Systematic evaluation of genome sequencing for the diagnostic assessment of autism spectrum disorder and fetal structural anomalies.

Short-read genome sequencing (GS) holds the promise of becoming the primary diagnostic approach for the assessment of autism spectrum disorder (ASD) and fetal structural anomalies (FSAs). However, few studies have comprehensively evaluated its performance against current standard-of-care diagnostic tests: karyotype, chromosomal microarray (CMA), and exome sequencing (ES). To assess the clinical utility of GS, we compared its diagnostic yield against these three tests in 1,612 quartet families including an individual with ASD and in 295 prenatal families. Our GS analytic framework identified a diagnostic variant in 7.8% of ASD probands, almost 2-fold more than CMA (4.3%) and 3-fold more than ES (2.7%). However, when we systematically captured copy-number variants (CNVs) from the exome data, the diagnostic yield of ES (7.4%) was brought much closer to, but did not surpass, GS. Similarly, we estimated that GS could achieve an overall diagnostic yield of 46.1% in unselected FSAs, representing a 17.2% increased yield over karyotype, 14.1% over CMA, and 4.1% over ES with CNV calling or 36.1% increase without CNV discovery. Overall, GS provided an added diagnostic yield of 0.4% and 0.8% beyond the combination of all three standard-of-care tests in ASD and FSAs, respectively. This corresponded to nine GS unique diagnostic variants, including sequence variants in exons not captured by ES, structural variants (SVs) inaccessible to existing standard-of-care tests, and SVs where the resolution of GS changed variant classification. Overall, this large-scale evaluation demonstrated that GS significantly outperforms each individual standard-of-care test while also outperforming the combination of all three tests, thus warranting consideration as the first-tier diagnostic approach for the assessment of ASD and FSAs.

Transcriptional and functional consequences of alterations to MEF2C and its topological organization in neuronal models.

Point mutations and structural variants that directly disrupt the coding sequence of MEF2C have been associated with a spectrum of neurodevelopmental disorders (NDDs). However, the impact of MEF2C haploinsufficiency on neurodevelopmental pathways and synaptic processes is not well understood, nor are the complex mechanisms that govern its regulation. To explore the functional changes associated with structural variants that alter MEF2C expression and/or regulation, we generated an allelic series of 204 isogenic human induced pluripotent stem cell (hiPSC)-derived neural stem cells and glutamatergic induced neurons. These neuronal models harbored CRISPR-engineered mutations that involved direct deletion of MEF2C or deletion of the boundary points for topologically associating domains (TADs) and chromatin loops encompassing MEF2C. Systematic profiling of mutation-specific alterations, contrasted to unedited controls that were exposed to the same guide RNAs for each edit, revealed that deletion of MEF2C caused differential expression of genes associated with neurodevelopmental pathways and synaptic function. We also discovered significant reduction in synaptic activity measured by multielectrode arrays (MEAs) in neuronal cells. By contrast, we observed robust buffering against MEF2C regulatory disruption following deletion of a distal 5q14.3 TAD and loop boundary, whereas homozygous loss of a proximal loop boundary resulted in down-regulation of MEF2C expression and reduced electrophysiological activity on MEA that was comparable to direct gene disruption. Collectively, these studies highlight the considerable functional impact of MEF2C deletion in neuronal cells and systematically characterize the complex interactions that challenge a priori predictions of regulatory consequences from structural variants that disrupt three-dimensional genome organization.

Tissue- and cell-type-specific molecular and functional signatures of 16p11.2 reciprocal genomic disorder across mouse brain and human neuronal models.

Chromosome 16p11.2 reciprocal genomic disorder, resulting from recurrent copy-number variants (CNVs), involves intellectual disability, autism spectrum disorder (ASD), and schizophrenia, but the responsible mechanisms are not known. To systemically dissect molecular effects, we performed transcriptome profiling of 350 libraries from six tissues (cortex, cerebellum, striatum, liver, brown fat, and white fat) in mouse models harboring CNVs of the syntenic 7qF3 region, as well as cellular, transcriptional, and single-cell analyses in 54 isogenic neural stem cell, induced neuron, and cerebral organoid models of CRISPR-engineered 16p11.2 CNVs. Transcriptome-wide differentially expressed genes were largely tissue-, cell-type-, and dosage-specific, although more effects were shared between deletion and duplication and across tissue than expected by chance. The broadest effects were observed in the cerebellum (2,163 differentially expressed genes), and the greatest enrichments were associated with synaptic pathways in mouse cerebellum and human induced neurons. Pathway and co-expression analyses identified energy and RNA metabolism as shared processes and enrichment for ASD-associated, loss-of-function constraint, and fragile X messenger ribonucleoprotein target gene sets. Intriguingly, reciprocal 16p11.2 dosage changes resulted in consistent decrements in neurite and electrophysiological features, and single-cell profiling of organoids showed reciprocal alterations to the proportions of excitatory and inhibitory GABAergic neurons. Changes both in neuronal ratios and in gene expression in our organoid analyses point most directly to calretinin GABAergic inhibitory neurons and the excitatory/inhibitory balance as targets of disruption that might contribute to changes in neurodevelopmental and cognitive function in 16p11.2 carriers. Collectively, our data indicate the genomic disorder involves disruption of multiple contributing biological processes and that this disruption has relative impacts that are context specific.

Dystonia-specific mutations in THAP1 alter transcription of genes associated with neurodevelopment and myelin.

Dystonia is a neurologic disorder associated with an increasingly large number of genetic variants in many genes, resulting in characteristic disturbances in volitional movement. Dissecting the relationships between these mutations and their functional outcomes is critical in understanding the pathways that drive dystonia pathogenesis. Here we established a pipeline for characterizing an allelic series of dystonia-specific mutations. We used this strategy to investigate the molecular consequences of genetic variation in THAP1, which encodes a transcription factor linked to neural differentiation. Multiple pathogenic mutations associated with dystonia cluster within distinct THAP1 functional domains and are predicted to alter DNA-binding properties and/or protein interactions differently, yet the relative impact of these varied changes on molecular signatures and neural deficits is unclear. To determine the effects of these mutations on THAP1 transcriptional activity, we engineered an allelic series of eight alterations in a common induced pluripotent stem cell background and differentiated these lines into a panel of near-isogenic neural stem cells (n = 94 lines). Transcriptome profiling followed by joint analysis of the most robust signatures across mutations identified a convergent pattern of dysregulated genes functionally related to neurodevelopment, lysosomal lipid metabolism, and myelin. On the basis of these observations, we examined mice bearing Thap1-disruptive alleles and detected significant changes in myelin gene expression and reduction of myelin structural integrity relative to control mice. These results suggest that deficits in neurodevelopment and myelination are common consequences of dystonia-associated THAP1 mutations and highlight the potential role of neuron-glial interactions in the pathogenesis of dystonia.

Changes in pulmonary artery pressure affect survival differently in lung transplant recipients who have pulmonary hypertension or chronic obstructive pulmonary disease.

OBJECTIVES: Pulmonary hypertension and right ventricular dysfunction can complicate lung transplant. Pulmonary artery pressures affect outcome are uncertain during wait list. We evaluated changes in wait list pulmonary artery pressures on survival after lung transplant. MATERIALS AND METHODS: We queried the United Network for Organ Sharing/Standard Transplant Analysis and Research registry from 1987 to 2012 for all lung transplants. Recipients with unique pulmonary artery pressure measurements upon listing and transplant were included. Mean pulmonary artery pressure was rated as increased (increase > 5 mm Hg), decreased (decrease > 5 mm Hg), or unchanged (variation < 5 mm Hg). RESULTS: There were 23 951 lung transplants and 1677 recipients were included. Diagnoses demonstrated significant changes in mean pulmonary artery pressure during the listing period (P ≤ .0001). In recipients with chronic obstructive pulmonary disease, survival was poorer when mean pulmonary artery pressure was increased than decreased (P ≤ .03). In recipients with primary pulmonary hypertension, survival was poorer when mean pulmonary artery pressure was decreased than increased (P ≤ .02). Proportional hazards analysis showed that increases in mean pulmonary artery pressure independently affected survival (hazard ratio, 0.78; 95% confidence interval, 0.62-0.96). CONCLUSIONS: Although the mechanism is unknown, an increase in mean pulmonary artery pressure in patients with chronic obstructive pulmonary disease is associated with poorer survival after lung transplant. In contrast, patients with primary pulmonary hypertension with decreased mean pulmonary artery pressure have poorer survival after lung transplant. In patients with primary pulmonary hypertension, changes in pulmonary artery pressure may be a surrogate for a failing right ventricular function. In chronic obstructive pulmonary disease, the change in pressure suggests an undetermined progressive process. Further study of right ventricular function is warranted to determine the effects of changes in pulmonary artery pressure on lung transplant recipients.

Early catastrophic stentless valve failure secondary to possible immune reaction.

Aortic valve replacement with stentless xenografts has become routine since their introduction in the early 1990s. Although concerns of structural valve deterioration and long-term durability have been voiced, the reports on the causes or pathology associated with early valve failure have been sparse. We report two unusual cases of failure leading to patient death within the first year after implantation of the aortic valve and root with the Freestyle prostheses (Medtronic Inc, Minneapolis, MN). We suggest an early immunologic reaction to the xenograft, leading to a fatal inflammatory process, as a mechanism for unusual early valve failure in these patients.

Bariatric surgery outcomes in patients aged 65 years and older at an American Society for Metabolic and Bariatric Surgery Center of Excellence.

BACKGROUND: Although morbid obesity rates in patients >or=65 years of age are increasing, few centers have reported weight loss surgery outcomes in elderly patients, resulting in a paucity of literature on perioperative mortality and morbidity. METHODS: A retrospective analysis was performed on 197 consecutive patients >or=65 years old who underwent weight loss surgery from January 2000 to December 2007. Primary data points included 30-day and 1-year mortality rates, length of stay (LOS), percent excess weight loss (EWL), change in daily medication use, and quality of life (QOL). RESULTS: The average patient's age was 67.3 years with 72.1% being female. Average preoperative weight and BMI were 131.9 kg and 48.1 kg/m(2), respectively. Average preoperative daily medication use was 8.04 +/- 3.67. Procedure types included Roux-en-Y gastric bypass (79.3%), adjustable gastric banding (17.2%), and vertical sleeve gastrectomy (3%). Ninety-seven percent of procedures were performed laparoscopically. Average LOS was 2.0 +/- 2.1 days. Average weight, BMI, and daily medication use were significantly reduced at 6 months and 1 year (p < 0.001), with patients achieving an average EWL of 44.5% and 55.3% at 6 months and 1 year, respectively. QOL scores improved at 6 months (p < 0.001) and 1 year (p = 0.049). In all patients, the 30-day mortality rate was 0%. The 1-year mortality rate for RYGB patients was 1.3%. Complication rates were acceptable, with 7% of RYGB patients experiencing a major postoperative complication. CONCLUSIONS: Weight loss surgery is effective in patients >or=65 years of age, producing significant EWL, reduction in daily medication use, and improvement in QOL. Surgery is also associated with a low mortality rate and an acceptable morbidity profile.

Upper extremity kinematic trends of fly-casting: establishing the effects of line length.

Fly-fishing is a popular form of recreation. Recent evidence has associated overhand fly-casting movements with upper extremity pain. However, little research exists on the motions and coordination common to fly-casting. The aim of this study was to establish upper extremity kinematic trends of fly-casting while casting greater line lengths. It was hypothesized that kinematic casting parameters would increase and time between peak angular velocities would decrease with greater line length. Eighteen males participated in the study. Three-dimensional motion capture was conducted to calculate shoulder, elbow, and wrist kinematics during casting conditions of 6.1, 12.2, 18.3, and 24.4 m of line. Multiple analyses of variance were used to assess the condition effect of line length on the kinematic variables (P = 0.05). Overall, total range of movement increased with increasing length of line cast. Peak angular velocity exhibited a proximal-to-distal trend: peak shoulder internal rotation followed by elbow extension, then wrist ulnar deviation. Time between peak shoulder and elbow angular velocities increased significantly as line length increased. Our findings indicate that specific changes in total range of movement accommodate the demands of casting greater lengths of line. Also, joint velocity coordination patterns of fly-casting appear to follow a proximal-to-distal pattern. These findings represent an initial foundation for connections between kinematics and upper extremity pain reported by fly-fisherman.