Epigenetic characterization of the FMR1 promoter in induced pluripotent stem cells from human fibroblasts carrying an unmethylated full mutation

Silencing of the FMR1 gene leads to fragile X syndrome, the most common cause of inherited intellectual disability. To study the epigenetic modifications of the FMR1 gene during silencing in time, we used fibroblasts and induced pluripotent stem cells (iPSCs) of an unmethylated full mutation (uFM) individual with normal intelligence. The uFM fibroblast line carried an unmethylated FMR1 promoter region and expressed normal to slightly increased FMR1 mRNA levels. The FMR1 expression in the uFM line corresponds with the increased H3 acetylation and H3K4 methylation in combination with a reduced H3K9 methylation. After reprogramming, the FMR1 promoter region was methylated in all uFM iPSC clones. Two clones were analyzed further and showed a lack of FMR1 expression, whereas the presence of specific histone modifications also indicated a repressed FMR1 promoter. In conclusion, these findings demonstrate that the standard reprogramming procedure leads to epigenetic silencing of the fully mutated FMR1 gene

Radiographic rib fracture nonunion and association with fracture classification in adults with multiple rib fractures without flail segment:A multicenter prospective cohort study

Background: Rib fracture nonunion is a probable cause of chronic pain following chest trauma, although its prevalence remains unknown. The aims of this study were to determine rib fracture nonunion prevalence following nonoperative management and to determine if presence of nonunion was associated with the number of rib fractures, or the rib fracture classification of anatomical location, type, and displacement. Methods: This multicenter prospective cohort study included trauma patients with three or more fractured ribs but without a flail segment, who participated in the nonoperative management group of the FixCon trial between January 2019 and June 2022. The number and classification of rib fractures were assessed on trauma chest CT. Chest CTs conducted six months post-trauma were evaluated for the presence of nonunion. Radiological characteristics of nonunions were compared with normally healed rib fractures using the Mann-Whitney U, χ2 test, and Fisher's exact test as appropriate. A generalized linear model adjusted for multiple observations per patient when assessing the associations between nonunion and fracture characteristics. Results: A total of 68 patients were included with 561 post-traumatic fractures in 429 ribs. Chest CT after six months revealed nonunions in 67 (12 %) rib fractures in 29 (43 %) patients with a median of 2 (P25-P75 1–3) nonunions per patient. Nonunion was most commonly observed in ribs seven to 10 (20–23 %, p &lt; 0.001, adjusted p = 0.006). Nonunion occurred in 14 (5 %) undisplaced, 22 (19 %) offset, and 20 (23 %) displaced rib fractures (p &lt; 0.001). No statistically significant association between rib fracture type and nonunion was found. Conclusions: Forty-three percent of patients with multiple rib fractures had radiographic nonunion six months after trauma. Fractures in ribs seven to 10 and dislocated fractures had an increased risk of rib fracture nonunion.</p

High prevalence of non-accidental trauma among deceased children presenting at Level I trauma centers in the Netherlands

PURPOSE: Between 0.1—3% of injured children who present at a hospital emergency department ultimately die as a result of their injuries. These events are typically reported as unnatural causes of death and may result from either accidental or non-accidental trauma (NAT). Examples of the latter include trauma that is inflicted directly or resulting from neglect. Although consultation with a forensic physician is mandatory for all deceased children, the prevalence of fatal inflicted trauma or neglect among children is currently unclear. METHODS: This is a retrospective study that included children (0–18 years) who presented and died at one of the 11 Level I trauma centers in the Netherlands between January 1, 2014, and January 1, 2019. Outcomes were classified based on the conclusions of the Child Abuse and Neglect team or those of forensic pathologists and/or the court in cases referred for legally mandated autopsies. Cases in which conclusions were unavailable and there was no clear accidental cause of death were reviewed by an expert panel. RESULTS: The study included 175 cases of childhood death. Seventeen (9.7%) of these children died due to inflicted trauma (9.7%), 18 (10.3%) due to neglect, and 140 (80%) due to accidents. Preschool children (< 5 years old) were significantly more likely to present with injuries due to inflicted trauma and neglect compared to older children (44% versus 6%, p < 0.001, odds ratio [OR] 5.80, 95% confidence interval [CI] 2.66–12.65). Drowning accounted for 14 of the 18 (78%) pediatric deaths due to neglect, representing 8% of the total cases. Postmortem radiological studies and autopsies were performed on 37 (21%) of all cases of childhood death. CONCLUSION: One of every five pediatric deaths in our nationwide Level I trauma center study was attributed to NAT; 44% of these deaths were the result of trauma experienced by preschool-aged children. A remarkable number of fatal drownings were due to neglect. Postmortem radiological studies and autopsies were performed in only one-fifth of all deceased children. The limited use of postmortem investigations may have resulted in missed cases of NAT, which will result in an overall underestimation of fatal NAT experienced by children. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12024-021-00416-7

Radiographic rib fracture nonunion and association with fracture classification in adults with multiple rib fractures without flail segment:A multicenter prospective cohort study

Background: Rib fracture nonunion is a probable cause of chronic pain following chest trauma, although its prevalence remains unknown. The aims of this study were to determine rib fracture nonunion prevalence following nonoperative management and to determine if presence of nonunion was associated with the number of rib fractures, or the rib fracture classification of anatomical location, type, and displacement. Methods: This multicenter prospective cohort study included trauma patients with three or more fractured ribs but without a flail segment, who participated in the nonoperative management group of the FixCon trial between January 2019 and June 2022. The number and classification of rib fractures were assessed on trauma chest CT. Chest CTs conducted six months post-trauma were evaluated for the presence of nonunion. Radiological characteristics of nonunions were compared with normally healed rib fractures using the Mann-Whitney U, χ2 test, and Fisher's exact test as appropriate. A generalized linear model adjusted for multiple observations per patient when assessing the associations between nonunion and fracture characteristics. Results: A total of 68 patients were included with 561 post-traumatic fractures in 429 ribs. Chest CT after six months revealed nonunions in 67 (12 %) rib fractures in 29 (43 %) patients with a median of 2 (P25-P75 1–3) nonunions per patient. Nonunion was most commonly observed in ribs seven to 10 (20–23 %, p &lt; 0.001, adjusted p = 0.006). Nonunion occurred in 14 (5 %) undisplaced, 22 (19 %) offset, and 20 (23 %) displaced rib fractures (p &lt; 0.001). No statistically significant association between rib fracture type and nonunion was found. Conclusions: Forty-three percent of patients with multiple rib fractures had radiographic nonunion six months after trauma. Fractures in ribs seven to 10 and dislocated fractures had an increased risk of rib fracture nonunion.</p

Genetic association study of QT interval highlights role for calcium signaling pathways in myocardial repolarization.

The QT interval, an electrocardiographic measure reflecting myocardial repolarization, is a heritable trait. QT prolongation is a risk factor for ventricular arrhythmias and sudden cardiac death (SCD) and could indicate the presence of the potentially lethal mendelian long-QT syndrome (LQTS). Using a genome-wide association and replication study in up to 100,000 individuals, we identified 35 common variant loci associated with QT interval that collectively explain ∼8-10% of QT-interval variation and highlight the importance of calcium regulation in myocardial repolarization. Rare variant analysis of 6 new QT interval-associated loci in 298 unrelated probands with LQTS identified coding variants not found in controls but of uncertain causality and therefore requiring validation. Several newly identified loci encode proteins that physically interact with other recognized repolarization proteins. Our integration of common variant association, expression and orthogonal protein-protein interaction screens provides new insights into cardiac electrophysiology and identifies new candidate genes for ventricular arrhythmias, LQTS and SCD

Hundreds of variants clustered in genomic loci and biological pathways affect human height

Most common human traits and diseases have a polygenic pattern of inheritance: DNA sequence variants at many genetic loci influence the phenotype. Genome-wide association (GWA) studies have identified more than 600 variants associated with human traits, but these typically explain small fractions of phenotypic variation, raising questions about the use of further studies. Here, using 183,727 individuals, we show that hundreds of genetic variants, in at least 180 loci, influence adult height, a highly heritable and classic polygenic trait. The large number of loci reveals patterns with important implications for genetic studies of common human diseases and traits. First, the 180 loci are not random, but instead are enriched for genes that are connected in biological pathways (P = 0.016) and that underlie skeletal growth defects (P < 0.001). Second, the likely causal gene is often located near the most strongly associated variant: in 13 of 21 loci containing a known skeletal growth gene, that gene was closest to the associated variant. Third, at least 19 loci have multiple independently associated variants, suggesting that allelic heterogeneity is a frequent feature of polygenic traits, that comprehensive explorations of already-discovered loci should discover additional variants and that an appreciable fraction of associated loci may have been identified. Fourth, associated variants are enriched for likely functional effects on genes, being over-represented among variants that alter amino-acid structure of proteins and expression levels of nearby genes. Our data explain approximately 10% of the phenotypic variation in height, and we estimate that unidentified common variants of similar effect sizes would increase this figure to approximately 16% of phenotypic variation (approximately 20% of heritable variation). Although additional approaches are needed to dissect the genetic architecture of polygenic human traits fully, our findings indicate that GWA studies can identify large numbers of loci that implicate biologically relevant genes and pathways.

New genetic loci implicated in fasting glucose homeostasis and their impact on type 2 diabetes risk.

Levels of circulating glucose are tightly regulated. To identify new loci influencing glycemic traits, we performed meta-analyses of 21 genome-wide association studies informative for fasting glucose, fasting insulin and indices of beta-cell function (HOMA-B) and insulin resistance (HOMA-IR) in up to 46,186 nondiabetic participants. Follow-up of 25 loci in up to 76,558 additional subjects identified 16 loci associated with fasting glucose and HOMA-B and two loci associated with fasting insulin and HOMA-IR. These include nine loci newly associated with fasting glucose (in or near ADCY5, MADD, ADRA2A, CRY2, FADS1, GLIS3, SLC2A2, PROX1 and C2CD4B) and one influencing fasting insulin and HOMA-IR (near IGF1). We also demonstrated association of ADCY5, PROX1, GCK, GCKR and DGKB-TMEM195 with type 2 diabetes. Within these loci, likely biological candidate genes influence signal transduction, cell proliferation, development, glucose-sensing and circadian regulation. Our results demonstrate that genetic studies of glycemic traits can identify type 2 diabetes risk loci, as well as loci containing gene variants that are associated with a modest elevation in glucose levels but are not associated with overt diabetes

Parent-of-origin-specific allelic associations among 106 genomic loci for age at menarche.

Age at menarche is a marker of timing of puberty in females. It varies widely between individuals, is a heritable trait and is associated with risks for obesity, type 2 diabetes, cardiovascular disease, breast cancer and all-cause mortality. Studies of rare human disorders of puberty and animal models point to a complex hypothalamic-pituitary-hormonal regulation, but the mechanisms that determine pubertal timing and underlie its links to disease risk remain unclear. Here, using genome-wide and custom-genotyping arrays in up to 182,416 women of European descent from 57 studies, we found robust evidence (P < 5 × 10(-8)) for 123 signals at 106 genomic loci associated with age at menarche. Many loci were associated with other pubertal traits in both sexes, and there was substantial overlap with genes implicated in body mass index and various diseases, including rare disorders of puberty. Menarche signals were enriched in imprinted regions, with three loci (DLK1-WDR25, MKRN3-MAGEL2 and KCNK9) demonstrating parent-of-origin-specific associations concordant with known parental expression patterns. Pathway analyses implicated nuclear hormone receptors, particularly retinoic acid and γ-aminobutyric acid-B2 receptor signalling, among novel mechanisms that regulate pubertal timing in humans. Our findings suggest a genetic architecture involving at least hundreds of common variants in the coordinated timing of the pubertal transition

Increasing tumoral 5-fluorouracil concentrations during a 5-day continuous infusion: a microdialysis study

Purpose: Response to anticancer therapy is believed to be directly related to the concentration of the anticancer drug in the tumor itself. Assessment of intra-tumor drug pharmacokinetics can be helpful to gain more insight into mechanisms involved in the (in)sensitivity of tumors to anticancer therapy. We explored the pharmacokinetics of 5-fluorouracil in both plasma and tumor tissue during a 5-day continuous infusion of 5-fluorouracil in patients with cancer. Sampling for measurement of 5-fluorouracil in tumor tissue was performed using microdialysis. Experimental design: In seven patients with an accessible (sub)cutaneous tumor treated with a continuous 5-fluorouracil infusion, plasma and microdialysate samples from tumor and normal adipose tissue were collected over a period of 5 days. Results: For six patients, drug concentrations in both tumor tissue and plasma were available. Concentration-time curves of unbound 5-fluorouracil were lower in tumor tissue compared to the curves in plasma, but exposure ratios of tumor tissue versus plasma increased during the 5-day infusion period. The presence of circadian rhythmicity of 5-fluorouracil pharmacokinetics in the tumor itself was demonstrated as 5-fluorouracil concentrations in tumor extracellular fluid were higher during the night than during daytime. Conclusion: Microdialysis was successfully employed in patients with cancer during a continuous 5-day 5-fluorouracil infusion. Plasma and tumor pharmacokinetics of 5-fluorouracil differed substantially with increasing 5-fluorouracil concentrations in tumor over time, possibly resulting from a lowered interstitial fluid pressure by 5-fluorouracil itself. This microdialysis 5-fluorouracil model might be useful to monitor the effect of drug delivery modulating strategies in future studies

Early fixation versus conservative therapy of multiple, simple rib fractures (FixCon): protocol for a multicenter randomized controlled trial

Background: Multiple rib fractures are common injuries in both the young and elderly. Rib fractures account for 10% of all trauma admissions and are seen in up to 39% of patients after thoracic trauma. With morbidity and mortality rates increasing with the number of rib fractures as well as poor quality of life at long-term follow-up, multiple rib fractures pose a serious health hazard. Operative fixation of flail chest is beneficial over nonoperative treatment regarding, among others, pneumonia and both intensive care unit (ICU) and hospital length of stay. With no high-quality evidence on the effects of multiple simple rib fracture treatment, the optimal treatment modality remains unknown. This study sets out to investigate outcome of operative fixation versus nonoperative treatment of multiple simple rib fractures. Methods: The proposed study is a multicenter randomized controlled trial. Patients will be eligible if they have three or more multiple simple rib fractures of which at least one is disl