Generation of Novel Immunocompetent Mouse Cell Lines to Model Experimental Metastasis of High-Risk Neuroblastoma.

NB, being a highly metastatic cancer, is one of the leading causes of cancer-related deaths in children. Increased disease recurrence and clinical resistance in patients with metastatic high-risk NBs (HR-NBs) result in poor outcomes and lower overall survival. However, the paucity of appropriate in vivo models for HR-NB metastasis has limited investigations into the underlying biology of HR-NB metastasis. This study was designed to address this limitation and develop suitable immunocompetent models for HR-NB metastasis. Here, we developed several highly metastatic immunocompetent murine HR-NB cell lines. Our newly developed cell lines show 100% efficiency in modeling experimental metastasis in C57BL6 mice and feature metastasis to the sites frequently observed in humans with HR-NB (liver and bone). In vivo validation demonstrated their specifically gained metastatic phenotype. The in vitro characterization of the cell lines showed increased cell invasion, acquired anchorage-independent growth ability, and resistance to MHC-I induction upon IFN-γ treatment. Furthermore, RNA-seq analysis of the newly developed cells identified a differentially regulated gene signature and an enrichment of processes consistent with their acquired metastatic phenotype, including extracellular matrix remodeling, angiogenesis, cell migration, and chemotaxis. The presented newly developed cell lines are, thus, suitable and promising tools for HR-NB metastasis and microenvironment studies in an immunocompetent system.

A novel metric to improve mismatched primer selection and quantification accuracy in amplifying DNA repeats for quantitative polymerase chain reactions.

In quantitative polymerase chain reaction (qPCR) experiments, primers containing mismatches with respect to the template are widely used in measuring repetitive DNA elements. Primer-template mismatches may lead to underestimation of the input sample quantity due to inefficient annealing and amplification. But how primer-template mismatches affect quantification accuracy has not been rigorously investigated. In this study, we performed a series of qPCR experiments in which we tested three pairs of mismatched telomere primers (tel1/tel2, tel1b/tel2b and telg/telc) and two pairs of perfect-match reference gene primers (36B4-F/-R and IFNB1-F/-R) at three different primer concentrations under four cycling conditions. Templates used were genomic DNA from two human cell lines and oligo duplexes which contained telomere sequences, reference gene sequences, or both. We demonstrated that the underestimation of input sample quantity from reactions containing mismatched primers was not due to lower amplification efficiency (E), but due to ineffective usage of the input sample. We defined a novel concept of amplification efficacy (f) which quantifies the effectiveness of input sample amplification by primers. We have modified the conventional qPCR kinetic formula to include f, which corrects the effects of primer mismatches. We demonstrated that reactions containing mismatched telomere primer pairs had similar efficiency (E), but varying degrees of reduced efficacy (f) in comparison to those with the perfect-match gene primer pairs. Using the quantitative parameter f, underestimation of initial target by telomere primers can be adjusted to provide a more accurate measurement. Additionally, we found that the tel1b/tel2b primer set at concentration of 500 nM and 900 nM exhibited the best amplification efficacy f. This study provides a novel way to incorporate an evaluation of amplification efficacy into qPCR analysis. In turn, it improves mismatched primer selection and quantification accuracy in amplifying DNA repeats using qPCR methods.

Salivary Epigenetic Measures of Body Mass Index and Social Determinants of Health Across Childhood and Adolescence.

Importance: Children who are socioeconomically disadvantaged are at increased risk for high body mass index (BMI) and multiple diseases in adulthood. The developmental origins of health and disease hypothesis proposes that early life conditions affect later-life health in a manner that is only partially modifiable by later-life experiences. Objective: To examine whether epigenetic measures of BMI developed in adults are valid biomarkers of childhood BMI and if they are sensitive to early life social determinants of health. Design, Setting, and Participants: This population-based study of over 3200 children and adolescents aged 8 to 18 years included data from 2 demographically diverse US pediatric cohort studies that combine longitudinal and twin study designs. Analyses were conducted from 2021 to 2022. Exposures: Socioeconomic status, marginalized groups. Main Outcome and Measure: Salivary epigenetic BMI, BMI. Analyses were conducted to validate the use of saliva epigenetic BMI as a potential biomarker of child BMI and to examine associations between epigenetic BMI and social determinants of health. Results: Salivary epigenetic BMI was calculated from 2 cohorts: (1) 1183 individuals aged 8 to 18 years (609 female [51%]; mean age, 13.4 years) from the Texas Twin Project and (2) 2020 children (1011 female [50%]) measured at 9 years of age and 15 years of age from the Future of Families and Child Well-Being Study. Salivary epigenetic BMI was associated with children's BMI (r = 0.36; 95% CI, 0.31-0.40 to r = 0.50; 95% CI, 0.42-0.59). Longitudinal analysis found that epigenetic BMI was highly stable across adolescence but remained both a leading and lagging indicator of BMI change. Twin analyses showed that epigenetic BMI captured differences in BMI between monozygotic twins. Moreover, children from more disadvantaged socioeconomic status (b = -0.13 to -0.15 across samples) and marginalized racial and ethnic groups (b = 0.08-0.34 across samples) had higher epigenetic BMI, even when controlling for concurrent BMI, pubertal development, and tobacco exposure. Socioeconomic status at birth relative to concurrent socioeconomic status best predicted epigenetic BMI in childhood and adolescence (b = -0.15; 95% CI, -0.20 to -0.09). Conclusion and Relevance: This study demonstrated that epigenetic measures of BMI calculated from pediatric saliva samples were valid biomarkers of childhood BMI and may be associated with early-life social inequalities. The findings are in line with the hypothesis that early-life conditions are especially important factors in epigenetic regulation of later-life health. Research showing that health later in life is linked to early-life conditions has important implications for the development of early-life interventions that could significantly extend healthy life span.

Sex-specific DNA methylation in saliva from the multi-ethnic Future of Families and Child Wellbeing Study.

The prevalence and severity of many diseases differs by sex, potentially due to sex-specific patterns in DNA methylation. Autosomal sex-specific differences in DNA methylation have been observed in cord blood and placental tissue but are not well studied in saliva or in diverse populations. We sought to characterize sex-specific DNA methylation on autosomal chromosomes in saliva samples from children in the Future of Families and Child Wellbeing Study, a multi-ethnic prospective birth cohort containing an oversampling of Black, Hispanic and low-income families. DNA methylation from saliva samples was analysed on 796 children (50.6% male) at both ages 9 and 15 with DNA methylation measured using the Illumina HumanMethylation 450k array. An epigenome-wide association analysis of the age 9 samples identified 8,430 sex-differentiated autosomal DNA methylation sites (P < 2.4 × 10-7), of which 76.2% had higher DNA methylation in female children. The strongest sex-difference was in the cg26921482 probe, in the AMDHD2 gene, with 30.6% higher DNA methylation in female compared to male children (P < 1 × 10-300). Treating the age 15 samples as an internal replication set, we observed highly consistent results between the ages 9 and 15 measurements, indicating stable and replicable sex-differentiation. Further, we directly compared our results to previously published DNA methylation sex differences in both cord blood and saliva and again found strong consistency. Our findings support widespread and robust sex-differential DNA methylation across age, human tissues, and populations. These findings help inform our understanding of potential biological processes contributing to sex differences in human physiology and disease.

Association between the timing of childhood adversity and epigenetic patterns across childhood and adolescence: findings from the Avon Longitudinal Study of Parents and Children (ALSPAC) prospective cohort.

BACKGROUND: Childhood adversity is a potent determinant of health across development and is associated with altered DNA methylation signatures, which might be more common in children exposed during sensitive periods in development. However, it remains unclear whether adversity has persistent epigenetic associations across childhood and adolescence. We aimed to examine the relationship between time-varying adversity (defined through sensitive period, accumulation of risk, and recency life course hypotheses) and genome-wide DNA methylation, measured three times from birth to adolescence, using data from a prospective, longitudinal cohort study. METHODS: We first investigated the relationship between the timing of exposure to childhood adversity between birth and 11 years and blood DNA methylation at age 15 years in the Avon Longitudinal Study of Parents and Children (ALSPAC) prospective cohort study. Our analytic sample included ALSPAC participants with DNA methylation data and complete childhood adversity data between birth and 11 years. We analysed seven types of adversity (caregiver physical or emotional abuse, sexual or physical abuse [by anyone], maternal psychopathology, one-adult households, family instability, financial hardship, and neighbourhood disadvantage) reported by mothers five to eight times between birth and 11 years. We used the structured life course modelling approach (SLCMA) to identify time-varying associations between childhood adversity and adolescent DNA methylation. Top loci were identified using an R2 threshold of 0·035 (ie, ≥3·5% of DNA methylation variance explained by adversity). We attempted to replicate these associations using data from the Raine Study and Future of Families and Child Wellbeing Study (FFCWS). We also assessed the persistence of adversity-DNA methylation associations we previously identified from age 7 blood DNA methylation into adolescence and the influence of adversity on DNA methylation trajectories from ages 0-15 years. FINDINGS: Of 13 988 children in the ALSPAC cohort, 609-665 children (311-337 [50-51%] boys and 298-332 [49-50%] girls) had complete data available for at least one of the seven childhood adversities and DNA methylation at 15 years. Exposure to adversity was associated with differences in DNA methylation at 15 years for 41 loci (R2 ≥0·035). Sensitive periods were the most often selected life course hypothesis by the SLCMA. 20 (49%) of 41 loci were associated with adversities occurring between age 3 and 5 years. Exposure to one-adult households was associated with differences in DNA methylation at 20 [49%] of 41 loci, exposure to financial hardship was associated with changes at nine (22%) loci, and physical or sexual abuse was associated with changes at four (10%) loci. We replicated the direction of associations for 18 (90%) of 20 loci associated with exposure to one-adult household using adolescent blood DNA methylation from the Raine Study and 18 (64%) of 28 loci using saliva DNA methylation from the FFCWS. The directions of effects for 11 one-adult household loci were replicated in both cohorts. Differences in DNA methylation at 15 years were not present at 7 years and differences identified at 7 years were no longer apparent by 15 years. We also identified six distinct DNA methylation trajectories from these patterns of stability and persistence. INTERPRETATION: These findings highlight the time-varying effect of childhood adversity on DNA methylation profiles across development, which might link exposure to adversity to potential adverse health outcomes in children and adolescents. If replicated, these epigenetic signatures could ultimately serve as biological indicators or early warning signs of initiated disease processes, helping identify people at greater risk for the adverse health consequences of childhood adversity. FUNDING: Canadian Institutes of Health Research, Cohort and Longitudinal Studies Enhancement Resources, EU's Horizon 2020, US National Institute of Mental Health.

Lower respiratory tract microbiome composition and community interactions in smokers.

The lung microbiome impacts on lung function, making any smoking-induced changes in the lung microbiome potentially significant. The complex co-occurrence and co-avoidance patterns between the bacterial taxa in the lower respiratory tract (LRT) microbiome were explored for a cohort of active (AS), former (FS) and never (NS) smokers. Bronchoalveolar lavages (BALs) were collected from 55 volunteer subjects (9 NS, 24 FS and 22 AS). The LRT microbiome composition was assessed using 16S rRNA amplicon sequencing. Identification of differentially abundant taxa and co-occurrence patterns, discriminant analysis and biomarker inferences were performed. The data show that smoking results in a loss in the diversity of the LRT microbiome, change in the co-occurrence patterns and a weakening of the tight community structure present in healthy microbiomes. The increased abundance of the genus Ralstonia in the lung microbiomes of both former and active smokers is significant. Partial least square discriminant and DESeq2 analyses suggested a compositional difference between the cohorts in the LRT microbiome. The groups were sufficiently distinct from each other to suggest that cessation of smoking may not be sufficient for the lung microbiota to return to a similar composition to that of NS. The linear discriminant analysis effect size (LEfSe) analyses identified several bacterial taxa as potential biomarkers of smoking status. Network-based clustering analysis highlighted different co-occurring and co-avoiding microbial taxa in the three groups. The analysis found a cluster of bacterial taxa that co-occur in smokers and non-smokers alike. The clusters exhibited tighter and more significant associations in NS compared to FS and AS. Higher degree of rivalry between clusters was observed in the AS. The groups were sufficiently distinct from each other to suggest that cessation of smoking may not be sufficient for the lung microbiota to return to a similar composition to that of NS.

Measuring the long arm of childhood in real-time: Epigenetic predictors of BMI and social determinants of health across childhood and adolescence.

Children who are socioeconomically disadvantaged are at increased risk for high body mass index (BMI) and multiple diseases in adulthood. The developmental origins of health and disease hypothesis proposes that early life conditions affect later-life health in a manner that is only partially modifiable by later-life experiences. Epigenetic mechanisms may regulate the influence of early life conditions on later life health. Recent epigenetic studies of adult blood samples have identified DNA-methylation sites associated with higher BMI and worse health (epigenetic-BMI). Here, we used longitudinal and twin study designs to examine whether epigenetic predictors of BMI developed in adults are valid biomarkers of child BMI and are sensitive to early life social determinants of health. Salivary epigenetic-BMI was calculated from two samples: (1) N=1,183 8-to-19-year-olds (609 female, mean age=13.4) from the Texas Twin Project (TTP), and (2) N=2,020 children (1,011 female) measured at 9 and 15 years from the Future of Families and Child Well-Being Study (FFCWS). We found that salivary epigenetic-BMI is robustly associated with children's BMI (r=0.36 to r=0.50). Longitudinal analysis suggested that epigenetic-BMI is highly stable across adolescence, but remains both a leading and lagging indicator of BMI change. Twin analyses showed that epigenetic-BMI captures differences in BMI between monozygotic twins. Moreover, children from more disadvantaged socioeconomic status (SES) and marginalized race/ethnic groups had higher epigenetic-BMI, even when controlling for concurrent BMI, pubertal development, and tobacco exposure. SES at birth relative to concurrent SES best predicted epigenetic-BMI in childhood and adolescence. We show for the first time that epigenetic predictors of BMI calculated from pediatric saliva samples are valid biomarkers of childhood BMI that are sensitive to social inequalities. Our findings are in line with the hypothesis that early life conditions are especially important factors in epigenetic regulation of later life health. Research showing that health later in life is linked to early life conditions have important implications for the development of early-life interventions that could significantly extend healthy life span.

Characteristics of salivary telomere length shortening in preterm infants.

OBJECTIVE: To examine the association between gestational age, telomere length (TL) and rate of shortening in newborns. STUDY DESIGN: Genomic DNA was isolated from buccal samples of 39 term infants at birth and one year and 32 preterm infants at birth, term-adjusted age (40 weeks post-conception) and age one-year corrected for gestational duration. Telomere length was measured by quantitative real-time PCR. Demographic and clinical data were collected during clinic or research visits and from hospital records. Socioeconomic status was estimated using the deprivation category (DEPCAT) scores derived from the Carstairs score of the subject's postal code. RESULTS: At birth, preterm infants had longer telomeres than infants born at term. However, there was no difference in telomere length between preterm infants and term infants at one year of age, implying that the rate of telomere shortening was greater in pre-term than term infants. Interestingly, TL at age 40 weeks post-conception in preterm infants was significantly longer than term infant TL at birth, suggesting that time since conception is not the only factor that affects rate of shortening. Several factors, including sex, fetal growth restriction, maternal age, maternal booking body mass index (BMI), mother education level and DEPCAT score, also differed between the preterm and term groups. CONCLUSIONS: Preterm infants have longer telomeres than term infants at birth. In the studied cohort, the rate of telomere shortening was greater in the premature group compared with the term infants. This finding agrees with previous studies using cord blood, suggesting that the longer TL in premature infants detected at birth do not persist and demonstrating that use of saliva DNA is acceptable for studies of telomere dynamics in infants. However, that the TL at age 40 weeks post-conception in preterm is longer than term infants at birth suggests that biological factors other than time since conception also affect rate of shortening.

Polymethylation scores for prenatal maternal smoke exposure persist until age 15 and are detected in saliva in the Fragile Families and Child Wellbeing cohort.

Prenatal maternal smoking is associated with low birthweight, neurological disorders, and asthma in exposed children. DNA methylation signatures can function as biomarkers of prenatal smoke exposure. However, the robustness of DNA methylation signatures across child ages, genetic ancestry groups, or tissues is not clear. Using coefficients from a meta-analysis of prenatal smoke exposure and DNA methylation in newborn cord blood, we created polymethylation scores of saliva DNA methylation from children at ages 9 and 15 in the Fragile Families and Child Wellbeing study. In the full sample at age 9 (n = 753), prenatal smoke exposure was associated with a 0.51 (95%CI: 0.35, 0.66) standard deviation higher polymethylation score. The direction and magnitude of the association was consistent in European and African genetic ancestry samples. In the full sample at age 15 (n = 747), prenatal smoke exposure was associated with a 0.48 (95%CI: 0.32, 0.63) standard deviation higher polymethylation score, and the association was attenuated among the European and Admixed-Latin genetic ancestry samples. The polymethylation score classified prenatal smoke exposure accurately (AUC age 9 = 0.77, age 15 = 0.76). Including the polymethylation score increased the AUC of base model covariates by 5 (95% CI: (2.1, 7.2)) percentage points, while including a single candidate site in the AHRR gene did not (P-value = 0.19). Polymethylation scores for prenatal smoking were portable across genetic ancestries and more accurate than an individual DNA methylation site. Polymethylation scores from saliva samples could serve as robust and practical biomarkers of prenatal smoke exposure.

Impact of paternal education on epigenetic ageing in adolescence and mid-adulthood: a multi-cohort study in the USA and Mexico.

BACKGROUND: Both parental and neighbourhood socio-economic status (SES) are linked to poorer health independently of personal SES measures, but the biological mechanisms are unclear. Our objective was to examine these influences via epigenetic age acceleration (EAA)-the discrepancy between chronological and epigenetic ages. METHODS: We examined three USA-based [Coronary Artery Risk Disease in Adults (CARDIA) study, Fragile Families and Child Wellbeing Study (FFCWS) and Programming Research in Obesity, Growth, Environment and Social Stressors (PROGRESS)] and one Mexico-based (Project Viva) cohort. DNA methylation was measured using Illumina arrays, personal/parental SES by questionnaire and neighbourhood disadvantage from geocoded address. In CARDIA, we examined the most strongly associated personal, parental and neighbourhood SES measures with EAA (Hannum's method) at study years 15 and 20 separately and combined using a generalized estimating equation (GEE) and compared with other EAA measures (Horvath's EAA, PhenoAge and GrimAge calculators, and DunedinPoAm). RESULTS: EAA was associated with paternal education in CARDIA [GEEs: ßsome college = -1.01 years (-1.91, -0.11) and ß

The Longitudinal Associations Between Paternal Incarceration and Family Well-Being: Implications for Ethnic/Racial Disparities in Health.

OBJECTIVE: Ethnic/racial minority children in the United States are more likely to experience father loss to incarceration than White children, and limited research has examined the health implications of these ethnic/racial disparities. Telomere length is a biomarker of chronic stress that is predictive of adverse health outcomes. This study examined whether paternal incarceration predicted telomere length shortening among offspring from childhood to adolescence, whether maternal depression mediated the link, and whether ethnicity/race moderated results. METHOD: Research participants included 2,395 families in the Fragile Families and Child Wellbeing study, a national and longitudinal cohort study of primarily low-income families from 20 large cities in the United States. Key constructs were measured when children were on average ages 9 (2007-2010) and 15 (2014-2017). RESULTS: Children who experienced paternal incarceration exhibited shorter telomere lengths between ages 9 and 15, and changes in maternal depression mediated this finding. Specifically, mothers who experienced a partner's incarceration were more likely to have depression between children's ages 9 and 15. In turn, increases in maternal depression between children's ages 9 and 15 predicted more accelerated telomere length shortening among children during this period. Paternal incarceration was more prevalent and frequent for ethnic/racial minority youth than for White youth. CONCLUSION: Paternal incarceration is associated with a biomarker of chronic stress among children in low-income families. Rates of paternal incarceration were more prevalent and frequent among Black American and multiethnic/multiracial families than among White Americans. As a result, the mass incarceration crisis of the criminal justice system is likely shaping intergenerational ethnic/racial health disparities.

Prenatal Particulate Matter Exposure Is Associated with Saliva DNA Methylation at Age 15: Applying Cumulative DNA Methylation Scores as an Exposure Biomarker.

Exposure in utero to particulate matter (PM2.5 and PM10) is associated with maladaptive health outcomes. Although exposure to prenatal PM2.5 and PM10 has cord blood DNA methylation signatures at birth, signature persistence into childhood and saliva cross-tissue applicability has not been tested. In the Fragile Families and Child Wellbeing Study, a United States 20-city birth cohort, average residential PM2.5 and PM10 during the three months prior to birth was estimated using air quality monitors with inverse distance weighting. Saliva DNA methylation at ages 9 (n = 749) and 15 (n = 793) was measured using the Illumina HumanMethylation 450 k BeadArray. Cumulative DNA methylation scores for particulate matter were estimated by weighting participant DNA methylation at each site by independent meta-analysis effect estimates and standardizing the sums. Using a mixed-effects regression analysis, we tested the associations between cumulative DNA methylation scores at ages 9 and 15 and PM exposure during pregnancy, adjusted for child sex, age, race/ethnicity, maternal income-to-needs ratio, nonmartial birth status, and saliva cell-type proportions. Our study sample was 50.5% male, 56.3% non-Hispanic Black, and 19.8% Hispanic, with a median income-to-needs ratio of 1.4. Mean exposure levels for PM2.5 were 27.9 µg/m3/day (standard deviation: 7.0; 23.7% of observations exceeded safety standards) and for PM10 were 15.0 µg/m3/day (standard deviation: 3.1). An interquartile range increase in PM2.5 exposure (10.73 µg/m3/day) was associated with a -0.0287 standard deviation lower cumulative DNA methylation score for PM2.5 (95% CI: -0.0732, 0.0158, p = 0.20) across all participants. An interquartile range increase in PM10 exposure (3.20 µg/m3/day) was associated with a -0.1472 standard deviation lower cumulative DNA methylation score for PM10 (95% CI: -0.3038, 0.0095, p = 0.06) across all participants. The PM10 findings were driven by the age 15 subset where an interquartile range increase in PM10 exposure was associated with a -0.024 standard deviation lower cumulative DNA methylation score for PM10 (95% CI: -0.043, -0.005, p = 0.012). Findings were robust to adjustment for PM exposure at ages 1 and 3. In utero PM10-associated DNA methylation differences were identified at age 15 in saliva. Benchmarking the timing and cell-type generalizability is critical for epigenetic exposure biomarker assessment.

Coming up short: Comparing venous blood, dried blood spots & saliva samples for measuring telomere length in health equity research.

BACKGROUND: Telomere length (TL) in peripheral blood mononuclear cells (PBMC) from fresh venous blood is increasingly used to estimate molecular impacts of accumulated social adversity on population health. Sometimes, TL extracted from saliva or dried blood spots (DBS) are substituted as less invasive and more scalable specimen collection methods; yet, are they interchangeable with fresh blood? Studies find TL is correlated across tissues, but have not addressed the critical question for social epidemiological applications: Do different specimen types show the same association between TL and social constructs? METHODS: We integrate expertise in social epidemiology, molecular biology, and the statistical impact of measurement error on parameter estimates. Recruiting a diverse sample of 132 Metro-Detroit women, we measure TL for each woman from fresh blood PBMC, DBS, and saliva. Using regression methods, we estimate associations between social characteristics and TL, comparing estimates across specimen types for each woman. RESULTS: Associations between TL and social characteristics vary by specimen type collected from the same woman, sometimes qualitatively altering estimates of the magnitude or direction of a theorized relationship. Being Black is associated with shorter TL in PBMC, but longer TL in saliva or DBS. Education is positively associated with TL in fresh blood, but negatively associated with TL using DBS. CONCLUSION: Findings raise concerns about the use of TL measures derived from different tissues in social epidemiological research. Investigators need to consider the possibility that associations between social variables and TL may be systematically related to specimen type, rather than be valid indicators of socially-patterned biopsychosocial processes.

Early childhood psychosocial family risks and cumulative dopaminergic sensitizing score: Links to behavior problems in U.S. 9-year-olds.

BACKGROUND: We examined, (a) whether in early childhood exposure to risky family environment in different domains (socioeconomic, mental, parenting practices, health behavior, and child-related risks) and accumulatively across various domains (cumulative risk) is associated with child's problem behavior at age 9, and (b) whether the association is more pronounced in children carrying cumulative dopaminergic sensitizing genotype or living in low-income families. METHODS: Participants were 2,860 9-year old children (48% females; 48% Black) and their mothers from the 'Fragile Families and Child Wellbeing Study', a probability birth cohort from large U.S. cities. Mothers responded to questions on child's problem behavior (CBCL). Children responded to questions about their vandalism and substance use. RESULTS: Cumulative family risk was associated with higher internalizing and externalizing behavior and higher vandalism and substance use. All domain-specific risk clusters were associated with higher internalizing behavior and, with the exception of child-related risk, with higher externalizing behavior. Mental health risks, risky parenting practices, and risky health behavior were associated with higher vandalism. Risky parenting practices were associated with higher substance use. The associations were robust to adjustment for cumulative dopaminergic sensitizing genotype. No G x E interactions with dopaminergic genotype and family SES were observed. LIMITATIONS: Sample size was relatively small for genetic analysis and polygenic risk scores were not available. CONCLUSIONS: Exposure to cumulative psychosocial family risks from early childhood is associated with early indicators of problem behavior in adolescence.

Early Puberty and Telomere Length in Preadolescent Girls and Mothers.

OBJECTIVE: To test the association between early puberty and telomere length in preadolescent girls and mothers from a large representative sample of US females. STUDY DESIGN: We analyzed data from 1194 preadolescent girls and 2421 mothers from the Fragile Families and Child Wellbeing Study. Participants were from a population-based birth cohort (1998-2000) born in large US cities. Telomere length was assessed by quantitative polymerase chain reaction from saliva samples provided by preadolescent girls and mothers of preadolescent youth. Mothers completed a questionnaire about their child's pubertal development to determine concurrent Tanner stages and provided self-reports of her own age at menarche. Linear regression models were used to estimate the association between pubertal development (status and timing) and telomere length. RESULTS: Early pubertal timing but not pubertal status was associated with shorter telomere length in preadolescent girls (P < .01). Early age at menarche was associated with shorter telomere length in a sample of mothers of preadolescent youth (P < .05). CONCLUSIONS: Results provide evidence for the association between early puberty and shorter telomeres evidenced by associations in both preadolescent girls and mothers. Future research should address the limitations of this study by using longitudinal measurements of pubertal development assessed through medical examinations and repeated assessments of telomere length to capture telomere attrition.

Demographic and developmental patterns in telomere length across adolescence.

Telomere length is often used in studies of adults as a biomarker of cellular aging and an indicator of stress exposure. However, we know little about how telomeres change over time, particularly over the course of the important developmental period of adolescence. We use data on telomere length collected at two points in time spanning adolescence (Years 9 and 15) from the Fragile Families and Child Wellbeing Study to examine longitudinal patterns (n = 1,654) in telomere length. We find a quantitatively small but significant average lengthening in telomere length across adolescence and little evidence of associations between telomere length and pubertal development.

Shared Reading at Age 1 Year and Later Vocabulary: A Gene-Environment Study.

OBJECTIVE: To assess the extent to which associations between shared reading at age 1 years and child vocabulary at age 3 years differ based on the presence of sensitizing alleles in the dopaminergic and serotonergic neurotransmitter systems. STUDY DESIGN: We conducted a secondary analysis of data from a national urban birth cohort using mother reports in conjunction with child assessments and salivary genetic data. Child vocabulary was assessed using the Peabody Picture Vocabulary Test. The primary exposure was mother-reported shared reading. We used data on gene variants that may affect the function of the dopaminergic and serotonergic systems. We examined associations between shared reading and Peabody Picture Vocabulary Test score using multiple linear regression. We then included interaction terms between shared reading and the presence of sensitizing alleles for each polymorphism to assess potential moderator effects adjusting for multiple comparisons. RESULTS: Of the 1772 children included (56% black, 52% male), 31% of their mothers reported reading with their child daily. Daily shared reading was strongly associated with child Peabody Picture Vocabulary Test scores in unadjusted (B = 7.9; 95% CI, 4.3-11.4) and adjusted models (B = 5.3; 95% CI, 2.0-8.6). The association differed based on the presence of sensitizing alleles in the dopamine receptor 2 and serotonin transporter genes. CONCLUSIONS: Among urban children, shared reading at age 1 years was associated with greater vocabulary at age 3 years. Although children with sensitizing alleles on the dopamine receptor 2 and serotonin transporter genes were at greater risk when not read to, they fared as well as children without these alleles when shared reading occurred.

Apolipoprotein E, cognitive function, and cognitive decline among older Taiwanese adults.

Apolipoprotein E (APOE) genotype is believed to play a role in the onset of dementia, though less is known about its relationship with non-pathogenic age-related cognitive decline. We assessed whether APOE was a risk factor for cognitive decline among older Taiwanese adults using nationally representative data. General cognition was measured longitudinally over eleven years; domain-specific cognitive assessments of working memory, declarative learning and three aspects of attention (executive function, alerting, and orientation) were performed once. Having at least one risky APOE allele was associated with more rapid longitudinal cognitive decline compared to those with no risky alleles. Some evidence from the cross-sectional analysis of domain-specific cognitive assessments suggested that APOE genotype may be more closely associated with working memory and declarative learning than with attention. Most genetic studies of cognition include only populations of European descent; extension is crucial. This study confirmed the association between APOE genotype and the rate of cognitive decline in a predominantly Han Chinese population. Additional studies on diverse populations are warranted.

Pseudomonas aeruginosa Regulated Intramembrane Proteolysis: Protease MucP Can Overcome Mutations in the AlgO Periplasmic Protease To Restore Alginate Production in Nonmucoid Revertants.

The progression of cystic fibrosis (CF) from an acute to a chronic disease is often associated with the conversion of the opportunistic pathogen Pseudomonas aeruginosa from a nonmucoid form to a mucoid form in the lung. This conversion involves the constitutive synthesis of the exopolysaccharide alginate, whose production is under the control of the AlgT/U sigma factor. This factor is regulated posttranslationally by an extremely unstable process and has been commonly attributed to mutations in the algT (algU) gene. By exploiting this unstable phenotype, we isolated 34 spontaneous nonmucoid variants arising from the mucoid strain PDO300, a PAO1 derivative containing the mucA22 allele commonly found in mucoid CF isolates. Complementation analysis using a minimal tiling path cosmid library revealed that most of these mutants mapped to two protease-encoding genes, algO, also known as prc or PA3257, and mucP Interestingly, our algO mutations were complemented by both mucP and algO, leading us to delete, clone, and overexpress mucP, algO, mucE, and mucD in both wild-type PAO1 and PDO300 backgrounds to better understand the regulation of this complex regulatory mechanism. Our findings suggest that the regulatory proteases follow two pathways for regulated intramembrane proteolysis (RIP), where both the AlgO/MucP pathway and MucE/AlgW pathway are required in the wild-type strain but where the AlgO/MucP pathway can bypass the MucE/AlgW pathway in mucoid strains with membrane-associated forms of MucA with shortened C termini, such as the MucA22 variant. This work gives us a better understanding of how alginate production is regulated in the clinically important mucoid variants of Pseudomonas aeruginosaIMPORTANCE Infection by the opportunistic pathogen Pseudomonas aeruginosa is the leading cause of morbidity and mortality seen in CF patients. Poor patient prognosis correlates with the genotypic and phenotypic change of the bacteria from a typical nonmucoid to a mucoid form in the CF lung, characterized by the overproduction of alginate. The expression of this exopolysaccharide is under the control an alternate sigma factor, AlgT/U, that is regulated posttranslationally by a series of proteases. A better understanding of this regulatory phenomenon will help in the development of therapies targeting alginate production, ultimately leading to an increase in the length and quality of life for those suffering from CF.