Evaluation of automated techniques for extraction of circulating cell-free DNA for implementation in standardized high-throughput workflows

AbstractAnalysis of circulating cell-free DNA (ccfDNA) is a suitable tool for detecting somatic mutations for the purpose of making decisions on treatment, monitoring treatment response, and predicting survival. High-throughput techniques for ccfDNA extraction are essential to implementing ccfDNA testing in the clinical setting. We set out to compare two automated techniques with regard to hands-on time, ccfDNA output and integrity, and circulating mitochondrial DNA (mtDNA). CcfDNA was isolated using the EZ1&amp;2 ccfDNA field test kit (EZ2 kit, QIAGEN) and the Maxwell RSC ccfDNA plasma kit (Maxwell kit, Promega). DNA was extracted from plasma of 30 breast cancer patients enrolled in the iMODE-B (#325_19B; 12.10.2020) study. Real-time PCR, fluorescence-based detection and automated electrophoresis were used to assess ccfDNA concentrations. The ccfDNA yield was significantly higher when extracted with the EZ2 kit. The EZ2 kit enabled the isolation of a higher proportion of short fragments and a lower proportion of long fragments, resulting in lower DNA integrity. Significantly lower mtDNA quantities were detected in the Maxwell eluate than in the EZ2 eluate. Thus, decisions on which extraction method to use should proceed on the basis of the required input for downstream applications, the anticipated fragment size and minimum hands-on time.</jats:p

Human TGF-β1 deficiency causes severe inflammatory bowel disease and encephalopathy

Transforming growth factor (TGF)-β1 (encoded by TGFB1) is the prototypic member of the TGF-β family of 33 proteins that orchestrate embryogenesis, development and tissue homeostasis1,2. Following its discovery3, enormous interest and numerous controversies have emerged about the role of TGF-β in coordinating the balance of pro- and anti-oncogenic properties4,5, pro- and anti-inflammatory effects6, or pro- and anti-fibrinogenic characteristics7. Here we describe three individuals from two pedigrees with biallelic loss-of-function mutations in the TGFB1 gene who presented with severe infantile inflammatory bowel disease (IBD) and central nervous system (CNS) disease associated with epilepsy, brain atrophy and posterior leukoencephalopathy. The proteins encoded by the mutated TGFB1 alleles were characterized by impaired secretion, function or stability of the TGF-β1–LAP complex, which is suggestive of perturbed bioavailability of TGF-β1. Our study shows that TGF-β1 has a critical and nonredundant role in the development and homeostasis of intestinal immunity and the CNS in humans

Cognitive-Motor Interference During Functional Mobility After Stroke: State of the Science and Implications for Future Research

Cognitive-motor interference (CMI) is evident when simultaneous performance of a cognitive task and a motor task results in deterioration in performance in one or both of the tasks, relative to performance of each task separately. The purpose of this review is to present a framework for categorizing patterns of CMI and to examine the specific patterns of CMI evident in published studies comparing single-task and dual-task performance of cognitive and motor tasks during gait and balance activities after stroke. We also examine the literature for associations between patterns of CMI and history of falls, as well as evidence for the effects of rehabilitation on CMI after stroke. Overall, this review suggests that during gait activities with an added cognitive task, people with stroke are likely to demonstrate significant decrements in motor performance only (cognitive-related motor interference) or decrements in both motor and cognitive performance (mutual interference). In contrast, patterns of CMI were variable among studies examining balance activities. Comparing people post-stroke with and without a history of falls, patterns and magnitude of CMI were similar for fallers and non-fallers. Longitudinal studies suggest that conventional rehabilitation has minimal effects on CMI during gait or balance activities. However, early phase pilot studies suggest that dual-task interventions may reduce CMI during gait performance in community-dwelling stroke survivors. It is our hope that this innovative and critical examination of the existing literature will highlight the limitations in current experimental designs and inform improvements in the design and reporting of dual-task studies in stroke