Georgia Chalkiadaki

Persistent organic pollutants exposure during pregnancy, maternal gestational Weight gain, and birth outcomes in the mother-child cohort in Crete, Greece (RHEA study)

BACKGROUND Persistent organic pollutants (POPs), including polychlorinated biphenyls (PCBs) and pesticides bioaccumulate through the food chain and cross the placenta. POPs are developmental toxicants in animals but the epidemiological evidence on pregnancy outcomes is inconsistent. Maternal gestational weight gain has been recently suggested as a key factor explaining the association between PCBs with lower birth weight. AIMS We examined whether in utero exposure to current low levels of different POPs is associated with fetal growth and gestational age in a mother-child cohort in Crete, Greece (Rhea study), and evaluated specifically whether maternal gestational weight gain may affect this association. METHODS We included 1117 mothers and their newborns from the Rhea study. Mothers were interviewed and blood samples collected during the first trimester of pregnancy. Information on birth outcomes was retrieved from medical records. Concentrations of several PCBs, other organochlorine compounds (dichlorodiphenyl dichloroethene [DDE], dichlorodiphenyl trichloroethane [DDT] and hexachlorobenzene [HCB]) and one polybrominated diphenyl ether congener (tetra-bromodiphenyl ether [BDE-47]), were determined in maternal serum by triple quadrupole mass spectrometry. Multiple linear regression models were used to investigate the associations of birth weight, gestational age, and head circumference with each compound individually on the log10 scale, and with combined exposures through the development of an exposure score. RESULTS In multivariate models, birth weight was negatively associated with increasing levels of HCB (β=-161.1g; 95% CI: -296.6, -25.7) and PCBs (β=-174.1g; 95% CI: -332.4, -15.9); after further adjustment for gestational weight gain these estimates were slightly reduced (β=-154.3g; 95% CI: -300.8, -7.9 for HCB and β=-135.7g; 95% CI: -315.4, 43.9 for PCBs). Furthermore, in stratified analysis, the association between POPs and birth weight was only observed in women with inadequate or excessive gestational weight gain. Small, negative associations were observed with head circumference while no association was observed with gestational age. CONCLUSIONS The findings suggest that prenatal exposure to PCBs and HCB impairs fetal growth and adds to the growing literature that demonstrates an association between low-level environmental pollutant exposure and fetal growth. Furthermore our results suggest that the association of POPs, maternal gestational weight gain and birth weight is probably more complex than that previously hypothesized.

Association of early life exposure to bisphenol A with obesity and cardiometabolic traits in childhood

BACKGROUND Bisphenol A (BPA) is a chemical used extensively worldwide in the manufacture of plastic polymers. The environmental obesogen hypothesis suggests that early life exposure to endocrine disrupting chemicals such as BPA may increase the risk for wt gain later in childhood but few prospective epidemiological studies have investigated this relationship. OBJECTIVES We examined the association of early life BPA exposure with offspring obesity and cardiometabolic risk factors in 500 mother-child pairs from the RHEA pregnancy cohort in Crete, Greece. METHODS BPA concentrations were measured in spot urine samples collected at the 1st trimester of pregnancy) and from children at 2.5 and 4 years of age. We measured birth wt, body mass index (BMI) from 6 months to 4 years of age, waist circumference, skinfold thickness, blood pressure, serum lipids, C-reactive protein, and adipokines at 4 years of age. BMI growth trajectories from birth to 4 years were estimated by mixed effects models with fractional polynomials of age. Adjusted associations were obtained via multivariable regression analyses. RESULTS The prevalence of overweight/obesity was 9% at 2, 13% at 3% and 17% at 4 years of age. Geometric mean BPA concentrations were 1.2μg/g creatinine±7.9 in 1st trimester, 5.1μg/g±13.3 in 2.5 years and 1.9μg/g±4.9 in 4 years. After confounder adjustment, each 10-fold increase in BPA at 4 years was associated with a higher BMI z-score (adj. β=0.2; 95% CI: 0.01, 0.4), waist circumference (adj. β=1.2; 95% CI: 0.1, 2.2) and sum of skinfold thickness (adj. β=3.7mm; 95% CI: 0.7, 6.7) at 4 years. Prenatal BPA was negatively associated with BMI and adiposity measures in girls and positively in boys. We found no associations of early life exposure to BPA with other offspring cardiometabolic risk factors. CONCLUSIONS Prenatal BPA exposure was not consistently associated with offspring growth and adiposity measures but higher early childhood BPA was associated with excess child adiposity.

Lumican regulates osteosarcoma cell adhesion by modulating TGFβ2 activity.

Human osteosarcoma cell lines were recently shown to express and secrete the small leucine rich proteoglycan (SLRP) lumican, with the ability to regulate the growth and motility of these cells. In this study, lumican-deficient Saos 2 cells were demonstrated to have increased adhesive capability onto fibronectin (FN) (p≤0.01). Upon neutralization of endogenous transforming growth factor β2 (TGF-β2) activity, no difference in the ability of lumican siRNA-transfected and scramble siRNA-transfected Saos 2 cells to adhere onto FN was detected (p=NS). Exogenous TGF-β2 was shown to stimulate Saos 2 cell adhesion to FN (p≤0.01). These results therefore, suggest that the inverse correlation existing between lumican expression and Saos 2 cell adhesion is dependent on active TGF-β2 signaling. Furthermore, the significant increase in Smad 2 activation present in lumican-deficient cells (p≤0.01) was annulled in the presence of the anti-TGF-β2 peptide, demonstrating that lumican is an upstream regulator of the TGF-β2/Smad 2 signaling cascade. Crucial to FN-dependent adhesion, β1 integrin expression and pFAK activation were likewise identified as downstream TGF-β2 effectors regulated by lumican expression. In conclusion, this study demonstrates a novel out-in signaling circuit in human osteosarcoma cells: secreted to extracellular matrix lumican is an endogenous inhibitor of TGF-β2 activity, resulting in downstream effector modulation including pSmad 2, integrin β1 and pFAK to regulate osteosarcoma adhesion.

Association of Prenatal Exposure to Persistent Organic Pollutants with Obesity and Cardiometabolic Traits in Early Childhood: The Rhea Mother-Child Cohort (Crete, Greece).

Background Prenatal exposure to endocrine-disrupting chemicals such as persistent organic pollutants (POPs) may increase risk of obesity later in life. Objective We examined the relation of in utero POPs exposure to offspring obesity and cardiometabolic risk factors at 4 years of age in the Rhea mother–child cohort in Crete, Greece (n = 689). Methods We determined concentrations of polychlorinated biphenyls (PCBs), dichlorodiphenyldichloroethylene (DDE), and hexachlorobenzene (HCB) in first-trimester maternal serum. We measured child weight, height, waist circumference, skinfold thicknesses, blood pressure (BP), blood levels of lipids, C-reactive protein, and adipokines at 4 years of age. Childhood obesity was defined using age- and sex-specific cut points for body mass index (BMI) as recommended by the International Obesity Task Force. Results On multivariable regression analyses, a 10-fold increase in HCB was associated with a higher BMI z-score (adjusted β = 0.49; 95% CI: 0.12, 0.86), obesity [relative risk (RR) = 8.14; 95% CI: 1.85, 35.81], abdominal obesity (RR = 3.49; 95% CI: 1.08, 11.28), greater sum of skinfold thickness (β = 7.71 mm; 95% CI: 2.04, 13.39), and higher systolic BP (β = 4.34 mmHg; 95% CI: 0.63, 8.05) at 4 years of age. Prenatal DDE exposure was associated with higher BMI z-score (β = 0.27; 95% CI: 0.04, 0.5), abdominal obesity (RR = 3.76; 95% CI: 1.70, 8.30), and higher diastolic BP (β = 1.79 mmHg; 95% CI: 0.13, 3.46). PCBs were not significantly associated with offspring obesity or cardiometabolic risk factors. Conclusions Prenatal exposure to DDE and HCB was associated with excess adiposity and higher blood pressure levels in early childhood. Citation Vafeiadi M, Georgiou V, Chalkiadaki G, Rantakokko P, Kiviranta H, Karachaliou M, Fthenou E, Venihaki M, Sarri K, Vassilaki M, Kyrtopoulos SA, Oken E, Kogevinas M, Chatzi L. 2015. Association of prenatal exposure to persistent organic pollutants with obesity and cardiometabolic traits in early childhood: the Rhea mother–child cohort (Crete, Greece). Environ Health Perspect 123:1015–1021; http://dx.doi.org/10.1289/ehp.1409062

Maternal and gestational factors and micronucleus frequencies in umbilical blood: the NewGeneris Rhea cohort in Crete.

Background: The use of cancer-related biomarkers in newborns has been very limited. Objective: We investigated the formation of micronuclei (MN) in full-term and preterm newborns and their mothers from the Rhea cohort (Crete), applying for the first time in cord blood a validated semiautomated analysis system, in both mono- and binucleated T lymphocytes. Methods: We assessed MN frequencies in peripheral blood samples from the mothers and in umbilical cord blood samples. We calculated MN in mononucleated (MNMONO) and binucleated (MNBN) T lymphocytes and the cytokinesis block proliferation index (CBPI) in 251 newborns (224 full term) and 223 mothers, including 182 mother–child pairs. Demographic and lifestyle characteristics were collected. Results: We observed significantly higher MNBN and CBPI levels in mothers than in newborns. In newborns, MNMONO and MNBN were correlated (r = 0.35, p < 0.001), and we found a moderate correlation between MNMONO in mothers and newborns (r = 0.26, p < 0.001). MNMONO frequencies in newborns were positively associated with the mother’s body mass index and inversely associated with gestational age and mother’s age, but we found no significant predictors of MNBN or CBPI in newborns. Conclusions: Although confirmation is needed by a larger study population, the results indicate the importance of taking into account both mono- and binucleated T lymphocytes for biomonitoring of newborns, because the first reflects damage expressed during in vivo cell division and accumulated in utero, and the latter includes additional damage expressed as MN during the in vitro culture step.

Low molecular weight heparin inhibits melanoma cell adhesion and migration through a PKCa/JNK signaling pathway inducing actin cytoskeleton changes

Low molecular weight heparin (LMWH) has significant antimetastatic capabilities and affects cancer progression in humans through, not fully defined mechanisms. Here we evaluated its activity at the intracellular level and how it is correlated with melanoma cell adhesion and migration. LMWH inhibited M5 and A375 melanoma cell adhesion and migration in a dose-dependent manner (p⩽0.01). Treatment of M5 melanoma cells with LMWH caused a marked down regulation of constitutive as well as the FN-induced phosphorylation (p⩽0.01) of protein kinase C alpha (PKCa). This was associated with a profound decrease in the cytoplasmic pPKCa (p⩽0.05) and a simultaneous enhancement of nuclear pPKCa localization (p⩽0.01). A significant decrease in the levels of pJNK (p⩽0.01), which is a downstream effector of PKCa, was also demonstrated in the LMWH-treated cells. Furthermore, LMWH-treated cells had disorganized actin stress fibers correlated to a strong decrease in cell-substratum interface area (p⩽0.05) and altered morphology. The decrease in the activation of PKCa, which is an important regulator of cell motility, was directly correlated to the reduced ability of the LMWH-treated melanoma cells to adhere onto and migrate towards the fibronectin (FN) substrate (p⩽0.01). The lineage activation of PKCa-JNK/p38 and their correlation to M5 cell adhesion was confirmed with the utilization of specific inhibitors. In conclusion, LMWH through the downregulation of pPKCa and redistribution to nuclear region attenuates JNK activation, which in turn induces cytoskeleton changes correlated to M5 cell decreased adhesion/migration. This may provide clues for the pharmacological targeting of melanoma.

Metabolic profile in early pregnancy is associated with offspring adiposity at 4 years of age: the rhea pregnancy cohort crete, Greece.

Context Maternal pre-pregnancy obesity may increase the risk of childhood obesity but it is unknown whether other metabolic factors in early pregnancy such as lipid profile and hypertension are associated with offspring cardiometabolic traits. Objective Our objective was to investigate whether fasting lipid, glucose, and insulin levels during early pregnancy and maternal pre-pregnancy weight status, are associated with offspring adiposity measures, lipid levels and blood pressure at preschool age. Design and Methods The study included 618 mother-child pairs of the pregnancy cohort “Rhea” study in Crete, Greece. Pregnant women were recruited at the first prenatal visit (mean: 12weeks, SD: 0.7). A subset of 348 women provided fasting serum samples for glucose and lipid measurements. Outcomes measures were body mass index, abdominal circumference, sum of skinfold thickness, and blood pressure measurements at 4 years of age. A subsample of 525 children provided non-fasting blood samples for lipid measurements. Results Pre-pregnancy overweight/obesity was associated with greater risk of offspring overweight/obesity (RR: 1.83, 95%CI: 1.19, 2.81), central adiposity (RR: 1.97, 95%CI: 1.11, 3.49), and greater fat mass by 5.10mm (95%CI: 2.49, 7.71) at 4 years of age. These associations were more pronounced in girls. An increase of 40mg/dl in fasting serum cholesterol levels in early pregnancy was associated with greater skinfold thickness by 3.30mm (95%CI: 1.41, 5.20) at 4 years of age after adjusting for pre-pregnancy BMI and several other confounders. An increase of 10mmHg in diastolic blood pressure in early pregnancy was associated with increased risk of offspring overweight/obesity (RR: 1.22, 95%CI: 1.03, 1.45), and greater skinfold thickness by 1.71mm (95% CI: 0.57, 2.86) at 4 years of age. Conclusions Metabolic dysregulation in early pregnancy may increase the risk of obesity at preschool age.

The Role of SLRP-Proteoglycans in Osteosarcoma Pathogenesis

The small leucine-rich repeat proteoglycans (SLRPs) constitute a group of structurally and functionally related molecules that participate in the organization of the extracellular matrix ECM and have important effects on cell behavior. Osteosarcomas are heterogeneous bone tumors whose common characteristic is the production of an abundant nonmineralized (ECM)–osteoid. The scope of this minireview is to briefly present the current state of knowledge on the role of the SLRPs in osteosarcoma pathogenesis, with special emphasis on the recently described in osteosarcoma, proteoglycan lumican.

Heparin plays a key regulatory role via a p53/FAK-dependent signaling in melanoma cell adhesion and migration

Heparin and its various derivatives affect cancer progression in humans. In this study, we show that heparin uptaken intracellularly by melanoma cells activated a signaling cascade, which in turn inhibited melanoma cell adhesion and migration. The reduced ability of M5 cells to adhere onto the fibronectin (FN) substrate was directly correlated to a decrease in the expression of focal adhesion kinase (FAK), which is a key regulator of melanoma motility. Cell treatment with heparin caused a marked downregulation in FAK expression (P ≤ 0.01). This is followed by an analogous inhibition of both constitutive and FN‐induced FAK Y397‐phosphorylation (P ≤ 0.01). Moreover, heparin stimulated the p53 expression (P ≤ 0.001) of M5 cells and its increased accumulation in the nucleus. This favors a decrease in FAK promoter activation and explains the reduced FAK transcript and protein levels. In conclusion, the results of this study clearly demonstrate that the action of heparin in the regulation of melanoma cell adhesion and migration involves a p53/FAK/signaling pathway, which may be of importance in molecular targeted therapy of the disease.

Outdoor air pollution exposures and micronuclei frequencies in lymphocytes from pregnant women and newborns in Crete, Greece (Rhea cohort)

BACKGROUND Micronuclei (MN) are biomarkers of early genetic effects that have been used to investigate the association between environmental exposures and cancer. However, few studies have examined the association between environmental exposures during pregnancy and MN in mothers and newborns. OBJECTIVES We examined MN frequency in maternal blood and in cord blood, in relation to maternal air pollution exposure, and the potential interaction with maternal vitamin C intake and maternal smoking. METHODS We used the cytokinesis-block micronucleus assay to assess MN frequency per 1000 bi-nucleated T-lymphocytes from 181 mothers and 183 newborns born in 2007-2008 in Heraklion (Crete, Greece). The ESCAPE land-use regression methods were used to estimate annual mean exposure to outdoor air pollution [particulate matter (PM), black carbon, nitrogen dioxide (NO2) and nitrogen oxides (NOx)] at maternal home addresses. Food frequency questionnaires were used to estimate maternal dietary vitamin C intake during pregnancy. Smoking habits were self-reported using questionnaires which were checked by measuring maternal urinary cotinine levels. RESULTS Exposure to PM2.5 was associated with increased MN frequencies in pregnant women [rate ratio [RR (95%CI)] per 5 µg/m(3)=1.53 (1.02, 2.29)]. This increase was considerably higher among women who did not fulfill the recommended vitamin C dietary allowances [RR=9.35 (2.77, 31.61); n=20]. Exposure to PM2.5-10, PM10, NO2 and NOx were also associated with a higher incidence of MN frequencies in smoker women (n=56). No associations were found for newborns. CONCLUSIONS We found an association between air pollution, particularly PM2.5, and MN frequency in mothers but not in newborns. This association was more pronounced among women with a lower dietary intake of vitamin C during pregnancy and among women who smoked during pregnancy. While results are clear in mothers, the association between maternal carcinogenic exposures during pregnancy and biomarkers of early biologic effect in the newborn remains poorly understood.