A Mass Spectrometry-Based Proteome Study of Twin Pairs Discordant for Incident Acute Myocardial Infarction within Three Years after Blood Sampling Suggests Novel Biomarkers.

Acute myocardial infarction (AMI) is a major cause of mortality and morbidity worldwide, yet biomarkers for AMI in the short- or medium-term are lacking. We apply the discordant twin pair design, reducing genetic and environmental confounding, by linking nationwide registry data on AMI diagnoses to a survey of 12,349 twins, thereby identifying 39 twin pairs (48-79 years) discordant for their first-ever AMI within three years after blood sampling. Mass spectrometry of blood plasma identified 715 proteins. Among 363 proteins with a call rate > 50%, imputation and stratified Cox regression analysis revealed seven significant proteins (FDR < 0.05): FGD6, MCAM, and PIK3CB reflected an increased level in AMI twins relative to their non-AMI co-twins (HR > 1), while LBP, IGHV3-15, C1RL, and APOC4 reflected a decreased level in AMI twins relative to their non-AMI co-twins (HR < 1). Additional 50 proteins were nominally significant (p < 0.05), and bioinformatics analyses of all 57 proteins revealed biology within hemostasis, coagulation cascades, the immune system, and the extracellular matrix. A protein-protein-interaction network revealed Fibronectin 1 as a central hub. Finally, technical validation confirmed MCAM, LBP, C1RL, and APOC3. We put forward novel biomarkers for incident AMI, a part of the proteome field where markers are surprisingly rare and where additional studies are highly needed.

Differences in DNA Methylation-Based Age Prediction Within Twin Pairs Discordant for Cancer.

DNA methylation-based age acceleration (DNAmAA) is associated with cancer, with both cancer tissue and blood showing increased DNAmAA. We aimed to investigate whether DNAmAA is associated with cancer risk within twin pairs discordant for cancer, and whether DNAmAA has the potential to serve as a biomarker for such. The study included 47 monozygotic and 48 same-sex-dizygotic cancer-discordant twin pairs from the Finnish Twin Cohort study with blood samples available between 17 and 31 years after the cancer diagnosis. We studied all cancers (95 pairs), then separately breast cancer (24 pairs) and all sites other than breast cancer (71 pairs). DNAmAA was calculated for seven models: Horvath, Horvath intrinsic epigenetic age acceleration, Hannum, Hannum intrinsic epigenetic age acceleration, Hannum extrinsic epigenetic age acceleration, PhenoAge and GrimAge. Within-pair differences in DNAmAA were analyzed by paired t tests and linear regression. Twin pairs sampled before cancer diagnosis did not differ significantly in DNAmAA. However, the within-pair differences in DNAmAA before cancer diagnosis increased significantly the closer the cancer diagnosis was, and this acceleration extended for years after the diagnosis. Pairs sampled after the diagnosis differed for DNAmAA with the Horvath models capturing cancer diagnosis-associated DNAmAA across all three cancer groupings. The results suggest that DNAmAA in blood is associated with cancer diagnosis. This may be due to epigenetic alterations in relation to cancer, its treatment or associated lifestyle changes. Based on the current study, the biomarker potential of DNAmAA in blood appears to be limited.

Microbial Species that Initially Colonize the Human Gut at Birth or in Early Childhood Can Stay in Human Body for Lifetime.

In recent years, many studies have described the composition and function of the human microbiome at different body sites and suggested a role for the microbiome in various diseases and health conditions. Some studies, using longitudinal samples, have also suggested how the microbiome changes over time due to disease, diet, development, travel, and other environmental factors. However, to date, no study has demonstrated whether the microorganisms established at birth or in early childhood, either transmitted from parents or obtained from the environment, can stay in the human body until adult or senior age. To directly answer this question is difficult, because microbiome samples at childhood and at later adulthood for the same individual will need to be compared and the field is not old enough to have allowed for that type of sample collection. Here, using a metagenomic approach, we analyzed 1004 gut microbiome samples from senior adults (65 ± 7.8 years) from the TwinsUK cohort. Our data indicate that many species in the human gut acquired in early childhood can stay for a lifetime until senior ages. We identified the rare genomic variants (single nucleotide variation and indels) for 27 prevalent species with enough sequencing coverage for confident genomic variant identification. We found that for some species, twin pairs, including both monozygotic (MZ) and dizygotic (DZ) twins, share significantly more rare variants than unrelated subject pairs. But no significant difference is found between MZ and DZ twin pairs. These observations strongly suggest that these species acquired in early childhood remained in these persons until senior adulthood.

A within-sibling pair analysis of lifestyle behaviours and BMI z-score in the multi-centre I.Family study.

BACKGROUND AND AIMS: By investigating differences in lifestyle behaviours and BMI in sibling pairs, family-level confounding is minimized and causal inference is improved, compared to cross-sectional studies of unrelated children. Thus, we aimed to investigate within-sibling pair differences in different lifestyle behaviours and differences in BMI z-scores in children and adolescents. METHODS AND RESULTS: We examined three groups of sibling pairs 1) all same-sex sibling pairs with maximum 4 years age difference (n = 1209 pairs from 1072 families in 8 countries, mean age 10.7 years, standard deviation 2.4 years), 2) sibling pairs discordant for overweight (n = 262) and 3) twin pairs (n = 85). Usual dietary intake was estimated by 24-h recalls and time spent in light (LPA) and moderate-to-vigorous physical activity (MVPA) was measured by accelerometers. Screen time, sleep and dieting for weight loss were assessed by questionnaires. Within all 3 groups of sibling pairs, more time in MVPA was associated with lower BMI z-score. Higher energy intake was associated with higher BMI z-score within twin pairs and within all sibling pairs who were not currently dieting for weight loss. Regarding LPA, screen time or sleep duration, no or inconsistent associations were observed for the three groups of sibling pairs. CONCLUSIONS: MVPA and energy intake were associated with BMI differences within sibling and twin pairs growing up in the same home, thus independent of family-level confounding factors. Future studies should explore whether genetic variants regulating appetite or energy expenditure behaviours account for weight differences in sibling pairs.

Copy Number Variation Analysis of 100 Twin Pairs Enriched for Neurodevelopmental Disorders.

Hundreds of penetrant risk loci have been identified across different neurodevelopmental disorders (NDDs), and these often involve rare (<1% frequency) copy number variations (CNVs), which can involve one or more genes. Monozygotic (MZ) twin pairs are long thought to share 100% of their genomic information. However, genetic differences in the form of postzygotic somatic variants have been reported recently both in typically developing (TD) and in clinically discordant MZ pairs. We sought to investigate the contribution of rare CNVs in 100 twin pairs enriched for NDD phenotypes with a particular focus on postzygotic CNVs in MZ pairs discordant for autism spectrum disorder (ASD) using the Illumina Infinium PsychArray. In our sample, no postzygotic de novo CNVs were found in 55 MZ twin pairs, including the 13 pairs discordant for ASD. We did detect a higher rate of CNVs overlapping genes involved in disorders of the nervous system, such as a rare deletion affecting HNRNPU, in MZ pairs discordant and concordant for ASD in comparison with TD pairs (p = .02). Our results are in concordance with earlier findings that postzygotic de novo CNV events are typically rare in genomic DNA derived from saliva or blood, and suggests that the discordance of NDDs in our sample of twins is not explained by discordant CNVs. Still, studies investigating postzygotic variation in MZ discordant twins using DNA from different tissues and single cells and higher resolution genomics are needed in the future.

Proteomic analysis of hair shafts from monozygotic twins: Expression profiles and genetically variant peptides.

Forensic association of hair shaft evidence with individuals is currently assessed by comparing mitochondrial DNA haplotypes of reference and casework samples, primarily for exclusionary purposes. Present work tests and validates more recent proteomic approaches to extract quantitative transcriptional and genetic information from hair samples of monozygotic twin pairs, which would be predicted to partition away from unrelated individuals if the datasets contain identifying information. Protein expression profiles and polymorphic, genetically variant hair peptides were generated from ten pairs of monozygotic twins. Profiling using the protein tryptic digests revealed that samples from identical twins had typically an order of magnitude fewer protein expression differences than unrelated individuals. The data did not indicate that the degree of difference within twin pairs increased with age. In parallel, data from the digests were used to detect genetically variant peptides that result from common nonsynonymous single nucleotide polymorphisms in genes expressed in the hair follicle. Compilation of the variants permitted sorting of the samples by hierarchical clustering, permitting accurate matching of twin pairs. The results demonstrate that genetic differences are detectable by proteomic methods and provide a framework for developing quantitative statistical estimates of personal identification that increase the value of hair shaft evidence.

Exploring Subclinical Phenotypic Features in Twin Pairs Discordant for Cleft Lip and Palate.

OBJECTIVE: Monozygotic twins of an individual with an orofacial cleft have a significantly elevated risk for orofacial cleft compared with the general population, but still the concordance rate for orofacial cleft in monozygotic twins is about 40% to 50%. The goal of this study was to determine whether unaffected cotwins have an increased frequency of orbicularis oris muscle defects, a subclinical form of orofacial cleft. The presence of such defects may reduce the overall rate of discordance. METHOD: A total of 63 discordant monozygotic and dizygotic twin pairs, 262 unaffected nontwin siblings, and 543 controls with no history of orofacial clefts were assessed for orbicularis oris defects by high-resolution ultrasound. Frequencies were compared by the Fisher exact test. RESULTS: Unaffected cotwins from discordant monozygotic pairs had a higher frequency of defects (12.5%) than the other test groups (6.38% to 6.99%), but the difference was not statistically significant (P = .74). CONCLUSIONS: In this study, orbicularis oris defects were not statistically significantly more common among the unaffected twins from orofacial cleft discordant twin pairs. The trends in the results warrant future studies with larger sample sizes and additional subclinical phenotypes.

Whole-Exome Sequencing in Nine Monozygotic Discordant Twins.

By definition, monozygotic (MZ) twins carry an identical set of genetic information. The observation of early post-twinning mutational events was shown to cause phenotypic discordance among MZ twin pairs. These mutational events comprise genomic alterations at different scales, ranging from single nucleotide changes to larger copy-number variations (CNVs) of varying sizes, as well as epigenetic changes. Here, we performed whole-exome sequencing (WES) in nine discordant MZ twins to identify somatic mutational events in the affected twin that might exert a dominant negative effect. Five of these MZ twin pairs were discordant for congenital heart defects (CHD), two for endocrine disorders, one for omphalocele, and one for congenital diaphragmatic hernia (CDH). Analysis of WES data from all nine MZ twin pairs using the de novo probability tool DeNovoGear detected only one apparent de novo variation in TMPRSS13 in one of the CHD-affected twins. Analysis of WES data from all nine MZ twin pairs by using standard filter criteria without the de novo probability tool DeNovoGear revealed a total of 6,657 variations in which both the twin pairs differed. After filtering for variations only present in the affected twins and absent in in-house controls, 722 variations remained. Visual inspection for read quality decreased this number to 12, present only in the affected twin. However, Sanger sequencing of the overall 13 variations failed to confirm the variation in the affected twin. These results suggest that somatic mutational events in coding regions do not seem to play a major role in the phenotypic expression of MZ discordant twin pairs.

DNA Methylation Changes in the IGF1R Gene in Birth Weight Discordant Adult Monozygotic Twins.

Low birth weight (LBW) can have an impact on health outcomes in later life, especially in relation to pre-disposition to metabolic disease. Several studies suggest that LBW resulting from restricted intrauterine growth leaves a footprint on DNA methylation in utero, and this influence likely persists into adulthood. To investigate this further, we performed epigenome-wide association analyses of blood DNA methylation using Infinium HumanMethylation450 BeadChip profiles in 71 adult monozygotic (MZ) twin pairs who were extremely discordant for birth weight. A signal mapping to the IGF1R gene (cg12562232, p = 2.62 × 10(-8)), was significantly associated with birth weight discordance at a genome-wide false-discovery rate (FDR) of 0.05. We pursued replication in three additional independent datasets of birth weight discordant MZ pairs and observed the same direction of association, but the results were not significant. However, a meta-analysis across the four independent samples, in total 216 birth-weight discordant MZ twin pairs, showed a significant positive association between birth weight and DNA methylation differences at IGF1R (random-effects meta-analysis p = .04), and the effect was particularly pronounced in older twins (random-effects meta-analysis p = .008, 98 older birth-weight discordant MZ twin pairs). The results suggest that severe intra-uterine growth differences (birth weight discordance >20%) are associated with methylation changes in the IGF1R gene in adulthood, independent of genetic effects.

Twin study dissects CXCR3+ memory B cells as non-heritable feature in multiple sclerosis.

BACKGROUND: In multiple sclerosis (MS), B cells are considered main triggers of the disease, likely as the result of complex interaction between genetic and environmental risk factors. Studies on monozygotic twins discordant for MS offer a unique way to reduce this complexity and reveal discrepant subsets. METHODS: In this study, we analyzed B cell subsets in blood samples of monozygotic twins with and without MS using publicly available data. We verified functional characteristics by exploring the role of therapy and performed separate analyses in unrelated individuals. FINDINGS: The frequencies of CXCR3+ memory B cells were reduced in the blood of genetically identical twins with MS compared to their unaffected twin siblings. Natalizumab (anti-VLA-4 antibody) was the only treatment regimen under which these frequencies were reversed. The CNS-homing features of CXCR3+ memory B cells were supported by elevated CXCL10 levels in MS cerebrospinal fluid and their in vitro propensity to develop into antibody-secreting cells. CONCLUSIONS: Circulating CXCR3+ memory B cells are affected by non-heritable cues in people who develop MS. This underlines the requirement of environmental risk factors such as Epstein-Barr virus in triggering these B cells. We propose that after CXCL10-mediated entry into the CNS, CXCR3+ memory B cells mature into antibody-secreting cells to drive MS. FUNDING: This work was supported by Nationaal MS Fonds (OZ2021-016), Stichting MS Research (19-1057 MS, 20-490f MS, and 21-1142 MS), the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program grant agreement no. 882424, and the Swiss National Science Foundation (733 310030_170320, 310030_188450, and CRSII5_183478).

Sex differences in ocular biometric measurements: A twin study.

Objective: Gender differences in ocular biometric measurements of opposite-sex and same-sex twin pairs are still unclear. We aimed to investigate the difference between ocular biometric measurements in adolescent twin pairs. Materials and methods: This retrospective study included a total of 64 eyes of 64 adolescents from 32 twins. The ocular biometric measurements and refractive prediction error (RE) were acquired from four groups of dizygotic (DZ) twins: boys from same-sex twin-pairs (SSM, n = 20), boys from opposite-sex twin-pairs (OSM, n = 8), girls from opposite-sex twin-pairs (OSF, n = 8), and girls from same-sex twin-pairs (SSF, n = 29). Results: The mean age of the patient was 9.92 ± 2.84 (range: 6-18) years. Overall, boys had higher height, AL, WTW, but lower Ks, and Kf than girls (p < 0.05). Specifically, SSF was found to have the lowest lens thickness (LT), anterior chamber depth (ACD), central corneal thickness (CCT), white to white (WTW), and axial length (AL) levels, while the highest keratometry readings in the flat (Kf) and steep (Ks) levels compared with OSM, OSF, and SSM adolescents (p < 0.05). Compared with the OSF adolescents, ACD levels of the SSF adolescents were significantly lower [(2.99 ± 0.35) and (3.26 ± 0.15) mm, p = 0.033)], but Kf indicator was significantly larger [(43.93 ± 1.64) and (42.91 ± 1.75), p = 0.016)]. Conclusion: Our study indicates that there was a significant difference in ocular biometric measurements between twin pairs, and sharing the uterus with a DZ twin SSF has smaller ocular indicator measurements. Our findings provide information on the eyeball and refractive development in adolescents.

Positive association and future perspectives of mitochondrial DNA copy number and telomere length - a pilot twin study.

INTRODUCTION: Recent experimental and population studies have highlighted the existence of telomere-mitochondria interplay. Besides studies revealing the molecular mechanisms underlying the associations of telomere defects and mitochondrial functions, investigations of mitochondrial DNA copy number (mtDNAcn) and telomere length (TL) in healthy and disease phenotypes have likewise begun, with the aim of gaining more insights about their relationship in humans. MATERIAL AND METHODS: A total of 142 asymptomatic adult twins, comprising 96 monozygotic (MZ) and 46 dizygotic (DZ) twins (mean age: 50.54 ±15.43 years), members of the Hungarian Twin Registry, were included in the analysis. Applying the qPCR standard curve method, we investigated the relationship of mtDNA copy number, telomere length and clinical data, besides assessing co-twin similarities of MZ and DZ twins for their mtDNAcn and TL measures. RESULTS: We found that twins were similar in their intraclass correlation coefficients irrespective of zygosity, suggesting a possibly more important role of common (shared) environmental factors compared to non-shared (unique) environmental and to a smaller degree also individual genetic influences. We confirmed a significant positive association between mtDNAcn and TL (r = 0.28, p < 0.01) in age- and sex-corrected analysis. Following bivariate estimates and correction with significant predictors, the independent positive associations were further verified. CONCLUSIONS: Our results extend the until now modest number of studies investigating mtDNAcn and TL simultaneously in humans. In addition, we are the first to examine the relationship between mtDNAcn and telomere length in MZ and DZ twin subjects.

Are the Relationships of Lean Mass and Fat Mass With Bone Microarchitecture Causal or Due to Familial Confounders? A Novel Study of Adult Female Twin Pairs.

It is not known whether the relationships of lean mass (LM) and fat mass (FM) with bone microarchitecture and geometry are causal and/or are because of confounders, including familial confounders arising from genetic and environment effects shared by relatives. We tested the hypotheses that: (i) LM is associated with cortical bone traits, (ii) FM is associated with trabecular bone traits, and (iii) these relationships of LM and FM with bone microarchitecture and geometry have a causal component. Total body composition was quantified for 98 monozygotic (MZ) and 54 dizygotic (DZ) white female twin pairs aged 31 to 77 years. Microarchitecture at the distal tibia and distal radius was quantified using HRpQCT and StrAx software. We applied the Inference about Causation through Examination of FAmiliaL CONfounding (ICE FALCON) method. Within-individuals, distal tibia total bone area, cortical area, cortical thickness, and trabecular number were positively associated with LM (standardized regression coefficient (ß) = 0.13 to 0.43; all p < 0.05); porosity of the inner transitional zone (ITZ) was negatively associated with LM (ß = -0.22; p < 0.01). Trabecular number was positively associated with FM (ß = 0.40; p < 0.001), and trabecular thickness was negatively associated with FM (ß = -0.27; p < 0.001). For porosity of ITZ and trabecular number, the cross-pair cross-trait association with LM was significant before and after adjustment for the within-individual association with LM (all ps < 0.05). For trabecular number, the cross-pair cross-trait association with FM was significant before and after adjustment for the within-individual association with FM (p < 0.01). There were no significant changes in these cross-pair cross-trait associations after adjustment for the within-individual association (p = 0.06 to 0.99). Similar results were found for distal radius measures. We conclude that there was no evidence that the relationships of LM and FM with bone microarchitecture and geometry are causal; they must in part due to by familial confounders affecting both bone architecture and body composition. © 2020 The Authors. JBMR Plus published by Wiley Periodicals LLC. on behalf of American Society for Bone and Mineral Research.

Twin studies on the effect of genetic factors on serum agalactosyl immunoglobulin G levels.

The level of immunoglobulin G (IgG) lacking the terminal galactose, referred to as agalactosyl IgG, was found to be increased in chronic inflammatory diseases, such as rheumatoid arthritis and inflammatory bowel disease (IBD), particularly in Crohn's disease, which is suggested to have a genetic component. This oligosaccharide modification of IgG is mainly regulated by the expression of glyco-genes; however, the association between genetic factors and changes in the IgG glycosylation has not been fully elucidated. The aim of the present study was to assess the role of genetics in this process by comparing the serum agalactosyl IgG levels between members of monozygotic and dizygotic twin pairs who underwent medical check-ups at the same time. The serum agalactosyl IgG level was assayed using high-performance liquid chromatography. Hematological and biochemical markers, including γ-glutamyltranspeptidase (γGTP), alanine aminotransferase (ALT) and white blood cell (WBC) count, were also measured. Although the serum γGTP levels (and, to a lesser extent, ALT and WBC levels) exhibited a correlation within monozygotic twin pairs, agalactosyl IgG levels were not found to be correlated between members of either type of twin pairs. Thus, the role of genetic factors in determining serum agalactosyl IgG levels may be less significant compared to the effect of environmental factors or the onset of inflammatory disease.

A comparison of anthropometric, metabolic, and reproductive characteristics of young adult women from opposite-sex and same-sex twin pairs.

BACKGROUND: Prenatal exposure to androgens has been linked to masculinization of several traits. We aimed to determine whether putative female intra-uterine exposure to androgens influences anthropometric, metabolic, and reproductive parameters using a twin design. METHODS: Two cohorts of Finnish twins born in 1975-1979 and 1983-1987 formed the basis for the longitudinal FinnTwin16 (FT16) and FinnTwin12 (FT12) studies. Self-reported anthropometric characteristics, disease status, and reproductive history were compared between 679 same-sex (SS) and 789 opposite-sex (OS) female twins (mean age ± SD: 34 ± 1.1) from the wave 5 of data collection in FT16. Serum lipid and lipoprotein subclass concentrations measured by nuclear magnetic resonance spectroscopy were compared in 226 SS and 169 OS female twins (mean age ± SD: 24 ± 2.1) from the wave 4 of data collection in FT12 and FT16. RESULTS: Anthropometric measures, the prevalence of hypertension and diabetes mellitus type 2 did not differ significantly between females from SS and OS twin pairs at age 34. Similarly, the prevalence of infertility, age at first pregnancy and number of induced and spontaneous abortions did not differ significantly between these two groups of women. The serum lipid and lipoprotein profile did not differ between females from SS and OS twins at age 24. CONCLUSION: We found no evidence that androgen overexposure of the female fetus affects obesity, metabolic profile, or reproductive health in young adult females. However, these results do not exclude the possibility that prenatal androgen exposure in females could be adversely associated with these phenotypes later in life.

Frontal white matter integrity as an endophenotype for schizophrenia: diffusion tensor imaging in monozygotic twins and patients' nonpsychotic relatives.

Diffusion tensor imaging (DTI) provides anatomical connectivity information by examining the directional organization of white matter microstructure. Anatomical connectivity and its abnormalities may be heritable traits associated with schizophrenia. To further examine this hypothesis, two studies were conducted to compare anatomical connectivity between (a) monozygotic (MZ) twin pairs and random pairings among twins and (b) first-degree relatives of schizophrenia patients and a healthy control group. Analyses focused on frontal regions of the brain following previous findings of anatomical connectivity abnormalities associated with schizophrenia. For Study 1, eighteen MZ twin pairs (11 female pairs, age: M = 25.44, SD = 5.69) were recruited. For Study 2, twenty-two first-degree relatives of schizophrenia patients (14 females, age: M = 48.50, SD = 8.22), and 30 healthy controls (12 females, age: M = 43.83, SD = 11.39) were recruited. Fractional anisotropy (FA), a white matter directional organization metric, was measured with DTI. In Study 1, FA values were more strongly correlated between MZ twin pairs than between randomly generated pairs in genu of corpus callosum, anterior cingulum and forceps minor. In Study 2, relatives of schizophrenia patients showed reduced FA values in medial frontal white matter (p < 0.05, corrected). The present study suggested that anatomical connectivity in medial prefrontal cortex appeared significantly heritable within MZ twin pairs, an important criterion in the development of an endophenotype. In addition, altered medial frontal white matter integrity found in non-affected relatives of schizophrenia patients seems to suggest that reduced white matter integrity in medial frontal regions of the brain might be associated with the genetic liability to schizophrenia.