Early-Life Exposure to Air Pollution and Childhood Asthma Cumulative Incidence in the ECHO CREW Consortium.

Importance: Exposure to outdoor air pollution contributes to childhood asthma development, but many studies lack the geographic, racial and ethnic, and socioeconomic diversity to evaluate susceptibility by individual-level and community-level contextual factors. Objective: To examine early life exposure to fine particulate matter (PM2.5) and nitrogen oxide (NO2) air pollution and asthma risk by early and middle childhood, and whether individual and community-level characteristics modify associations between air pollution exposure and asthma. Design, Setting, and Participants: This cohort study included children enrolled in cohorts participating in the Children's Respiratory and Environmental Workgroup consortium. The birth cohorts were located throughout the US, recruited between 1987 and 2007, and followed up through age 11 years. The survival analysis was adjusted for mother's education, parental asthma, smoking during pregnancy, child's race and ethnicity, sex, neighborhood characteristics, and cohort. Statistical analysis was performed from February 2022 to December 2023. Exposure: Early-life exposures to PM2.5 and NO2 according to participants' birth address. Main Outcomes and Measures: Caregiver report of physician-diagnosed asthma through early (age 4 years) and middle (age 11 years) childhood. Results: Among 5279 children included, 1659 (31.4%) were Black, 835 (15.8%) were Hispanic, 2555 (48.4%) where White, and 229 (4.3%) were other race or ethnicity; 2721 (51.5%) were male and 2596 (49.2%) were female; 1305 children (24.7%) had asthma by 11 years of age and 954 (18.1%) had asthma by 4 years of age. Mean values of pollutants over the first 3 years of life were associated with asthma incidence. A 1 IQR increase in NO2 (6.1 µg/m3) was associated with increased asthma incidence among children younger than 5 years (HR, 1.25 [95% CI, 1.03-1.52]) and children younger than 11 years (HR, 1.22 [95% CI, 1.04-1.44]). A 1 IQR increase in PM2.5 (3.4 µg/m3) was associated with increased asthma incidence among children younger than 5 years (HR, 1.31 [95% CI, 1.04-1.66]) and children younger than 11 years (OR, 1.23 [95% CI, 1.01-1.50]). Associations of PM2.5 or NO2 with asthma were increased when mothers had less than a high school diploma, among Black children, in communities with fewer child opportunities, and in census tracts with higher percentage Black population and population density; for example, there was a significantly higher association between PM2.5 and asthma incidence by younger than 5 years of age in Black children (HR, 1.60 [95% CI, 1.15-2.22]) compared with White children (HR, 1.17 [95% CI, 0.90-1.52]). Conclusions and Relevance: In this cohort study, early life air pollution was associated with increased asthma incidence by early and middle childhood, with higher risk among minoritized families living in urban communities characterized by fewer opportunities and resources and multiple environmental coexposures. Reducing asthma risk in the US requires air pollution regulation and reduction combined with greater environmental, educational, and health equity at the community level.

Socioeconomic Status at Birth and Breast Tissue Composition in Adolescence and Adulthood.

BACKGROUND: Socioeconomic status (SES) at birth is associated with breast cancer risk. Whether this association is driven by changes in breast tissue composition (BTC) prior to adulthood remains unclear. METHODS: We used multivariable linear regression models to examine whether SES at birth is associated with BTC in adolescence and adulthood using data from a New York City cohort of daughters (n = 165, 11-20 years) and mothers (n = 160, 29-55 years). We used maternal-reported data on daughters' household income and maternal education at birth, analyzed individually and in combination (SES index). Women also reported their own mothers' education at birth. We used optical spectroscopy to evaluate BTC measures that positively (water content, collagen content, optical index) and negatively (lipid content) correlate with mammographic breast density, a recognized breast cancer risk factor. RESULTS: Being in the highest versus lowest category of the SES index was associated with lower lipid content [ßadjusted (ßadj) = -0.80; 95% confidence interval (CI), -1.30 to -0.31] and higher collagen content (ßadj = 0.54; 95% CI, 0.09-0.99) in adolescence. In women with a body mass index (BMI) <30 kg/m2, higher maternal education at birth (≥ vs. < high school degree) was associated with lower lipid content (ßadj = -0.57; 95% CI, -0.97 to -0.17), higher water content (ßadj = 0.70; 95% CI, 0.26-1.14), and higher optical index (ßadj = 0.53; 95% CI, 0.10-0.95). CONCLUSIONS: This study supports that SES at birth is associated with BTC in adolescence and adulthood, although the latter association may depend on adult BMI. IMPACT: Further research is needed to identify the socially patterned early life factors influencing BTC.

Prenatal airborne polycyclic aromatic hydrocarbon exposure, altered regulation of peroxisome proliferator-activated receptor gamma (Ppar)γ, and links with mammary cancer.

Environmental exposure to polycyclic aromatic hydrocarbons (PAH) has been shown to be associated with chronic disease outcomes through multiple mechanisms including altered regulation of the transcription factor peroxisome proliferator-activated receptor gamma (Ppar) γ. Because PAH exposure and Pparγ each have been associated with mammary cancer, we asked whether PAH would induce altered regulation of Pparγ in mammary tissue, and whether this association may underlie the association between PAH and mammary cancer. Pregnant mice were exposed to aerosolized PAH at proportions that mimic equivalent human exposures in New York City air. We hypothesized that prenatal PAH exposure would alter Pparγ DNA methylation and gene expression and induce the epithelial to mesenchymal transition (EMT) in mammary tissue of offspring (F1) and grandoffspring (F2) mice. We also hypothesized that altered regulation of Pparγ in mammary tissue would associate with biomarkers of EMT, and examined associations with whole body weight. We found that prenatal PAH exposure lowered Pparγ mammary tissue methylation among grandoffspring mice at postnatal day (PND) 28. However, PAH exposure did not associate with altered Pparγ gene expression or consistently with biomarkers of EMT. Finally, lower Pparγ methylation, but not gene expression, was associated with higher body weight among offspring and grandoffspring mice at PND28 and PND60. Findings suggest additional evidence of multi-generational adverse epigenetic effects of prenatal PAH exposure among grandoffspring mice.

Characteristics of peak exposure to black carbon pollution in school, commute and home environments among school children in an urban community.

Although real-time personal exposure monitoring devices have the ability to capture a wealth of data regarding fluctuations in pollutant levels, only a few studies have defined 'peaks' in black carbon (BC) exposure utilizing high-resolution data. Furthermore, studies to assess and characterize various features of peak exposure are very limited especially among children. A better understanding of characteristics of BC peak exposure would improve our understanding of health risks associated with BC. By capturing personal BC exposure at 5-min intervals using a real-time monitor during 24-hr monitoring periods among children in New York City (NYC), we defined 'peak characteristics' in 4 different ways across three major microenvironments (school vs. commute vs. home): 1) mean concentrations of BC across the 3 microenvironments, 2) 'peak duration' or time spent above the peak threshold (i.e., ≥1.5 µg/m3), 3) 'peak intensity' or the rate of exposure, defined as time spent above the threshold within each microenvironment divided by the total time spent in the microenvironment and 4) a novel metric of 'peak variability', defined as frequency of peaks (i.e., data points with +50% and -50% changes compared to the preceding and the subsequent data points), divided by the total time spent in the microenvironment. While peak duration was greatest at home, the intensity of peak exposure was greatest during commute hours, despite the short time spent in commute (p < 0.05). Peak variability was highest during commute, yet lowest in home environments (p < 0.05), particularly during non-sleeping hours. Children residing in a high-density urban setting spent on average, 5.4 hr per day above our peak threshold (≥1.5 µg/m3) in their everyday environments. Policies that limit children's exposure during high traffic periods and improved efforts to increase the number of vehicles using clean air technology could reduce the intensity of peaks and peak variability in children's BC exposure.

African-specific alleles modify risk for asthma at the 17q12-q21 locus in African Americans.

BACKGROUND: Asthma is the most common chronic disease in children, occurring at higher frequencies and with more severe disease in children with African ancestry. METHODS: We tested for association with haplotypes at the most replicated and significant childhood-onset asthma locus at 17q12-q21 and asthma in European American and African American children. Following this, we used whole-genome sequencing data from 1060 African American and 100 European American individuals to identify novel variants on a high-risk African American-specific haplotype. We characterized these variants in silico using gene expression and ATAC-seq data from airway epithelial cells, functional annotations from ENCODE, and promoter capture (pc)Hi-C maps in airway epithelial cells. Candidate causal variants were then assessed for correlation with asthma-associated phenotypes in African American children and adults. RESULTS: Our studies revealed nine novel African-specific common variants, enriched on a high-risk asthma haplotype, which regulated the expression of GSDMA in airway epithelial cells and were associated with features of severe asthma. Using ENCODE annotations, ATAC-seq, and pcHi-C, we narrowed the associations to two candidate causal variants that are associated with features of T2 low severe asthma. CONCLUSIONS: Previously unknown genetic variation at the 17q12-21 childhood-onset asthma locus contributes to asthma severity in individuals with African ancestries. We suggest that many other population-specific variants that have not been discovered in GWAS contribute to the genetic risk for asthma and other common diseases.

Exposure to polycyclic aromatic hydrocarbons during pregnancy and breast tissue composition in adolescent daughters and their mothers: a prospective cohort study.

BACKGROUND: Polycyclic aromatic hydrocarbons (PAH), which are found in air pollution, have carcinogenic and endocrine disrupting properties that might increase breast cancer risk. PAH exposure might be particularly detrimental during pregnancy, as this is a time when the breast tissue of both the mother and daughter is undergoing structural and functional changes. In this study, we tested the hypothesis that ambient PAH exposure during pregnancy is associated with breast tissue composition, measured one to two decades later, in adolescent daughters and their mothers. METHODS: We conducted a prospective analysis using data from a New York City cohort of non-Hispanic Black and Hispanic mother-daughter dyads (recruited 1998-2006). During the third trimester of pregnancy, women wore backpacks containing a continuously operating air sampling pump for two consecutive days that measured ambient exposure to eight carcinogenic higher molecular weight nonvolatile PAH compounds (Σ8 PAH) and pyrene. When daughters (n = 186) and mothers (n = 175) reached ages 11-20 and 29-55 years, respectively, optical spectroscopy (OS) was used to evaluate measures of breast tissue composition (BTC) that positively (water content, collagen content, optical index) and negatively (lipid content) correlate with mammographic breast density, a recognized risk factor for breast cancer. Multivariable linear regression was used to evaluate associations between ambient PAH exposure and BTC, overall and by exposure to household tobacco smoke during pregnancy (yes/no). Models were adjusted for race/ethnicity, age, and percent body fat at OS. RESULTS: No overall associations were found between ambient PAH exposure (Σ8 PAH or pyrene) and BTC, but statistically significant additive interactions between Σ8 PAH and household tobacco smoke exposure were identified for water content and optical index in both daughters and mothers (interaction p values < 0.05). Σ8 PAH exposure was associated with higher water content (ßdaughters = 0.42, 95% CI = 0.15-0.68; ßmothers = 0.32, 95% CI = 0.05-0.61) and higher optical index (ßdaughters = 0.38, 95% CI = 0.12-0.64; ßmothers = 0.38, 95% CI = 0.12-0.65) in those exposed to household tobacco smoke during pregnancy; no associations were found in non-smoking households (interaction p values < 0.05). CONCLUSIONS: Exposure to ambient Σ8 PAH and tobacco smoke during pregnancy might interact synergistically to impact BTC in mothers and daughters. If replicated in other cohorts, these findings might have important implications for breast cancer risk across generations.

Childhood Asthma Incidence, Early and Persistent Wheeze, and Neighborhood Socioeconomic Factors in the ECHO/CREW Consortium.

Importance: In the United States, Black and Hispanic children have higher rates of asthma and asthma-related morbidity compared with White children and disproportionately reside in communities with economic deprivation. Objective: To determine the extent to which neighborhood-level socioeconomic indicators explain racial and ethnic disparities in childhood wheezing and asthma. Design, Setting, and Participants: The study population comprised children in birth cohorts located throughout the United States that are part of the Children's Respiratory and Environmental Workgroup consortium. Cox proportional hazard models were used to estimate hazard ratios (HRs) of asthma incidence, and logistic regression was used to estimate odds ratios of early and persistent wheeze prevalence accounting for mother's education, parental asthma, smoking during pregnancy, child's race and ethnicity, sex, and region and decade of birth. Exposures: Neighborhood-level socioeconomic indicators defined by US census tracts calculated as z scores for multiple tract-level variables relative to the US average linked to participants' birth record address and decade of birth. The parent or caregiver reported the child's race and ethnicity. Main Outcomes and Measures: Prevalence of early and persistent childhood wheeze and asthma incidence. Results: Of 5809 children, 46% reported wheezing before age 2 years, and 26% reported persistent wheeze through age 11 years. Asthma prevalence by age 11 years varied by cohort, with an overall median prevalence of 25%. Black children (HR, 1.47; 95% CI, 1.26-1.73) and Hispanic children (HR, 1.29; 95% CI, 1.09-1.53) were at significantly increased risk for asthma incidence compared with White children, with onset occurring earlier in childhood. Children born in tracts with a greater proportion of low-income households, population density, and poverty had increased asthma incidence. Results for early and persistent wheeze were similar. In effect modification analysis, census variables did not significantly modify the association between race and ethnicity and risk for asthma incidence; Black and Hispanic children remained at higher risk for asthma compared with White children across census tracts socioeconomic levels. Conclusions and Relevance: Adjusting for individual-level characteristics, we observed neighborhood socioeconomic disparities in childhood wheeze and asthma. Black and Hispanic children had more asthma in neighborhoods of all income levels. Neighborhood- and individual-level characteristics and their root causes should be considered as sources of respiratory health inequities.

Cancer Risk Reduction Through Education of Adolescents: Development of a Tailored Cancer Risk-Reduction Educational Tool.

Growing evidence links adolescent exposures to cancer risk later in life, particularly for common cancers like breast. The adolescent time period is also important for cancer risk reduction as many individual lifestyle behaviors are initiated including smoking and alcohol use. We developed a cancer risk-reduction educational tool tailored for adolescents that focused on five modifiable cancer risk factors. To contextualize risk factors in adolescents' social and physical environments, the intervention also focused on structural barriers to individual- and community-level change, with an emphasis on environmental justice or the fair treatment and meaningful involvement of all people regardless of race, color, national origin, or income with respect to the development, implementation, and enforcement of environmental laws, regulations, and policies. The educational tool consisted of a 50-min module that included an introduction to cancer biology including genetic susceptibility and environmental interactions, cancer burden in the local community, and risk reduction strategies. The module also included an interactive activity in which adolescent students identify cancer risk factors and brainstorm strategies for risk reduction at both the individual and community level. We administered the module to 12 classes of over 280 high school and college students in New York City. Cancer risk reduction strategies identified by the students included family- or peer-level strategies such as team physical activity and community-level action including improving parks and taxing sugary foods. We developed a novel and interactive cancer risk-reduction education tool focused on multiple cancers that can be adopted by other communities and educational institutions.

Polycyclic Aromatic Hydrocarbons and Mammary Cancer Risk: Does Obesity Matter too?

Breast cancer risk remains incompletely explained, and higher incidence rates of breast cancer over recent times and in urban and industrialized areas suggest environmental causes. Polycyclic aromatic hydrocarbons (PAH) are ubiquitous in the environment and epidemiological and rodent studies have shown associations between exposure to PAH and breast cancer incidence as well as mammary tumorigenesis. In addition, in vitro and rodent studies have implicated alterations in estrogen receptor alpha (Erα) signaling pathways following PAH exposure in limited experimental studies. However, our understanding of these mechanisms is incomplete. Sahay et al. addressed this gap by examining the effect of PAH exposure on epigenetic and transcriptional regulation of genes in the Erα pathway in a mouse cohort exposed to aerosolized PAH at proportions measured in urban air. In addition to alterations in the Erα signaling pathway in the pregnant mice and in their offspring and grandoffspring, the investigators observed higher body weights in mice exposed to PAH compared to the control. Given that associations between mammary tissue adiposity, systemic adiposity, and breast cancer risk have been observed previously, the finding of higher body weight in the PAH exposure group raises the possibility that body weight might influence the association between PAH exposure and breast cancer risk. Along with new analyses, we discuss the possibility that body weight may modify the association between PAH exposure, mammary cellular proliferation, and mammary gland ductal hyperplasia in offspring and grandoffspring mice and future research that may be needed to delineate these associations.

Th2/Th1 Cytokine Imbalance Is Associated With Higher COVID-19 Risk Mortality.

A major component of COVID-19 severe respiratory syndrome is the patient's immune response to the SARS-CoV-2 virus and the consequential multi-organ inflammatory response. Several studies suggested a potential role of CD4+ T cells in COVID-19 severe respiratory syndrome. We first hypothesized that there is a type 2 helper (Th2)/type 1 helper (Th1) imbalance in older age, male, asthma, smokers, and high ACE2 expression phenotype in the airway of non-infected patients. Next, we hypothesized that a Th2/Th1 imbalance may predict higher mortality in COVID-19 infected hospitalized patients with and without patient reported current asthma. We first analyzed publicly available gene expression from the sputum of 118 moderate-to-severe asthma patients and 21 healthy controls, and from nasal epithelium of 26 healthy current smokers and 21 healthy never smokers. Secondly, we profiled 288 new serum proteomics samples measured at admission from patients hospitalized within the Mount Sinai Health System with positive SARS-CoV-2 infection. We first computed Th1 and Th2 pathway enrichment scores by gene set variation analysis and then compared the differences in Th2 and Th1 pathway scores between patients that died compared to those that survived, by linear regression. The level of Th2/Th1 imbalance, as determined by the enrichment score, was associated with age, sex, and ACE2 expression in sputum, and with active smoking status in nasal epithelium (p < 0.05). Th2/Th1 imbalance at hospital admission in sera of patients was not significantly associated with death from COVID-19 (p = 0.11), unless evaluated in the asthmatic strata (p = 0.01). Using a similar approach we also observed a higher Th17/Th1 cytokine imbalance in all deceased patients compared to those that survived (p < 0.001), as well as in the asthmatic strata only (p < 0.01). Th2/Th1 imbalance is higher in the sera of asthma patients at admission that do not survive COVID-19, suggesting that the Th2/Th1 interplay may affect patient outcomes in SARS-CoV2 infection. In addition, we report that Th17/Th1 imbalance is increased in all patients that die of COVID-19.

Prenatal polycyclic aromatic hydrocarbons, altered ERα pathway-related methylation and expression, and mammary epithelial cell proliferation in offspring and grandoffspring adult mice.

BACKGROUND: Airborne polycyclic aromatic hydrocarbons (PAH) possess carcinogenic and endocrine disrupting properties linked to mammary tumorigenesis. These effects may be initiated during a prenatal period of susceptibility to PAH activation of the aryl hydrocarbon receptor (Ahr) and through downstream effects on estrogen receptor (Er) α. PURPOSE: We hypothesized prenatal airborne PAH exposure induces sustained effects in female adult wild type BALB/cByj mice detected in the offspring (F1) and grandoffspring (F2) generation. We hypothesized these effects would include altered expression and epigenetic regulation of Erα and altered expression of aryl hydrocarbon receptor repressor (Ahrr, Ahrr/aryl hydrocarbon receptor nuclear translocator (Arnt), and breast cancer type 1 susceptibility (Brca1). Further, we hypothesized that PAH would induce precancerous outcomes such as epithelial cell proliferation and epithelial cell hyperplasia in mammary glands of adult female offspring and grandoffspring. RESULTS: Prenatal ambient PAH exposure lowered Erα mRNA expression (F1 and F2: p<0.001 for each) and induced methylation in the Erα promoter in mammary tissue in offspring and grandoffspring mice on postnatal day (PND) 60. Prenatal PAH lowered Brca1 mRNA (F1: p=0.002, F2: p=0.02); Erα mRNA was correlated with Brca1 (F1: r=0.42, p=0.02; F2: r=0.53, p=0.005). Prenatal PAH lowered Ahrr (F1: p=0.03, F2: p=0.009) and raised Arnt mRNA expression (F1: p=0.01, F2: p=0.03). Alterations in Erα mRNA (F2: p<0.0001) and Ahrr (F2: p=0.02) in the grandoffspring mice also occured by PND 28, and similarly occurred in the dam on postpartum day (PPD) 28. Finally, prenatal PAH was associated with higher mammary epithelial cell proliferation in the offspring (p=0.02), but not grandoffspring mice, without differences in the frequency of mammary cell hyperplasia. These results did not differ after adjustment by each candidate gene expression level. CONCLUSIONS: Prenatal PAH exposure induces DNA methylation and alters gene expression in the Erα-mediated pathway across generations, and suggests that functional outcomes such as mammary cell proliferation also may occur in offspring as a result.

Associations of prenatal exposure to polycyclic aromatic hydrocarbons with pubertal timing and body composition in adolescent girls: Implications for breast cancer risk.

BACKGROUND: While animal data support an association between prenatal exposure to endocrine disrupting chemicals (EDCs) and altered mammary gland development and tumorigenesis, epidemiologic studies have only considered a few classes of EDCs in association with pubertal growth and development in girls. Polycyclic aromatic hydrocarbons (PAH) are a class of EDCs that have not been rigorously evaluated in terms of prenatal exposure and pubertal growth and development in girls. OBJECTIVE: In a New York City birth cohort of Black and Hispanic girls (n = 196; recruited 1998-2006), we examined associations of prenatal PAH exposure with self-reported age at growth spurt onset, breast development onset and menarche, and clinical measures of adolescent body composition including body mass index, waist-to-hip ratio, and body fat measured at ages 11-20 years. METHODS: We measured prenatal exposure to PAH using personal air monitoring data collected from backpacks worn by mothers during the third trimester of pregnancy (data available for all 196 girls) and biomarkers of benzo[α]pyrene-DNA adducts in umbilical cord blood (data available for 106 girls). We examined associations of prenatal PAH with the timing of pubertal milestones and adolescent body composition (11-20 years) using multivariable linear regression models adjusted for race/ethnicity, household public assistance status at birth, and age at outcome assessment. We also fit models further adjusted for potential mediators, including birthweight and childhood body size (BMI-for-age z-score measured at 6-8 years). RESULTS: Girls in the highest versus lowest tertile of ambient exposure to PAH, based on a summary measure of eight carcinogenic higher-molecular weight non-volatile PAH compounds (Σ8 PAH), had a 0.90 year delay in growth spurt onset (95% confidence interval (CI) = 0.25, 1.55; n = 196), a 0.35 year delay in breast development onset (95% CI = -0.26, 0.95; n = 193), and a 0.59 year delay in menarche (95% CI = 0.06, 1.11; n = 191) in models adjusted for race/ethnicity and household public assistance at birth. The statistically significant associations for age at growth spurt onset and menarche were not impacted by adjustment for birthweight or childhood body size. No differences in BMI-for-age z-score, waist-to-hip ratio, or percent body fat were found between girls in the highest versus lowest tertile of ambient Σ8 PAH. Results were similar when we evaluated benzo[α]pyrene-DNA adduct levels. DISCUSSION: Our results suggest that prenatal exposure to PAH might delay pubertal milestones in girls, but findings need to be replicated in other cohorts using prospectively collected data on pubertal outcomes.

Expression quantitative trait locus fine mapping of the 17q12-21 asthma locus in African American children: a genetic association and gene expression study.

BACKGROUND: African ancestry is associated with a higher prevalence and greater severity of asthma than European ancestries, yet genetic studies of the most common locus associated with childhood-onset asthma, 17q12-21, in African Americans have been inconclusive. The aim of this study was to leverage both the phenotyping of the Children's Respiratory and Environmental Workgroup (CREW) birth cohort consortium, and the reduced linkage disequilibrium in African Americans, to fine map the 17q12-21 locus. METHODS: We first did a genetic association study and meta-analysis using 17q12-21 tag single-nucleotide polymorphisms (SNPs) for childhood-onset asthma in 1613 European American and 870 African American children from the CREW consortium. Nine tag SNPs were selected based on linkage disequilibrium patterns at 17q12-21 and their association with asthma, considering the effect allele under an additive model (0, 1, or 2 effect alleles). Results were meta-analysed with publicly available summary data from the EVE consortium (on 4303 European American and 3034 African American individuals) for seven of the nine SNPs of interest. Subsequently, we tested for expression quantitative trait loci (eQTLs) among the SNPs associated with childhood-onset asthma and the expression of 17q12-21 genes in resting peripheral blood mononuclear cells (PBMCs) from 85 African American CREW children and in upper airway epithelial cells from 246 African American CREW children; and in lower airway epithelial cells from 44 European American and 72 African American adults from a case-control study of asthma genetic risk in Chicago (IL, USA). FINDINGS: 17q12-21 SNPs were broadly associated with asthma in European Americans. Only two SNPs (rs2305480 in gasdermin-B [GSDMB] and rs8076131 in ORMDL sphingolipid biosynthesis regulator 3 [ORMDL3]) were associated with asthma in African Americans, at a Bonferroni-corrected threshold of p<0·0055 (for rs2305480_G, odds ratio [OR] 1·36 [95% CI 1·12-1·65], p=0·0014; and for rs8076131_A, OR 1·37 [1·13-1·67], p=0·0010). In upper airway epithelial cells from African American children, genotype at rs2305480 was the most significant eQTL for GSDMB (eQTL effect size [ß] 1·35 [95% CI 1·25-1·46], p<0·0001), and to a lesser extent showed an eQTL effect for post-GPI attachment to proteins phospholipase 3 (ß 1·15 [1·08-1·22], p<0·0001). No SNPs were eQTLs for ORMDL3. By contrast, in PBMCs, the five core SNPs were associated only with expression of GSDMB and ORMDL3. Genotype at rs12936231 (in zona pellucida binding protein 2) showed the strongest associations across both genes (for GSDMB, eQTLß 1·24 [1·15-1·32], p<0·0001; and for ORMDL3 (ß 1·19 [1·12-1·24], p<0·0001). The eQTL effects of rs2305480 on GSDMB expression were replicated in lower airway cells from African American adults (ß 1·29 [1·15-1·44], p<0·0001). INTERPRETATION: Our study suggests that SNPs regulating GSDMB expression in airway epithelial cells have a major role in childhood-onset asthma, whereas SNPs regulating the expression levels of 17q12-21 genes in resting blood cells are not central to asthma risk. Our genetic and gene expression data in African Americans and European Americans indicated GSDMB to be the leading candidate gene at this important asthma locus. FUNDING: National Institutes of Health, Office of the Director.

Environmental exposures during windows of susceptibility for breast cancer: a framework for prevention research.

BACKGROUND: The long time from exposure to potentially harmful chemicals until breast cancer occurrence poses challenges for designing etiologic studies and for implementing successful prevention programs. Growing evidence from animal and human studies indicates that distinct time periods of heightened susceptibility to endocrine disruptors exist throughout the life course. The influence of environmental chemicals on breast cancer risk may be greater during several windows of susceptibility (WOS) in a woman's life, including prenatal development, puberty, pregnancy, and the menopausal transition. These time windows are considered as specific periods of susceptibility for breast cancer because significant structural and functional changes occur in the mammary gland, as well as alterations in the mammary micro-environment and hormone signaling that may influence risk. Breast cancer research focused on these breast cancer WOS will accelerate understanding of disease etiology and prevention. MAIN TEXT: Despite the plausible heightened mechanistic influences of environmental chemicals on breast cancer risk during time periods of change in the mammary gland's structure and function, most human studies of environmental chemicals are not focused on specific WOS. This article reviews studies conducted over the past few decades that have specifically addressed the effect of environmental chemicals and metals on breast cancer risk during at least one of these WOS. In addition to summarizing the broader evidence-base specific to WOS, we include discussion of the NIH-funded Breast Cancer and the Environment Research Program (BCERP) which included population-based and basic science research focused on specific WOS to evaluate associations between breast cancer risk and particular classes of endocrine-disrupting chemicals-including polycyclic aromatic hydrocarbons, perfluorinated compounds, polybrominated diphenyl ethers, and phenols-and metals. We outline ways in which ongoing transdisciplinary BCERP projects incorporate animal research and human epidemiologic studies in close partnership with community organizations and communication scientists to identify research priorities and effectively translate evidence-based findings to the public and policy makers. CONCLUSIONS: An integrative model of breast cancer research is needed to determine the impact and mechanisms of action of endocrine disruptors at different WOS. By focusing on environmental chemical exposure during specific WOS, scientists and their community partners may identify when prevention efforts are likely to be most effective.

HIV Diversity and Genetic Compartmentalization in Blood and Testes during Suppressive Antiretroviral Therapy.

HIV's ability to persist during suppressive antiretroviral therapy is the main barrier to cure. Immune-privileged tissues, such as the testes, may constitute distinctive sites of HIV persistence, but this has been challenging to study in humans. We analyzed the proviral burden and genetics in the blood and testes of 10 individuals on suppressive therapy who underwent elective gender-affirming surgery. HIV DNA levels in matched blood and testes were quantified by quantitative PCR, and subgenomic proviral sequences (nef region) were characterized from single templates. HIV diversity, compartmentalization, and immune escape burden were assessed using genetic and phylogenetic approaches. Diverse proviruses were recovered from the blood (396 sequences; 354 nef-intact sequences) and testes (326 sequences; 309 nef-intact sequences) of all participants. Notably, the frequency of identical HIV sequences varied markedly between and within individuals. Nevertheless, proviral loads, within-host unique HIV sequence diversity, and the immune escape burden correlated positively between blood and testes. When all intact nef sequences were evaluated, 60% of participants exhibited significant blood-testis genetic compartmentalization, but none did so when the evaluation was restricted to unique sequences per site, suggesting that compartmentalization, when present, is attributable to the clonal expansion of HIV-infected cells. Our observations confirm the testes as a site of HIV persistence and suggest that individuals with larger and more diverse blood reservoirs will have larger and more diverse testis reservoirs. Furthermore, while the testis microenvironment may not be sufficiently unique to facilitate the seeding of unique viral populations therein, differential clonal expansion dynamics may be at play, which may complicate HIV eradication.IMPORTANCE Two key questions in HIV reservoir biology are whether immune-privileged tissues, such as the testes, harbor distinctive proviral populations during suppressive therapy and, if so, by what mechanism. While our results indicated that blood-testis HIV genetic compartmentalization was reasonably common (60%), it was always attributable to differential frequencies of identical HIV sequences between sites. No blood-tissue data set retained evidence of compartmentalization when only unique HIV sequences per site were considered; moreover, HIV immune escape mutation burdens were highly concordant between sites. We conclude that the principal mechanism by which blood and testis reservoirs differ is not via seeding of divergent HIV sequences therein but, rather, via differential clonal expansion of latently infected cells. Thus, while viral diversity and escape-related barriers to HIV eradication are of a broadly similar magnitude across the blood and testes, clonal expansion represents a challenge. The results support individualized analysis of within-host reservoir diversity to inform curative approaches.

HIV Subtype and Nef-Mediated Immune Evasion Function Correlate with Viral Reservoir Size in Early-Treated Individuals.

The HIV accessory protein Nef modulates key immune evasion and pathogenic functions, and its encoding gene region exhibits high sequence diversity. Given the recent identification of early HIV-specific adaptive immune responses as novel correlates of HIV reservoir size, we hypothesized that viral factors that facilitate the evasion of such responses-namely, Nef genetic and functional diversity-might also influence reservoir establishment and/or persistence. We isolated baseline plasma HIV RNA-derived nef clones from 30 acute/early-infected individuals who participated in a clinical trial of early combination antiretroviral therapy (cART) (<6 months following infection) and assessed each Nef clone's ability to downregulate CD4 and human leukocyte antigen (HLA) class I in vitro We then explored the relationships between baseline clinical, immunological, and virological characteristics and the HIV reservoir size measured 48 weeks following initiation of suppressive cART (where the reservoir size was quantified in terms of the proviral DNA loads as well as the levels of replication-competent HIV in CD4+ T cells). Maximal within-host Nef-mediated downregulation of HLA, but not CD4, correlated positively with post-cART proviral DNA levels (Spearman's R = 0.61, P = 0.0004) and replication-competent reservoir sizes (Spearman's R = 0.36, P = 0.056) in univariable analyses. Furthermore, the Nef-mediated HLA downregulation function was retained in final multivariable models adjusting for established clinical and immunological correlates of reservoir size. Finally, HIV subtype B-infected persons (n = 25) harbored significantly larger viral reservoirs than non-subtype B-infected persons (2 infected with subtype CRF01_AE and 3 infected with subtype G). Our results highlight a potentially important role of viral factors-in particular, HIV subtype and accessory protein function-in modulating viral reservoir establishment and persistence.IMPORTANCE While combination antiretroviral therapies (cART) have transformed HIV infection into a chronic manageable condition, they do not act upon the latent HIV reservoir and are therefore not curative. As HIV cure or remission should be more readily achievable in individuals with smaller HIV reservoirs, achieving a deeper understanding of the clinical, immunological, and virological determinants of reservoir size is critical to eradication efforts. We performed a post hoc analysis of 30 participants of a clinical trial of early cART who had previously been assessed in detail for their clinical, immunological, and reservoir size characteristics. We observed that the HIV subtype and autologous Nef-mediated HLA downregulation function correlated with the viral reservoir size measured approximately 1 year post-cART initiation. Our findings highlight virological characteristics-both genetic and functional-as possible novel determinants of HIV reservoir establishment and persistence.

Breast cancer family history and allele-specific DNA methylation in the legacy girls study.

Family history, a well-established risk factor for breast cancer, can have both genetic and environmental contributions. Shared environment in families as well as epigenetic changes that also may be influenced by shared genetics and environment may also explain familial clustering of cancers. Epigenetic regulation, such as DNA methylation, can change the activity of a DNA segment without a change in the sequence; environmental exposures experienced across the life course can induce such changes. However, genetic-epigenetic interactions, detected as methylation quantitative trait loci (mQTLs; a.k.a. meQTLs) and haplotype-dependent allele-specific methylation (hap-ASM), can also contribute to inter-individual differences in DNA methylation patterns. To identify differentially methylated regions (DMRs) associated with breast cancer susceptibility, we examined differences in white blood cell DNA methylation in 29 candidate genes in 426 girls (ages 6-13 years) from the LEGACY Girls Study, 239 with and 187 without a breast cancer family history (BCFH). We measured methylation by targeted massively parallel bisulfite sequencing (bis-seq) and observed BCFH DMRs in two genes: ESR1 (Δ4.9%, P = 0.003) and SEC16B (Δ3.6%, P = 0.026), each of which has been previously implicated in breast cancer susceptibility and pubertal development. These DMRs showed high inter-individual variability in methylation, suggesting the presence of mQTLs/hap-ASM. Using single nucleotide polymorphisms data in the bis-seq amplicon, we found strong hap-ASM in SEC16B (with allele specific-differences ranging from 42% to 74%). These findings suggest that differential methylation in genes relevant to breast cancer susceptibility may be present early in life, and that inherited genetic factors underlie some of these epigenetic differences.

Combined effects of prenatal exposure to polycyclic aromatic hydrocarbons and material hardship on child ADHD behavior problems.

IMPORTANCE: Polycyclic aromatic hydrocarbons (PAH) are carcinogenic and neurotoxic combustion by-products commonly found in urban air. Exposure to PAH is disproportionately high in low income communities of color who also experience chronic economic stress. OBJECTIVE: In a prospective cohort study in New York City (NYC) we previously found a significant association between prenatal PAH exposure and Attention Deficit Hyperactivity Disorder (ADHD) behavior problems at age 9. Here, we have evaluated the joint effects of prenatal exposure to PAH and prenatal/childhood material hardship on ADHD behavior problems. MATERIALS AND METHODS: We enrolled nonsmoking African-American and Dominican pregnant women in New York City between 1998 and 2006 and followed their children through 9 years of age. As a biomarker of prenatal PAH exposure, PAH-DNA adducts were measured in maternal blood at delivery and were dichotomized at the limit of detection (to indicate high vs. low exposure). Maternal material hardship (lack of adequate food, housing, utilities, and clothing) was self-reported prenatally and at multiple time points through child age 9. Latent variable analysis identified four distinct patterns of hardship. ADHD behavior problems were assessed using the Conners Parent Rating Scale- Revised. Analyses adjusted for relevant covariates. RESULTS: Among 351 children in our sample, across all hardship groups, children with high prenatal PAH exposure (high adducts) generally had more symptoms of ADHD (higher scores) compared to those with low PAH exposure. The greatest difference was seen among the children with hardship persisting from pregnancy through childhood. Although the interactions between high PAH exposure and hardship experienced at either period ("persistent" hardship or "any" hardship) were not significant, we observed significant differences in the number of ADHD symptoms between children with high prenatal PAH exposure and either persistent hardship or any hardship compared to the others. These differences were most significant for combined high PAH and persistent hardship: ADHD Index (p < 0.008), DSM-IV Inattentive (p = 0.006), DSM-IV Hyperactive Impulsive problems (p = 0.033), and DSM-IV Index Total (p = 0.009). CONCLUSION: The present findings add to existing evidence that co-exposure to socioeconomic disadvantage and air pollution in early life significantly increases the risk of adverse neurodevelopmental outcomes. They suggest the need for multifaceted interventions to protect pregnant mothers and their children.

Effect of personal exposure to black carbon on changes in allergic asthma gene methylation measured 5 days later in urban children: importance of allergic sensitization.

BACKGROUND: Asthma gene DNA methylation may underlie the effects of air pollution on airway inflammation. However, the temporality and individual susceptibility to environmental epigenetic regulation of asthma has not been fully elucidated. Our objective was to determine the timeline of black carbon (BC) exposure, measured by personal sampling, on DNA methylation of allergic asthma genes 5 days later to capture usual weather variations and differences related to changes in behavior and activities. We also sought to determine how methylation may vary by seroatopy and cockroach sensitization and by elevated fractional exhaled nitric oxide (FeNO). METHODS: Personal BC levels were measured during two 24-h periods over a 6-day sampling period in 163 New York City children (age 9-14 years), repeated 6 months later. During home visits, buccal cells were collected as noninvasive surrogates for lower airway epithelial cells and FeNO measured as an indicator of airway inflammation. CpG promoter loci of allergic asthma genes (e.g., interleukin 4 (IL4), interferon gamma (IFNγ), inducible nitric oxide synthase (NOS2A)), arginase 2 (ARG2)) were pyrosequenced at the start and end of each sampling period. RESULTS: Higher levels of BC were associated with lower methylation of IL4 promoter CpG-48 5 days later. The magnitude of association between BC exposure and demethylation of IL4 CpG-48 and NOS2A CpG+5099 measured 5 days later appeared to be greater among seroatopic children, especially those sensitized to cockroach allergens (RR [95% CI] 0.55 [0.37-0.82] and 0.67 [0.45-0.98] for IL4 CpG-48 and NOS2A CpG+5099, respectively), compared to non-sensitized children (RR [95% CI] 0.87 [0.65-1.17] and 0.95 [0.69-1.33] for IL4 CpG-48 and NOS2A CpG+5099, respectively); however, the difference was not statistically different. In multivariable linear regression models, lower DNA methylation of IL4 CpG-48 and NOS2A CpG+5099 were associated with increased FeNO. CONCLUSIONS: Our results suggest that exposure to BC may exert asthma proinflammatory gene demethylation 5 days later that in turn may link to airway inflammation. Our results further suggest that seroatopic children, especially those sensitized to cockroach allergens, may be more susceptible to the effect of acute BC exposure on epigenetic changes.

Reduced mouse allergen is associated with epigenetic changes in regulatory genes, but not mouse sensitization, in asthmatic children.

Chronic exposure to mouse allergen may contribute greatly to the inner-city asthma burden. We hypothesized that reducing mouse allergen exposure may modulate the immunopathology underlying symptomatic pediatric allergic asthma, and that this occurs through epigenetic regulation. To test this hypothesis, we studied a cohort of mouse sensitized, persistent asthmatic inner-city children undergoing mouse allergen-targeted integrated pest management (IPM) vs education in a randomized controlled intervention trial. We found that decreasing mouse allergen exposure, but not cockroach, was associated with reduced FOXP3 buccal DNA promoter methylation, but this was unrelated to mouse specific IgE production. This finding suggests that the environmental epigenetic regulation of an immunomodulatory gene may occur following changing allergen exposures in some highly exposed cohorts. Given the clinical and public health importance of inner-city pediatric asthma and the potential impact of environmental interventions, further studies will be needed to corroborate changes in epigenetic regulation following changing exposures over time, and determine their impact on asthma morbidity in susceptible children.