Physical activity and epigenetic biomarkers in maternal blood during pregnancy.

AIM: Investigate associations of leisure time physical activity (LTPA) with DNA methylation and miRNAs during pregnancy. Patients & methods: LTPA, candidate DNA methylation and circulating miRNAs were measured (average 15 weeks gestation) in pregnant women (n = 92). RESULTS: Each additional hour of prepregnancy LTPA duration was associated with hypermethylation in C1orf212 (ß = 0.137, 95% CI: 0.004-0.270) and higher circulating miR-146b-5p (ß = 0.084, 95% CI: 0.017-0.151). Each additional metabolic equivalent hour of early-pregnancy LTPA energy expenditure was associated with higher circulating miR-21-3p (ß = 0.431, 95% CI: 0.089-0.772) in women carrying female offspring, and lower circulating miR-146b-5p (ß = -0.285, 95% CI: -0.528 to -0.043) and miR-517-5p (ß = -0.406, 95% CI: -0.736 to -0.076) in women carrying male offspring. CONCLUSION: Our findings suggest that LTPA may influence maternal epigenetic biomarkers, possibly in an offspring sex-specific manner.

Genetic determinants of susceptibility to silver nanoparticle-induced acute lung inflammation in mice.

Silver nanoparticles (AgNPs) are employed in a variety of consumer products; however, in vivo rodent studies indicate that AgNPs can cause lung inflammation and toxicity in a strain- and particle type-dependent manner, but mechanisms of susceptibility remain unclear. The aim of this study was to assess the variation in AgNP-induced lung inflammation and toxicity across multiple inbred mouse strains and to use genome-wide association (GWA) mapping to identify potential candidate susceptibility genes. Mice received doses of 0.25 mg/kg of either 20-nm citrate-coated AgNPs or citrate buffer using oropharyngeal aspiration. Neutrophils in bronchoalveolar lavage fluid (BALF) served as markers of inflammation. We found significant strain- and treatment-dependent variation in neutrophils in BALF. GWA mapping identified 10 significant single-nucleotide polymorphisms (false discovery rate, 15%) in 4 quantitative trait loci on mouse chromosomes 1, 4, 15, and 18, and Nedd4l (neural precursor cell expressed developmentally downregulated gene 4-like; chromosome 18), Ano6 (anocatmin 6; chromosome 15), and Rnf220 (Ring finger protein 220; chromosome 4) were considered candidate genes. Quantitative RT-PCR revealed significant inverse associations between mRNA levels of these genes and neutrophil influx. Nedd4l, Ano6, and Rnf220 are candidate susceptibility genes for AgNP-induced lung inflammation that warrant additional exploration in future studies.-Scoville, D. K., Botta, D., Galdanes, K., Schmuck, S. C., White, C. C., Stapleton, P. L., Bammler, T. K., MacDonald, J. W., Altemeier, W. A., Hernandez, M., Kleeberger, S. R., Chen, L.-C., Gordon, T., Kavanagh, T. J. Genetic determinants of susceptibility to silver nanoparticle-induced acute lung inflammation in mice.

Genetics, Diet, and Season Are Associated with Serum 25-Hydroxycholecalciferol Concentration in a Yup'ik Study Population from Southwestern Alaska.

BACKGROUND: Low blood vitamin D concentration is a concern for people living in circumpolar regions, where sunlight is insufficient for vitamin D synthesis in winter months and the consumption of traditional dietary sources of vitamin D is decreasing. OBJECTIVE: The objective was to characterize the effects of diet, genetic variation, and season on serum 25-hydroxycholecalciferol [25(OH)D3] concentrations in Yup'ik Alaska Native people living in rural southwest Alaska. METHODS: This study was a cross-sectional design that assessed the associations of traditional diet (via a biomarker, the RBC δ(15)N value), age, gender, body mass index (BMI), community location, and genotype of select single nucleotide polymorphisms (SNPs) in cytochrome P450 family 2, subfamily R, peptide 1 (CYP2R1), 7-dehydrocholesterol reductase (DHCR7), and vitamin D binding protein (GC) with serum 25(OH)D3 concentrations in 743 Yup'ik male and female participants, aged 14-93 y, recruited between September 2009 and December 2013. RESULTS: Yup'ik participants, on average, had adequate concentrations of serum 25(OH)D3 (31.1 ± 1.0 ng/mL). Variations in diet, BMI, age, gender, season of sample collection, and inland or coastal community geography were all significantly associated with serum 25(OH)D3 concentration. In models not adjusting for other covariates, age, diet, and seasonal effects explained 33.7%, 20.7%, and 9.8%, respectively, of variability in serum 25(OH)D3 concentrations. Of the 8 SNPs interrogated in CYP2R1 and DHCR7, only rs11023374 in CYP2R1 was significantly associated with serum 25(OH)D3, explaining 1.5% of variability. The GC haplotype explained an additional 2.8% of variability. Together, age, diet, gender, season of sample collection, BMI, geography of the community, and genotype at rs11023374 explained 52.5% of the variability in serum 25(OH)D3 concentrations. CONCLUSIONS: Lower consumption of the traditional diet was associated with lower serum concentrations of 25(OH)D3. Younger adults and youth in this community may be at increased risk of adverse outcomes associated with vitamin D insufficiency compared with older members of the community, especially during seasons of low sunlight exposure, because of lower consumption of dietary sources of vitamin D.

Differential Gene Expression in Liver, Gill, and Olfactory Rosettes of Coho Salmon (Oncorhynchus kisutch) After Acclimation to Salinity.

Most Pacific salmonids undergo smoltification and transition from freshwater to saltwater, making various adjustments in metabolism, catabolism, osmotic, and ion regulation. The molecular mechanisms underlying this transition are largely unknown. In the present study, we acclimated coho salmon (Oncorhynchus kisutch) to four different salinities and assessed gene expression through microarray analysis of gills, liver, and olfactory rosettes. Gills are involved in osmotic regulation, liver plays a role in energetics, and olfactory rosettes are involved in behavior. Between all salinity treatments, liver had the highest number of differentially expressed genes at 1616, gills had 1074, and olfactory rosettes had 924, using a 1.5-fold cutoff and a false discovery rate of 0.5. Higher responsiveness of liver to metabolic changes after salinity acclimation to provide energy for other osmoregulatory tissues such as the gills may explain the differences in number of differentially expressed genes. Differentially expressed genes were tissue- and salinity-dependent. There were no known genes differentially expressed that were common to all salinity treatments and all tissues. Gene ontology term analysis revealed biological processes, molecular functions, and cellular components that were significantly affected by salinity, a majority of which were tissue-dependent. For liver, oxygen binding and transport terms were highlighted. For gills, muscle, and cytoskeleton-related terms predominated and for olfactory rosettes, immune response-related genes were accentuated. Interaction networks were examined in combination with GO terms and determined similarities between tissues for potential osmosensors, signal transduction cascades, and transcription factors.

Inducible nitric oxide synthase gene methylation and parkinsonism in manganese-exposed welders.

INTRODUCTION: Neurologist-assessed parkinsonism signs are prevalent among workers exposed to manganese (Mn)-containing welding fume. Neuroinflammation may possibly play a role. Inducible nitric oxide synthase, coded by NOS2, is involved in inflammation, and particulate exposure increases the gene's expression through methylation of CpG sites in the 5' region. METHODS: We assessed DNA methylation at three CpG sites in the NOS2 exon 1 from blood from 201 welders. All were non-Hispanic Caucasian men 25-65 years old who were examined by a neurologist specializing in movement disorders. We categorized the workers according to their Unified Parkinson Disease Rating Scale motor subsection 3 (UPDRS3) scores as parkinsonism cases (UPDRS3 ≥ 15; n = 49), controls (UPDRS3 < 6; n = 103), or intermediate (UPDRS3 ≥ 6 to < 15; n = 49). RESULTS: While accounting for age, examiner and experimental plate, parkinsonism cases had lower mean NOS2 methylation than controls (p-value for trend = 0.04), specifically at CpG site 8329 located in an exonic splicing enhancer of NOS2 (p-value for trend = 0.07). These associations were not observed for the intermediate UPDRS3 group (both p-value for trend ≥ 0.59). CONCLUSIONS: Inflammation mediated by inducible nitric oxide synthase may possibly contribute to the association between welding fume and parkinsonism, but requires verification in a longitudinal study.

Transcriptional impact of organophosphate and metal mixtures on olfaction: copper dominates the chlorpyrifos-induced response in adult zebrafish.

Chemical exposures in fish have been linked to loss of olfaction leading to an inability to detect predators and prey and decreased survival. However, the mechanisms underlying olfactory neurotoxicity are not well characterized, especially in environmental exposures which involve chemical mixtures. We used zebrafish to characterize olfactory transcriptional responses by two model olfactory inhibitors, the pesticide chlorpyrifos (CPF) and mixtures of CPF with the neurotoxic metal copper (Cu). Microarray analysis was performed on RNA from olfactory tissues of zebrafish exposed to CPF alone or to a mixture of CPF and Cu. Gene expression profiles were analyzed using principal component analysis and hierarchical clustering, whereas gene set analysis was used to identify biological themes in the microarray data. Microarray results were confirmed by real-time PCR on genes serving as potential biomarkers of olfactory injury. In addition, we mined our previously published Cu-induced zebrafish olfactory transcriptional response database (Tilton et al., 2008) for the purposes of discriminating pathways of olfaction impacted by either the individual agents or the CPF-Cu mixture transcriptional signatures. CPF exposure altered the expression of gene pathways associated with cellular morphogenesis and odorant binding, but not olfactory signal transduction, a known olfactory pathway for Cu. The mixture profiles shared genes from the Cu and CPF datasets, whereas some genes were altered only by the mixtures. The transcriptional signature of the mixtures was more similar to that in zebrafish exposed to Cu alone than for CPF. In conclusion, exposure to a mixture containing a common environmental metal and pesticide causes a unique transcriptional signature that is heavily influenced by the metal, even when organophosphate predominates.

Sulforaphane- and phenethyl isothiocyanate-induced inhibition of aflatoxin B1-mediated genotoxicity in human hepatocytes: role of GSTM1 genotype and CYP3A4 gene expression.

Primary cultures of human hepatocytes were used to investigate whether the dietary isothiocyanates, sulforaphane (SFN), and phenethyl isothiocyanate (PEITC) can reduce DNA adduct formation of the hepatocarcinogen aflatoxin B(1) (AFB). Following 48 h of pretreatment, 10 and 50 microM SFN greatly decreased AFB-DNA adduct levels, whereas 25muM PEITC decreased AFB-DNA adducts in some but not all hepatocyte preparations. Microarray and quantitative reverse transcriptase (RT)-PCR analyses of gene expression in SFN and PEITC-treated hepatocytes demonstrated that SFN greatly decreased cytochrome P450 (CYP) 3A4 mRNA but did not induce the expression of either glutathione S-transferase (GST) M1 or GSTT1. The protective effects of SFN required pretreatment; cotreatment of hepatocytes with SFN and AFB in the absence of pretreatment had no effect on AFB-DNA adduct formation. When AFB-DNA adduct formation was evaluated by GST genotype, the presence of one or two functional alleles of GSTM1 was associated with a 75% reduction in AFB-DNA adducts, compared with GSTM1 null. In conclusion, these results demonstrate that the inhibition of AFB-DNA adduct formation by SFN is dependent on changes in gene expression rather than direct inhibition of catalytic activity. Transcriptional repression of genes involved in AFB bioactivation (CYP3A4 and CYP1A2), but not transcriptional activation of GSTs, may be responsible for the protective effects of SFN. Although GSTM1 expression was not induced by SFN, the presence of a functional GSTM1 allele can afford substantial protection against AFB-DNA damage in human liver. The downregulation of CYP3A4 by SFN may have important implications for drug interactions.

Modulation of aflatoxin B1-mediated genotoxicity in primary cultures of human hepatocytes by diindolylmethane, curcumin, and xanthohumols.

This study employed cultured human primary hepatocytes to investigate the ability of the putative chemopreventive phytochemicals curcumin (CUR), 3,3'-diindolylmethane (DIM), isoxanthohumol (IXN), or 8-prenylnaringenin (8PN) to reduce DNA adduct formation of the hepatocarcinogen aflatoxin B1 (AFB). Following 48 h of pretreatment, DIM and 8PN significantly increased AFB-DNA adduct levels, whereas CUR and IXN had no effect. DIM greatly enhanced the transcriptional expression of cytochrome P450 (CYP) 1A1 and CYP1A2 mRNA. Glutathione S-transferase mRNAs were not increased by any of the treatments. In vitro enzyme activity assays demonstrated that 8PN and DIM, but not CUR or IXN, inhibited human CYP1A1, CYP1A2, and CYP3A4 activities. To distinguish between treatment effects on transcription versus direct effects on enzyme activity for DIM, we evaluated the effects of pretreatment alone (transcriptional activation) versus cotreatment alone (enzyme inhibition). The results demonstrated that effects on gene expression, but not catalytic activity, are responsible for the observed effects of DIM on AFB-DNA adduct formation. The increase in AFB-DNA damage following DIM treatment may be explained through its substantial induction of CYP1A2 and/or its downregulation of GSTM1, both of which were significant. The increase in DNA damage by DIM raises potential safety risks for dietary supplements of DIM and its precursor indole-3-carbinol.

Gene expression profiles in zebrafish brain after acute exposure to domoic acid at symptomatic and asymptomatic doses.

Domoic acid (DA) is a neuroexcitatory amino acid that is naturally produced by some marine diatom species of the genus Pseudo-nitzschia. Ingestion of DA-contaminated seafood by humans results in a severe neurotoxic disease known as amnesic shellfish poisoning (ASP). Clinical signs of ASP include seizures and neuronal damage from activation of ionotropic glutamate receptors. However, the impacts of DA exposure at levels below those known to induce outward signs of neurobehavioral exicitotoxicity have not been well characterized. To further understand the mechanisms of neurotoxic injury associated with DA exposure, we examined the transcriptome of whole brains from zebrafish (Danio rerio) receiving intracoelomic (IC) injection of DA at both symptomatic and asymptomatic doses. A majority of zebrafish exposed to high-dose DA (1.2 microg DA/g) exhibited clinical signs of neuroexcitotoxicity (EC(50) of 0.86 microg DA/g) within 5-20 min of IC injection. All zebrafish receiving low-dose DA (0.47 microg DA/g) or vehicle only maintained normal behavior. Microarray analysis of symptomatic and asymptomatic exposures collectively yielded 306 differentially expressed genes (1.5-fold, p

Role of cytochrome P450 2C8 and 2J2 genotypes in calcineurin inhibitor-induced chronic kidney disease.

OBJECTIVES: The calcineurin inhibitors (CNIs) cyclosporine A (CsA) and tacrolimus (Tac) help prevent allograft rejection but are associated with nephrotoxicity. Cytochrome P450 2C8 (CYP2C8) and CYP2J2 are polymorphic enzymes expressed in the kidney that metabolize arachidonic acid (AA) to epoxyeicosatrienoic acids, promoting kidney homeostasis. This study examined the association between CNI-induced nephrotoxicity in liver transplant patients and CYP2C8 and CYP2J2 polymorphisms. METHODS: Liver transplantation patients receiving CNIs for at least 3 years were genotyped for CYP2C8*3, CYP2C8*4, CYP2C8 Haplotypes B and C, and CYP2J2*7 and evaluated for nephrotoxicity (serum creatinine > or = 1.6 mg/dl) 3-year post-transplantation. CYP2C8 proteins were also engineered in E. coli and their activity towards AA and inhibition by CNIs was investigated in vitro. RESULTS: The risk of kidney disease post-transplantation was positively associated with CYP2C8*3 genotype. Odds ratios for all participants carrying at least one CYP2C8*3 allele were significant [odds ratio=2.38 (1.19-4.78)]. Stratification by CNI indicated a significant association between CYP2C8*3 and nephrotoxicity among patients receiving Tac but not CsA. The risk of renal dysfunction was not significantly influenced by CYP2C8*4, CYP2J2*7, or CYP2C8 haplotype B genotypes although inheritance of haplotype C seems to be protective. In vitro, the gene products of CYP2C8*3 and CYP2C8*4 were deficient in AA epoxidation, retaining 26 and 18% of wild-type activity, respectively. Circulating plasma concentrations of CsA and Tac inhibited CYP2C8 wild-type in vitro epoxidation of AA by 17 and 35%, respectively. CONCLUSION: Inheritance of CYP2C8*3 is associated with a higher risk of developing renal toxicity in patients treated chronically with CNIs, and especially Tac, possibly by reducing formation of kidney protecting vasodilatory epoxyeicosatrienoic acids.

Hepatic expression profiling in smolting and adult coho salmon (Onchorhynchus kisutch).

Coho salmon are a critical Pacific salmon species that undergo complex physiological transformations as they migrate towards seawater and enter adult life stages. During these periods, coho may receive exposure to waterborne pollutants that coincide with outmigration through contaminated waterways and return to natal streams. However, little is known regarding the ontogenic modulation of gene expression during these critical life stages. Accordingly, the purpose of the present study was to characterize the hepatic transcriptome of smolting coho, adult males, and adult females by carrying out microarray analysis with a commercially available 16,000 cDNA element platform. Quantitative PCR (Q-PCR) analysis of genes involved in chemical biotransformation (cytochrome P450 isoforms 1A, and 2M1, glutathione S-transferase pi, microsomal GST), defense against metal exposure (metallothionein-A), and reproductive function (vitellogenin receptor) were developed for the purpose of analyzing specific genes of interest and to validate the microarray data. Microarray analysis identified 842 genes that were differentially expressed between smolts and adult males or females (p<0.001 and more than 2-fold difference). These 842 genes were not differentially expressed between adult males and females and, therefore, can be interpreted as a smolt-specific transcriptional profile. Of these 842 genes, 275 were well annotated and formed the basis for further bioinformatics analysis. Many of the differentially expressed genes were involved in basic cellular processes related to protein biosynthesis and degradation (24%), ion transport (12%), transcription (8%), cell structure (8%) and cellular energetics (6%). The majority of differentially expressed genes involved in signal transduction and energy metabolism were expressed at higher levels in adult coho relative to smolts. However, genes associated with cellular protection against chemical injury (i.e. biotransformation, DNA damage repair, and protection against oxidative stress) did not generally differ among the groups. Q-PCR studies revealed extensive interindividual variation in mRNA expression, but were consistent with the microarray results (R(2)=0.74). Collectively, our results indicate differences in liver gene expression in young smolting coho salmon relative to adults and extensive interindividual variation in biotransformation gene expression. However, we did not find a global lack of hepatic biotransformation capacity or poor cellular detoxification response capacity in smolting cohos based on mRNA profiles.

Transcriptional biomarkers and mechanisms of copper-induced olfactory injury in zebrafish.

Metals such as copper disrupt olfactory function in fish. Unfortunately, little is understood of the molecular consequences of copper olfactory impairment, thus hindering the development of relevant diagnostic tools of olfactory injury. To address this critical data gap, we analyzed gene expression within olfactory tissues of adult zebrafish exposed to CuCl2 (6, 16, 40 ppb) for 24 h. Transcriptional markers of copper impairment within the entire olfactory system were identified and specific genes of interest (e.g., S100a, parvalbumin 8, olfactory marker protein, and calbindin 2-like protein) were confirmed with quantitative real-time PCR. In addition, we performed gene set analysis (GSA) using both a priori and custom pathways of gene sets specifically targeting the olfactory signal transduction (OST) pathway. These analyses revealed down-regulated gene sets related to calcium channels and ion transport, g-proteins, and olfactory receptors. Collectively, these data demonstrate that copper causes a depression of transcription of key genes within the OST pathway and elsewhere within olfactory tissues, likely resulting in an olfactory system less responsive to odorants. Further, these data provide a mechanistic explanation in support of earlier studies of functional olfactory impairment in fish following copper exposure.

Association of estrogen-related polymorphisms with age at menarche, age at final menstrual period, and stages of the menopausal transition.

OBJECTIVE: To explore the association of estrogen-related polymorphisms with age at menarche, age at onset and duration of stages of the menopausal transition, and age at final menstrual period (FMP). DESIGN: A total of 152 white women were genotyped for CYP17, CYP19 3-untranslated region, CYP19 TTTA7-13, HSDB1, CYP1A1, CYP1B1, and ESR1 polymorphisms. Analysis of variance was used to test a nonspecific model for differences among genotypes associated with each polymorphism. RESULTS: Five of the 84 associations tested were significant at P < 0.05, which could be expected by chance. Women with two CYP19 7r alleles had menarche earlier (11.5 y) than those with one 7r allele (13.1 y). Women with two 11r alleles were 2 years older at onset of late stage than those with one 11r allele (50.7 y vs 48.6 y). Those with two 7r(-3) alleles were 2 years older at FMP than those without this allele (53.9 y vs 51.3 y). Women with the homozygous wild-type allele for HSDB1 (rs2830) were younger at FMP by 2 years than those with the heterozygous allele (50.8 y vs 52.9 y). Women with the heterozygous allele for CYP1B1*2 had a later age at menarche compared with women with the homozygous wild type (13 y vs 12.5 y). CONCLUSIONS: Age at onset of late stage and FMP and age at menarche are associated with specific genetic polymorphisms in the estrogen biosynthesis and metabolism genes. However, because of the number of comparisons, these associations may be false positives. These findings should be confirmed with a larger sample of white women.

Characterization of Phase I biotransformation enzymes in coho salmon (Oncorhynchus kisutch).

Wild stocks of Pacific salmon in the Northwestern United States have declined in recent years, and the major factors contributing to these losses include water pollution and loss of habitat. In salmon, sublethal chemical exposures may impact critical behaviors (such as homing, feeding, predator-avoidance) that are important for species survival. Therefore, understanding the potential for these species to biotransform organic compounds within sensitive target tissues such as liver, gills and olfactory region can help estimate or predict their susceptibility to pollutants. In this study, we used real-time quantitative polymerase chain reaction (Q-PCR), Western blotting, and catalytic assays to characterize the expression of Phase I biotransformation enzymes in coho salmon (Oncorhynchus kisutch), a sensitive species in the Pacific Northwest. Gene expression analysis using Q-PCR assays developed for coho genes revealed the presence of the predominant cytochrome P450 mRNAs (CYP1A, CYP2K1, CYP2M1, CYP3A27) in the olfactory rosettes and provided quantitative mRNA expression levels in coho liver and gills. Q-PCR analysis revealed relatively high expression of the major CYP isoforms in the liver and olfactory rosettes, which was generally confirmed by Western blotting. Extrahepatic CYP expression was generally higher in the olfactory rosettes as compared to the gills. Catalytic studies demonstrated functional CYP1A-dependent ethoxyresorufin-O-deethylase, CYP2-dependent pentoxyresorufin-O-dealkylase, CYP2K1-dependent testosterone 16beta-hydroxylase, and CYP3A27-dependent testosterone 6beta-hydroxylase activities in liver, but not at detectable levels in gills. In contrast, flavin-containing monooxygenase (FMO)-dependent thiourea S-oxidase activity was readily observed in the gills and was substantially higher than that observed in liver. Collectively, the results of this study suggest that the olfactory rosettes are important sites of extrahepatic biotransformation in coho salmon, and that tissue specific-differences in Phase I metabolism may lead to contrasting tissue-specific biotransformation capabilities in this species.

Cytochrome p450 and glutathione s-transferase mRNA expression in human fetal liver hematopoietic stem cells.

During fetal development, the liver serves as the primary hematopoietic organ in which hematopoietic stem cells (HSC) capable of initiating long-term hematopoiesis comprise a large proportion of the hepatic cell population. Although HSC are potential targets for transplacental chemicals, little is known regarding their xenobiotic biotransformation ability. We quantitated the steady-state mRNA expression of six cytochrome P450 (P450) and 11 glutathione S-transferase (GST) isoforms in CD34(+)-selected HSC isolated from second trimester human fetal liver donors, genotyped donors for polymorphic hGSTM1 and hGSTT1 status, and analyzed gene expression in HSC relative to total liver from donors of similar gestational ages. Several P450 isoforms, including CYP1A1, CYP2E1, CYP3A4, and CYP3A5, were expressed at low levels in HSC (relative mRNA expression CYP3A5 > CYP1A1 > CYP2E1 > CYP3A4). CYP1A2 and CYP3A7 were not detected in HSC. The CYP3A4/5 mRNA expression in HSC was accompanied by detectable CYP3A protein and low midazolam oxidation activity. Several GST isoforms, including hGSTM1, hGSTM2, hGSTM4, and hGSTP1, were significantly higher in HSC as compared with total fetal liver. With the exception of hGSTA4, alpha class GST were not detected in HSC. GST expression in HSC was accompanied by substantial GST catalytic activity toward 1-chloro-2,4-dinitrobenzene. In summary, our data indicate that fetal liver CD34(+)-derived HSC constitutively express several P450 isoforms at low levels relative to total hepatic cell populations but have a higher capacity for GST conjugation reactions through mu and pi class isoforms. The functional ramifications of these observations are discussed relative to the sensitivity of human fetal HSC to transplacental chemical injury.

Polymorphisms in the estrogen synthesis and metabolism pathways and symptoms during the menopausal transition: observations from the Seattle Midlife Women's Health Study.

OBJECTIVE: The purpose of this study was to determine whether polymorphisms in the estrogen synthesis and metabolism pathways are associated with women's vasomotor symptom experiences during the menopausal transition. DESIGN: In 2002, a subset of women enrolled in the Seattle Midlife Women's Health Study since 1990 (N = 174) provided a buccal smear for genotyping. Women were recruited by complete ascertainment of selected multiethnic neighborhoods in 1990. Participants were midlife women with a mean age of 53 years in 2005, well educated, employed, married, and represented a multiethnic population. Genotyping was done for the following polymorphisms: CYP1A1m2; CYP1B1*2 and CYP1B1*3; CYP17 5'UTR; CYP19 3'UTR; CYP19 (TTTA)n; including CYP19 7r and CYP19 7(r-3); CYP19 8r and CYP19 11r; and ESR1PvuII and ESR1XbaI. Women rated their vasomotor symptom severity in diaries on days 5, 6, and 7 of the menstrual cycle or on a constant date each month for women skipping periods. Menopausal transition stage was determined from daily menstrual calendars. First voided urine specimens provided several times each year were assayed for estrone glucuronide. RESULTS: Women with the CYP19 11r polymorphism reported more severe and frequent hot flashes during the middle and late menopausal transition stages and postmenopause and higher E1G levels during middle and late stages. None of the other polymorphisms studied were related to hot flashes or to estrone glucuronide levels. CONCLUSIONS: These findings suggest a possible role for CYP19 polymorphisms in estrogen levels and in vasomotor symptoms during the menopausal transition that warrants further study in larger and more diverse populations of women.

The effect of CYP19 and COMT polymorphisms on exercise-induced fat loss in postmenopausal women.

OBJECTIVE: To examine whether genetic polymorphisms in CYP19 [intron 4 (TTTA)n; n = 7 to 13 and a 3-base pair deletion, which is in strong linkage disequilibrium with the seven repeat] and COMT (Val108/158Met) modified the change in BMI, total and percentage body fat, or subcutaneous and intra-abdominal fat during a year-long exercise intervention trial. These genes metabolize estrogens and androgens, which are important in body fat regulation. RESEARCH METHODS AND PROCEDURES: A randomized intervention trial was used, with an intervention goal of 225 min/wk of moderate-intensity exercise for one year. Participants (n = 173) were postmenopausal, 50 to 75 years old, sedentary, overweight or obese, and not taking hormone therapy at baseline. RESULTS: Exercisers with two vs. no CYP19 11-repeat alleles had a larger decrease in total fat (-3.1 kg vs. -0.5 kg, respectively, p = 0.01) and percentage body fat (-2.4% vs. -0.6%, respectively, p = 0.001). Exercisers with the COMT Met/Met vs. Val/Val genotype had a smaller decrease in percentage fat (-0.7% vs. -1.9%, respectively, p = 0.05). Among exercisers, women with the COMT Val/Val genotype and at least one copy of the CYP19 11-repeat allele vs. those with neither genotype/allele had a significantly larger decrease in BMI (-1.0 vs. +0.1 kg/m2, respectively, p = 0.009), total fat (-2.9 vs. -0.5 kg, respectively, p = 0.004), and percentage body fat (-2.6% vs. -0.4%, respectively, p < 0.001). DISCUSSION: Genetic polymorphisms in CYP19 and COMT may be important for body fat regulation and possibly modify the effect of exercise on fat loss in postmenopausal women.

Association of CYP17, CYP19, CYP1B1, and COMT polymorphisms with serum and urinary sex hormone concentrations in postmenopausal women.

Women with high circulating estrogen concentrations have an increased risk of breast cancer; thus, it is important to understand factors, including genetic variability, that influence estrogen concentrations. Several genetic polymorphisms that may influence sex hormone concentrations have been identified, including CYP17 (5'-untranslated region T-->C), CYP19 [intron 4 (TTTA)(n = 7-13) and a 3-bp deletion (-3)], CYP1B1 (Val(432)Leu), and COMT (Val(108/158)Met). We examined associations between these polymorphisms and serum concentrations of estrogens, androgens, and sex hormone-binding globulin and urinary concentrations of 2- and 16alpha-hydroxyestrone in 171 postmenopausal women, using data from the prerandomization visit of an exercise clinical trial. Participants were sedentary, not taking hormone therapy, and had a body mass index >24.0. Compared with noncarriers, women carrying two CYP19 7r(-3) alleles had 26% lower estrone (P < 0.001), 19% lower estradiol (P = 0.01), 23% lower free estradiol (P = 0.01), and 22% higher sex hormone-binding globulin concentrations (P = 0.06). Compared with noncarriers, women carrying at least one CYP19 8r allele had 20% higher estrone (P = 0.003), 18% higher estradiol (P = 0.02), and 21% higher free estradiol concentrations (P = 0.01). Women with the COMT Met/Met genotype had 28% higher 2-hydroxyestrone (P = 0.08) and 31% higher 16alpha-hydroxyestrone concentrations (P = 0.02), compared with Val/Val women. Few associations were found for CYP17 and CYP1B1 or with serum androgen concentrations. This study provides further evidence that genetic variation may appreciably alter sex hormone concentrations in postmenopausal women not taking hormone therapy.

Glutathione S-transferase M1, T1, and P1 polymorphisms and survival among lung cancer patients.

Glutathione S-transferase (GST) enzymes detoxify therapeutic drugs and reactive oxidants, so GST polymorphisms may influence survival after diagnosis of cancer. We evaluated survival according to GST polymorphisms in a population-based series of lung cancer patients. The study subjects (n = 274) were men diagnosed with lung cancer from 1993 through 1996 who participated in a case control study and provided a blood sample for genotyping. The presence of the GSTM1 and GSTT1 genes were assayed by multiplex PCR. Genotype at the GSTP1 Ile(105)Val substitution was determined by PCR and oligonucleotide ligation assay. The study subjects were followed for vital status through 2000, and overall survival was evaluated in Kaplan-Meier survival functions and Cox proportional hazards models. Subjects with the GSTM1 null genotype had shorter survival; the proportion of GSTM1 null subjects surviving at 5 years was 0.20 [95% confidence interval (CI) 0.14-0.27], compared with 0.29 (95% CI 0.22-0.37) for GSTM1 present subjects. The relative risk of death associated with GSTM1 null genotype, adjusted for stage at diagnosis and histology, was 1.36, 95% CI 1.04-1.80. There was no association between GSTT1 or GSTP1 genotype and survival in the overall study population, nor in a subgroup of patients treated with chemotherapy (n = 130). For GSTM1, our results are consistent with a previous study, which also observed that the GSTM1-null genotype, which confers susceptibility to lung cancer, was associated with shorter survival. Future studies of lung cancer survival should take into account GSTM1 genotype as well as investigate underlying mechanisms.

Glutathione S-transferase M1, T1, and P1 polymorphisms and Parkinson's disease.

Oxidative stress is widely thought to contribute significantly to the pathogenesis Parkinson's disease (PD). Given the role of glutathione S-transferases (GSTs) in the conjugation of electrophiles and protection against reactive oxygen species, genes encoding the GSTs have been considered candidates for association studies of PD. We tested for associations between genotypes of GSTM1(homozygous deletion vs. non-deleted), GSTT1(homozygous deletion vs. non-deleted), and GSTP1 (Ile104Val and Ala113Val) and PD in a case-control study of 214 idiopathic PD cases and 330 age- and gender-matched, unrelated controls of Caucasian ethnicity. No significant associations with any of the GST genotypes were observed. However, there was a marginally significant difference in the distribution of GSTP1 104 genotypes between cases and controls (P=0.07), with an excess of Ile104Val heterozygotes found among cases (odds ratio (OR)=1.43; 95% Confidence Interval (CI): 0.98-2.08). This difference in the genotype distribution was strongest among smokers (OR for heterozygote=1.92; 95% CI: 1.12-3.29) versus non-smokers and among males (OR for heterozygote=1.99; 95% CI: 1.24-3.19) versus females. The distribution of GSTP1 Ile104Val and Ala113Val haplotypes did not differ between cases and controls. Taken together, these results suggest a potentially minor role of GSTP1 in PD, but do not give evidence for associations with either GSTM1 or GSTT1.