High-level multiplex genotyping of polymorphisms involved in folate or homocysteine metabolism by matrix-assisted laser desorption/ionization mass spectrometry

BACKGROUND: Increased plasma total homocysteine (tHcy), a risk factor for cardiovascular disease, is related to genetic, environmental, and nutritional factors, in particular folate status. Future large epidemiologic studies of the genetic basis of hyperhomocysteinemia will require high-throughput assays for polymorphisms of genes related to folate and Hcy metabolism. METHOD: We developed a high-level multiplex genotyping method based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the detection of 12 polymorphisms in 8 genes involved in folate or Hcy metabolism. The assay includes methylenetetrahydrofolate reductase (MTHFR) 677C>T and 1298A>C, methionine synthase (MTR) 2756A>G, methionine synthase reductase (MTRR) 66A>G, cystathionine beta-synthase (CBS) 844ins68 and 699C>T, transcobalamin II (TCII) 776C>G and 67A>G, reduced folate carrier-1 (RFC1) 80G>A, paraoxonase-1 (PON1) 575A>G and 163T>A, and betaine homocysteine methyltransferase (BHMT) 742G>A. RESULTS: The failure rate of the assay was < or = 1.7% and was attributable to unsuccessful DNA purification, nanoliter dispensing, and spectrum calibration. Most errors were related to identification of heterozygotes as homozygotes. The mean error rate was 0.26%, and error rates differed for the various single-nucleotide polymorphisms. Identification of CBS 844ins68 was carried out by a semiquantitative approach. The throughput of the MALDI-TOF MS assay was 1152 genotypes within 20 min. CONCLUSIONS: This high-level multiplex method is able to genotype 12 polymorphisms involved in folate or Hcy metabolism. The method is rapid and reproducible and could facilitate large-scale studies of the genetic basis of hyperhomocysteinemia and associated pathologies.     

Plasma vitamin B6 vitamers before and after oral vitamin B6 treatment: a randomized placebo-controlled study

BACKGROUND: Vitamin B(6) has attracted renewed interest because of its role in homocysteine metabolism and its possible relation to cardiovascular risk. We examined the plasma B(6) vitamers, pyridoxal 5'-phosphate (PLP), pyridoxal (PL), pyridoxine (PN), and 4-pyridoxic acid (4-PA) before and after vitamin B(6) supplementation. METHODS: Patients (n = 90; age range, 38-80 years) undergoing coronary angiography (part of the homocysteine-lowering Western Norway B-Vitamin Intervention Trial) were allocated to the following daily oral treatment groups: (A), vitamin B(12) (0.4 mg), folic acid (0.8 mg), and vitamin B(6) (40 mg); (B), vitamin B(12) and folic acid; (C), vitamin B(6); or (D), placebo. EDTA blood was obtained before treatment and 3, 14, 28, and 84 days thereafter. RESULTS: Before treatment, PLP (range, 5-111 nmol/L) and 4-PA (6-93 nmol/L) were the predominant B(6) vitamers identified in plasma. During the 84-day study period, the intraindividual variation (CV) in patients not treated with vitamin B(6) (groups B and D) was 45% for PLP and 67% for 4-PA. Three days after the start of treatment, the increases in concentration were approximately 10-, 50-, and 100-fold for PLP, 4-PA, and PL, respectively. No significant additional increase was observed at the later time points. The PLP concentration correlated to the concentrations of 4-PA and PL before treatment, but not after treatment. The PL concentration correlated with 4-PA before and after treatment. CONCLUSIONS: Vitamin B(6) treatment has an immediate effect on the concentrations and the forms of B(6) vitamers present in plasma, and the changes remain the same during prolonged treatment. Our results suggest that the B(6) vitamers in plasma reflect vitamin B(6) intake.     

Biological monitoring of benzene exposure for process operators during ordinary activity in the upstream petroleum industry

This study characterized the exposure of crude oil process operators to benzene and related aromatics during ordinary activity and investigated whether the operators take up benzene at this level of exposure. We performed the study on a fixed, integrated oil and gas production facility on Norway's continental shelf. The study population included 12 operators and 9 referents. We measured personal exposure to benzene, toluene, ethylbenzene and xylene during three consecutive 12-h work shifts using organic vapour passive dosimeter badges. We sampled blood and urine before departure to the production facility (pre-shift), immediately after the work shift on Day 13 of the work period (post-shift) and immediately before the following work shift (pre-next shift). We also measured the exposure to hydrocarbons during short-term tasks by active sampling using Tenax tubes. The arithmetic mean exposure over the 3 days was 0.042 ppm for benzene (range <0.001-0.69 ppm), 0.05 ppm for toluene, 0.02 ppm for ethylbenzene and 0.03 ppm for xylene. Full-shift personal exposure was significantly higher when the process operators performed flotation work during the shift versus other tasks. Work in the flotation area was associated with short-term (6-15 min) arithmetic mean exposure to benzene of 1.06 ppm (range 0.09-2.33 ppm). The concentrations of benzene in blood and urine did not differ between operators and referents at any time point. When we adjusted for current smoking in regression analysis, benzene exposure was significantly associated with the post-shift concentration of benzene in blood (P = 0.01) and urine (P = 0.03), respectively. Although these operators perform tasks with relatively high short-term exposure to benzene, the full-shift mean exposure is low during ordinary activity. Some evidence indicates benzene uptake within this range of exposure.     

Life course determinants for early disability pension: a follow-up of Norwegian men and women born 1967–1976

Background Most studies on disability pension (DP) have focused on work conditions, socio-economic status and other contemporary factors. We wanted to study possible determinants of an early DP with a life course perspective within a large register-based cohort, with a main focus on the biological and social factors from childhood. Methods We established a longitudinal, population-based cohort of all persons liveborn in Norway between 1967 and 1976. Through linkage between several national registers we obtained personal data on biological/health related as well as social background factors. After excluding persons who died, emigrated or were granted a DP before age 20 years (at which age follow-up started) and persons who did not become gainfully employed during the study period, the study population consisted of 595,393 persons. They were categorized into four strata according to gender and educational attainment. Adjusted hazard ratios (HR) for granting a DP until the end of 2003 and the corresponding population attributable risks (PAR) were computed. Results A total of 9,649 persons (1.6%) were granted a DP during follow-up. The disability risk was slightly higher among women than among men (1.7% vs. 1,5%). The following PARs were found: birth weight below the mean 5.7%, chronic childhood disease 6.8%, maternal marital status 4.4% and parental disability 8.8%. Low educational achievement was highly associated with DP, with a PAR more than twice as high as the overall PAR for the childhood factors. Conclusions Early DP is associated with several biological and social background factors from childhood. It also shows a strong dependency on educational achievement.     

Organic Food Consumption during Pregnancy and Hypospadias and Cryptorchidism at Birth: The Norwegian Mother and Child Cohort Study (MoBa)

Background

The etiologies of the male urogenital anomalies hypospadias and cryptorchidism remain unclear. It has been suggested that maternal diet and environmental contaminants may affect the risk of these anomalies via placental or hormonal disturbances.

Objectives

We examined associations between organic food consumption during pregnancy and prevalence of hypospadias and cryptorchidism at birth.

Methods

Our study includes 35,107 women participating in the Norwegian Mother and Child Cohort Study (MoBa) who delivered a singleton male infant. Information about use of six groups of organically produced food (vegetables, fruit, bread/cereal, milk/dairy products, eggs, and meat) during pregnancy was collected by a food frequency questionnaire. Women who indicated that they sometimes, often, or mostly consumed organic foods in at least one of the six food groups were classified as organic food consumers in analyses. Hypospadias and cryptorchidism diagnoses were retrieved from the Medical Birth Registry of Norway. We estimated odds ratios (ORs) and 95% confidence intervals (CIs) using multiple logistic regression.

Results

Seventy-four male newborns were diagnosed with hypospadias (0.2%), and 151 with cryptorchidism (0.4%). Women who consumed any organic food during pregnancy were less likely to give birth to a boy with hypospadias (OR = 0.42; 95% CI: 0.25, 0.70, based on 21 exposed cases) than women who reported they never or seldom consumed organic food. Associations with specific organic foods were strongest for vegetable (OR = 0.36; 95% CI: 0.15, 0.85; 10 exposed cases) and milk/dairy (OR = 0.43; 95% CI: 0.17, 1.07; 7 exposed cases) consumption. No substantial association was observed for consumption of organic food and cryptorchidism.

Conclusions

Consumption of organically produced foods during pregnancy was associated with a lower prevalence of hypospadias in our study population. These findings were based on small numbers of cases and require replication in other study populations.

Citation

Brantsæter AL, Torjusen H, Meltzer HM, Papadopoulou E, Hoppin JA, Alexander J, Lieblein G, Roos G, Holten JM, Swartz J, Haugen M. 2016. Organic food consumption during pregnancy and hypospadias and cryptorchidism at birth: the Norwegian Mother and Child Cohort Study (MoBa). Environ Health Perspect 124:357–364; http://dx.doi.org/10.1289/ehp.1409518     

Actinobacillus equuli subsp. equuli associated with equine valvular endocarditis

Microbiological and pathological data from a case of equine valvular endocarditis are reported. Limited information is available on the pathogenic potential of equine Actinobacillus species as several strains originate from apparently healthy horses. After the establishment of two subspecies within this species, this seems to be the first report of an etiological association between A. equuli subsp. equuli and equine endocarditis. Furthermore, new information on some phenotypical characteristics of this subspecies is reported, compared to previous findings.     

Syndecan-1 and angiogenic cytokines in multiple myeloma: correlation with bone marrow angiogenesis and survival

Angiogenesis is a complex process involved in the proliferation and metastasis of malignant tumours, and partly triggered by the secretion of various angiogenic factors by tumour cells or cells in the stromal environment. We investigated the correlation between bone marrow angiogenesis, estimated as microvessel density (MVD), and interleukin-6 (IL-6), basic fibroblastic growth factor (bFGF), hepatocyte growth factor (HGF) and syndecan-1 in 67 patients with newly diagnosed multiple myeloma, and evaluated the prognostic value of these parameters. Circulating levels of IL-6, bFGF, HGF and syndecan-1 were significantly higher in patients than in controls. Moreover, in patients, bone marrow levels of bFGF, HGF and syndecan-1 were higher than peripheral blood levels. Positive correlations were found between MVD and syndecan-1 blood levels (r = 0.33, P = 0.017), syndecan-1 bone marrow levels (r = 0.49, P = 0.046) and HGF blood levels (r = 0.36, P = 0.008) respectively. High MVD and high blood levels of IL-6, HGF and syndecan-1 were predictive of a shorter survival. In a multivariate survival analysis MVD and blood levels of IL-6 retained independent prognostic significance, while in a survival analysis without MVD the peripheral blood levels of HGF and syndecan-1 were strong independent prognostic factors.     

Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications

The purpose of this study was to classify breast carcinomas based on variations in gene expression patterns derived from cDNA microarrays and to correlate tumor characteristics to clinical outcome. A total of 85 cDNA microarray experiments representing 78 cancers, three fibroadenomas, and four normal breast tissues were analyzed by hierarchical clustering. As reported previously, the cancers could be classified into a basal epithelial-like group, an ERBB2-overexpressing group and a normal breast-like group based on variations in gene expression. A novel finding was that the previously characterized luminal epithelial/estrogen receptor-positive group could be divided into at least two subgroups, each with a distinctive expression profile. These subtypes proved to be reasonably robust by clustering using two different gene sets: first, a set of 456 cDNA clones previously selected to reflect intrinsic properties of the tumors and, second, a gene set that highly correlated with patient outcome. Survival analyses on a subcohort of patients with locally advanced breast cancer uniformly treated in a prospective study showed significantly different outcomes for the patients belonging to the various groups, including a poor prognosis for the basal-like subtype and a significant difference in outcome for the two estrogen receptor-positive groups.