Disease Modeling and Disease Gene Discovery in Cardiomyopathies: A Molecular Study of Induced Pluripotent Stem Cell Generated Cardiomyocytes.

The in vitro modeling of cardiac development and cardiomyopathies in human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs) provides opportunities to aid the discovery of genetic, molecular, and developmental changes that are causal to, or influence, cardiomyopathies and related diseases. To better understand the functional and disease modeling potential of iPSC-differentiated CMs and to provide a proof of principle for large, epidemiological-scale disease gene discovery approaches into cardiomyopathies, well-characterized CMs, generated from validated iPSCs of 12 individuals who belong to four sibships, and one of whom reported a major adverse cardiac event (MACE), were analyzed by genome-wide mRNA sequencing. The generated CMs expressed CM-specific genes and were highly concordant in their total expressed transcriptome across the 12 samples (correlation coefficient at 95% CI =0.92 ± 0.02). The functional annotation and enrichment analysis of the 2116 genes that were significantly upregulated in CMs suggest that generated CMs have a transcriptomic and functional profile of immature atrial-like CMs; however, the CMs-upregulated transcriptome also showed high overlap and significant enrichment in primary cardiomyocyte (p-value = 4.36 × 10-9), primary heart tissue (p-value = 1.37 × 10-41) and cardiomyopathy (p-value = 1.13 × 10-21) associated gene sets. Modeling the effect of MACE in the generated CMs-upregulated transcriptome identified gene expression phenotypes consistent with the predisposition of the MACE-affected sibship to arrhythmia, prothrombotic, and atherosclerosis risk.

Role of miRNA-mRNA Interaction in Neural Stem Cell Differentiation of Induced Pluripotent Stem Cells.

miRNA regulates the expression of protein coding genes and plays a regulatory role in human development and disease. The human iPSCs and their differentiated progenies provide a unique opportunity to identify these miRNA-mediated regulatory mechanisms. To identify miRNA-mRNA regulatory interactions in human nervous system development, well characterized NSCs were differentiated from six validated iPSC lines and analyzed for differentially expressed (DE) miRNome and transcriptome by RNA sequencing. Following the criteria, moderated t statistics, FDR-corrected p-value ≤ 0.05 and fold change-absolute (FC-abs) ≥2.0, 51 miRNAs and 4033 mRNAs were found to be significantly DE between iPSCs and NSCs. The miRNA target prediction analysis identified 513 interactions between 30 miRNA families (mapped to 51 DE miRNAs) and 456 DE mRNAs that were paradoxically oppositely expressed. These 513 interactions were highly enriched in nervous system development functions (154 mRNAs; FDR-adjusted p-value range: 8.06 × 10-15-1.44 × 10-4). Furthermore, we have shown that the upregulated miR-10a-5p, miR-30c-5p, miR23-3p, miR130a-3p and miR-17-5p miRNA families were predicted to down-regulate several genes associated with the differentiation of neurons, neurite outgrowth and synapse formation, suggesting their role in promoting the self-renewal of undifferentiated NSCs. This study also provides a comprehensive characterization of iPSC-generated NSCs as dorsal neuroepithelium, important for their potential use in in vitro modeling of human brain development and disease.

Rare DEGS1 variant significantly alters de novo ceramide synthesis pathway.

The de novo ceramide synthesis pathway is essential to human biology and health, but genetic influences remain unexplored. The core function of this pathway is the generation of biologically active ceramide from its precursor, dihydroceramide. Dihydroceramides have diverse, often protective, biological roles; conversely, increased ceramide levels are biomarkers of complex disease. To explore the genetics of the ceramide synthesis pathway, we searched for deleterious nonsynonymous variants in the genomes of 1,020 Mexican Americans from extended pedigrees. We identified a Hispanic ancestry-specific rare functional variant, L175Q, in delta 4-desaturase, sphingolipid 1 (DEGS1), a key enzyme in the pathway that converts dihydroceramide to ceramide. This amino acid change was significantly associated with large increases in plasma dihydroceramides. Indexes of DEGS1 enzymatic activity were dramatically reduced in heterozygotes. CRISPR/Cas9 genome editing of HepG2 cells confirmed that the L175Q variant results in a partial loss of function for the DEGS1 enzyme. Understanding the biological role of DEGS1 variants, such as L175Q, in ceramide synthesis may improve the understanding of metabolic-related disorders and spur ongoing research of drug targets along this pathway.

Pleiotropy of cardiometabolic syndrome with obesity-related anthropometric traits determined using empirically derived kinships from the Busselton Health Study.

Over two billion adults are overweight or obese and therefore at an increased risk of cardiometabolic syndrome (CMS). Obesity-related anthropometric traits genetically correlated with CMS may provide insight into CMS aetiology. The aim of this study was to utilise an empirically derived genetic relatedness matrix to calculate heritabilities and genetic correlations between CMS and anthropometric traits to determine whether they share genetic risk factors (pleiotropy). We used genome-wide single nucleotide polymorphism (SNP) data on 4671 Busselton Health Study participants. Exploiting both known and unknown relatedness, empirical kinship probabilities were estimated using these SNP data. General linear mixed models implemented in SOLAR were used to estimate narrow-sense heritabilities (h 2) and genetic correlations (r g) between 15 anthropometric and 9 CMS traits. Anthropometric traits were adjusted by body mass index (BMI) to determine whether the observed genetic correlation was independent of obesity. After adjustment for multiple testing, all CMS and anthropometric traits were significantly heritable (h 2 range 0.18-0.57). We identified 50 significant genetic correlations (r g range: - 0.37 to 0.75) between CMS and anthropometric traits. Five genetic correlations remained significant after adjustment for BMI [high density lipoprotein cholesterol (HDL-C) and waist-hip ratio; triglycerides and waist-hip ratio; triglycerides and waist-height ratio; non-HDL-C and waist-height ratio; insulin and iliac skinfold thickness]. This study provides evidence for the presence of potentially pleiotropic genes that affect both anthropometric and CMS traits, independently of obesity.

Quantitative physical activity assessment of children and adolescents in a rural population from Eastern Nepal.

OBJECTIVES: We report cross-sectional, objectively measured physical activity data for 399 children and adolescents aged 6 to 18 years. We evaluated physical activity of children and adolescents, considered time spent in each activity intensity category, and explored the impact of growth disruption (stunting and wasting) on physical activity patterns. METHODS: Participants wore an Actical (Mini-Mitter, Bend, OR) omnidirectional accelerometer for one week as part of their annual visit to the Jiri Growth Study. The percentage of time spent in standard activity intensities were computed using standard metabolic equivalents (METS) cutpoints and compared by chronological age, sex, and school versus non-school days. RESULTS: Primary findings include (1) children are more active on non-school days and adolescents are more active during the school week; (2) Jirel children do not exhibit the reduction in physical activity that most Western populations experience during the transition from childhood to adolescence; and (3) Jirel children and adolescents routinely meet the suggested one hour/day MVPA threshold; (4) Stunting is prevalent and factors leading to this growth disruption may contribute to the amount of time in sedentary or light physical activity. CONCLUSIONS: We report child and adolescent physical activity patterns from the Jirel population of eastern Nepal. In this rural context, children and adolescents are more active than populations reported from Western contexts. This key finding has important biomedical implications for the maintenance of healthy body composition, skeletal health, and other health traits.

Lipidomic risk score independently and cost-effectively predicts risk of future type 2 diabetes: results from diverse cohorts.

BACKGROUND: Detection of type 2 diabetes (T2D) is routinely based on the presence of dysglycemia. Although disturbed lipid metabolism is a hallmark of T2D, the potential of plasma lipidomics as a biomarker of future T2D is unknown. Our objective was to develop and validate a plasma lipidomic risk score (LRS) as a biomarker of future type 2 diabetes and to evaluate its cost-effectiveness for T2D screening. METHODS: Plasma LRS, based on significantly associated lipid species from an array of 319 lipid species, was developed in a cohort of initially T2D-free individuals from the San Antonio Family Heart Study (SAFHS). The LRS derived from SAFHS as well as its recalibrated version were validated in an independent cohort from Australia--the AusDiab cohort. The participants were T2D-free at baseline and followed for 9197 person-years in the SAFHS cohort (n = 771) and 5930 person-years in the AusDiab cohort (n = 644). Statistically and clinically improved T2D prediction was evaluated with established statistical parameters in both cohorts. Modeling studies were conducted to determine whether the use of LRS would be cost-effective for T2D screening. The main outcome measures included accuracy and incremental value of the LRS over routinely used clinical predictors of T2D risk; validation of these results in an independent cohort and cost-effectiveness of including LRS in screening/intervention programs for T2D. RESULTS: The LRS was based on plasma concentration of dihydroceramide 18:0, lysoalkylphosphatidylcholine 22:1 and triacyglycerol 16:0/18:0/18:1. The score predicted future T2D independently of prediabetes with an accuracy of 76%. Even in the subset of initially euglycemic individuals, the LRS improved T2D prediction. In the AusDiab cohort, the LRS continued to predict T2D significantly and independently. When combined with risk-stratification methods currently used in clinical practice, the LRS significantly improved the model fit (p < 0.001), information content (p < 0.001), discrimination (p < 0.001) and reclassification (p < 0.001) in both cohorts. Modeling studies demonstrated that LRS-based risk-stratification combined with metformin supplementation for high-risk individuals was the most cost-effective strategy for T2D prevention. CONCLUSIONS: Considering the novelty, incremental value and cost-effectiveness of LRS it should be used for risk-stratification of future T2D.

Statistical Genetic Approaches to Investigate Genotype-by-Environment Interaction: Review and Novel Extension of Models.

Statistical genetic models of genotype-by-environment (G×E) interaction can be divided into two general classes, one on G×E interaction in response to dichotomous environments (e.g., sex, disease-affection status, or presence/absence of an exposure) and the other in response to continuous environments (e.g., physical activity, nutritional measurements, or continuous socioeconomic measures). Here we develop a novel model to jointly account for dichotomous and continuous environments. We develop the model in terms of a joint genotype-by-sex (for the dichotomous environment) and genotype-by-social determinants of health (SDoH; for the continuous environment). Using this model, we show how a depression variable, as measured by the Beck Depression Inventory-II survey instrument, is not only underlain by genetic effects (as has been reported elsewhere) but is also significantly determined by joint G×Sex and G×SDoH interaction effects. This model has numerous applications leading to potentially transformative research on the genetic and environmental determinants underlying complex diseases.

Role of Neurocellular Endoplasmic Reticulum Stress Response in Alzheimer's Disease and Related Dementias Risk.

Currently, more than 55 million people around the world suffer from dementia, and Alzheimer's Disease and Related Dementias (ADRD) accounts for nearly 60-70% of all those cases. The spread of Alzheimer's Disease (AD) pathology and progressive neurodegeneration in the hippocampus and cerebral cortex is strongly correlated with cognitive decline in AD patients; however, the molecular underpinning of ADRD's causality is still unclear. Studies of postmortem AD brains and animal models of AD suggest that elevated endoplasmic reticulum (ER) stress may have a role in ADRD pathology through altered neurocellular homeostasis in brain regions associated with learning and memory. To study the ER stress-associated neurocellular response and its effects on neurocellular homeostasis and neurogenesis, we modeled an ER stress challenge using thapsigargin (TG), a specific inhibitor of sarco/endoplasmic reticulum Ca2+ ATPase (SERCA), in the induced pluripotent stem cell (iPSC)-derived neural stem cells (NSCs) of two individuals from our Mexican American Family Study (MAFS). High-content screening and transcriptomic analysis of the control and ER stress-challenged NSCs showed that the NSCs' ER stress response resulted in a significant decline in NSC self-renewal and an increase in apoptosis and cellular oxidative stress. A total of 2300 genes were significantly (moderated t statistics FDR-corrected p-value ≤ 0.05 and fold change absolute ≥ 2.0) differentially expressed (DE). The pathway enrichment and gene network analysis of DE genes suggests that all three unfolded protein response (UPR) pathways, protein kinase RNA-like ER kinase (PERK), activating transcription factor-6 (ATF-6), and inositol-requiring enzyme-1 (IRE1), were significantly activated and cooperatively regulated the NSCs' transcriptional response to ER stress. Our results show that IRE1/X-box binding protein 1 (XBP1) mediated transcriptional regulation of the E2F transcription factor 1 (E2F1) gene, and its downstream targets have a dominant role in inducing G1/S-phase cell cycle arrest in ER stress-challenged NSCs. The ER stress-challenged NSCs also showed the activation of C/EBP homologous protein (CHOP)-mediated apoptosis and the dysregulation of synaptic plasticity and neurotransmitter homeostasis-associated genes. Overall, our results suggest that the ER stress-associated attenuation of NSC self-renewal, increased apoptosis, and dysregulated synaptic plasticity and neurotransmitter homeostasis plausibly play a role in the causation of ADRD.

Reconstructing parent genomes using siblings and other relatives.

Reconstructing the DNA of ancestors from their descendants has the potential to empower phenotypic analyses (including association and genetic nurture studies), improve pedigree reconstruction, and shed light on the ancestral population and phenotypes of ancestors. We developed HAPI-RECAP, a method that reconstructs the DNA of parents from full siblings and their relatives. This tool leverages HAPI2's output, a new phasing approach that applies to siblings (and optionally one or both parents) and reliably infers parent haplotypes but does not link the ungenotyped parents' DNA across chromosomes or between segments flanking ambiguities. By combining IBD between the reconstructed parents and the relatives, HAPI-RECAP resolves the source parent of these segments. Moreover, the method exploits crossovers the children inherited and sex-specific genetic maps to infer the reconstructed parents' sexes. We validated these methods on research participants from both 23andMe, Inc. and the San Antonio Mexican American Family Studies. Given data for one parent, HAPI2 reconstructs large fractions of the missing parent's DNA, between 77.6% and 99.97% among all families, and 90.3% on average in three- and four-child families. When reconstructing both parents, HAPI-RECAP inferred between 33.2% and 96.6% of the parents' genotypes, averaging 70.6% in four-child families. Reconstructed genotypes have average error rates < 10-3, or comparable to those from direct genotyping. HAPI-RECAP inferred the parent sexes 100% correctly given IBD-linked segments and can also reconstruct parents without any IBD. As datasets grow in size, more families will be implicitly collected; HAPI-RECAP holds promise to enable high quality parent genotype reconstruction.

Pre-Infection Innate Immunity Attenuates SARS-CoV-2 Infection and Viral Load in iPSC-Derived Alveolar Epithelial Type 2 Cells.

A large portion of the heterogeneity in coronavirus disease 2019 (COVID-19) susceptibility and severity of illness (SOI) remains poorly understood. Recent evidence suggests that SARS-CoV-2 infection-associated damage to alveolar epithelial type 2 cells (AT2s) in the distal lung may directly contribute to disease severity and poor prognosis in COVID-19 patients. Our in vitro modeling of SARS-CoV-2 infection in induced pluripotent stem cell (iPSC)-derived AT2s from 10 different individuals showed interindividual variability in infection susceptibility and the postinfection cellular viral load. To understand the underlying mechanism of the AT2's capacity to regulate SARS-CoV-2 infection and cellular viral load, a genome-wide differential gene expression analysis between the mock and SARS-CoV-2 infection-challenged AT2s was performed. The 1393 genes, which were significantly (one-way ANOVA FDR-corrected p ≤ 0.05; FC abs ≥ 2.0) differentially expressed (DE), suggest significant upregulation of viral infection-related cellular innate immune response pathways (p-value ≤ 0.05; activation z-score ≥ 3.5), and significant downregulation of the cholesterol- and xenobiotic-related metabolic pathways (p-value ≤ 0.05; activation z-score ≤ -3.5). Whilst the effect of post-SARS-CoV-2 infection response on the infection susceptibility and postinfection viral load in AT2s is not clear, interestingly, pre-infection (mock-challenged) expression of 238 DE genes showed a high correlation with the postinfection SARS-CoV-2 viral load (FDR-corrected p-value ≤ 0.05 and r2-absolute ≥ 0.57). The 85 genes whose expression was negatively correlated with the viral load showed significant enrichment in viral recognition and cytokine-mediated innate immune GO biological processes (p-value range: 4.65 × 10-10 to 2.24 × 10-6). The 153 genes whose expression was positively correlated with the viral load showed significant enrichment in cholesterol homeostasis, extracellular matrix, and MAPK/ERK pathway-related GO biological processes (p-value range: 5.06 × 10-5 to 6.53 × 10-4). Overall, our results strongly suggest that AT2s' pre-infection innate immunity and metabolic state affect their susceptibility to SARS-CoV-2 infection and viral load.

Metabolic syndrome traits exhibit genotype-by-environment interaction in relation to socioeconomic status in the Mexican American family heart study.

Background: Socioeconomic Status (SES) is a potent environmental determinant of health. To our knowledge, no assessment of genotype-environment interaction has been conducted to consider the joint effects of socioeconomic status and genetics on risk for metabolic disease. We analyzed data from the Mexican American Family Studies (MAFS) to evaluate the hypothesis that genotype-by-environment interaction (GxE) is an essential determinant of variation in risk factors for metabolic syndrome (MS). Methods: We employed a maximum likelihood estimation of the decomposition of variance components to detect GxE interaction. After excluding individuals with diabetes and individuals on medication for diabetes, hypertension, or dyslipidemia, we analyzed 12 MS risk factors: fasting glucose (FG), fasting insulin (FI), 2-h glucose (2G), 2-h insulin (2I), body mass index (BMI), waist circumference (WC), leptin (LP), high-density lipoprotein-cholesterol (HDL-C), triglycerides (TG), total serum cholesterol (TSC), systolic blood pressure (SBP), and diastolic blood pressure (DBP). Our SES variable used a combined score of Duncan's socioeconomic index and education years. Heterogeneity in the additive genetic variance across the SES continuum and a departure from unity in the genetic correlation coefficient were taken as evidence of GxE interaction. Hypothesis tests were conducted using standard likelihood ratio tests. Results: We found evidence of GxE for fasting glucose, 2-h glucose, 2-h insulin, BMI, and triglycerides. The genetic effects underlying the insulin/glucose metabolism component of MS are upregulated at the lower end of the SES spectrum. We also determined that the household variance for systolic blood pressure decreased with increasing SES. Conclusion: These results show a significant change in the GxE interaction underlying the major components of MS in response to changes in socioeconomic status. Further mRNA sequencing studies will identify genes and canonical gene pathways to support our molecular-level hypotheses.

Nonsyndromic brachydactyly type D and type E mapped to 7p15 in healthy children and adults from the Jirel ethnic group in eastern Nepal.

OBJECTIVES: There is phenotypic overlap between Brachydactyly Type D (BDD) and Brachydactyly Type E (BDE) that suggests a possible common underlying etiology. We seek to understand the genetic underpinnings of, and relationship between, these skeletal anomalies. METHODS: The Jirel ethnic group of eastern Nepal participates in various genetic epidemiologic studies, including those in which hand-wrist radiographs have been taken to examine skeletal development. Nearly 2,130 individuals (969 males; 1,161 females) were phenotyped for BDD/BDE. Of these, 1,722 individuals (773 males; 949 females) were genotyped for 371 STR markers spanning the autosomal genome. Variance components-based linkage analysis was used to conduct a genome-wide linkage scan for QTL influencing the BDD/BDE phenotype. RESULTS: BDD was present in 3.55%, and BDE was present in 0.39%, of the study sample. Because of the phenotypic overlap between two traits, affecteds of either type were considered as affected by a single combined phenotype (BDD/BDE) having a prevalence of 3.94%. The additive genetic heritability of BDD/BDE was highly significant (h(2) ± SE = 0.89 ± 0.13; P = 1.7 × 10(-11) ). Significant linkage of BDD/BDE was found to markers on chromosome 7p21-7p14 (peak LOD score = 3.74 at 7p15 between markers D7S493 and D7S516). CONCLUSIONS: Possible positional candidate genes in the one-lod support interval of this QTL include TWIST and the HOXA1-A13 cluster. This is the first study to report significant linkage results for BDD/BDE using a large extended pedigree, and the first to suggest that mutations in TWIST and/or the HOXA1-A13 cluster may contribute to these specific skeletal anomalies.

Frailty index in the Colonias of the Rio Grande Valley: health related quality of life and resilience.

Background: Frailty is characterized by an accumulation of deficits that lead to vulnerability to adverse health outcomes. The Frailty Index (FI) quantifies frailty by measuring deficits that increase susceptibility to stressors. This study focused on a population of Mexican Americans living in vulnerable communities in the Rio Grande Valley of south Texas. We used a Frailty Index developed based on common health-related data--the Patient Health Questionnaire (PHQ-9) and a Health-related Quality of Life survey (Duke Health Profile). Quality of life, resilience, and frailty are interrelated and influenced by chronic illness, mental illness, poverty, cognitive impairment, and community support. Methods: We used Logistic regression analysis, factor component analysis, receiver operating characteristic curves, and odds ratios to identify potential associations between clinical variables and candidate predictor variables and seven physiological health variables, and two survey instruments. We analyzed data obtained from participants (894) that live in two Colonias located on the Texas-Mexico border. We calculated the FI with seven physiological variables, PHQ-9 score, and the 11 domain-specific Duke Profile scores, for a total of 19 health deficits. We then dichotomized FI (>0.25) and determined ROC curves through model selection to determine best predictors of frailty. Results: Females (n = 622) had a higher starting frailty, and males (n = 272) had a significantly greater change rate with age. Women score higher in anxiety, depression, anxiety/depression, and pain. The frailty index and quality of life markers are strongly inversely related; poorer quality of life leads to greater frailty independent physiological health variables, the PHQ 9, sex, and age. Conclusion: The study highlights the importance of addressing modifiable mental health and social stressors to reduce frailty. Furthermore, it suggests that factors supporting resilience and well-being, such as physical and mental health, social support, and perceived health, play a crucial role in frailty development. The findings have implications for interventions targeting vulnerable populations and emphasize the need for further research on the relationship between health-related quality of life and frailty.

Gene-by-Environment Interaction in Non-Alcoholic Fatty Liver Disease and Depression: The Role of Hepatic Transaminases.

Non-alcoholic fatty liver disease (NAFLD) encompasses a range of liver conditions, from benign fatty accumulation to severe fibrosis. The global prevalence of NAFLD has risen to 25-30%, with variations across ethnic groups. NAFLD may advance to hepatocellular carcinoma, increases cardiovascular risk, is associated with chronic kidney disease, and is an independent metabolic disease risk factor. Assessment methods for liver health include liver biopsy, magnetic resonance imaging, ultrasound, and vibration-controlled transient elastography (VCTE by FibroScan). Hepatic transaminases are cost-effective and minimally invasive liver health assessment methods options. This study focuses on the interaction between genetic factors underlying the traits (hepatic transaminases and the FibroScan results) on the one hand and the environment (depression) on the other. We examined 525 individuals at risk for metabolic disorders. We utilized variance components models and likelihood-based statistical inference to examine potential GxE interactions in markers of NAFLD, including aspartate aminotransferase (AST), alanine aminotransferase (ALT), and the AST/ALT ratio, and Vibration-Controlled Transient Elastography (VCTE by FibroScan). We calculated the Fibroscan-AST (FAST) score (a score that identifies the risk of progressive non-alcoholic steatohepatitis (NASH) and screened for depression using the Beck Depression Inventory-II (BDI-II). We identified significant G × E interactions for AST/ALT ratio × BDI-II, but not AST, ALT, or the FAST score. Our findings support that genetic factors play a role in hepatic transaminases, especially the AST/ALT ratio, with depression influencing this relationship. These insights contribute to understanding the complex interplay of genetics, environment, and liver health, potentially guiding future personalized interventions.

Genotype-by-socioeconomic status interaction influences heart disease risk scores and carotid artery thickness in Mexican Americans: the predominant role of education in comparison to household income and socioeconomic index.

Background: Socioeconomic status (SES) is a potent environmental determinant of health. To our knowledge, no assessment of genotype-environment interaction has been conducted to consider the joint effects of socioeconomic status and genetics on risk for cardiovascular disease (CVD). We analyzed Mexican American Family Studies (MAFS) data to evaluate the hypothesis that genotype-by-environment interaction (GxE) is an important determinant of variation in CVD risk factors. Methods: We employed a linear mixed model to investigate GxE in Mexican American extended families. We studied two proxies for CVD [Pooled Cohort Equation Risk Scores/Framingham Risk Scores (FRS/PCRS) and carotid artery intima-media thickness (CA-IMT)] in relation to socioeconomic status as determined by Duncan's Socioeconomic Index (SEI), years of education, and household income. Results: We calculated heritability for FRS/PCRS and carotid artery intima-media thickness. There was evidence of GxE due to additive genetic variance heterogeneity and genetic correlation for FRS, PCRS, and CA-IMT measures for education (environment) but not for household income or SEI. Conclusion: The genetic effects underlying CVD are dynamically modulated at the lower end of the SES spectrum. There is a significant change in the genetic architecture underlying the major components of CVD in response to changes in education.

A Scan of Pleiotropic Immune Mediated Disease Genes Identifies Novel Determinants of Baseline FVIII Inhibitor Status in Hemophilia-A.

Hemophilia-A (HA) is caused by heterogeneous loss-of-function factor (F)VIII gene (F8)-mutations and deficiencies in plasma-FVIII-activity that impair intrinsic-pathway-mediated coagulation-amplification. The standard-of-care for severe-HA-patients is regular infusions of therapeutic-FVIII-proteins (tFVIIIs) but ~30% develop neutralizing-tFVIII-antibodies called "FVIII-inhibitors (FEIs)" and become refractory. We used the PATH study and ImmunoChip to scan immune-mediated-disease (IMD)-genes for novel and/or replicated genomic-sequence-variations associated with baseline-FEI-status while accounting for non-independence of data due to genetic-relatedness and F8-mutational-heterogeneity. The baseline-FEI-status of 450 North American PATH subjects-206 with black-African-ancestry and 244 with white-European-ancestry-was the dependent variable. The F8-mutation-data and a genetic-relatedness matrix were incorporated into a binary linear-mixed model of genetic association with baseline-FEI-status. We adopted a gene-centric-association-strategy to scan, as candidates, pleiotropic-IMD-genes implicated in the development of either ³2 autoimmune-/autoinflammatory-disorders (AADs) or ³1 AAD and FEIs. Baseline-FEI-status was significantly associated with SNPs assigned to NOS2A (rs117382854; p=3.2E-6) and B3GNT2 (rs10176009; p=5.1E-6), which have functions in anti-microbial-/-tumoral-immunity. Among IMD-genes implicated in FEI-risk previously, we identified strong associations with CTLA4 assigned SNPs (p=2.2E-5). The F8-mutation-effect underlies ~15% of the total heritability for baseline-FEI-status. Additive genetic heritability and SNPs in IMD-genes account for >50% of the patient-specific variability in baseline-FEI-status. Race is a significant determinant independent of F8-mutation-effects and non-F8-genetics.

Evaluation of qualitative methods for phenotyping brachymesophalangia-V from radiographs of children.

OBJECTIVE: Brachymesophalangia-V (BMP-V), the general term for a short and broad middle phalanx of the 5th digit, presents both alone and in a large number of complex brachydactylies and developmental disorders. Past anthropological and epidemiological studies of growth and development have examined the prevalence of BMP-V because small developmental disorders may signal more complex disruptions of skeletal growth and development. Historically, however, consensus on qualitative phenotype methodology has not been established. In large-scale, non-clinical studies such as the Fels Longitudinal Study and the Jiri Growth Study, quantitative assessment of the hand is not always the most efficient manner of screening for skeletal dysmorphologies. The current study evaluates qualitative phenotyping techniques for BMP-V used in past anthropological studies of growth and development to establish a useful and reliable screening method for large study samples. METHODS: A total of 1,360 radiographs from Jiri Growth Study participants aged 3-18 years were evaluated. BMP-V was assessed using three methods: (1) subjective evaluation of length and width of the bone; (2) comparison with skeletal age-matched radiographs; and (3) subjective evaluation of the length of the middle 4th and 5th phalanges. RESULTS: We found that the method that uses skeletal age-matched reference radiographs is the better tool for assessing BMP-V because it considers the shape, rather than solely the length and width of the bone, which can be difficult to judge accurately without measurement. This study highlights the complexity of phenotypic assessment of BMP-V and by extension other brachydactylies.

Efficient Generation of Functional Hepatocytes from Human Induced Pluripotent Stem Cells for Disease Modeling and Disease Gene Discovery.

In vitro hepatocyte cell models are being used to study the pathogenesis of liver disease and in the discovery and preclinical stages of drug development. The culture of hepatic cell lines and primary hepatocytes as in vitro cell models has been carried out for several decades. However, hepatic cell lines (hepatic carcinoma generated or immortalized) have limited accuracy when recapitulating complex physiological functions of the liver. Additionally, primary hepatocytes sourced from human cadavers or medical biopsies are difficult to obtain due to sourcing limitations, particularly for large-scale population studies or in applications requiring large number of cells. Hepatocyte cultures differentiated from human embryonic stem cells (ESCs) and induced pluripotent stem cell (iPSCs) overcome in large part the limitations of traditional hepatocyte in vitro models. In this chapter, we described an efficient protocol routinely used in our laboratory to differentiate human iPSCs into functional hepatocyte cultures for in vitro modeling of liver function and disease. The protocol uses a three-stage differentiation strategy to generate functional hepatocytes from human iPSCs. The differentiated cells show characteristic hepatocyte morphology including flat and polygonal shape, distinct round nuclei, and presence of biliary canaliculi and they express hepatic markers alpha-fetoprotein (AFP), albumin (ALB), E-cadherin (CHD1), hepatocyte nuclear factor 4 alpha (HNF4α), and actin.

Non-alcoholic Fatty Liver Disease and Depression: Evidence for Genotype × Environment Interaction in Mexican Americans.

This study examines the impact of G × E interaction effects on non-alcoholic fatty liver disease (NAFLD) among Mexican Americans in the Rio Grande Valley (RGV) of South Texas. We examined potential G × E interaction using variance components models and likelihood-based statistical inference in the phenotypic expression of NAFLD, including hepatic steatosis and hepatic fibrosis (identified using vibration controlled transient elastography and controlled attenuation parameter measured by the FibroScan Device). We screened for depression using the Beck Depression Inventory-II (BDI-II). We identified significant G × E interactions for hepatic fibrosis × BDI-II. These findings provide evidence that genetic factors interact with depression to influence the expression of hepatic fibrosis.

Genetic influences on dentognathic morphology in the Jirel population of Nepal.

Patterns of genetic variation and covariation impact the evolution of the craniofacial complex and contribute to clinically significant malocclusions in modern human populations. Previous quantitative genetic studies have estimated the heritabilities and genetic correlations of skeletal and dental traits in humans and nonhuman primates, but none have estimated these quantitative genetic parameters across the dentognathic complex. A large and powerful pedigree from the Jirel population of Nepal was leveraged to estimate heritabilities and genetic correlations in 62 maxillary and mandibular arch dimensions, incisor and canine lengths, and post-canine tooth crown areas (N ≥ 739). Quantitative genetic parameter estimation was performed using maximum likelihood-based variance decomposition. Residual heritability estimates were significant for all traits, ranging from 0.269 to 0.898. Genetic correlations were positive for all trait pairs. Principal components analyses of the phenotypic and genetic correlation matrices indicate an overall size effect across all measurements on the first principal component. Additional principal components demonstrate positive relationships between post-canine tooth crown areas and arch lengths and negative relationships between post-canine tooth crown areas and arch widths, and between arch lengths and arch widths. Based on these findings, morphological variation in the human dentognathic complex may be constrained by genetic relationships between dental dimensions and arch lengths, with weaker genetic correlations between these traits and arch widths allowing for variation in arch shape. The patterns identified are expected to have impacted the evolution of the dentognathic complex and its genetic architecture as well as the prevalence of dental crowding in modern human populations.