viability and these are related to outcome, especially at 24-25 weeks.

Organisation of obstetric services for very preterm births in Europe: results from the MOSAIC project.

OBJECTIVE: To study the impact of the organisation of obstetric services on the regionalisation of care for very preterm births. DESIGN: Cohort study. SETTING: Ten European regions covering 490 000 live births. POPULATION: All children born in 2003 between 24 and 31 weeks of gestation. METHOD: The rate of specialised maternity units per 10 000 total births, the proportion of total births in specialised units and the proportion of very preterm births by referral status in specialised units were compared. MAIN OUTCOME MEASURE: Birth in a specialised maternity unit (level III unit or unit with a large neonatal unit (at least 50 annual very preterm admissions). RESULTS: The organisation of obstetric care varied in these regions with respect to the supply of level III units (from 2.3 per 10 000 births in the Portuguese region to 0.2 in the Polish region), their characteristics (annual number of deliveries, 24 hour presence of a trained obstetrician) and the proportion of all births (term and preterm) that occur in these units. The proportion of very preterm births in level III units ranged from 93 to 63% in the regions. Different approaches were used to obtain a high level of regionalisation: high proportions of total deliveries in specialised units, high proportions of in utero transfers or high proportions of high-risk women who were referred to a specialised unit during pregnancy. CONCLUSION: Consensus does not exist on the optimal characteristics of specialised units but regionalisation may be achieved in different models of organisation of obstetric services.

Potential effect of freshwater virus on the structure and activity of bacterial communities in the Marennes-Oléron Bay (France).

Batch culture experiments using viral enrichment were conducted to test the response of a coastal bacterial community to autochthonous (i.e., co-existing) or allochthonous riverine viruses. The effects of viral infections on bacterial dynamics and activity were assessed by epifluorescence microscopy and thymidine incorporation, respectively, whereas the effect of viral infection on bacterial community composition was examined by polymerase chain reaction-single strand conformation polymorphism 16S ribosomal RNA fingerprinting. The percentages of high nucleic acid-containing cells, evaluated by flow cytometry, were significantly correlated (r2=0.91, n=12, p<0.0001) to bacterial production, making this value a good predictor of active cell dynamics along the study. While confinement and temperature were the two principal experimental factors affecting bacterial community composition and dynamics, respectively, additions of freshwater viruses had significant effects on coastal bacterial communities. Thus, foreign viruses significantly reduced net bacterial population increase as compared to the enrichment treated with inactivated virus. Moreover, freshwater viruses recurrently and specifically affected bacterial community composition, as compared to addition of autochthonous viruses. In most cases, the combined treatment viruses and freshwater dissolved organic matter helped to maintain or even enhance species richness in coastal bacterial communities in agreement to the 'killing the winner' hypothesis. Thus, riverine virus input could potentially influence bacterial community composition of the coastal bay albeit with modest modification of bulk bacterial growth.

Termination of pregnancy among very preterm births and its impact on very preterm mortality: results from ten European population-based cohorts in the MOSAIC study.

OBJECTIVE: To study the impact of terminations of pregnancy (TOP) on very preterm mortality in Europe. DESIGN: European prospective population-based cohort study. SETTING: Ten regions from nine European countries participating in the MOSAIC (Models of OrganiSing Access to Intensive Care for very preterm babies) study. These regions had different policies on screening for congenital anomalies (CAs) and on pregnancy termination. POPULATION OR SAMPLE: Births 22-31 weeks gestational age. METHODS: The analysis compares the proportion of TOP among very preterm births and assesses differences in mortality between the regions. MAIN OUTCOME MEASURES: Pregnancy outcomes (termination, antepartum death, intrapartum death and live birth) and reasons for termination, presence of CAs and causes of death for stillbirths and live births in 2003. RESULTS: Pregnancy terminations constituted between 1 and 21.5% of all very preterm births and between 4 and 53% of stillbirths. Most terminations were for CAs, although some were for obstetric indications (severe pre-eclampsia, growth restriction, premature rupture of membranes). TOP contributed substantially to overall fetal mortality rates in the two regions with late second-trimester screening. There was no clear association between policies governing screening and pregnancy termination and the proportion of CAs among stillbirths and live births, except in Poland, where neonatal deaths associated with CAs were more frequent, reflecting restrictive pregnancy termination policies. CONCLUSION: Proportions of TOP among very preterm births varied widely between European regions. Information on terminations should be reported when very preterm live births and stillbirths are compared internationally since national policies related to screening for CAs and the legality and timing of medical terminations differ.

Dynamic of virioplankton abundance and its environmental control in the Charente estuary (France).

The Charente River provides nutrient- and virus-rich freshwater input to the Marennes Oléron Basin, the largest oyster-producing region in Europe. To evaluate virioplankton distribution in the Charente Estuary and identify which environmental variables control dynamic of virioplankton abundance, five stations defined by a salinity gradient (0-0.5, 0.6-5, 13-17, 20-24, and higher than 30 PSU) were surveyed over a year. Viral abundance was related to bacterioplankton abundance and activities, photosynthetic pigments, nutrient concentration, and physical parameters (temperature and salinity). On a spatial scale, virus displayed a decreasing pattern seaward with abundance ranging over the sampling period from 1.4x10(7) to 20.8x10(7) viruses mL-1 making virioplankton the most abundant component of planktonic microorganisms in the Charente Estuary. A good correlation was found between viral and bacterial abundance (rs=0.85). Furthermore, bacterial abundance was the most important predictor of viral abundance explaining alone between 66% (winter) and 76% (summer) of viral variability. However, no relation existed between viral abundance and chlorophyll a. Temporal variations in viral distributions were mainly controlled by temperature through the control of bacterial dynamics. Spatial variations of viral abundance were influenced by hydrodynamic conditions especially during the winter season where virioplankton distribution was entirely driven by mixing processes.

Human hepatic cell uptake of resveratrol: involvement of both passive diffusion and carrier-mediated process.

This work reports significant advances on the transport in hepatic cells of resveratrol, a natural polyphenol with potent protective properties. First, we describe a new simple technique to qualitatively follow resveratrol cell uptake and intracellular distribution, based on resveratrol fluorescent properties. Second, the time-course study and the quantification of (3)H-labelled resveratrol uptake have been performed using human hepatic derived cells (HepG2 tumor cells) and hepatocytes. The temperature-dependence of the kinetics of uptake as well as the cis-inhibition experiments agree with the involvement of a carrier-mediated transport in addition to passive diffusion. The decrease of passive uptake resulting from resveratrol binding to serum proteins brings to light a mediated mechanism in physiological situation.

Inhibitory effect of resveratrol on the proliferation of human and rat hepatic derived cell lines.

Resveratrol is a polyphenolic compound especially produced by grapevine and consequently found in wine. Based on epidemiological studies resveratrol may act as a cancer chemopreventive compound. The ability of resveratrol to inhibit cell proliferation was studied in rat hepatoma Fao cell line and human hepatoblastoma HepG2 cell line. The results show that resveratrol strongly inhibits cell proliferation at the micromolar range in a time- and dose-dependent manner. Concentrations higher than 50 microM become toxic. Fao cells are more sensitive than HepG2 cells. Interestingly, the presence of ethanol lowers the threshold of resveratrol effect. Resveratrol appears to prevent or to delay the entry to mitosis since no inhibition of [3H]thymidine incorporation is observed, while there is an increase of cell number in S and G2/M phases. In conclusion, resveratrol shows a strong inhibition of hepatic cell proliferation where alcohol may act as an enhancing agent.

[Regulation of bacterial proteolytic activity at natural concentrations in dissolved organic matter in a maritime pond]. / Régulation de l'activité exoprotéolytique bactérienne aux concentrations naturelles en matière organique dissoute dans un marais maritime.

The regulation of the bacterial exoproteolytic activity, at natural substrate concentrations, was studied during the survey of an Atlantic coastal marine pond (France). The regulation of this activity occurs at two different levels: on the one hand, at the cellular level, the ectoenzyme synthesis is regulated by hydrolysis substrates, dissolved combined amino acids (DCAA), and end products, dissolved free amino acids (DFAA), in terms of the relative amounts available to the cell, and on the other hand, at the ecosystem level, i.e. the hydrolytic activity, by the total amounts of DCAA and DFAA in situ. The DFAA acts as an inhibitor in enzymatic synthesis; in contrast, dissolved proteins induce the enzymatic synthesis and the exoproteolytic activity. These results, obtained in natural concentration conditions, confirm the functioning in situ of the ectoenzymatic activity regulation model of Chróst, until now only validated in an enriched experimental medium.