Fixation and Spread of Somatic Mutations in Adult Human Colonic Epithelium

We investigated the means and timing by which mutations become fixed in the human colonic epithelium by visualizing somatic clones and mathematical inference. Fixation requires two sequential steps. First, one of approximately seven active stem cells residing within each colonic crypt has to be mutated. Second, the mutated stem cell has to replace neighbors to populate the entire crypt in a process that takes several years. Subsequent clonal expansion due to crypt fission is infrequent for neutral mutations (around 0.7% of all crypts undergo fission in a single year). Pro-oncogenic mutations subvert both stem cell replacement to accelerate fixation and clonal expansion by crypt fission to achieve high mutant allele frequencies with age. The benchmarking of these behaviors allows the advantage associated with different gene-specific mutations to be compared irrespective of the cellular mechanisms by which they are conferred

Antisense-induced exon skipping for duplications in Duchenne muscular dystrophy

<p>Abstract</p> <p>Background</p> <p>Antisense-mediated exon skipping is currently one of the most promising therapeutic approaches for Duchenne muscular dystrophy (DMD). Using antisense oligonucleotides (AONs) targeting specific exons the DMD reading frame is restored and partially functional dystrophins are produced. Following proof of concept in cultured muscle cells from patients with various deletions and point mutations, we now focus on single and multiple exon duplications. These mutations are in principle ideal targets for this approach since the specific skipping of duplicated exons would generate original, full-length transcripts.</p> <p>Methods</p> <p>Cultured muscle cells from DMD patients carrying duplications were transfected with AONs targeting the duplicated exons, and the dystrophin RNA and protein were analyzed.</p> <p>Results</p> <p>For two brothers with an exon 44 duplication, skipping was, even at suboptimal transfection conditions, so efficient that both exons 44 were skipped, thus generating, once more, an out-of-frame transcript. In such cases, one may resort to multi-exon skipping to restore the reading frame, as is shown here by inducing skipping of exon 43 and both exons 44. By contrast, in cells from a patient with an exon 45 duplication we were able to induce single exon 45 skipping, which allowed restoration of wild type dystrophin. The correction of a larger duplication (involving exons 52 to 62), by combinations of AONs targeting the outer exons, appeared problematic due to inefficient skipping and mistargeting of original instead of duplicated exons.</p> <p>Conclusion</p> <p>The correction of DMD duplications by exon skipping depends on the specific exons targeted. Its options vary from the ideal one, restoring for the first time the true, wild type dystrophin, to requiring more 'classical' skipping strategies, while the correction of multi-exon deletions may need the design of tailored approaches.</p

The role of multiple marks in epigenetic silencing and the emergence of a stable bivalent chromatin state

We introduce and analyze a minimal model of epigenetic silencing in budding yeast, built upon known biomolecular interactions in the system. Doing so, we identify the epigenetic marks essential for the bistability of epigenetic states. The model explicitly incorporates two key chromatin marks, namely H4K16 acetylation and H3K79 methylation, and explores whether the presence of multiple marks lead to a qualitatively different systems behavior. We find that having both modifications is important for the robustness of epigenetic silencing. Besides the silenced and transcriptionally active fate of chromatin, our model leads to a novel state with bivalent (i.e., both active and silencing) marks under certain perturbations (knock-out mutations, inhibition or enhancement of enzymatic activity). The bivalent state appears under several perturbations and is shown to result in patchy silencing. We also show that the titration effect, owing to a limited supply of silencing proteins, can result in counter-intuitive responses. The design principles of the silencing system is systematically investigated and disparate experimental observations are assessed within a single theoretical framework. Specifically, we discuss the behavior of Sir protein recruitment, spreading and stability of silenced regions in commonly-studied mutants (e.g., sas2, dot1) illuminating the controversial role of Dot1 in the systems biology of yeast silencing.Comment: Supplementary Material, 14 page

La Relación Entre la Motivación Docente y Variables de la Organización: Revisión de la Literatura

Abstract Teacher motivation plays a central role in education because ofitsimpacton student motivation. Previous reviews of teacher motivation have focused on individual variables and psychopathology indicators. However, it is also important to understand the effect of organizational variableson teacher motivationbecause these highlightthe contextthat the teacher is a part of(i.e.,the school). The literature review in this paper analysed studies related to teacher motivation and a pre-defined group of organizational variablesthat werepublished between 1990 and 2014 in several electronic databases.The study found that organizational culture was the most studied variable associated with teacher motivationand most studies in this area were published between 2010 and 2014.Further,there was a prevalence of quantitative studies. This paper concludes with the theoreticaland practical implications of the results,as well assuggestions for future research directions

Muscle biopsies in clinical trials for Duchenne muscular dystrophy - Patients' and caregivers' perspective

The number of clinical trials for Duchenne muscular dystrophy is increasing. Many trials require muscle biopsies, which involve an invasive surgical procedure. Little is known about short- and long-term impacts of muscle biopsies as perceived by patients and caregivers. Therefore a survey was held among patients and their caregivers who participated in trials involving muscle biopsies, in seven countries. Seventy-eight responses were received. Analysis revealed that many patients and parents had significant anxiety before the biopsy. The main concern of caregivers was the required general anaesthesia. In most cases biopsies caused pain and temporarily hampered daily activities. The main long-term impact was scarring, although large variation in size was reported. Seventy-nine percent of caregivers were little bothered and 21% were moderately or severely bothered by the scar. Willingness to consider another biopsy in future protocols was higher for open-label studies than for placebo-controlled trials. Caregivers stressed the importance of knowing the results of biopsy analyses; only a minority actually received this information. Recommendations are made on the informed consent procedure regarding risks and consequences of muscle biopsies, and communication of results. Furthermore, efforts should be made to minimise the impact of biopsies through pain management and by considering plastic surgery

Episiotomy and perineal trauma during childbirth in primiparous women: associations with anxiety, quality of life, vaginal and sexual symptoms in the first year postpartum

Introduction Childbirth-related pelvic floor trauma is prevalent among primiparous women and can lead to significant physical and psychological consequences. While the impact of pelvic floor trauma on physical outcomes has been studied, the relationship between anxiety caused by such trauma and long-term patient-reported outcomes (PROs) such as vaginal symptoms, sexual function, and quality of life (QoL) remains underexplored. This study aims to fill this gap by investigating the association between anxiety induced by pelvic floor trauma during childbirth and these key PROs. Methods This prospective longitudinal cohort study analyzed data from 175 nulliparous women who delivered at term a singleton fetus in cephalic presentation and sustained some form of perineal trauma. Anxiety levels were assessed at two time points: during labor and at 12 months postpartum, using a single-item 10-point Likert scale. The other PROs were measured using the International Consultation on Incontinence Questionnaire-Vaginal Symptoms tool (ICIQ-VS). Results Findings revealed that higher anxiety scores at birth were associated with elevated anxiety levels at 12 months postpartum and correlated significantly with increased vaginal symptoms, sexual symptoms, and QoL. Notably, while anxiety was linked to negative physical outcomes, higher anxiety scores were also associated with improved perceived QoL, suggesting the potential role of coping mechanisms in response to childbirth trauma as well as the need for future studies using more specialized anxiety tools. Conclusion The study underscores the intricate relationship between psychological distress and physical health outcomes in postpartum women. Addressing both anxiety and physical symptoms through personalized care strategies may enhance recovery and overall wellbeing. Future research should explore effective interventions to mitigate anxiety, evaluate resilience and improve PROs in this population

Dynamics of Co-Transcriptional Pre-mRNA Folding Influences the Induction of Dystrophin Exon Skipping by Antisense Oligonucleotides

Antisense oligonucleotides (AONs) mediated exon skipping offers potential therapy for Duchenne muscular dystrophy. However, the identification of effective AON target sites remains unsatisfactory for lack of a precise method to predict their binding accessibility. This study demonstrates the importance of co-transcriptional pre-mRNA folding in determining the accessibility of AON target sites for AON induction of selective exon skipping in DMD. Because transcription and splicing occur in tandem, AONs must bind to their target sites before splicing factors. Furthermore, co-transcriptional pre-mRNA folding forms transient secondary structures, which redistributes accessible binding sites. In our analysis, to approximate transcription elongation, a “window of analysis” that included the entire targeted exon was shifted one nucleotide at a time along the pre-mRNA. Possible co-transcriptional secondary structures were predicted using the sequence in each step of transcriptional analysis. A nucleotide was considered “engaged” if it formed a complementary base pairing in all predicted secondary structures of a particular step. Correlation of frequency and localisation of engaged nucleotides in AON target sites accounted for the performance (efficacy and efficiency) of 94% of 176 previously reported AONs. Four novel insights are inferred: (1) the lowest frequencies of engaged nucleotides are associated with the most efficient AONs; (2) engaged nucleotides at 3′ or 5′ ends of the target site attenuate AON performance more than at other sites; (3) the performance of longer AONs is less attenuated by engaged nucleotides at 3′ or 5′ ends of the target site compared to shorter AONs; (4) engaged nucleotides at 3′ end of a short target site attenuates AON efficiency more than at 5′ end