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ObjectiveTo determine whether mitochondrial haplogroups function as disease-modifiers or as susceptibility factors in preeclampsia using a traditional haplogroup association model.MethodsThis retrospective study haplotyped 235 control and 78 preeclamptic pregnancies from Denmark using either real-time PCR or Sanger sequencing depending on the rarity of the haplogroup.ResultsNo significant association between haplogroups and the risk of preeclampsia was found, nor was any role for haplogroups in disease severity uncovered.ConclusionMitochondrial haplogroups are not associated with preeclampsia or the severity of preeclampsia in the Danish population. However, this study cannot exclude a role for less common mtDNA variation. Models that can examine these should be applied in preeclamptic patients.Graphical Abstract

Background and purposeCongenital clubfoot is a common musculoskeletal anomaly, with a suspected multifactorial etiopathogenesis. To begin examining its etiopathogenesis in a nationwide Danish population, liveborn infants with clubfoot were ascertained to classify co-occurring congenital anomalies, estimate annual prevalence, and compare occurrence with maternal smoking rates, a commonly reported risk factor.Patients and methodsThis case-cohort study used data from the Danish National Patient Register and Danish Civil Registration System to identify 1,315,282 liveborn infants delivered during 1994–2021 in Denmark to Danish parents. Among these, 2,358 infants (65.1% male) were ascertained with clubfoot and classified as syndromic (co-occurring chromosomal, genetic, or teratogenic syndromes) and nonsyndromic (isolated or co-occurring multiple congenital anomalies [MCA]). Annual prevalence estimates and corresponding 95% confidence intervals (CIs) for children with nonsyndromic clubfoot were estimated using Poisson regression and compared with population-based, maternal annual smoking rates obtained from publicly available resources.ResultsInfants most often presented with nonsyndromic clubfoot (isolated=84.6%; MCA=10.9%); limb and heart anomalies were the most frequently identified MCAs. Prevalence (per 1,000 liveborn infants) was 1.52 (CI 1.45 – 1.58) for isolated and 0.19 (CI 0.17 – 0.22) for MCA clubfoot. Prevalence estimates for both isolated and MCA clubfoot remained stable during the study period, despite marked decreases in population-based maternal smoking rates.InterpretationFrom 1994-2021, prevalence of nonsyndromic clubfoot in Denmark was reasonably stable. Reduction in population-level maternal smoking rates did not seem to impact prevalence estimates. Ascertainment and characterization of this cohort provides a population-based clinical and biological resource for etiopathogenic investigations.

Numerous circulating microRNAs (miRNAs) have been detected in maternal blood. Initial studies in third trimester demonstrated differential miRNA expression profiles between uncomplicated pregnancies and pregnancies complicated by pre-eclampsia (PE). Recently, studies in first trimester have shown similar differential profiles, however, these studies were often under-powered. We conducted a nested case-control study, in which serum samples, taken between 10-14 weeks gestation, were obtained from 413 singleton pregnant women, 126 of which later developed PE. Total RNAs were purified and a selection of 46 miRNAs plus two miRNA controls were quantitated by real time quantitative PCR. Seven of the miRNAs, hsa-miR-181b-5p, -323a-3p, -518b, -363-3p, -20a-5p, -29a-3p, and -142-3p, could differentiate between uncomplicated pregnancies and pregnancies which develop PE, but only a single miRNA, hsa-miR-363-3p, could differentiate between mild and severe PE. A combination of all seven differentiating miRNAs was the best at discriminating between PE and uncomplicated pregnancies (AUC= 0.879). In conclusion, first trimester maternal serum miRNA expression profile could differentiate between uncomplicated pregnancies and pregnancies complicated by PE. These circulating miRNA markers have the potential to improve risk assessment of PE in the first trimester, weeks before the onset of symptoms.

BackgroundMaternal obesity has been positively associated with increased risk of congenital heart defects in the offspring. However, none of the large studies have included the considerable proportion of congenital heart defects that are identified due to improvements in prenatal diagnostics and terminated in pregnancy. The mechanism behind the association is poorly understood, and a relation to interpregnancy weight changes is to be investigated.ObjectivesTo evaluate the association between maternal obesity and congenital heart defects in the offspring when including all pregnancies and to investigate if interpregnancy weight change between the first and second pregnancy influences risk of fetal congenital heart defects.Study DesignA nationwide cohort study of all singleton pregnancies in Denmark from 2008 to 2018. All data on maternal and offspring characteristics were retrieved from the Danish Fetal Medicine Database. The database included data on postnatal diagnoses of congenital heart defects in live births and prenatal diagnoses of congenital heart defects from ultrasound examinations during pregnancy resulting in live birth, stillbirth, spontaneous abortion after gestational week 12 or termination of pregnancies after gestational week 12. As this cohort encompassed all pregnancies over a 10-year period, it was possible for women to experience multiple pregnancies. Congenital heart defects and severe congenital heart defects were grouped according to European Surveillance of Congenital Anomalies’ definitions. Children or fetuses with chromosomal aberrations were excluded. Relative risks were calculated using log-linear Poisson models for congenital heart defects overall, severe congenital heart defects and for five of the most prevalent subtypes of congenital heart defects.ResultsOf the 547 178 pregnancies included in the cohort, 5 498 had congenital heart defects (1.0%). Risk of congenital heart defects became gradually higher with higher maternal BMI; for BMI 30-34.9 kg/m2, adjusted relative risk = 1.23 (95% confidence interval 1.12-1.36), for BMI 35-39.9 kg/m2, adjusted relative risk = 1.26 (95% confidence interval 1.09-1.46) and for BMI ≥ 40 kg/m2, adjusted relative risk = 1.81 (95% confidence interval 1.50-2.15). Data was adjusted for maternal age, smoking status and year of estimated due date. The same pattern was seen for the subgroup of severe congenital heart defects. Among the atrioventricular septal defects (n = 245), a particularly strong association with maternal BMI ≥ 40 kg/m2was seen, adjusted relative risk = 4.19 (95% confidence interval 2.13-7.42). 107 627 women were identified with their first and second pregnancies in the cohort. Interpregnancy BMI change was positively, albeit not statistically significant, associated with risk of congenital heart defects in the second pregnancy when adjusting for maternal age and BMI, with an adjusted relative risk = 1.27 (95% confidence interval 0.96-1.64) among persons with a BMI increase of ≥ 4 kg/m2.ConclusionsWhen including both pre- and postnatally diagnosed congenital heart defects, this study showed a positive dose-response association between maternal BMI and risk of congenital heart defects in the offspring. However, only a non-significant trend was seen between interpregnancy BMI changes and risk of congenital heart defects in the second pregnancies.CondensationTweetable statement: The risk of fetal congenital heart defect is associated with high maternal BMI, and it may also be affected by a substantial weight gain between pregnancies.AJOG at a GlanceA. Why was this study conducted?None of the large cohort studies on the association between maternal BMI and congenital heart defects (CHDs) have included the proportion of CHDs that are identified in terminated pregnancies.No previous studies have investigated the association between interpregnancy weight changes and CHDs.B. What are the key findings?Risk of CHDs became gradually higher with higher maternal BMI when including live births, stillbirths, abortions and terminated pregnancies.A trend was found between interpregnancy weight gain and risk of CHDs.C. What does this study add to what is already known?The association between high maternal BMI and risk of CHDs are not caused by a lower detection rate of fetal CHDs in pregnant women with obesity.

Using provisional or opportunistic data, three nationwide studies (The Netherlands, the USA and Denmark) have identified a reduction in preterm or extremely preterm births during periods of COVID-19 restrictions. However, none of the studies accounted for perinatal deaths. To determine whether the reduction in extremely preterm births, observed in Denmark during the COVID-19 lockdown, could be the result of an increase in perinatal deaths and to assess the impact of extended COVID-19 restrictions, we performed a nationwide Danish register-based prevalence proportion study. We examined all singleton pregnancies delivered in Denmark during the COVID-19 strict lockdown calendar periods (March 12–April 14, 2015-2020, N = 31,164 births) and the extended calendar periods of COVID-19 restrictions (February 27–September 30, 2015-2020, N = 214,862 births). The extremely preterm birth rate was reduced (OR 0.27, 95% CI 0.07 to 0.86) during the strict lockdown period in 2020, while perinatal mortality was not significantly different. During the extended period of restrictions in 2020, the extremely preterm birth rate was marginally reduced, and a significant reduction in the stillbirth rate (OR 0.69, 0.50 to 0.95) was observed. No changes in early neonatal mortality rates were found.Conclusion: Stillbirth and extremely preterm birth rates were reduced in Denmark during the period of COVID-19 restrictions and lockdown, respectively, suggesting that aspects of these containment and control measures confer an element of protection. The present observational study does not allow for causal inference; however, the results support the design of studies to ascertain whether behavioural or social changes for pregnant women may improve pregnancy outcomes. What is Known:• The aetiologies of preterm birth and stillbirth are multifaceted and linked to a wide range of socio-demographic, medical, obstetric, foetal, psychosocial and environmental factors.• The COVID-19 lockdown saw a reduction in extremely preterm births in Denmark and other high-income countries. An urgent question is whether this reduction can be explained by increased perinatal mortality. What is New:• The reduction in extremely preterm births during the Danish COVID-19 lockdown was not a consequence of increased perinatal mortality, which remained unchanged during this period.• The stillbirth rate was reduced throughout the extended period of COVID-19 restrictions.

The serum adiponectin-leptin ratio (A/L ratio) is a surrogate marker of insulin sensitivity. Pre-eclampsia (PE) is associated with maternal metabolic syndrome and occasionally impaired fetal growth. We assessed whether the A/L ratio in first-trimester maternal serum was associated with PE and/or birth weight. Adiponectin and leptin were quantitated in first-trimester blood samples (gestational week 10+3-13+6) from 126 women who later developed PE with proteinuria, (98 mild PE; 21 severe PE; 7 HELLP syndrome), and 297 controls, recruited from the Copenhagen First-Trimester Screening Study. The A/L ratio was reduced in PE pregnancies, median 0.17 (IQR: 0.12-0.27) compared to controls, median 0.32 (IQR: 0.19-0.62), (pβ= −0.144, 95% CI = −9.944 to −0.093), independent of maternal BMI. After correction for maternal BMI, leptin was significantly associated with relative birth weight, (β= 0.197, 95 % CI = 2.361 to 14.353), while adiponectin was not significantly associated. Our findings suggest that an impairment of the A/L ratio (as seen in metabolic syndrome) in first-trimester is characteristic of PE, while aberrant fetal growth in PE is not dependent on insulin sensitivity but rather on leptin associated pathways.

In Denmark, a nationwide COVID-19 lockdown was implemented on March 12, 2020 and eased on April 14, 2020. The COVID-19 lockdown featured reduced prevalence of extremely preterm or extremely low birth weight births. Here we explore the impact of this COVID-19 lockdown on term birth weights in Denmark.We conducted a nationwide register-based cohort study on 27,870 live singleton infants, born at term (weeks 37-41), between March 12 and April 14, 2015-2020, using data from the Danish Neonatal Screening Biobank. Primary outcomes, corrected for confounders, were birth weight, small-for-gestational-age (SGA), and large-for-gestational-age (LGA), comparing the COVID-19 lockdown to the previous five years. Data were analysed using linear regression to assess associations with birth weight. Multinomial logistic regression was used to assess associations with size-for-gestational-age (xGA) categories.Adjusted mean birth weight was significantly increased by 16.9 g (95% CI = 4.1 – 31.3) during the lockdown period. A dip in mean birth weight was found in gestational weeks 37 and 38 balanced by an increase in weeks 40 and 41. The 2020 lockdown period was associated with an increased LGA prevalence (aOR 1.13, 95% CI = 1.05 - 1.21). No significant changes in proportions of xGA groups were found between 2015 and 2019.ConclusionThe nationwide COVID-19 lockdown resulted in a small but significant increase in birth weight and proportion of LGA infants, driven by an increase in birth weight in gestational weeks 40 and 41.What is knownThe COVID-19 lockdown periods have had profound, albeit variable, effects on pregnant women in various countries.Extremely preterm and stillbirth rates were reduced during COVID-19 lockdown/mitigation periods in DenmarkWhat is newDuring COVID-19 lockdown a moderate, but statistically significant, increase in birth weight was seen.The proportion of large-for-gestational age pregnancies was statistically significantly elevated in the lockdown period.

Denmark’s response to the COVID-19 pandemic was to issue guidelines on containment, isolation, and surveillance. Subsequently, Denmark entered a period with variable mitigation measures including closures of schools and workplaces, travel restrictions, and restrictions on public gatherings. A Danish study covering the lockdown period (March 12 – April 14, 2020) as well as a period of mitigation measures (February 27 – September 30, 2020) showed that the reduction in extremely preterm births was not associated with an increase in singleton stillbirth rate. Subsequent studies examining the impacts of COVID-19 mitigation measures on preterm birth have combined singleton with multifetal pregnancies. However, multifetal pregnancies have a different set of risk factors for preterm birth, as well as, increased risk of adverse outcomes, and a shorter expected pregnancy length. We assessed the impact of the Danish lockdown period, or the mitigation period on multifetal births and found no significant difference in the proportion of preterm twin births among all twin births, either within gestational age groups or in total, for either period.

Hydrocephalus is a genetically and phenotypically heterogenous condition with complex etiology. Ciliary dysfunction has been shown to play a role, either through interference with signaling functions in primary cilia, cerebrospinal fluid flow by motile cilia, or both. Ciliary function is highly energy-dependent, consequently, variation in mitochondrial OXPHOS function might be a susceptibility factor for hydrocephalus. Furthermore, familial hydrocephalus exhibits preferential maternal inheritance. Mitochondrial DNA (mtDNA) haplogroups, have been associated with different characteristics of OXPHOS function as well as susceptibility to autism spectrum disorders, a frequent co-morbidity of hydrocephalus. This nested case-cohort study, a substudy of the iPSYCH study, used mtDNA data from 191 hydrocephalus cases and 24,831 population controls and found no association between hydrocephalus and any mtDNA haplogroup. Likewise, the distribution of European macro-haplogroups, HV, JT, and UK, did not differ between 172 hydrocephalus cases and 21,850 population controls. Thus, mtDNA haplogroups are not susceptibility factors for hydrocephalus.

IntroductionPre-eclampsia (PE) is a leading cause of perinatal morbidity and mortality worldwide. Low-dose aspirin can prevent PE in high risk pregnancies if started early. However, despite intense research into the area, first-trimester screening for PE risk is still not a routine part of pregnancy care. Several studies have described the application of artificial intelligence (AI) and machine learning (ML) in risk prediction of PE and its subtypes. A systematic review of available literature is necessary to catalogue the current applications of AI/ML methods in early pregnancy screening for PE, in order to better inform the development of clinically relevant risk prediction algorithms which will enable timely intervention and the development of new treatment strategies. The aim of this systematic review is to identify and assess studies regarding the application of AI/ML methods in first-trimester screening for PE.MethodsA systematic review of peer-reviewed as well as the grey literature cohort or case-control studies will be conducted. Relevant information will be accessed from the following databases; PubMed, Google Scholar, Web of Science, Arxiv, BioRxiv, and MedRxiv. The studies will be evaluated by two reviewers in a parallel, blind assessment of the literature, a third reviewer will assess any studies in which the first two reviewers did not agree. The free online tool Rayyan, will be used in this literature assessment stage. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 checklist will be used to guide the review process and the methods of the studies will be assessed using the Newcastle-Ottawa scale. Narrative synthesis will be conducted for all included studies. Meta-analysis will also be conducted where data quality and availability allow.Ethics and disseminationThe review will not require ethical approval and the findings will be published in a peer-reviewed journal using the PRISMA guidelines.

ObjectivesPre-eclampsia (PE) is an important cause of perinatal morbidity and mortality. Despite an elusive pathophysiology, PE has been associated with changes in maternal serum concentrations of adipokines in early pregnancy. We hypothesized, that symbolic regression might identify interactions between adipokines and PE, which may have eluded regression and Bayesian models.MethodsIn this nested case-control sub-study, of the Copenhagen First Trimester Screening Study, data regarding maternal weight and serum concentrations of PAPP-A, leptin (Lp), soluble leptin receptor (sLR), adiponectin, and resistin (Re) was available from 423 first trimester pregnancies (gestational week 10+3– 13+6), 126 of which developed PE. Symbolic regression with QLattice/Feyn 2.1 was used to identify models comprising two-interactions between up-to three markers.ResultsThe optimal mathematical model exhibited a non-linear relation between Re and the combined effect of sLR and Lp. The model was dependent, in a Gaussian way, on the level of Re. The receiver operating characteristic (ROC) curve of the model viz. identification of PE cases in first trimester had an AUC of 0.81 and a modelled DR of 40 % for a FPR of 4 %. Symbolic regression outperformed logistic regression of the same parameters with a ROC with AUC = 0.77, and a DR of 7 % for a 3 % FPR.ConclusionsSymbolic regression identified non-linear interactions between Lp, sLR and Re concentrations in first trimester pregnancy serum of pregnancies which later developed PE. Non-linear interactions suggest new pathophysiological pathways and may help in designing more efficient screening protocols for PE.

Background and aims: Pediatric obesity associates with both low-grade inflammation and cardiometabolic risk on the population level. Yet on an individual patient level, overweight/obesity does not always equal increased cardiometabolic risk. In this study, we examine whether low-grade inflammation associates with cardiometabolic risk in Danish children, independent of degree of adiposity. We further assess the value of integrating multiple inflammation markers to identify children with very-high cardiometabolic risk profiles.Method and results: We studied 2192 children and adolescents aged 6-18 years from an obesity clinic cohort and a population-based cohort, in a cross-sectional study design. Anthropometry, blood pressure, pubertal stage and body composition by dual-energy X-ray absorptiometry were assessed, and biomarkers including fasting serum high sensitivity C-reactive protein (hsCRP), white blood cells (WBC), resistin, lipid profile and glucose metabolism were measured. Adjusted correlation analysis and odds ratios were calculated. We found that, independent of degree of adiposity, having high-normal inflammation marker concentrations associated with increased cardiometabolic risk: for girls, hsCRP >0.57-9.98 mg/L (mid/upper tertile) associated with similar to 2-fold higher odds of dyslipidemia and hepatic steatosis (vs. lower tertile). For both sexes, WBC >7.0-12.4 10(9) /L (upper tertile) associated with 2.5-fold higher odds of insulin resistance. Lastly, children with multiple inflammation markers in the high-normal range exhibited the most severe cardiometabolic risk profile.Conclusion: Low-grade inflammation associates with cardiometabolic risk in children independent of degree of adiposity. The associations vary with sex and inflammation marker measured. Finally, integrating multiple low-grade inflammation markers identifies a very-high-risk subgroup of children with overweight/obesity and may have clinical value. (C) 2020 The Italian Diabetes Society, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

Mitochondrial DNA (mtDNA) haplogroup (hg) H has been reported as a susceptibility factor for hypertrophic cardiomyopathy (HCM). This was established in genetic association studies, however, the SNP or SNP’s that are associated with the increased risk have not been identified. Hg H is the most frequent European mtDNA hg with greater than 80 subhaplogroups (subhgs) each defined by specific SNPs. We tested the hypothesis that the distribution of H subhgs might differ between HCM patients and controls. The subhg H distribution in 55 HCM index cases was compared to that of two Danish mtDNA hg H control groups (n = 170 and n = 908, respectively). In the HCM group, H and 12 different H subhgs were found. All these, except subhgs H73, were also found in both control groups. The HCM group was also characterized by a higher proportion of H3 compared to H2. In the HCM group the H3/H2 proportion was 1.7, whereas it was 0.45 and 0.54 in the control groups. This tendency was replicated in an independent group of Hg H HCM index cases (n = 39) from Queensland, Australia, where the H3/H2 ratio was 1.5. In conclusion, the H subhgs distribution differs between HCM cases and controls, but the difference is subtle, and the understanding of the pathogenic significance is hampered by the lack of functional studies on the subhgs of H.

Introduction Pre-eclampsia (PE) is a leading cause of perinatal morbidity and mortality worldwide. Low-dose aspirin can prevent PE in high risk pregnancies if started early. However, despite intense research into the area, early pregnancy screening for PE risk is still not a routine part of pregnancy care. Several studies have described the application of artificial intelligence (AI) and machine learning (ML) in risk prediction of PE and its subtypes. A systematic review of available literature is necessary to catalogue the current applications of AI/ML methods in early pregnancy screening for PE, in order to better inform the development of clinically relevant risk prediction algorithms which will enable timely intervention and the development of new treatment strategies. The aim of this systematic review is to identify and assess studies regarding the application of AI/ML methods in early pregnancy screening for PE. Methods A systematic review of peer-reviewed as well as the pre-published cohort, case-control, or cross-sectional studies will be conducted. Relevant information will be accessed from the following databases; PubMed, Google Scholar, Scopus, Embase, Web of Science, Cochrane Library, Arxiv, BioRxiv, and MedRxiv. The studies will be evaluated by two reviewers in a parallel, blind assessment of the literature, a third reviewer will assess any studies in which the first two reviewers did not agree. The free online tool Rayyan, will be used in this literature assessment stage. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 checklist will be used to guide the review process and the methods of the studies will be assessed using the Newcastle-Ottawa scale. Narrative synthesis will be conducted for all included studies. Meta-analysis will also be conducted where data quality and availability allow. Ethics and dissemination The review will not require ethical approval and the findings will be published in a peer-reviewed journal using the PRISMA guidelines. Trial registration Trial registration: The protocol for this systematic review has been registered in PROSPERO [CRD42022345786]. https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42022345786.

ImportanceUsing provisional or opportunistic data, three nationwide studies (The Netherlands, USA, and Denmark) have identified a reduction in preterm or extremely preterm births during periods of COVID-19 restrictions. However, these findings have been challenged as none of the studies accounted for perinatal deaths.ObjectiveTo determine whether the reduction in extremely preterm births, observed in Denmark during the COVID-19 lockdown, could be a result of an increase in number of perinatal deaths; and to assess the impact of extended COVID-19 restrictions on the prevalence of preterm birth and perinatal death.DesignThe study is a nationwide Danish register-based prevalence proportion study using detailed data to capture all births (induced abortions were excluded) throughout Denmark. We assessed the prevalence of stillbirth, preterm birth, and early neonatal death.SettingPopulation-based studyParticipantsAll singleton pregnancies delivered in Denmark, between February 27, and September 30, 2015-2020. COVID-19 lockdown was studied in 31,164 births and the extended period of COVID-19 restrictions in 214,862 births.ExposureCOVID-19 restrictions broadly (February 27, – September 30, 2020) or COVID-19 lockdown specifically (March 12, – April 14, 2020).Main OutcomePrevalence of preterm births, stillbirths and early neonatal deaths across the periods under study.ResultsThe extremely preterm birth rate was reduced (OR 0.27, 95% CI 0.07 to 0.86) during the strict lockdown period, while perinatal mortality was not significantly different. During the extended period of restrictions, the extremely preterm birth rate was marginally reduced, and a significant reduction in the stillbirth rate (OR 0.69, 0.50 to 0.95) was observed. No changes in early neonatal mortality rates were found.Conclusion and RelevanceStillbirth and extremely preterm birth rates were reduced in Denmark during the period of COVID-19 restrictions and lockdown, respectively, suggesting that aspects of these containment and control measures confer an element of protection. The present observational study does not allow for causal inference; however, the results support design of studies to ascertain whether behavioural or social changes for pregnant women may improve pregnancy outcomes.FundingNoneKey pointsQuestionCan changes in stillbirth and early neonatal mortality rates during the COVID-19 lockdown explain the reduction in extremely preterm birth seen in Denmark?FindingsIn this nationwide register-based study that included data pertaining to 245,999 pregnancies, the statistically significant reduction in extremely preterm birth during the COVID-19 lockdown was confirmed. The stillbirth and early neonatal mortality rates were unchanged during lockdown, while the stillbirth rate was reduced over the extended period of COVID-19 restrictions.MeaningThe reduction in extremely preterm births seen during the COVID-19 lockdown in Denmark is not a result of increased stillbirth or early neonatal death.

Background The association between autism spectrum disorder and hydrocephalus is not well understood, despite demonstrated links between autism spectrum disorder and cerebrospinal fluid abnormalities. Based on the hypothesis that autism spectrum disorder and hydrocephalus may, at least in some cases, be two manifestations of a shared congenital brain pathology, we investigated the potential association between autism spectrum disorder and hydrocephalus in a large Danish population-based cohort. Methods Patients and controls were obtained from the Lundbeck Foundation Initiative for Integrative Psychiatric Research iPSYCH2012 case-cohort, which includes all patients with selected psychiatric disorders born in Denmark 1981–2005 along with randomly selected population controls (end of follow-up, December 31, 2016). The associations between individual psychiatric disorders and hydrocephalus were estimated using binary logistic regression with adjustment for age and sex. Results The cohort consisted of 86,571 individuals, of which 14,654 were diagnosed with autism spectrum disorder, 28,606 were population controls, and the remaining were diagnosed with other psychiatric disorders. We identified 201 hydrocephalus cases; 68 among autism spectrum disorder patients and 40 among controls (OR 3.77, 95% CI 2.48–5.78), which corresponds to an absolute risk of 0.46 % (i.e. approximately one in 217 children with autism spectrum disorder had co-occurring hydrocephalus). The autism spectrum disorder-hydrocephalus association was significant over the entire subgroup spectrum of autism spectrum disorder. Conclusions Given the considerable risk of hydrocephalus among patients with autism spectrum disorder, we suggest that patients with autism spectrum disorder should be evaluated for co-occurring hydrocephalus on a routine basis as timely neurosurgical intervention is important. Likewise, attention must be paid to traits of autism spectrum disorder in children with hydrocephalus. The results of this study call for future investigations on a potential shared aetiology between hydrocephalus and autism spectrum disorder, including the role abnormal CSF dynamics in the pathogenesis of autism spectrum disorder.

Background The association between autism spectrum disorder and hydrocephalus is not well understood, despite demonstrated links between autism spectrum disorder and cerebrospinal fluid abnormalities. Based on the hypothesis that autism spectrum disorder and hydrocephalus may, at least in some cases, be two manifestations of a shared congenital brain pathology, we investigated the potential association between autism spectrum disorder and hydrocephalus in a large Danish population-based cohort, and whether the polygenic risk scores for autism spectrum disorder changed as a function of the presence of hydrocephalus. Methods Patients and controls were obtained from the Lundbeck Foundation Initiative for Integrative Psychiatric Research iPSYCH2012 case-cohort, which includes all patients with selected psychiatric disorders born in Denmark 1981–2005 along with randomly selected population controls (end of follow-up, December 31, 2016). The associations between individual psychiatric disorders and hydrocephalus were estimated using binary logistic regression with adjustment for age and sex. Polygenic risk scores for autism spectrum disorder were used to compare the genetic architecture of autism spectrum disorder as a function of the presence of hydrocephalus. Results The cohort consisted of 86,571 individuals, of which 14,654 were diagnosed with autism spectrum disorder, 28,606 were population controls, and the remaining were diagnosed with other psychiatric disorders. We identified 201 hydrocephalus cases; 68 among autism spectrum disorder patients and 40 among controls (OR 3.77, 95% CI 2.48–5.78). The autism spectrum disorder-hydrocephalus association was significant over the entire subgroup spectrum of autism spectrum disorder. The presence of hydrocephalus did not markedly influence the polygenic risk scores in patients with autism spectrum disorder, which may indicate overlapping genetic architectures or other common aetiology. Conclusions Given the very strong association, we suggest that patients with autism spectrum disorder should be evaluated for co-occurring hydrocephalus on a routine basis as timely neurosurgical intervention is important. Further clarification of the genetic aetiology of both diseases, may help in elucidating shared genetic pathways between autism spectrum disorder and hydrocephalus, and it may elucidate the role of abnormal CSF dynamics in the pathogenesis of autism spectrum disorders.

BackgroundCongenital heart defects (CHDs) are the most common congenital malformations. The aetiology of CHDs is complex. Large cohort studies and systematic reviews and meta-analyses based on these have reported an association between higher risk of CHDs in the offspring and individual maternal metabolic disorders such as obesity, diabetes, hypertension, and preeclampsia, all conditions that can be related to insulin resistance or hyperglycaemia. However, the clinical reality is that these conditions often occur simultaneously. The aim of this review is, in consequence, both to evaluate the existing evidence on the association between maternal metabolic disorders, defined as obesity, diabetes, hypertension, preeclampsia, dyslipidaemia and CHDs in the offspring, as well as the significance of combinations, such as metabolic syndrome, as risk factors.MethodsA systematic literature search of papers published between January 1, 1990 and January 14, 2021 was conducted using PubMed and Embase. Studies were eligible if they were published in English and were case-control or cohort studies. The exposures of interest were maternal overweight or obesity, hypertension, preeclampsia, diabetes, dyslipidaemia, and/or metabolic syndrome, and the outcome of interest was CHDs in the offspring. Furthermore, the studies were included according to a quality assessment score.ResultsOf the 2,250 identified studies, 32 qualified for inclusion. All but one study investigated only the individual metabolic disorders. Some disorders (obesity, gestational diabetes, and hypertension) increased risk of CHDs marginally whereas pre-gestational diabetes and early-onset preeclampsia were strongly associated with CHDs, without consistent differences between CHD subtypes. A single study suggested a possible additive effect of maternal obesity and gestational diabetes.ConclusionsFuture studies of the role of aberrations of the glucose-insulin homeostasis in the common aetiology and mechanisms of metabolic disorders, present during pregnancy, and their association, both as single conditions and–particularly–in combination, with CHDs are needed.

Objectives To explore the impact of COVID-19 lockdown on premature birth rates in Denmark Design Nationwide register-based prevalence proportion study. Participants 31,180 live singleton infants born in Denmark between March 12, and April 14, from 2015 to 2020 Main outcome measures The Main outcome measure was the odds ratio of premature birth, per preterm category, during the lockdown period compared with the calendar match period in the five previous years. Results A total of 31 180 newborns were included in the study period, of these 58 were born extremely premature (gestational age below 28 weeks). The distribution of gestational ages was significantly different (p = 0.004) during the lockdown period compared to the previous five years. The extremely premature birth rate during the lockdown was significantly lower than the corresponding mean rate for the same dates in the previous years (odds ratio 0.09 [95 % CI 0.01 - 0.04], p < 0.001). No significant difference between the lockdown and previous years was found for other gestational age categories. Conclusions The birth rate of extremely premature infants decreased significantly (~90 % reduction) during the Danish nationwide lockdown from a stable rate in the preceding five years. The reasons for this decrease are unclear. Identification of possible causal mechanisms might stimulate changes in clinical practice. Ideally, some cases of extreme prematurity are preventable which may decrease infant morbidity and mortality.

Objective Preeclampsia (PE) is a serious complication of pregnancy, the pathogenesis of which is largely unknown. We hypothesize that adipocytokines may play a role in the pathogenesis of PE, particularly in obese women, and evaluate leptin and adiponectin as potential first trimester markers for predicting PE. Study design A cohort of 2503 pregnancies, containing 93 PE pregnancies, was divided into women with normal weight, moderate, or severe obesity. All pregnancies had serum adiponectin and leptin measured in first trimester. Logistic regression was used to model PE with maternal characteristics and concentrations of the biomarkers. Results In obese women a lower concentration of adiponectin was found in PE pregnancies; the concentration was lowest in the severely obese (p = 0.005). No association was found in normal weight women (p = 0.72). Leptin concentration had no association with PE in normal weight and moderately obese (p = 0.175–0.072), however in women with severe obesity a lower level of leptin was found (p = 0.049). The AUC was 0.73 for the ROC curve of combined maternal characteristics and adiponectin. Using adiponectin in women with moderate to severe obesity the sensitivity was 72.9% and the specificity was 49%. Conclusions In severely obese women, PE is associated with low serum adiponectin and leptin concentrations in first trimester. This indicates that the inability of adipokine regulation to adapt to severe obesity may play a role in the pathogenesis of PE. Adipocytokines may contribute in identification of risk pregnancies among severe obese.

The aim of this study was to determine whether COVID-19 restrictions had an impact on Chlamydia trachomatis infections compared with 2018 and 2019. A retrospective nationwide observational study was performed using monthly incidences of laboratory-confirmed chlamydia cases and number of tests, obtained from Danish national surveillance data. Testing rates and positivity rates were compared using Poisson and logistic regression. The first Danish COVID-19 lockdown (12 March to 14 April 2020) resulted in a reduction in the number of chlamydia tests performed (rate ratio 0.72, 95% confidence interval 0.71–0.73) and a consequent reduction in the number of laboratory-identified cases (66.5 vs 88.3 per 100,000 population during the same period in 2018 to 2019). This period was followed by a return of testing and test positivity close to the level seen in 2018 to 2019. The second Danish COVID-19 lockdown (17 December to 31 March 2021) resulted in crude incidence rates of laboratory-confirmed chlamydia infection that were similar to the crude incidence rates seen during same period in 2018 to 2019. In conclusion, the Danish COVID-19 restrictions have had negligible effects on laboratory-confirmed C. trachomatis transmission.

BACKGROUND: The genetic risk factors of schizophrenia (SCZ), a severe psychiatric disorder, are not yet fully understood. Multiple lines of evidence suggest mitochondrial dysfunction may play a role in SCZ, but comprehensive association studies are lacking. We hypothesized that variants in nuclear-encoded mitochondrial genes influence susceptibility to SCZ. METHODS: We conducted gene-based and gene-set analyses using summary association results from the Schizophrenia Psychiatric Genomics Consortium GWAS (PGC-SCZ2) comprising 35,476 cases and 46,839 controls. We applied the MAGMA method to three sets of nuclear-encoded mitochondrial genes: oxidative phosphorylation genes, other nuclear-encoded mitochondrial genes, and genes involved in nucleus-mitochondria crosstalk. Furthermore, we conducted a replication study using the iPSYCH SCZ sample of 2,290 cases and 21,621 controls. RESULTS: In the PGC-SCZ2 sample, 1,186 mitochondrial genes were analyzed among which 159 had p-values

Introduction Both women with polycystic ovary syndrome (PCOS) and women with twin pregnancies have increased risk of adverse pregnancy outcome. The aim of this study was to investigate the impact of PCOS and maternal androgen levels on the outcome of dichorionic twin pregnancy. Material and methods A retrospective study of 360 women with dichorionic twin pregnancies: 72 women with PCOS from a fertility clinic (years 1997–2010) and 288 women without PCOS from a hospital cohort (years 2005–2007). The obstetrical outcome was extracted from Danish National registers and supplemented by patient file data. In all, 65% of the PCOS group had a registered prepregnancy androgen level and these were stratified into normoandrogenic and hyperandrogenic women. The groups were compared by multiple regression analysis adjusting for mode of conception and prepregnancy body mass index. Results We found no overall impact of PCOS on the pregnancy outcome; the risks of preeclampsia, gestational diabetes and preterm delivery were comparable within the groups. However, five deliveries in the PCOS group compared with two in the control group occurred before gestational week 28. No difference in the obstetrical outcome between hyperandrogenic and normoandrogenic women was found. The body mass index in the PCOS population was lower than in the non-PCOS, possibly reflecting a higher socioeconomic status and a healthier lifestyle, which may underestimate the impact of a PCOS diagnosis. Conclusion Neither PCOS nor maternal androgen levels confer additional risks to the outcome of dichorionic twin pregnancies of normal weight women.

Background Syncope is a common condition in the general population causing frequent hospitalisation and visits to the emergency department. Family aggregation and twin studies have previously indicated that syncope and collapse has a heritable component. Purpose We investigated whether common genetic variants predispose to syncope and collapse. Methods We used genome-wide association data on syncope and collapse for 408,961 individuals with European ancestry from the UK Biobank study. In a replication study, the Integrative Psychiatric Research Consortium (iPSYCH) cohort (n=86,189) was used to investigate the risk of incident syncope stratified by genotype carrier status. Results We report on a genome-wide significant locus on chromosome 2q32.1 with the lead SNP rs12465214 (odds ratio [OR] = 1.13, 95% confidence interval [CI] = 1.10–1.17, P=5.8x10–15; Figure 1a). This association was replicated in the iPSYCH cohort, where homozygous carriers of the C allele conferred an increased hazard ratio (HR=1.30, CI: 1.15–1.46, P=1.68x10–5; Figure 1b). LD score regression demonstrated a significant genetic correlation (rg) with coronary artery disease (rg=0.41, P=6.99x10–15) and related phenotypes such as angina and hypertension (Figure 1c). Analyses of eQTL (P=4x10–8) and epigenetic chromatin states revealed that variation in this locus likely affects expression of the gene ZNF804A, which resides in its proximity (Figure 1d). A qPCR analysis showed that ZNF804A was mostly expressed in the brain. A lower level of ZNF804A expression was also detected in the cerebral arteries. ZNF804A was not expressed in heart tissue. Figure 1 Conclusion rs12465214 is associated with syncope and collapse. Variation in this locus likely modulates the expression of the nearby gene ZNF804A through eQTLs and chromatin interactions. ZNF804A is mainly expressed in the brain and cerebral arteries. However, the precise function of ZNF804A is unknown. Furthermore, syncope and collapse is a polygenetic trait and share a significant genetic overlap with coronary artery disease, angina and hypertension. Acknowledgement/Funding This work was supported by grants from The John and Birthe Meyer Foundation, The Research Foundation of the Heart Centre, Rigshospitalet, The Research

Background: Adipokines are biologically active, low-molecular weight peptides, which play a major role in metabolic homeostasis in humans. Leptin has gained increasing attention in pediatrics as a biomarker for various metabolic pathologies. Yet, its usefulness is hampered by the relative lack of reference values from pediatric settings. Accordingly, this study aims to evaluate serum concentrations of leptin, soluble leptin receptor (sOB-R), and free leptin index (FLI) in healthy Danish schoolchildren aged 6–18 years and subsequently to establish reference intervals across sex and age groups. Methods: A total of 1193 healthy, non-obese Danish schoolchildren (730 girls, 463 boys) aged 6–18 years (median 11.9) were examined by trained medical staff. Serum leptin and sOB-R concentrations in venous fasting blood samples were quantitated by immunoassay. Percentile curves of leptin, sOB-R, and free leptin index were calculated using the General Additive Model for Location Scale and Shape (GAMLSS). Results: Significant age and sex-dependent differences in circulating leptin levels were found. In boys, the median leptin concentration for all ages combined was 3.35 μg/L (95%-interval: 0.71–22.47) and in girls, it was 9.89 ng/L (95%-interval: 2.06–41.49). For SOB-R, no sex-specific difference was found, and the median sOB-R concentration was 8.24 μg/L (IQR: 3.58–23.74; range: < 1.56–744.15). Conclusion: We demonstrated an age-dependent correlation with both serum leptin concentration and free leptin index with a gradual and significant increase in girls throughout childhood and adolescence and a significantly higher leptin concentration and free leptin index bell-shaped peak in early adolescence in boys.

Background Leptin and adiponectin are two key adipocyte secreted hormones and both are involved in several essential physiological mechanisms. Due to their central role in energy homeostasis their ratio, the leptin/adiponectin ratio, is believed to be a marker of metabolic derangement. Pediatric reference values are needed for the risk stratification of individual-measured ratios. Methods Blood samples were drawn from healthy, Danish schoolchildren following an overnight fast. A ratio was calculated from serum leptin and adiponectin quantifications done using commercially available ELISA Kits. Results Nine hundred eighty-three participants (583 girls) aged 6–18 years were included. Smoothed percentile curves and age-group specific percentiles were calculated. A correlation with age was demonstrated, with a gradual increase with age in girls and a negative parabolic relation, with a peak in age group 10–14, in boys. The leptin/adiponectin ratio was positively correlated to the body mass index standard deviation score for both girls and boys (p Conclusion The leptin/adiponectin ratio is correlated to age and differs between the sexes in healthy children and adolescents.

See Article by Lin et al Genome-wide association studies (GWAS) have revolutionized the study of disease biology, the genetics of common complex diseases, and have greatly improved our understanding of the myriads of pathways amenable to pharmacological intervention. Gradually, we are witnessing the translation of GWAS findings into patient care through the definition of risk profiles and identification of biomarkers.1 The heritability of cardiovascular traits, such as electrocardiograph (ECG) parameters, is of particular interest in cardiology. ECG is a valuable tool in determining the presence of many cardiovascular conditions, including structural derangements, arrhythmias, infarctions, and conduction disturbances. Better understanding of the genetics behind cardiovascular conditions will enhance the understanding of the pathophysiological basis of these diseases, help identify novel pharmacological targets, as well as contribute to risk assessment and clinical management of patients. Since Arking et al2 identified a common NOS1AP variant, which modulates cardiac repolarization, over a decade ago, GWAS has been applied to several ECG parameters. However, an examination of heritability of ECG traits determined that although 35% to 55% of the variance, seen in twin and family studies, can be explained by genetics, only 10% of the total trait variability have been identified using GWAS.3 This is despite increasing sample sizes and the concomitant increase in discoveries of common variants scattered throughout the genome.4 Furthermore, GWAS data are largely generated in populations of European ancestry. It is important to expand our knowledge of genetic variation and disease association in multiple populations along with examining rare variants, both in a bid to combat the phenomenon of missing heterozygosity, which plagues GWAS, but also to target genes and genetic …

Objective: To examine whether resistin levels in first trimester maternal serum are associated with insulin resistance or preeclampsia (PE). Methods: A case-control study of maternal serum resistin concentration conducted using 285 normal pregnancies and 123 PE pregnancies matched for gestational age, parity and maternal age. Samples were taken in gestational weeks 10+0–13+6. Results: There was a negative correlation between resistin and clinical severity of PE, but no correlation with IS, TNF-α, body mass index, birth weight and pregnancy length. Conclusions: Resistin is reduced in first trimester of PE pregnancies, particularly in severe PE. Inflammation and IS cannot explain this phenomenon.

Mitochondria play a significant role in human diseases. However, disease associations with mitochondrial DNA (mtDNA) SNPs have proven difficult to replicate. A reanalysis of eight schizophrenia-associated mtDNA SNPs, in 23,743 normal Danes and 2,538 schizophrenia patients, revealed marked inter-allelic differences in haplogroup affiliation and nuclear ancestry, genogeophraphic affinity (GGA). This bi-genomic linkage disequilibrium (2GLD) could entail population stratification. Only two mitochondrial SNPs, m. 15043A and m. 15218G, were significantly associated with schizophrenia. However, these associations disappeared when corrected for haplogroup affiliation. The extensive 2GLD documented is a major concern when interpreting historic as well as designing future mtDNA association studies.

Background: Placental protein-13 (PP13) is involved in placental invasion and has been suggested as a maternal serum marker of preeclampsia (PE) development. However, the discriminatory ability of PP13 in first trimester has not been completely clarified. Methods: PP13 was measured in first trimester (week 10+3–13+6) maternal serum from 120 PE pregnancies and 267 control pregnancies and was correlated with clinical parameters. The population screening performance of PP13 in combination with the PE markers pregnancy associated plasma protein A (PAPPA) and free leptin index (fLI) was assessed by Monte Carlo simulation. Results: In severe PE (including HELLP) cases (n=26) the median PP13 concentration was 35.8 pg/mL (range: 17.8–85.5 pg/mL) and in PE pregnancies (n=10) with birth prior to week 34, the median PP13 concentration was 30.6 pg/mL (13.1–50.1 pg/mL), compared to controls with a median of 54.8 pg/mL (range: 15.4–142.6 pg/mL) (p Conclusions: PP13 is a marker of severe PE and HELLP syndrome. The screening performance of PP13 can be markedly improved by combining it with fLI and PAPP-A.

Background Mitochondrial DNA (mtDNA) haplogroups (hgs) are evolutionarily conserved sets of mtDNA SNPs. Associations of hgs with geographical origin, disease and physiological characteristics have been reported, but have frequently not been reproducible. We assessed, using 418 mtDNA SNPs on the PsychChip (Illumina), the spatio-temporal distribution of mtDNA hgs in DNA isolated from geographically un-biased dried blood spots (DBS), collected from 1981 to 2005 through the Danish National Neonatal Screening program. Methods As part of the iPSYCH (www.iPSYCH.au.dk) recruitment protocol, 24,651 singletons (47.1% female), born between May 1 1981 and Dec 31 2005 were selected at random from the Danish Central Person Registry. The Samples were extracted from the Danish National Biobank and Genotyped at the Broad Institute as part of iPSYCH and PGC. Haplotyping of mtDNA was performed using the defining SNPs reported in www.phylotree.org. First the affiliation to macro-hg i.e. L0 – L6, M, N, R, and then to hgs – units more distal in the cladogram was established. Ancestry estimation was done using ADMIXTURE 1.3.052. Briefly, a reference population consisting of Human Genome Diversity Project (HGDP) ( http://www.hagsc.org/hgdp/ ) genotyping SNP data set, supplemented with representative samples of danes (716 individuals) and greenlanders (592 individuals) available at SSI, was used. Results The hg distribution was typically Northern European, and hgs were highly variable based on median-joining analysis, suggesting multiple founder events. Considerable heterogeneity and variation in autosomal geno-geographic affinity (ancestry) was observed. Thus, individuals with hg H exhibited 95%, and U hgs 38.2% - 92.5%, Danish ancestry. Significant clines between geographical regions and rural and metropolitan populations were found. Over 25 years, macro-hg L increased from 0.2% to 1.2% (p=1.1⁎E-10), and M from 1% to 2.4% (p=3.7⁎E-8). Hg U increased among the R macro-hg from 14.1% to 16.5% (p=1.9⁎E-3). Geno-geographic affinity, geographical skewedness, and sub-hg distribution suggested that the L, M and U increases are due to immigration. Discussion This study shows that the distribution of mtDNA hgs in Denmark is highly dynamic and complex. It comprises 1.6% of the Danish population over a 25-year period, and is by far the largest ever performed of the distribution of mtDNA hgs in any country. The method of collecting stored DBS from the PKU biobank, where the coverage is ~99%, enabled us to survey a true population based sample, where the time and place of birth was known from national electronic registries. This is in contrast to most other population genetic studies of adults sampled in a specific bias-prone context, e.g. hospitalized patients or localized samplings. The mtDNA haplotyping was based on array data with only 418 mtDNA SNPs and as a consequence not all sub-hgs could be called. The stringent adherence to specific SNPs left a number of persons, normally in the order of 1–2%, not haplotypable. An advantage of only using a limited set of markers is that confounding due to private variants is avoided.

Background Disturbed mitochondrial function has been implicated in psychiatric disease. All mitochondria contain a small maternally inherited DNA (mtDNA) of 16.6 kb. Through evolution and genetic drift, the mtDNA sequence has become fixed into haplogroups (hgs) with a characteristic population and geographical distribution. Different hgs exhibit variable functional capacity and have been demonstrated to be susceptibility factors for various, predominantly degenerative, diseases. Associations with Parkinsons and Alzheimers diseases, as well as cardiac, metabolic and psychiatric disorders have been reported. These findings have prompted the suggestion that variations in mitochondrial function might be susceptibility factors for disease. However, reports on association between mtDNA hgs and disease have frequently been conflicting. This is partly due to population stratification in examined populations and low-powered study designs. We analysed the association between mtDNA hgs and psychiatric disease using a very large geographically unbiased cohort, iPSYCH, comprising 2% of Danes as controls and 50,000+ psychiatric patients. Methods Using DNA extracted from dried blood spots from 50,567 Danish psychiatric patients (ADHD: 13,103; ASD: 12,891; Anorexia: 2,124; BD: 1,242; SCZ: 2,438, and AD: 16,225) and 24,196 controls we examined the association between psychiatric disease and mtDNA haplogroup in the Danish population. Psychiatric patients were identified from the Danish Psychiatric Central Registry. DNA was obtained from the Danish Neonatal Screening Biobank. Haplotyping was performed using the PGC developed PsychChip v.1.0. containing 418 SNPs in mtDNA. The SNP data were manually analysed and haplotyped according to haplogroup defining SNPs in Phylotree. Results Hg M was associated with affective disorder (n = 17260) with an OR of 0.47 (cfi.95%: 0.39 – 0.58) (p = 1*10–14), ADHD (n = 13395) with an OR of 0.50 (0.40 – 0.62)(p = 2*10–11). Among patients belonging to the macro-hg N, patients with schizophrenia (n = 2589), had a high proportion of hg A (n = 45), OR: 4.52 (cfi.95%: 3.0 – 6.7)(p = 1.2*10–12). No significant associations between mtDNA hgs and BD or ASD were found. In a mitoGWAS for all six conditions, three mitoSNPs were highly associated with affective disorder (p-values ranging from 2*10E-25 – 2*10E-21), ADHD (p-values: 2*10E-11 – 1*10E-10), and anorexia (p-values: 6*10E -8 - 6*10E-7) and another mitoSNP with schizophrenia (p = 2*10–11). Discussion Psychiatric disease seems to be bigenomic diseases with contributions from both the mitochondrial and nuclear genome. Haplogroup A is a risk factor for schizophrenia and haplogroup M is a protective factor for ADHD and affective disorder. However, both hgs A and M are infrequent in the Danish population and markers of Greenlandic and Asian origin, respectively. So it will be important through studies of interaction between mtDNA and the genomic DNA to precisely define the role of each of these factors in the etiology of psychiatric disease.

Background Schizophrenia (SCZ) is a severe psychiatric disorder wherein the genetic risk factors are far from being fully understood. Multiple lines of evidence have suggested mitochondrial dysfunction in SCZ, but association of mitochondrial variants with SCZ has not been extensively investigated. Methods We conducted gene-based and gene-set analyses to test the hypothesis that variants in nuclear-encoded mitochondrial genes influence susceptibility to SCZ, using results from the most recent release of the Schizophrenia Psychiatric Genomics Consortium GWAS (35,476 cases and 46,839 controls). We applied the MAGMA statistical tool to three gene sets: oxidative phosphorylation genes, other nuclear-encoded mitochondrial genes, and genes involved in nucleus-mitochondria communication signaling. Results For gene-based analysis, a total of 1,187 mitochondrial genes were analyzed and 19 were significant after correction for multiple testing. We applied the method for an independent sample (iPSYCH study, 2,290 SCZ cases and 21,621 controls) and 12 genes had replication p-values Discussion Overall, our study showed evidence that mitochondria play a role in SCZ. Results from our study, combined with future availability of mitochondrial DNA data, will assist researchers to evaluate the role of mitochondrial genomics, potentially across all complex genetic disorders.

Long QT syndrome (LQTS) is a genetic cardiac condition associated with prolonged ventricular repolarization, primarily a result of perturbations in cardiac ion channels, which predisposes individuals to life-threatening arrhythmias. Using DNA screening and sequencing methods, over 700 different LQTS-causing mutations have been identified in 13 genes worldwide. Despite this, the genetic cause of 30-50% of LQTS is presently unknown. MicroRNAs (miRNAs) are small (∼ 22 nucleotides) noncoding RNAs which post-transcriptionally regulate gene expression by binding complementary sequences within messenger RNAs (mRNAs). The human genome encodes over 1800 miRNAs, which target about 60% of human genes. Consequently, miRNAs are likely to regulate many complex processes in the body, indeed aberrant expression of various miRNA species has been implicated in numerous disease states, including cardiovascular diseases. MiR-1 and MiR-133A are the most abundant miRNAs in the heart and have both been reported to regulate cardiac ion channels. We hypothesized that, as a consequence of their role in regulating cardiac ion channels, genetic variation in the genes which encode MiR-1 and MiR-133A might explain some cases of LQTS. Four miRNA genes (miR-1-1, miR-1-2, miR-133a-1 and miR-133a-2), which encode MiR-1 and MiR-133A, were sequenced in 125 LQTS probands. No genetic variants were identified in miR-1-1 or miR-133a-1; but in miR-1-2 we identified a single substitution (n.100A> G) and in miR-133a-2 we identified two substitutions (n.-19G> A and n.98C> T). None of the variants affect the mature miRNA products. Our findings indicate that sequence variants of miR-1-1, miR-1-2, miR-133a-1 and miR-133a-2 are not a cause of LQTS in this cohort.

BACKGROUND: Low-grade chronic inflammation is a characteristic feature of obesity, the most important lifestyle risk factor for hypertension. Elevated plasma concentrations of the inflammatory biomarker C-reactive protein (CRP) are associated with an increased risk of hypertension, but elevated plasma CRP concentrations are also closely associated with obesity. It is uncertain whether CRP is directly involved in the pathogenesis of hypertension or is only a marker of other pathogenic processes closely related to obesity. METHODS: We studied 103 obese men (body mass index (BMI) ≥ 30.0 kg/m(2)); 63 of these men had 24-hour ambulatory blood pressure (ABP) ≥ 130/80 mm Hg and comprised the obese hypertensive (OHT) group. The 40 remaining obese men had 24-hour ABP < 130/80 mm Hg and comprised the obese normotensive (ONT) group. Our control group comprised 27 lean normotensive (LNT) men. All participants were medication-free. We measured plasma CRP concentrations with a high-sensitivity assay and determined body composition by dual energy x-ray absorptiometry scanning. RESULTS: There were no differences in anthropometric measures (BMI, waist circumference, or total fat mass percentage) between OHT and ONT groups (P ≥ 0.08). The obese groups had higher CRP concentrations than the LNT group (OHT: median = 2.30, interquartile range (IQR) = 1.10-4.10mg/L; ONT: median = 2.55, IQR = 1.25-4.80 mg/L; LNT: median = 0.60, IQR = 0.30-1.00 mg/L; P < 0.001), but there was no difference in CRP concentrations between OHT and ONT groups (P = 1.00). In the obese men, CRP was not correlated with either 24-hour systolic (r = 0.04; P = 0.71) or 24-hour diastolic ABP (r = -0.03; P = 0.78). CONCLUSIONS: Obese hypertensive men, matched for anthropometric measurements, have plasma CRP concentrations similar to those of obese normotensive men.

Objective To study the risk of adverse pregnancy outcomes in women with polycystic ovary syndrome (PCOS), and to examine the role of hyperandrogenaemia. Design Cohort study. Setting Singleton pregnancies in women with PCOS identified at a private fertility clinic during 1997–2010 and a background population including all singleton deliveries at Hvidovre Hospital, Denmark, in 2005. Population A cohort of 459 women with PCOS and a background population of 5409 women. Methods Obstetric outcomes were extracted from national Danish registries and odds ratios (ORs) were calculated by multiple logistic regression analysis, adjusting for age, parity, and body mass index. Main outcome measures Risk of pre-eclampsia, preterm delivery, and small for gestational age offspring in the entire PCOS population and in a subsample with hyperandrogenaemia. Results Women with PCOS had an increased risk of preterm delivery

Several studies have shown that obese persons have lower circulating natriuretic peptide (NP) concentrations. The cause of the relative NP deficiency seen in obese persons is poorly understood, although variation in body composition and metabolic abnormalities has been suggested to play a role. Thus, the aim of this study was to assess whether variation in circulating NP concentrations would be associated with differences in metabolic disturbances rather than with differences in body composition.In 27 normal weight men (body mass index (BMI) = 20.0-24.9kg/m(2)) and 103 obese men (BMI ≥ 30kg/m(2)), we determined body composition (total, android, and gynoid fat mass) by dual energy x-ray absorptiometry scanning, and we measured fasting serum concentrations of midregional proatrial NP (MR-proANP) and insulin, as well as fasting plasma glucose concentrations.Mean weight ± SD was 74.9±6.7kg in the normal weight men and 106.1±10.8kg in obese men. Applying multiple regressions, adjusting for age and weight status (normal weight vs. obese), serum MR-proANP concentrations were significantly inversely associated with serum insulin concentrations (β = -0.39; P0.0001) and plasma glucose concentrations (β = -0.21; P = 0.02) but not with total (β = 0.00), android (β = -0.01), or gynoid (β = 0.03) fat mass percentage (P0.76). No significant interaction effects between metabolic measurements or body composition measurements and weight status on MR-proANP concentrations were found (P0.08).In normal weight and obese men, lower circulating NP concentrations are associated with higher insulin and glucose concentrations and not with the proportion of total fat mass or the distribution of fat mass.

Congenital long QT syndrome (cLQTS) is a genetic disorder predisposing to ventricular arrhythmia, syncope and sudden death. Over 700 different cLQTS-causing mutations in 13 genes are known. The genetic spectrum of LQTS in 44 South African cLQTS patients (23 known to carry the South African founder mutation p.A341V in KCNQ1) was established by screening for mutations in the coding regions of KCNQ1, KCNH2, KCNE1, KCNE2 and SCN5A, the most frequently implicated cLQTS-causing genes (five-gene screening). Fourteen disease-causing mutations were identified, eight (including the founder mutation) in KCNQ1, five in KCNH2 and one in KCNE1. Two mutations were novel. Two double heterozygotes were found among the 23 families (8.5%) carrying the founder mutation. In conclusion, cLQTS in South Africa reflects both a strong founder effect and a genetic spectrum similar to that seen in other populations. Consequently, five-gene screening should be offered as a standard screening option, as is the case internationally. This will disclose compound and double heterozygotes. Fivegene screening will most likely be even more informative in other South African sub-populations with a greater genetic diversity.

Hypertrophic cardiomyopathy (HCM) is a myocardial disorder, with complications including heart failure, thromboemboli and sudden death. Human and feline HCM (fHCM) are clinically comparable, thus fHCM may serve as a spontaneous animal model. fHCM in Maine Coon (MC) cats is associated with the p.A31P mutation in the cMyBP-C protein. The mutation is located in the cMyBP-C C0-domain which is known to interact with actin. The presence and levels of the wild type and mutated protein in heart tissue from mutant and wild type MC cats were examined by SDS-PAGE and mass spectrometry (MS). Quantitative yeast-2-hybrid (Y2H) protein-protein interaction analysis was used to assess the effect of the mutation on C0C1/actin interaction. The NMR-based structure of the C0 domain was used to calculate the energetic consequence of replacing alanine with a proline residue. In the homozygous MC cat, the mutated cMyBP-C protein was present, and cMyBPC-C levels were not reduced compared to that of the wild type cat. However, the interaction of actin with mutant cMyBP-C C0C1 was reduced compared to that of wild type. This may be because the substitution of the alanine with proline in position 31 was energetically highly unfavorable and resulted in only one hydrogen bond within the anti-parallel beta-strand compared to two hydrogen-bonds for alanine, possibly destabilizing the structure of the actin-interacting domain. The p.A31P mutation is present in cardiac tissue and the most likely pathogenic mechanism is interference with contractility by reducing binding of the C0C1 domain of cMyBP-C to actin.

Vertical transmission of human immunodeficiency virus (HIV) remains a major global health problem. We assessed the association of mannose binding lectin (MBL) deficiency and vertical transmission of HIV. Novel diagnostics would be a major breakthrough in this regard. MBL is a liver-derived protein and a key component of the innate immune system. MBL levels may be classified as normal, intermediate, or deficient in the plasma and can use MBL2 haplotypes as a proxy. These haplotypes comprise polymorphisms in the MBL2 gene and promoter region and are known to result in varying levels of MBL deficiency. MBL deficiency can be defined as presence of A/O and O/O genotypes in the mothers and their children. MBL deficiency leads to defective opsonization activities of the innate immune system and increased susceptibility to several infections, including HIV-1. We determined the prevalence of MBL deficiency, using MBL2 haplotypes among 622 HIV-positive Zimbabwean mothers and their children aged 9-18 months old, in relation to the HIV-1 vertical transmission risk. The median age of the mothers was 30 (26-34, interquartile range [IQR]) years, and the babies' median age was 13 (11-15, IQR) months old at the time of enrollment. From the sample of 622 mothers who were HIV-1 infected, 574 babies were HIV negative and 48 were HIV-1-positive babies, giving a transmission rate of 7.7%. MBL2 normal structural allele A and variants B (codon 5 AG), C (codon 57 AG), and promoter region SNPs -550(H/L) and -221(X/Y) were detected. Prevalence of haplotype-predicted MBL deficiency was 34% among the mothers and 32% among the children. We found no association between maternal MBL2 deficiency and HIV-1 transmission to their children. We found no difference in the distribution of HIV-1 infected and uninfected children between the MBL2 genotypes of the mothers and those of the children. Taken together, the present study in a large sample of mother-infant pairs in Zimbabwe adds to the emerging literature and the hypothesis that MBL2 variation as predicted by haplotypes does not influence the vertical transmission risk for HIV. Research from other populations from the African continent is called for to test this hypothesis further.

The implementation of new methods of treating and preventing disease raises many question of both technical and moral character. Currently, many studies focus on developing a screening test for preeclampsia (PE), a disease complicating 2-8% of pregnancies, potentially causing severe consequences for pregnant women and their fetuses. The purpose is to develop a test that can identify pregnancies at high risk for developing PE sufficiently early in pregnancy to allow for prophylaxis. However, the question of implementing a screening test for PE does not only involve an evaluation of technical feasibility and clinical efficacy, it also requires an analysis of how the test influences the conditions and choices for those tested. This study evaluates state-of-the-art techniques for preeclampsia screening in an ethical framework, pointing out the central areas of moral relevance within the context of such screening activity. Furthermore, we propose ethical guidelines that a screening programme for PE should meet in order to become an uncontroversial addition to prenatal health care.

Background— Atrial fibrillation (AF) is the most common cardiac arrhythmia. The cardiac sodium channel, Na V 1.5, plays a pivotal role in setting the conduction velocity and the initial depolarization of the cardiac myocytes. We hypothesized that early-onset lone AF was associated with genetic variation in SCN5A . Methods and Results— The coding sequence of SCN5A was sequenced in 192 patients with early-onset lone AF. Eight nonsynonymous mutations (T220I, R340Q, T1304M, F1596I, R1626H, D1819N, R1897W, and V1951M) and 2 rare variants (S216L in 2 patients and F2004L) were identified. Of 11 genopositive probands, 6 (3.2% of the total population) had a variant previously associated with long QT syndrome type 3 (LQTS3). The prevalence of LQTS3-associated variants in the patients with lone AF was much higher than expected, compared with the prevalence in recent exome data (minor allele frequency, 1.6% versus 0.3%; P =0.003), mainly representing the general population. The functional effects of the mutations were analyzed by whole cell patch clamp in HEK293 cells; for 5 of the mutations previously associated with LQTS3, patch-clamp experiments showed an increased sustained sodium current, suggesting a mechanistic overlap between LQTS3 and early-onset lone AF. In 9 of 10 identified mutations and rare variants, we observed compromised biophysical properties affecting the transient peak current. Conclusions— In a cohort of patients with early-onset lone AF, we identified a high prevalence of SCN5A mutations previously associated with LQTS3. Functional investigations of the mutations revealed both compromised transient peak current and increased sustained current.

Background Mitochondria play a significant role in human diseases. Genetic variants in mitochondrial DNA (mtDNA) – and in nuclear genes coding for mitochondrial function - have been associated with disease. Mitochondrial disease is clinically characterised by complex metabolic, neurological, muscular and psychiatric symptoms. SNPs in mtDNA have been proposed as potential disease modifiers. This has been reported in neurological degenerative diseases such as Alzheimer's disease and Parkinson's disease, metabolic diseases and cancers, as well as psychiatric diseases, notably schizophrenia (SZ) and bipolar disease. However, disease associations with mitochondrial DNA (mtDNA) SNPs have proven difficult to replicate. We identified eight mtDNA SNPs that had previously been associated with SZ and re-analysed the association with SZ in the Danish iPSYCH cohort, and examined the effect of mtDNA variation and variation in nuclear genogeographic affinity, or ancestry, on the association. Methods Persons were recruited through the iPSYCH program, which is a case-cohort study. DNA was isolated from dried blood spots obtained through the Danish Neonatal Screening program. Genotyping was performed using the PsychChip array, comprising 418 SNPs in mtDNA. Supervised ancestry estimation of the nuclear genome was performed using a reference population of 2,248 persons assigned to one of nine population Groups. The estimation was done with 103,268 nuclear SNPs using ADMIXTURE 1.3.050. Results A reanalysis of eight schizophrenia-associated mtDNA SNPs, in 23,743 normal Danes and 2,538 schizophrenia patients, revealed marked inter-allelic differences both in haplogroup affiliation and nuclear ancestry, genogeophraphic affinity (GGA). Only two mitochondrial SNPs, m.15043 A and m.15218 G, were significantly associated with schizophrenia. However, these associations disappeared when corrected for haplogroup affiliation. Discussion The combined variation in mtDNA affiliation and nuclear genomic ancestry, bi-genomic linkage disequilibrium (2GLD) could entail population stratification. This might result in both false positive and negative associations between mtDNA SNPs and disease. The extensive 2GLD documented for the eight SNPs examined here is a major concern when interpreting historic as well as designing future mtDNA association studies. The fact that careful control, as here, of mtDNA affiliation and the 2GLD, results in none of eight previously SZ associated mtDNA SNPs being associated with SZ in the very large Danish iPSYCH cohort, suggests that previously reported associations could indeed be spurious findings due to cryptic population stratification.

Mitochondrial DNA (mtDNA) haplogroups (hgs) are evolutionarily conserved sets of mtDNA SNP-haplotypes with characteristic geographical distribution. Associations of hgs with disease and physiological characteristics have been reported, but have frequently not been reproducible. Using 418 mtDNA SNPs on the PsychChip (Illumina), we assessed the spatio-temporal distribution of mtDNA hgs in Denmark from DNA isolated from 24,642 geographically un-biased dried blood spots (DBS), collected from 1981 to 2005 through the Danish National Neonatal Screening program. ADMIXTURE was used to establish the genomic ancestry of all samples using a reference of 100K+ autosomal SNPs in 2,248 individuals from nine populations. Median-joining analysis determined that the hgs were highly variable, despite being typically Northern European in origin, suggesting multiple founder events. Furthermore, considerable heterogeneity and variation in nuclear genomic ancestry was observed. Thus, individuals with hg H exhibited 95%, and U hgs 38.2% - 92.5%, Danish ancestry. Significant clines between geographical regions and rural and metropolitan populations were found. Over 25 years, macro-hg L increased from 0.2% to 1.2% (p = 1.1*E-10), and M from 1% to 2.4% (p = 3.7*E-8). Hg U increased among the R macro-hg from 14.1% to 16.5% (p = 1.9*E-3). Genomic ancestry, geographical skewedness, and sub-hg distribution suggested that the L, M and U increases are due to immigration. The complex spatio-temporal dynamics and genomic ancestry of mtDNA in the Danish population reflect repeated migratory events and, in later years, net immigration. Such complexity may explain the often contradictory and population-specific reports of mito-genomic association with disease.

Background ADAM12 has been shown to be an efficient maternal serum marker for Down syndrome (DS) in the first trimester; but recent studies, using a second generation assay, have not confirmed these findings. We examined the efficiency of a second generation assay for ADAM12. Materials and Methods ADAM12 concentrations were determined in 28 first trimester DS and 503 control pregnancies using a novel Research DelfiaR ADAM12 kit. Log10MoM distributions of ADAM12 and correlations with other markers were established. Population performance of screening was estimated by Monte Carlo simulation. Results ADAM12 was significantly reduced in the first trimester in DS pregnancies with a log10MoM of −0.1621 (equivalent to 0.68 MoM) (p < 0.001). The reduction decreased with advancing gestational age. ADAM12 used with PAPP-A + hCGβ + NT (CUB screening) increased the detection rate (DR) from 86% to 89% for a false positive rate (FPR) of 5%. When used for a fixed DR of 90%, the addition of ADAM12 resulted in a 25% reduction of the FPR. Conclusion ADAM12 is a moderately effective DS marker. It is not a cost-effective addition to CUB screening, but may be used to reduce the FPR in selected high-risk cases. Copyright © 2009 John Wiley & Sons, Ltd.

Background Prophylaxis with low-dose aspirin may reduce the risk of pre-eclampsia (PE) if introduced in first trimester. The performance of first trimester maternal serum screening for PE using free leptin index (fLI) and PAPP-A, where fLI = leptin/leptin soluble receptor was studied. Methods First trimester serum samples from 126 PE pregnancies and 289 control pregnancies were studied. fLI and PAPP-A were converted into gestational age and maternal weight independent log MoM values of PAPP-A and fLI. The screening performance of markers was studied by receiver-operator-characteristics curves. The performance of population screening was estimated by Monte Carlo simulation. Results fLI was significantly (p < 0.001) elevated [mean log MoM 0.2165 (SD: 0.2604)] compared to controls [mean log MoM −0.0368 (SD: 0.3132)] and PAPP-A was significantly (p < 0.001) reduced [mean log MoM −0.0133 (SD: 0.2661)] compared to controls [mean log MoM 0.0474 (SD: 0.2521)] in PE pregnancies. There was no correlation between fLI and PAPP-A in control or PE pregnancies. Combined fLI and PAPP-A screening for PE had estimated population detection rates of 22% and 35% for false positives rates of 6% and 12%, respectively. Conclusion Combining PAPP-A and fLI improves screening performance for PE compared to single marker screening. Copyright © 2009 John Wiley & Sons, Ltd.