Cholesterol Interference in the Assessment of Vitamin D Status: A Canadian Health Measures Survey Biobank Project.

BACKGROUND: Matrix effects are a known problem with immunoassays measuring serum 25-hydroxyvitamin D [25(OH)D]. OBJECTIVES: To determine if the inverse association between serum 25(OH)D and serum cholesterol concentrations is a function of assay method: Diasorin Liaison 25(OH) Vitamin D Total Assay (Liaison Total Assay), an immunoassay, compared with liquid chromatography tandem mass spectrometry (LC-MS/MS). METHODS: Canadian Health Measures Survey data and biobank serum (White males aged 20-79 y, n = 392) were evaluated for bias in serum 25(OH)D using Bland-Altman plots. Differences in serum 25(OH)D (Liaison Total Assay - LC-MS/MS) were compared among non-HDL-cholesterol <4.2 (n = 295) compared with ≥4.2 (n = 97) mmol/L and total cholesterol groups <5.2 (n = 256) compared with ≥5.2 (n = 136) mmol/L, and associations tested between 25(OH)D and non-HDL-cholesterol or total cholesterol concentrations, using regression. RESULTS: Serum 25(OH)D measured using Liaison Total Assay ranged from 10.7 to 137.0 nmol/L and 14.4 to 137.9 nmol/L by LC-MS/MS. Liaison Total Assay - LC-MS/MS showed a negative bias of 5.5 (95% limits of agreement -23.8, 12.7) nmol/L. Differences in 25(OH)D were -4.0 ± 9.0 (±SD) nmol/L if non-HDL-cholesterol was <4.2 mmol/L and -10.2 ± 8.7 nmol/L if ≥4.2 mmol/L (P < 0.0001). Differences in 25(OH)D, if total cholesterol was <5.2 mmol/L, were -3.4 ± 8.6 nmol/L and -9.6 ± 9.3 nmol/L if ≥5.2 mmol/L (P < 0.0001). Serum non-HDL-cholesterol (beta -3.17, P = 0.0014) and total cholesterol (beta -2.77, P = 0.0046) were inversely associated with Liaison Total Assay 25(OH)D (adjusted for age, fasting, and body mass index), but not with LC-MS/MS measured 25(OH)D. Interference by these lipoproteins was not eliminated by standardization of the Liaison Total Assay. Similar associations were observed with triglycerides as for the lipoproteins. CONCLUSIONS: Total cholesterol inversely associates with 25(OH)D, which is likely due to elevated non-HDL-cholesterol lipoprotein or triglyceride interference with the Liaison Total Assay. This is important as elevated cholesterol is common, and an underestimation of vitamin D status could be an unnecessary cause for concern.

Longitudinal Assessment of Vitamin D Status across Trimesters of Pregnancy.

BACKGROUND: The evolution of vitamin D status across pregnancy trimesters and its association with prepregnancy body mass index (ppBMI; in kg/m2) remain unclear. OBJECTIVES: We aimed to 1) assess trimester-specific serum total 25-hydroxyvitamin D [25(OH)D] concentrations, 2) compare those concentrations between ppBMI categories, and 3) examine associations between 25(OH)D concentrations, ppBMI, and vitamin D intake. METHODS: As part of a prospective cohort study, 79 pregnant women with a mean age of 32.1 y and ppBMI of 25.7 kg/m2 were recruited in their first trimester (average 9.3 weeks of gestation). Each trimester, vitamin D intake was assessed by 3 Web-based 24-h recalls and a Web questionnaire on supplement use. Serum total 25(OH)D was measured by LC-tandem MS. Repeated-measures ANOVA was performed to assess the evolution of 25(OH)D concentrations across trimesters of pregnancy and comparisons of 25(OH)D concentrations between ppBMI categories were assessed by 1-factor ANOVAs. Stepwise regression analyses were used to identify determinants of 25(OH)D concentrations in the third trimester. RESULTS: Mean ± SD serum total 25(OH)D concentrations increased across trimesters, even after adjustments for ppBMI, seasonal variation, and vitamin D intake from supplements (67.5 ± 20.4, 86.5 ± 30.9, and 88.3 ± 29.0 nmol/L at mean ± SD 12.6 ± 0.8, 22.5 ± 0.8, and 33.0 ± 0.6 weeks of gestation, respectively; P < 0.0001). In the first and third trimesters, women with a ppBMI ≥30 had lower serum total 25(OH)D concentrations than women with a ppBMI <25 (P < 0.05); however, most had concentrations >40nmol/L by the second trimester. Vitamin D intake from supplements was the strongest determinant of third-trimester serum total 25(OH)D concentrations (r2 = 0.246, ß = 0.51; P < 0.0001). CONCLUSIONS: There was an increase in serum total 25(OH)D concentrations across trimesters, independent of ppBMI, seasonal variation, and vitamin D intake from supplements. Almost all women had serum total 25(OH)D concentrations over the 40- and 50-nmol/L thresholds, thus our study supports the prenatal use of a multivitamin across pregnancy.

Cardiovascular and inflammatory mechanisms in healthy humans exposed to air pollution in the vicinity of a steel mill.

BACKGROUND: There is a paucity of mechanistic information that is central to the understanding of the adverse health effects of source emission exposures. To identify source emission-related effects, blood and saliva samples from healthy volunteers who spent five days near a steel plant (Bayview site, with and without a mask that filtered many criteria pollutants) and at a well-removed College site were tested for oxidative stress, inflammation and endothelial dysfunction markers. METHODS: Biomarker analyses were done using multiplexed protein-array, HPLC-Fluorescence, EIA and ELISA methods. Mixed effects models were used to test for associations between exposure, biological markers and physiological outcomes. Heat map with hierarchical clustering and Ingenuity Pathway Analysis (IPA) were used for mechanistic analyses. RESULTS: Mean CO, SO2 and ultrafine particles (UFP) levels on the day of biological sampling were higher at the Bayview site compared to College site. Bayview site exposures "without" mask were associated with increased (p < 0.05) pro-inflammatory cytokines (e.g IL-4, IL-6) and endothelins (ETs) compared to College site. Plasma IL-1ß, IL-2 were increased (p < 0.05) after Bayview site "without" compared to "with" mask exposures. Interquartile range (IQR) increases in CO, UFP and SO2 were associated with increased (p < 0.05) plasma pro-inflammatory cytokines (e.g. IL-6, IL-8) and ET-1(1-21) levels. Plasma/saliva BET-1 levels were positively associated (p < 0.05) with increased systolic BP. C-reactive protein (CRP) was positively associated (p < 0.05) with increased heart rate. Protein network analyses exhibited activation of distinct inflammatory mechanisms after "with" and "without" mask exposures at the Bayview site relative to College site exposures. CONCLUSIONS: These findings suggest that air pollutants in the proximity of steel mill site can influence inflammatory and vascular mechanisms. Use of mask and multiple biomarker data can be valuable in gaining insight into source emission-related health impacts.

Infant birth weight and third trimester maternal plasma markers of vascular integrity: the MIREC study.

BACKGROUND: There is paucity of information on mechanisms constituting adverse birth outcomes. We assessed here the relationship between vascular integrity and adverse birth effects. METHODS AND RESULTS: Third trimester maternal plasma (n = 144) from the Maternal-Infant Research on Environmental Chemicals Study (MIREC) was analysed for vascular, inflammatory and oxidative stress markers by HPLC-fluorescence, protein array and EIA method. Analysis of the <25th and >75th percentile birth weight subgroups revealed markers associated with birth weight (ETs, MMP-9, VEGF, and 8-isoPGF-2α) and gestational age (ET-1, MMP-2, and VEGF). CONCLUSIONS: Mechanistic insights into adverse birth outcome pathways can be achieved by integrating information on multiple biomarkers, physiology using systems biology approach.

Nitrative stress, oxidative stress and plasma endothelin levels after inhalation of particulate matter and ozone.

BACKGROUND: While exposure to ambient air contaminants is clearly associated with adverse health outcomes, disentangling mechanisms of pollutant interactions remains a challenge. OBJECTIVES: We aimed at characterizing free radical pathways and the endothelinergic system in rats after inhalation of urban particulate matter, ozone, and a combination of particles plus ozone to gain insight into pollutant-specific toxicity mechanisms and any effect modification due to air pollutant mixtures. METHODS: Fischer 344 rats were exposed for 4 h to a 3 × 3 concentration matrix of ozone (0, 0.4, 0.8 ppm) and EHC-93 particles (0, 5, 50 mg/m(3)). Bronchoalveolar lavage fluid (BALF), BAL cells, blood and plasma were analysed for biomarkers of effects immediately and 24 h post-exposure. RESULTS: Inhalation of ozone increased (p < 0.05) lipid oxidation products in BAL cells immediately post-exposure, and increased (p < 0.05) total protein, neutrophils and mature macrophages in the BALF 24 h post-exposure. Ozone increased (p < 0.05) the formation of reactive oxygen species (ROS), assessed by m-, p-, o-tyrosines in BALF (Ozone main effects, p < 0.05), while formation of reactive nitrogen species (RNS), indicated by 3-nitrotyrosine, correlated with dose of urban particles (EHC-93 main effects or EHC-93 × Ozone interactions, p < 0.05). Carboxyhemoglobin levels in blood exhibited particle exposure-related increase (p < 0.05) 24 h post recovery. Plasma 3-nitrotyrosine and o-tyrosine were increased (p < 0.05) after inhalation of particles; the effect on 3-nitrotyrosine was abrogated after exposure to ozone plus particles (EHC-93 × Ozone, p < 0.05). Big endothelin-1 (BET-1) and ET-1 were increased in plasma after inhalation of particles or ozone alone, but the effects appeared to be attenuated by co-exposure to contaminants (EHC-93 × Ozone, p < 0.05). Plasma ET levels were positively correlated (p < 0.05) with BALF m- and o-tyrosine levels. CONCLUSIONS: Pollutant-specific changes can be amplified or abrogated following multi-pollutant exposures. Oxidative and nitrative stress in the lung compartment may contribute to secondary extra-pulmonary ROS/RNS formation. Nitrative stress and endothelinergic imbalance emerge as potential key pathways of air pollutant health effects, notably of ambient particulate matter.

Estrogen receptor-ß mediates the inhibition of DLD-1 human colon adenocarcinoma cells by soy isoflavones.

To understand the relationship between the role of soy isoflavones and estrogen receptor (ER)-ß in colon tumorigenesis, we investigated the cellular effects of soy isoflavones (composed of genistein, daidzein, and glycitein) in DLD-1 human colon adenocarcinoma cells with or without ER-ß gene silencing by RNA interference (RNAi). Soy isoflavones decreased the expression of proliferating cell nuclear antigen (PCNA), extracellular signal-regulated kinase (ERK)-1/2, AKT, and nuclear factor (NF)-κB. Soy isoflavones dose-dependently caused G2/M cell cycle arrest and downregulated the expression of cyclin A. This was associated with inhibition of cyclin dependent kinase (CDK)-4 and up-regulation of its inhibitor p21(cip1) expressions. ER-ß gene silencing lowered soy isoflavone-mediated suppression of cell viability and proliferation. ERK-1/2 and AKT expressions were unaltered and NF-κB was modestly upregulated by soy isoflavones after transient knockdown of ER-ß expression. Soy isoflavone-mediated arrest of cells at G2/M phase and upregulation of p21(cip1) expression were not observed when ER-ß gene was silenced. These findings suggest that maintaining the expression of ER-ß is crucial in mediating the growth-suppressive effects of soy isoflavones against colon tumors. Thus upregulation of ER-ß status by specific food-borne ER-ligands such as soy isoflavones could potentially be a dietary prevention or therapeutic strategy for colon cancer.

Soy isoflavones modulate azoxymethane-induced rat colon carcinogenesis exposed pre- and postnatally and inhibit growth of DLD-1 human colon adenocarcinoma cells by increasing the expression of estrogen receptor-beta.

We studied the effects of lifetime exposure to dietary soy isoflavones in an azoxymethane (AOM)-induced rat colon cancer model. Male pups born to Sprague-Dawley rats exposed (including during pregnancy and lactation) to soy isoflavones at either no (0 mg = control), low (40 mg), or high (1000 mg) doses/kg diet were weaned and continued receiving their respective parental diets until the end of the study. Weaned rats received 2 subcutaneous injections (15 mg/kg body weight) of AOM 1 wk apart. After 26 wk, rats were killed and the coordinates of colon aberrant crypt foci (ACF) and tumors were determined. Expression of estrogen receptor (ER)-beta was assessed in rat colon tumors and in DLD-1 human colon adenocarcinoma cells exposed to soy isoflavones. Compared with the control, soy isoflavones did not affect incidences or multiplicities of colon ACF or tumors. Low-dose soy isoflavones decreased tumor burden and size compared with the control (P < 0.05). Expression of ERbeta increased in colon tumors of soy isoflavone-treated groups compared with the control. Soy isoflavones dose-dependently arrested the growth of DLD-1 cells and at subcytotoxic levels increased the expression of ERbeta. Our results suggest that pre- and postnatal exposure to dietary soy isoflavones suppresses the growth of colon tumors in male rats. The overexpression of ERbeta in both rat colon tumors and DLD-1 cells caused by soy isoflavones suggests that ERbeta is a critical mediator in mitigating its cancer-preventive effects.

Characterization of maternal plasma biomarkers associated with delivery of small and large for gestational age infants in the MIREC study cohort.

OBJECTIVE: Neonatal morbidity and mortality can be influenced by maternal health status. Information on maternal and fetal biomarkers of adverse health outcomes is limited. This work aims at identifying maternal biomarkers associated with low and high birth weight for gestational age groups. DESIGN AND SETTINGS: Population-based prospective cohort study of the potential adverse health effects of exposure to environmental contaminants on pregnancy and infant health. METHODS: Third trimester maternal plasma samples (n = 1588) from a pregnancy cohort (Maternal-Infant Research on Environmental Chemicals Study, MIREC) were analyzed for changes in a target spectrum of biomarkers of vascular health (e.g., matrix metalloproteinases MMPs, vascular endothelial cell growth factor VEGF), inflammation (e.g. cellular adhesion molecules CAMs, cytokines, chemokines) by affinity-based multiplex protein array analyses. Multivariate logistic regression analyses were done to examine associations between target plasma biomarkers, maternal-infant characteristics, and birth weight outcomes assessed as small for gestational age (SGA) ≤10th percentile and large for gestational age (LGA) ≥90th percentile groups. RESULTS AND OUTCOMES: Our results revealed that maternal plasma biomarkers monocyte chemoattractant protein-1 MCP-1 (p<0.05, +ve) and VEGF (p<0.05, -ve) along with parity = 1 (p<0.01, -ve) and gestational hypertension (p<0.05, +ve) were associated with SGA births. Meanwhile, LGA was associated with maternal plasma VEGF (p<0.05, +ve) and MMP-9 (p<0.05, -ve) and gestational hypertension (p<0.01, +ve), pre-pregnancy body mass index (p<0.01, +ve), parity (p<0.05, +ve) and education (p<0.05, -ve). CONCLUSIONS: Third trimester maternal plasma biomarkers in combination with maternal health and socioeconomic characteristics can be useful in predicting SGA and LGA outcomes. Maternal vascular health and inflammatory status may contribute to both SGA and LGA births through distinct molecular mechanisms.

Blood metal levels and third trimester maternal plasma matrix metalloproteinases (MMPs).

While it is known that in utero exposure to environmental toxicants, namely heavy metals, can adversely affect the neonate, there remains a significant paucity of information on maternal biological changes specific to metal exposures during pregnancy. This study aims at identifying associations between maternal metal exposures and matrix metalloproteinases (MMPs) that are known to be engaged in pregnancy process. Third trimester maternal plasma (n = 1533) from a pregnancy cohort (Maternal-Infant Research on Environmental Chemicals Study, MIREC) were analyzed for MMP-1,-2,-7,-9 and -10 by affinity-based multiplex protein array analyses. Maternal metal concentrations (mercury, cadmium, lead, arsenic and manganese) in 1st and 3rd trimesters exhibited strong correlations (p < 0.05). Multivariate regression models were used to estimate odds ratio (OR) for the association between metal concentrations in quartiles and high (90%) and low (10%) maternal MMP levels. Significant (p < 0.05) metal exposure-related effects were observed with the different MMP isoform responses. MMP profiles were specific to the trimester at which the maternal blood metals were analyzed. Our findings suggest that the profiles of these MMP isoforms vary with the type of metal exposure, blood metal concentrations and the trimester at which metal levels were determined. These new findings on maternal metal-MMP relationships can guide future explorations on toxicity mechanisms relevant to metal exposure-mediated adverse birth outcomes.