Reassessing the evidence of a survival advantage in Type 2 diabetes treated with metformin compared with controls without diabetes: a retrospective cohort study.

BACKGROUND: Previous research has suggested that individuals with Type 2 diabetes and initiated on metformin monotherapy present with a survival advantage compared with the general population without diabetes. This finding has generated considerable interest in the prophylactic use of metformin against age-related morbidity. METHODS: Utilizing Danish National Health Registers, we assessed differences in survival associated with metformin monotherapy for Type 2 diabetes compared with no diagnosis of diabetes in both singleton and discordant twin populations between 1996 and 2012. Data were analysed in both nested case-control and matched cohort study designs, with incidence rate ratios (IRRs) and hazard ratios estimated using conditional logistic regression and Cox proportional hazards regression, respectively. RESULTS: In case-control pairs matched on birth year and sex or co-twin (sex, birth year and familial factors), incident Type 2 diabetes with treatment by metformin monotherapy initiation compared with no diagnosis of diabetes was associated with increased mortality in both singletons (IRR = 1.52, 95% CI: 1.37, 1.68) and discordant twin pairs (IRR = 1.90, 95% CI: 1.35, 2.67). After adjusting for co-morbidities and social indicators, these associations were attenuated to 1.32 (95% CI: 1.16, 1.50) and 1.64 (95% CI: 1.10, 2.46), respectively. Increased mortality was observed across all levels of cumulative use and invariant to a range of study designs and sensitivity analyses. CONCLUSIONS: Treatment initiation by metformin monotherapy in Type 2 diabetes was not associated with survival equal or superior to that of the general population without diabetes. Our contrasting findings compared with previous research are unlikely to be the result of differences in epidemiological or methodological parameters.

Impact of Cytomegalovirus Infection and Genetic Background on the Frequencies of Peripheral Blood Suppressor Cells in Human Twins.

Frequencies and proportions of regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs) in peripheral blood may be informative biomarkers for certain disease states. The influence of genetics and lifetime pathogen exposures on Treg and MDSC frequencies is largely unexplored. Cytomegalovirus (CMV) establishes a latent infection and causes an accumulation of late-differentiated CD8+ memory T cells, commonly associated with a lower frequency of naive cells. Here, analyzing peripheral blood mononuclear cells by multicolor flow cytometry, we found a tendency towards lower frequencies of CD4+CD25+FoxP3+ Tregs in CMV-seropositive than -seronegative middle-aged individuals (p = 0.054), whereas frequencies of lineage-negative CD14+HLA-DR-MDSCs were significantly lower in CMV-seropositive participants (p = 0.005). Assessing associations with the presence of antibodies against different CMV structural proteins, rather than merely assigning seropositivity or seronegativity, failed to yield any closer associations. Examining Treg subsets revealed at most a minor role of the individual's genetic background, based on an analysis of monozygotic (MZ, n = 42) versus dizygotic (DZ, n = 39) twin pairs from the Danish Twin Registry. The same was true for MDSCs. These initial results suggest that an immunological history of exposures is more important than genetics in determining overall human suppressor cell levels.

Genetic Influence on the Peripheral Differentiation Signature of Vδ2+ γδ and CD4+ αß T Cells in Adults.

Adaptive as well as innate immune traits are variously affected by environmental and genetic influences, but little is known about the impact of genetics on the diversity, differentiation and functionality of γδ T cells in humans. Here, we analyzed a cohort of 95 middle-aged twins from the Danish Twin Registry. The differentiation status of peripheral αß and γδ T cells was assessed by flow cytometry based on the surface expression of CD27, CD28 and CD45RA. Our data confirm the established associations of latent cytomegalovirus (CMV) infection with an accumulation of late differentiated memory T cells in the αß compartment as well as in the Vδ1+ γδ T cell subset. A comparison of differentiation phenotypes of γδ and αß T cells that were not affected by CMV seropositivity identified a significant correlation of early differentiated (ED) Vδ2+ and intermediate differentiated (ID) CD4+ T cells in monozygotic (MZ), but not in dizygotic (DZ) co-twins. Thus, our data suggest a genetic influence on the differentiation of γδ and αß T cell subsets.

Clonal hematopoiesis in elderly twins: concordance, discordance, and mortality.

Clonal hematopoiesis (CH) of indeterminate potential (CHIP) is defined by mutations in myeloid cancer-associated genes with a variant allele frequency of at least 2%. Recent studies have suggested a possible genetic predisposition to CH. To further explore this phenomenon, we conducted a population-based study of 594 twins from 299 pairs aged 73 to 94 years, all with >20 years' follow-up. We sequenced DNA from peripheral blood with a customized 21-gene panel at a median coverage of 6179X. The casewise concordance rates for mutations were calculated to assess genetic predisposition. Mutations were identified in 214 (36%) of the twins. Whereas 20 twin pairs had mutations within the same genes, the exact same mutation was only observed in 2 twin pairs. No significant difference in casewise concordance between monozygotic and dizygotic twins was found for any specific gene, subgroup, or CHIP mutations overall, and no significant heritability could be detected. In pairs discordant for CHIP mutations, we tested if the affected twin died before the unaffected twin, as a direct measurement of the association of having CH when controlling for familial factors. A total of 127 twin pairs were discordant for carrying a mutation, and in 61 (48%) cases, the affected twin died first (P = .72). Overall, we did not find a genetic predisposition to CHIP mutations in this twin study. The previously described negative association of CHIP mutations on survival could not be confirmed in a direct comparison among twin pairs that were discordant for CHIP mutations.

The Danish Twin Registry: An Updated Overview.

The Danish Twin Registry (DTR) was established in the 1950s, when twins born from 1870 to 1910 were ascertained, and has since been extended to include twins from birth cohorts until 2009. The DTR currently comprises of more than 175,000 twins from the 140 birth cohorts. This makes the DTR the oldest nationwide twin register and among the largest in the world. The combination of data from several surveys, including biological samples and repeated measurements on the same individuals, and data from Danish national registers provides a unique resource for a wide range of twin studies. This article provides an updated overview of the data in the DTR: First, we provide a summary of the establishment of the register, the different ascertainment methods and the twins included; then follows an overview of major surveys conducted in the DTR since 1994 and a description of the DTR biobank, including a description of the molecular data created so far; finally, a short description is given of the linkage to Danish national registers at Statistics Denmark and some recent examples of studies using the various data resources in the DTR are highlighted.

Sex differences in the 1-year risk of dying following all-cause and cause-specific hospital admission after age 50 in comparison with a general and non-hospitalised population: a register-based cohort study of the Danish population.

OBJECTIVES: We examine the mortality of men and women within the first year after all-cause and cause-specific hospital admission to investigate whether the sex differences in mortality after hospitalisation are higher than in the corresponding general and non-hospitalised population. DESIGN: This is a population-based, longitudinal study with nationwide coverage. The study population was identified by linking the National Patient Register with the Central Population Register using a 5% random sample of the Danish population. SETTING: The population born between 1898 and 1961, who was alive and residing in Denmark after 1977, was followed up between 1977 and 2011 with respect to hospital admissions and mortality while aged 50-79. PRIMARY OUTCOME MEASURES: The absolute sex differences in the 1-year risk of dying after all-cause and cause-specific hospital admission. The hospitalised population sex differentials were then compared with the sex differences in a general and a non-hospitalised population, randomly matched by age, sex and hospitalisation status. RESULTS: The risk of dying was consistently higher for hospitalised men and women. At all ages, the absolute sex differences in mortality were largest in the hospitalised population, were smaller in the general population and were smallest in the non-hospitalised population. This pattern was consistent across all-cause admissions, and with respect to admissions for neoplasms, circulatory diseases and respiratory diseases. For all-cause hospital admissions, absolute sex differences in the 1-year risk of dying resulted in 43.8 excess male deaths per 1,000 individuals within the age range 50-79, while the levels were lower in the general and the non-hospitalised population, at levels of 13.5 and 6.6, respectively. CONCLUSIONS: This study indicates a larger male disadvantage in mortality following hospitalisation, pointing towards an association between the health status of a population and the magnitude of the female advantage in mortality.

Comparison of Late Mortality Among Twins Versus Singletons With Congenital Heart Defects.

In 2014, in the United States, nearly 7% of newborns were twins. Congenital heart defects (CHDs) are more frequent in both monozygotic and dizygotic twins than in singletons. Still, the longer-term prognosis for CHD twins is unknown. Here we assess the mortality pattern for CHD twins up to age 36 years and compare it with that for non-CHD twins, non-CHD co-twins, and CHD singletons. We identified all twins and a 5% random sample of all singletons born in Denmark from 1977 to 2009 by linking Danish national population and health registers. CHD cases were defined as subjects having a primary inpatient diagnosis of CHD (excluding preterm ductus) within the first year of life, and mortality was assessed through 2013. Among 63,362 live-born twin individuals, a total of 373 twins (0.59%) had a CHD diagnosis, whereas the corresponding numbers for singletons were 383 of 98,647 (0.39%). During the follow-up, 82 (22.0%) CHD twins died compared with 91 (23.8%) CHD singletons (p = 0.56). Despite a 5 times higher proportion of prematurity, CHD twins had a tendency toward only a moderately increased neonatal mortality compared with CHD singletons (hazard ratio 1.5, 95% confidence interval 0.94 to 2.5), and after the neonatal period up to age 36 the tendency was reversed (hazard ratio 0.8, 95% confidence interval 0.5 to 1.2). A potential underlying mechanism for this mortality pattern is selective intrauterine and neonatal mortality of twins with the most severe CHD. In conclusion, the study indicates that the overall survival prognosis for CHD twins is similar to that of CHD singletons.

Early-life mortality risks in opposite-sex and same-sex twins: a Danish cohort study of the twin testosterone transfer hypothesis.

PURPOSE: To investigate the twin testosterone transfer (TTT) hypothesis by comparing early-life mortality risks of opposite-sex (OS) and same-sex (SS) twins during the first 15 years of life. METHODS: We performed a population-based cohort study to compare mortality in OS and SS twins. We included 68,629 live-born Danish twins from 1973 to 2009 identified through the Danish Twin Registry and performed piecewise stratified Cox regression and log-binomial regression. RESULTS: Among 1933 deaths, we found significantly higher mortality for twin boys than for twin girls. For both sexes, OS twins had lower mortality than SS twins; the difference persisted for the first year of life for boys and for the first week of life for girls. CONCLUSIONS: Although the mortality risk for OS boys was in the expected direction according to the TTT hypothesis, the results for OS girls pointed in the opposite direction, providing no clear evidence for the TTT hypothesis.

Mitral valve regurgitation in twins: Concordance and survival.

BACKGROUND: Smaller observational studies have suggested familial clustering of mitral regurgitation (MR). Using a large twin cohort, the aims were to assess MR concordance rates and assess mortality in MR twins and unaffected cotwins. METHODS: Through the Danish Twin Registry, twins with an International Classification of Diseases, Eighth Revision and Tenth Revision diagnosis code of MR born 1880-1989 were identified and proband-wise concordance rates were calculated. To assess whether having a cotwin with MR affected survival, 10 matched twins without MR (n = 5,575) were selected for each MR twin (n = 562), and all-cause mortality rates were assessed. RESULTS: Among the 87,432 twins alive January 1, 1977, or later, 494 (0.57%) MR individuals were identified. Six MR concordant pairs were found, of which 3 were monozygotic. Proband-wise concordance rate when accounting for right censoring and competing risk of death was 0.12 (95% CI 0.04-0.32) for monozygotic twins and 0.03 (95% CI 0.01-0.09) for dizygotic twins. Mortality was significantly higher among the affected twins with MR within discordant twin pairs where both were alive at the date of MR diagnosis (sex-adjusted hazard ratio [HR] 2.57, 95% CI 1.86-3.54). Overall there was no increased mortality risk for unaffected cotwins to MR cases compared with matched controls (HR 1.02, 95% CI 0.90-1.17) except for first year of life (HR 1.92, 95% CI 1.10-3.36) and for monozygotic twins older than 65 years (HR 1.49, 95% CI 1.07-2.08). CONCLUSION: Although there is a familial aggregation of MR, the absolute risk is low, even for monozygotic cotwins to affected twins. The study found increased mortality in MR twins compared with their unaffected cotwins. Overall no excess mortality was observed in the unaffected cotwins except for first year of life and for monozygotic cotwins older than 65 years.

Genetic Influence on the Peripheral Blood CD4+ T-cell Differentiation Status in CMV Infection.

A latent infection with cytomegalovirus (CMV), a ubiquitous beta herpesvirus, is associated with an accumulation of late-differentiated memory T-cells, often accompanied by a reciprocal reduced frequency of early-differentiated cells (commonly also referred to as "naïve"). However, this impact of CMV on T-cell phenotypes is variable between individuals. Our previous findings in a subgroup of participants in the Leiden familial Longevity Study indicated an important role of genetics. For further testing, we have analyzed middle-aged monozygotic (MZ, n = 42) and dizygotic (DZ, n = 39) twin pairs from the Danish Twin Registry for their T-cell differentiation status, assessed by surface expression of CD27, CD28, CD57, and KLRG-1. We observed a significant intraclass correlation between cotwins of MZ, but not DZ pairs for the differentiation status of CD4+ and CD8+ subsets. Classical heritability analysis confirmed a substantial contribution of genetics to the differentiation status of T-cells in CMV infection. The humoral (IgG) response to different CMV antigens also seems to be genetically influenced, suggesting that a similar degree of immune control of the virus in MZ twins might be responsible for their similar T-cell differentiation status. Thus, the way T-cells differentiate in the face of a latent CMV infection, and the parallel humoral responses, both controlling the virus, are genetically influenced.

Mortality is Written on the Face.

BACKGROUND: It is unknown whether facial or surrounding (eg, hair and clothing) cues have the strongest influence on the perceived age of subjects in photographic images, and which drives links between perceived age and survival. METHODS: In 2001, 187 Danish twin pairs (n = 374) aged 70+ years were photographed generating passport-type images. The faces of the twins in these images were swapped creating two new images per twin pair (748 images in total). Ten nurses rated the perceived age of the twin from the original and swapped facial images. The survival of the twins was determined through to the end of 2013. RESULTS: Changing the face or its surrounding significantly changed the perceived age of the images, with only a marginal difference between their effect sizes (difference of 0.5 years, 95% confidence interval CI -0.1 to 1.1). Perceived age, adjusting for chronological age, and sex, was a predictor of survival up to 7 years (hazard ratio 1.17, 95% CI 1.10-1.25) and also 7-12 years (hazard ratio 1.06, 95% CI 1.00-1.12) after the photographs were taken. Where the older looking twin died first they had a significantly older looking face (1.4 years older, 95% CI 0.3-2.6) but not surrounding (0.3 years older, 95% CI -0.8 to 1.4) compared to where the older looking twin died second. CONCLUSIONS: Facial visual cues but not hair or clothing cues drive the link between perceived age and survival.

Genetic and environmental influence on DNA strand break repair: a twin study.

Accumulation of DNA damage deriving from exogenous and endogenous sources has significant consequences for cellular survival, and is implicated in aging, cancer, and neurological diseases. Different DNA repair pathways have evolved in order to maintain genomic stability. Genetic and environmental factors are likely to influence DNA repair capacity. In order to gain more insight into the genetic and environmental contribution to the molecular basis of DNA repair, we have performed a human twin study, where we focused on the consequences of some of the most abundant types of DNA damage (single-strand breaks), and some of the most hazardous lesions (DNA double-strand breaks). DNA damage signaling response (Gamma-H2AX signaling), relative amount of endogenous damage, and DNA-strand break repair capacities were studied in peripheral blood mononuclear cells from 198 twins (94 monozygotic and 104 dizygotic). We did not detect genetic effects on the DNA-strand break variables in our study.