Xenobiotic Metabolism Genes and Clubfoot

Idiopathic or isolated clubfoot is a common orthopedic birth defect that affects approximately 135,000 children worldwide. It is characterized by equinus, varus and adductus deformities of the ankle and foot. Correction of clubfoot involves months of serial manipulations, castings and bracing, with surgical correction needed in forty percent of cases. Multifactorial etiology has been suggested in numerous studies with both environmental and genetic factors playing an etiologic role. Maternal smoking during pregnancy is the only common environmental factor that has consistently been shown to increase the risk for clubfoot. Moreover, a positive family history of clubfoot and maternal smoking increases the risk of clubfoot twenty fold. These findings suggest that genetic variation in smoking metabolism genes may increase susceptibility to clubfoot. Based on this reasoning, we interrogated eight candidate genes, chosen based on their involvement in phase 1 and 2 cigarette smoke metabolism. Twenty-two SNPs and two null alleles in eight genes (CYP1A1, CYP1A2, CYP1B1, CYP2A6, EPHX1, NAT2, GSTM1 and GSTT1) were genotyped in a dataset composed of nonHispanic white and Hispanic multiplex and simplex families. Only one SNP in CYP1A1, rs1048943, had significantly altered transmission in the aggregate and multiplex NHW datasets (p=0.003 and p=0.009). Perturbation of CYP1A1 by rs1048943 polymorphism causes an increase in the amount of harmful, adduct forming metabolic intermediates. A significant gene interaction between EPHX1 and NAT2 was also found (p=0.007). This interaction may affect the metabolism of harmful metabolic intermediates. Additionally, marginal interactions were found for other xenobiotic genes and these interactions may play a contributory role in clubfoot. Importantly, for CYP1A2, significant maternal (p=0.03; RR=1.24; 95% CI: 1.04-1.44) and fetal (p=0.01; RR=1.33; 95% CI: 1.13-1.54) genotypic effects were identified suggesting that both maternal and fetal genotypes impact normal limb development. No association was found for maternal smoking status and tobacco metabolism genes. Together, these results suggest that xenobiotic metabolism genes may play a contributory role in the etiology of clubfoot regardless of maternal smoking status and may impact foot development through perturbation of tobacco metabolic pathways.

Multiple independent genetic factors at NOS1AP modulate the QT interval in a multi-ethnic population.

Extremes of electrocardiographic QT interval are associated with increased risk for sudden cardiac death (SCD); thus, identification and characterization of genetic variants that modulate QT interval may elucidate the underlying etiology of SCD. Previous studies have revealed an association between a common genetic variant in NOS1AP and QT interval in populations of European ancestry, but this finding has not been extended to other ethnic populations. We sought to characterize the effects of NOS1AP genetic variants on QT interval in the multi-ethnic population-based Dallas Heart Study (DHS, n = 3,072). The SNP most strongly associated with QT interval in previous samples of European ancestry, rs16847548, was the most strongly associated in White (P = 0.005) and Black (P = 3.6 x 10(-5)) participants, with the same direction of effect in Hispanics (P = 0.17), and further showed a significant SNP x sex-interaction (P = 0.03). A second SNP, rs16856785, uncorrelated with rs16847548, was also associated with QT interval in Blacks (P = 0.01), with qualitatively similar results in Whites and Hispanics. In a previously genotyped cohort of 14,107 White individuals drawn from the combined Atherosclerotic Risk in Communities (ARIC) and Cardiovascular Health Study (CHS) cohorts, we validated both the second locus at rs16856785 (P = 7.63 x 10(-8)), as well as the sex-interaction with rs16847548 (P = 8.68 x 10(-6)). These data extend the association of genetic variants in NOS1AP with QT interval to a Black population, with similar trends, though not statistically significant at P<0.05, in Hispanics. In addition, we identify a strong sex-interaction and the presence of a second independent site within NOS1AP associated with the QT interval. These results highlight the consistent and complex role of NOS1AP genetic variants in modulating QT interval.

Fluktuierende Kopfschmerzen und Wesensveränderung bei einem 63-jährigen Patienten

We present a 63 year old man with new onset of fluctuating headache and behavioural changes showing marked inhibition and disorientation. After non invasive and invasive diagnostics an isolated cerebral vasculitis was found. Key results have been found in cerebral MRI and cerebral digital subtraction angiography with irregularities of vessel calibre of the intracerebral arteries. During treatment with high-dose corticosteroid therapy and Cyclophosphamid pulse therapy qualitative disorders and headache rapidly regressed. We discuss differential diagnosis of secondary headache, etiology of cerebral vasculitides, diagnostic challenge and therapy in isolated cerebral vasculitis.

Status epilepticus and multiple organ system dysfunction

Recent reports have suggested the possible association of status epilepticus and multiple organ system failure. The purpose of this case control study was to investigate this association and to identify factors that predispose individuals with status epilepticus (SE) or aborted status epilepticus (ASE) to develop multiple organ system failure (MOSF) or multiple organ system dysfunction (MODS).^ For the purpose of the study, definitions of SE, ASE, MOSF, and MODS were operationalized as follows: SE was defined as any seizure lasting for a duration of $\ge$30 minutes or intermittent seizures lasting for $\ge$30 minutes from which the patient does not regain consciousness. ASE was defined as any seizure lasting for a duration of $\ge$10 minutes but $<$30 minutes and which was aborted as a result of a medical intervention. MOSF was defined as the failure of $\ge$2 organ systems in the same patient; organ system failure was said to be present whenever standard MOSF criteria were met. MODS was defined as the dysfunction of $\ge$2 organ systems in the same patient; organ system dysfunction was said to be present, whenever the monitor(s) of that organ's function exceeded the normal range for the physiological or laboratory parameters.^ Medical records of 686 individuals between the age of 5 and 44 years, with history of seizures needing hospitalization at the Texas Children's Hospital or Methodist Hospital, Houston, Texas, between 1991-95 were reviewed and 100 individuals with SE/ASE were identified. Of these 100 individuals with SE/ASE, 45 developed MOSF/MODS during their hospitalization and 9 of these individuals died. Using multivariate analyses, it was found that adult individuals who had an "acute" etiology of their seizure disorder (OR = 5.23; 95%CI: 0.41, 66.24) and children who had a "remote" etiology of their seizure disorder (OR = 3.92; 95%CI: 0.53, 29.22), were more likely to develop MOSF/MODS compared with those who had other etiologies of the seizure disorder. Individuals with SE lasting more than one hour were more likely to develop MOSF/MODS compared with individuals with SE lasting less than 1 hour (OR = 6.51; 95%CI: 1.63, 25.92). Individuals who presented with the SE/ASE episode as their first seizure episode were more likely to develop MOSF/MODS compared to those with a previous history of seizure episodes (OR = 1.78; 95%CI: 0.36, 8.82).^ The major limitations of this study includes the relatively small sample size and the study being performed in only two institutions. However, this is the first study of this kind and should therefore be viewed as largely exploratory in nature. Future studies should investigate the relationship of the risk factors identified in this study using a larger number of institutions and patients. ^

Emergency referrals for syncopes - is an ambulatory treatment safe?

Patients after syncopy arrive frequently in an emergency unit. Two scoring systems have been validated for clinical decision making, use of diagnostic methods and need for hospitalisation. Goal of the study was quality control of ambulatory treatment of syncope patients in a University Emergency Department. 200 consecutive patients with syncope were documented, 109 of whom followed by phone-call during two years. The decision for hospitalisation or ambulatory treatment was up to the treating doctor. Age-distribution was biphasic: female sex mainly below the age 25, from 55 to 75 predominantly men. Etiology of syncope remained unclear for the majority of cases, a few neurologic (n=3) or cardiac (n=5) reasons were found with treatment consequences.

Multiple Sclerosis: Current Knowledge and Future Outlook

Background: Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system characterized by demyelination and axonal loss. The etiology of MS is unknown; however, environmental and genetic factors play a key role in the development of MS. Diagnostic criteria have been adapted to facilitate earlier diagnosis with increased sensitivity and specificity. Our understanding of the pathophysiology of MS has deepened considerably in recent years, resulting in different therapies to modify the disease course. Furthermore, several drugs have lately shown efficacy in phase III studies and their approval is expected in the near future. As treatment options expand, a future challenge will be to find the optimal treatment for the individual patient. Summary: This mini-review gives an overview of the current knowledge of MS with emphasis on the latest diagnostic criteria and both current and upcoming treatment options. Key Messages: Treatment of MS changes rapidly as the knowledge and therapeutic options in MS expand. Clinical Impact: Diagnosis of MS is based on McDonald criteria. MS therapy can be divided into relapse, disease-modifying and symptomatic treatment. Relapses are commonly treated with intravenous methylprednisolone. First-line therapy consists of either interferon-β, glatiramer acetate or teriflunomide. In general, agents used as escalation therapies (natalizumab, fingolimod and mitoxantrone) are more potent than the agents used for first-line therapy; however, these have potentially serious side effects and should be used with care.

A Multi-Breed Genome-Wide Association Analysis for Canine Hypothyroidism Identifies a Shared Major Risk Locus on CFA12.

Hypothyroidism is a complex clinical condition found in both humans and dogs, thought to be caused by a combination of genetic and environmental factors. In this study we present a multi-breed analysis of predisposing genetic risk factors for hypothyroidism in dogs using three high-risk breeds-the Gordon Setter, Hovawart and the Rhodesian Ridgeback. Using a genome-wide association approach and meta-analysis, we identified a major hypothyroidism risk locus shared by these breeds on chromosome 12 (p = 2.1x10-11). Further characterisation of the candidate region revealed a shared ~167 kb risk haplotype (4,915,018-5,081,823 bp), tagged by two SNPs in almost complete linkage disequilibrium. This breed-shared risk haplotype includes three genes (LHFPL5, SRPK1 and SLC26A8) and does not extend to the dog leukocyte antigen (DLA) class II gene cluster located in the vicinity. These three genes have not been identified as candidate genes for hypothyroid disease previously, but have functions that could potentially contribute to the development of the disease. Our results implicate the potential involvement of novel genes and pathways for the development of canine hypothyroidism, raising new possibilities for screening, breeding programmes and treatments in dogs. This study may also contribute to our understanding of the genetic etiology of human hypothyroid disease, which is one of the most common endocrine disorders in humans.

Socioeconomic Status and Childhood Leukemia Incidence in Switzerland.

Socioeconomic status (SES) discrepancies exist for child and adult cancer morbidity and are a major public health concern. In this Swiss population-based matched case-control study on the etiology of childhood leukemia, we selected the cases from the Swiss Childhood Cancer Registry diagnosed since 1991 and the controls randomly from census. We assigned eight controls per case from the 1990 and 2000 census and matched them by the year of birth and gender. SES information for both cases and controls was obtained from census records by probabilistic record linkage. We investigated the association of SES with childhood leukemia in Switzerland, and explored whether it varied with different definitions of socioeconomic status (parental education, living condition, area-based SES), time period, and age. In conditional logistic regression analyses of 565 leukemia cases and 4433 controls, we found no consistent evidence for an association between SES and childhood leukemia. The odds ratio comparing the highest with the lowest SES category ranged from 0.95 (95% CI: 0.71-1.26; P trend = 0.73) for paternal education to 1.37 (1.00-1.89; P trend = 0.064) for maternal education. No effect modification was found for time period and age at diagnosis. Based on this population-based study, which avoided participation and reporting bias, we assume the potential association of socioeconomic status and childhood leukemia if existing to be small. This study did not find evidence that socioeconomic status, of Switzerland or comparable countries, is a relevant risk factor or strong confounder in etiological investigations on childhood leukemia.

Parental demographic risk factors and occupational exposure to ionizing radiation for achondroplasia, thanatophoric and autosomal deletions in Texas, 1996--2002

Little is known about the etiology of Achondroplasia (AC), Thanatophoric Dwarfism (TD), and autosomal deletions (CD). These syndromes are due to fully penetrate genetic mutations, yet arise de novo, instead of being inherited. We examined the association between parental demographic characteristics and parental occupations with exposure to ionizing radiation and these birth defects. ^ We conducted a cross-sectional study and two case-control studies using a large database that was created by linking records from Texas Birth Defects Registry, Texas birth certificates and Texas fetal death certificates from 1996 to 2002. The first case-control study was matched on paternal age and examined 73 cases of AC and 43 cases of TD. The second case-control study was unmatched and examined 343 cases of autosomal deletion syndromes. ^ We used a job exposure matrix (JEM) to measure exposures to ionizing radiation in the workplace. This gives an estimate of the intensity and probability of exposure to ionizing radiation for each occupation and industry. ^ The prevalence rate of Achondroplasia, Thanatophoric Dwarfism and autosomal deletions was 0.36 per 10,000, 0.21 per 10,000, and 1.68 per 10,000 births respectively in Texas 1996–2002. ^ Older fathers had a strong increase in the risk of having offspring with AC or TD and a modest increase in the risk of CD. Fathers who were Black or Hispanic were less likely to have infants with AC or TD compared to Whites (adjusted POR=0.61; 95% CI 0.30, 1.26 and 0.44; 95% CI 0.27, 0.88, respectively). Black fathers and Hispanic mothers were also less likely to have infants with CD (adjusted POR=0.54; 95% CI 0.22, 1.35 and 0.62; 95% CI 0.39, 0.97). ^ After adjusting for other parental demographic factors, there was no significant relation between fathers exposure to ionizing radiation in the work place and AC or TD (adjusted OR=0.48; 95% CI 0.19, 1.25) and no significant relation between parental exposure to ionizing radiation in the work place and CD (adjusted OR=1.16; 95% CI 0.73, 1.85). ^ This is the first study to find an association between father's age and TD and CD and paternal race and AC or CD. Parental exposure to radiation for therapeutic or diagnostic indications was not measured, thus it can not be excluded as a cause of these birth defects. ^

Immune and genetic risk factors in glioma

Despite extensive research, the etiology of adult glioma remains largely unknown. We sought to further explore the role of immune and genetic factors in glioma etiology using data from the Harris County Brain Tumor Study and the first U.S. genome-wide association study of glioma. First, using a case-control study design, we examined the association between adult glioma risk and surrogates of the timing and frequency of common early childhood infections, birth order and sibship size, respectively. We found that each one-unit increase in birth order was associated with a 12% decreased risk of glioma development in adulthood (OR=0.88, 95% CI=0.81-0.96); however, sibship size was not associated with adult glioma risk (OR=0.96, 95% CI=0.91-1.02). Second, we used a multi-strategic approach to explore the relationships between glioma risk, history of asthma/allergies, and 23 functional SNPs in 11 inflammation genes. We found three inflammation gene SNPs to be significantly associated with glioma risk (COX2/PTGS2 rs20417 [OR=1.41]; IL10 rs1800896 [OR=1.57]; and IL13 rs20541 [OR=0.39]). Joint effects analysis of the risk-conferring alleles of these three SNPs revealed a trend of increasing risk with increasing number of adverse alleles among those without asthma/allergies (p<0.0001). Finally, we conducted a case-only study to explore pairwise SNP-SNP interactions in immune-related pathways among a population of 1304 non-Hispanic white glioma cases. After correction for multiple comparisons, we found 279 significant SNP-SNP interactions among polymorphisms of immune-related genes, many of which have not been previously examined. Our results, cumulatively, suggest an important role for immune and genetic factors in glioma etiology and provide several new hypotheses for future studies.^

Single nucleotide polymorphisms (SNPs) associated with TGF-beta pathway and their significance in systemic sclerosis - A multilevel analysis

Systemic sclerosis (SSc) or Scleroderma is a complex disease and its etiopathogenesis remains unelucidated. Fibrosis in multiple organs is a key feature of SSc and studies have shown that transforming growth factor-β (TGF-β) pathway has a crucial role in fibrotic responses. For a complex disease such as SSc, expression quantitative trait loci (eQTL) analysis is a powerful tool for identifying genetic variations that affect expression of genes involved in this disease. In this study, a multilevel model is described to perform a multivariate eQTL for identifying genetic variation (SNPs) specifically associated with the expression of three members of TGF-β pathway, CTGF, SPARC and COL3A1. The uniqueness of this model is that all three genes were included in one model, rather than one gene being examined at a time. A protein might contribute to multiple pathways and this approach allows the identification of important genetic variations linked to multiple genes belonging to the same pathway. In this study, 29 SNPs were identified and 16 of them located in known genes. Exploring the roles of these genes in TGF-β regulation will help elucidate the etiology of SSc, which will in turn help to better manage this complex disease. ^

DIETARY RELATIONSHIPS TO FATAL BREAST CANCER AMONG SEVENTH-DAY ADVENTISTS

Laboratory experiments in animals, correlational and migrant studies in humans suggest a role for diet in the etiology of breast cancer. Data gathered from individuals via case-control studies are less consistent. Seventh-day Adventist women experience lower mortality from breast cancer than comparable U.S. populations and this decrease is thought to be, at least in part, related to dietary practices (half are vegetarian). In 1960, 25,000 California Seventh-day Adventists completed a questionnaire which included a 21 item food frequency section. Cancer mortality in this population was monitored between 1960 and 1980 and the relationship of high fat food intake and fatal breast cancer was evaluated. Although established risk factors for breast cancer were observed in this population (e.g. age at menarche, age at first pregnancy, age at menopause and obesity) consumption of high fat foods were not observed to exert a strong influence on fatal breast cancer risk. Odds ratios (O.R.) for fatal breast cancer among non-vegetarians was 1.2. Increasing meat consumption bore little relation to risk; O.R. = 1.0, 1.2, 1.1 for consumption categories of none/occasional, 1-3 days/week and 4+ days/week respectively. Nor did the consumption of other high fat foods of animal origin (e.g. butter, cheese, milk, eggs) show any relationship to risk. These results remained unchanged after simultaneously controlling for the effect of other, potentially confounding variables (menstrual characteristics, obesity) via logistic regression analysis. ^

IDENTIFYING GENETIC VARIANTS AND CHARACTERIZING THEIR ROLE IN CLUBFOOT

Clubfoot is a common, complex birth defect affecting 4,000 newborns in the United States and 135,000 world-wide each year. The clubfoot deformity is characterized by inward and rigid downward displacement of one or both feet, along with persistent calf muscle hypoplasia. Despite strong evidence for a genetic liability, there is a limited understanding of the genetic and environmental factors contributing to the etiology of clubfoot. The studies described in this dissertation were performed to identify variants and/or genes associated with clubfoot. Genome-wide linkage scan performed on ten multiplex clubfoot families identified seven new chromosomal regions that provide new areas to search for clubfoot genes. Troponin C (TNNC2) the strongest candidate gene, located in 20q12-q13.11, is involved in muscle contraction. Exon sequencing of TNNC2 did not identify any novel coding variants. Interrogation of fifteen muscle contraction genes found strong associations with SNPs located in potential regulatory regions of TPM1 (rs4075583 and rs3805965), TPM2 (rs2025126 and rs2145925) and TNNC2 (rs383112 and rs437122). In previous studies, a strong association was found with rs3801776 located in the basal promoter of HOXA9, a gene also involved in muscle development and patterning. Altogether, this data suggests that SNPs located in potential regulatory regions of genes involved in muscle development and function could alter transcription factor binding leading to changes in gene expression. Functional analysis of 3801776/HOXA9, rs2025126/TPM2 and rs2145925/TPM2 showed altered protein binding, which significantly influenced promoter activity. Although the ancestral allele (G) of rs4075583/TPM1 creates a DNA-protein complex, it did not affect TPM1 promoter activity. However and importantly, in the context of a haplotype, rs4075583/G significantly decreased TPM1 promoter activity. These results suggest dysregulation of multiple skeletal muscle genes, TPM1, TPM2, TNNC2 and HOXA9, working in concert may contribute to clubfoot. However, specific allelic combinations involving these four regulatory SNPs did not confer a significantly higher risk for clubfoot. Other combinations of these variants are being evaluated. Moreover, these variants may interact with yet to be discovered variants in other genes to confer a higher clubfoot risk. Collectively, we show novel evidence for the role of skeletal muscle genes in clubfoot indicating that there are multiple genetic factors contributing to this complex birth defect.

The association between parity and selected structural birth defects

BACKGROUND: Parity is a risk factor in neonatal morbidity and mortality. This dissertation examined the association between first births and selected birth defects. The first aim was to assess the risk of 66 birth defects among first births and third or greater births. The second aim was to determine if maternal race, maternal age, infant sex or infant birth weight modify the association between first births and selected birth defects. METHODS: The Texas Birth Defects Registry provided data for 1999-2009. For the first aim, odds ratios were calculated for each birth defect. For the second aim, analysis was restricted to the ten birth defects significantly associated with first births. Stratified analyses were conducted and interaction terms were added to logistic regression models to assess whether differences in the odds ratios for the effect of first birth were statistically significant across strata. RESULTS: Findings for the first aim showed that first births had significantly increased odds of having an infant with 24 of the 66 birth defects. Third or greater births had significantly increased odds of having four of the 66 birth defects. For the second aim, a number of significant effect modifiers were observed. For patent ductus arteriosis, obstructive urinary defects and gastroschisis, the effect of first births was significantly modified by black or U.S.-born Hispanic mothers. The effect of first birth was also significantly modified among mothers ≥30 years for mitral valve insufficiency, atrial septal defect and congenital hip dislocation. The effect of first births was significantly modified among infants with low birth weight for hypospadias, congenital hip dislocation and gastroschisis. CONCLUSIONS: First births were associated with an elevated risk of 24 categories of birth defects. For some of the birth defects studied, the effect of first birth is modified by maternal age, maternal race and low birth weight. Knowledge of the increased risk for birth defects among women having their first birth allows physicians and midwives to provide better patient care and spur further research into the etiology of associated birth defects. This knowledge may bring about interventions prior to conception in populations most likely to conceive.^

Maternal exposure to hazardous air pollutants and oral clefts among offspring: Texas, 1999-2008

Birth defects are a leading cause of infant mortality in the United States. About one in 33 births in the United States is diagnosed with birth defects. Common birth defects include neural tube defects, Down syndrome and oral clefts. The present study focused on oral clefts. ^ Oral clefts refer to the malformation of lip, mouth or both. Birth prevalence of oral clefts in Texas is about 11 per 10,000 births. Etiologically, oral clefts have been classified into two groups, cleft lip with or without cleft palate (CL±P) and isolated cleft palate (CP). In spite of their high prevalence and clinical significance, the etiology of oral clefts in humans has not been well understood. Though a number of risk factors have been identified in epidemiological studies, most of them do not explain the majority of the cases. The need to identify novel risk factors associated with oral clefts provided the motivation for this study. The present study focused on maternal exposure to several hazardous air pollutants. A common subgroup of hazardous air pollutants is the volatile organic compounds found in petroleum derivatives. Four important hydrocarbons in this group are benzene, toluene, ethyl benzene and xylenes (BTEX). ^ The specific aim of this study was to evaluate the association between maternal exposure to environmental levels of BTEX and oral clefts among offspring in Texas for the period 1999-2008. ^ A case-control study design was used to assess if maternal exposure to BTEX increased the risk of oral clefts. The Texas Birth Defects Registry provided data on cases of non-syndromic oral clefts delivered in Texas during the period 1999-2008. Census tract level maternal exposure to BTEX concentrations were obtained from the Hazardous Air Pollutant Exposure Model (HAPEM) developed by the U.S. Environmental Protection Agency. Unconditional logistic regression was used to assess the relationship between maternal exposure to BTEX levels and risk of oral clefts in offspring. ^ In the selected population, mothers who had high estimated exposure to any of the BTEX compounds were not more likely to deliver an offspring with oral clefts. Future research efforts will focus on additional birth defects and thorough assessment of additional potential confounders.^