Arrayed CRISPR libraries for the genome-wide activation, deletion and silencing of human protein-coding genes.

Arrayed CRISPR libraries extend the scope of gene-perturbation screens to non-selectable cell phenotypes. However, library generation requires assembling thousands of vectors expressing single-guide RNAs (sgRNAs). Here, by leveraging massively parallel plasmid-cloning methodology, we show that arrayed libraries can be constructed for the genome-wide ablation (19,936 plasmids) of human protein-coding genes and for their activation and epigenetic silencing (22,442 plasmids), with each plasmid encoding an array of four non-overlapping sgRNAs designed to tolerate most human DNA polymorphisms.

Health-related quality of life among pediatric patients with acute lymphoblastic leukemia: An exploratory cross-sectional study.

Background: Improved survival rates broadened the pediatric oncology focus to include health-related quality of life (HRQoL). This cross-sectional study aimed to examine HRQoL by treatment phase and disease risk level in pediatric patients with B-cell acute lymphoblastic leukemia (B-ALL), including those in early survivorship.

Procedure: A subset of data from a larger prospective cohort study was analyzed.

Distressing Discussions in Pediatric Interpreted Medical Encounters: A Qualitative Study of Medical Interpreter Perspectives on Clinician Communication Practices.

Introduction: This study explores pediatric medical interpreters' perspectives on clinician communication practices in medical encounters characterized by distressing content and difficult discussions.

Method: In this interpretative phenomenological analysis, 13 Spanish-English interpreters at a midwestern pediatric hospital were purposively recruited and, in 2021-2022, completed a demographic survey and semistructured interview on communication in distressing interpreted medical encounters.

Results: Participants described clinician practices for effective cross-cultural interpreted communication.

The impact of the COVID-19 pandemic on families of children with medical complexity: A qualitative study of caregivers' experiences.

Background: Families of children with medical complexity (CMC) have been negatively affected by the COVID-19 pandemic, experiencing challenges such as decreased access to services, increased financial hardship and increased isolation. However, there are few qualitative studies which explore parental experiences. The aim of the present study was to describe the impact of the COVID-19 pandemic on families of CMC.

Drug screen in iPSC-Neurons identifies nucleoside analogs as inhibitors of (GC) expression in C9orf72 ALS/FTD.

An intronic (GC) expansion in C9orf72 causes amyotrophic lateral sclerosis and frontotemporal dementia primarily through gain-of-function mechanisms: the accumulation of sense and antisense repeat RNA foci and dipeptide repeat (DPR) proteins (poly-GA/GP/GR/PA/PR) translated from repeat RNA. To therapeutically block this pathway, we screen a library of 1,430 approved drugs and known bioactive compounds in patient-derived induced pluripotent stem cell-derived neurons (iPSC-Neurons) for inhibitors of DPR expression. The clinically used guanosine/cytidine analogs decitabine, entecavir, and nelarabine reduce poly-GA/GP expression, with decitabine being the most potent.

"It Has Added Another Layer of Stress": COVID-19's Impact in the PICU.

Objective: The novel coronavirus disease 2019 (COVID-19) pandemic has dramatically changed health care delivery and impacted health care providers. However, little is known about the impact of the pandemic in PICUs. In this qualitative study, we aimed to assess pediatric critical care providers' perspectives on the impact of the COVID-19 pandemic on the experiences of patients and families in the PICU and on their personal and professional lives.

Identification of Chemical and Pharmacological Chaperones for Correction of Trafficking-Deficient Mutant Cyclic Nucleotide-Gated A3 Channels.

Trafficking deficiency caused by missense mutations is a well known phenomenon that occurs for mutant, misfolded proteins. Typically, the misfolded protein is retained by the protein quality-control system and degraded by the endoplasmic reticulum-associated protein degradation pathway and thus does not reach its destination, although residual function of the protein may be preserved. Chemical and pharmacological chaperones can improve the targeting of trafficking-deficient proteins and thus may be promising candidates for therapeutic applications.

Automated Production of Human Induced Pluripotent Stem Cell-Derived Cortical and Dopaminergic Neurons with Integrated Live-Cell Monitoring.

Manual culture and differentiation protocols for human induced pluripotent stem cells (hiPSC) are difficult to standardize, show high variability and are prone to spontaneous differentiation into unwanted cell types. The methods are labor-intensive and are not easily amenable to large-scale experiments. To overcome these limitations, we developed an automated cell culture system coupled to a high-throughput imaging system and implemented protocols for maintaining multiple hiPSC lines in parallel and neuronal differentiation.

Emotionally evocative patients in the emergency department: a mixed methods investigation of providers' reported emotions and implications for patient safety.

Background: Emergency department (ED) physicians and nurses frequently interact with emotionally evocative patients, which can impact clinical decision-making and behaviour. This study introduces well-established methods from social psychology to investigate ED providers' reported emotional experiences and engagement in their own recent patient encounters, as well as perceived effects of emotion on patient care.

Methods: Ninety-four experienced ED providers (50 physicians and 44 nurses) vividly recalled and wrote about three recent patient encounters (qualitative data): one that elicited anger/frustration/irritation (angry encounter), one that elicited happiness/satisfaction/appreciation (positive encounter), and one with a patient with a mental health condition (mental health encounter).

Helping parents prepare for their child's end of life: A retrospective survey of cancer-bereaved parents.

Background: Most parents vividly recall the weeks, days, and moments preceding their child's death for years to come. Dissatisfaction with communication about their child's condition and lack of guidance can contribute to stress prior to a child's death. Based on findings from a study assessing the degree of preparation bereaved parents received and our collective clinical experience, the authors provide suggestions on end-of-life communication and guidance for parents.

Participation in Online Research Examining End-of-Life Experiences: Is It Beneficial, Burdensome, or Both for Parents Bereaved by Childhood Cancer?

It is important for the health care community to understand the impact of a child's death on parent functioning. Yet involving bereaved parents in research that enquires about such a stressful time in their life can potentially bring harm to them. The current study examines the perceived benefit and burden of parents participating in a survey exploring their perceptions of their child's end-of-life (EoL) and bereavement experiences.

An early nonsense mutation facilitates the expression of a short isoform of CNGA3 by alternative translation initiation.

The cyclic nucleotide-gated (CNG) channel - composed of CNGA3 and CNGB3 subunits - mediates the influx of cations in cone photoreceptors after light stimulation and thus is a key element in cone phototransduction. Mutations in CNGA3 and CNGB3 are associated with achromatopsia, a rare autosomal recessive retinal disorder. Here, we demonstrate that the presence of an early nonsense mutation in CNGA3 induces the usage of a downstream alternative translation initiation site giving rise to a short CNGA3 isoform.

Personalized and yet standardized: An informed approach to the integration of bereavement care in pediatric oncology settings.

Objective: The death of a child has been associated with adverse parental outcomes, including a heightened risk for psychological distress, poor physical health, loss of employment income, and diminished psychosocial well-being. Psychosocial standards of care for centers serving pediatric cancer patients recommend maintaining at least one meaningful contact between the healthcare team and bereaved parents to identify families at risk for negative psychosocial sequelae and to provide resources for bereavement support. This study assessed how this standard is being implemented in current healthcare and palliative care practices, as well as barriers to its implementation.

A Summary of Pediatric Palliative Care Team Structure and Services as Reported by Centers Caring for Children with Cancer.

Background: Little is known about the composition, availability, integration, communication, perceived barriers, and work load of pediatric palliative care (PPC) providers serving children and adolescents with cancer.

Objective: To summarize the structure and services of programs to better understand successes and gaps in implementing palliative care as a standard of care.

Methods: Cross-sectional online survey about the palliative care domains determined by the Psychosocial Care of Children with Cancer and Their Families Workgroup.

Non-viral genetic transfection of rat Schwann cells with FuGENE HD© lipofection and AMAXA© nucleofection is feasible but impairs cell viability.

Purpose: To determine transfection efficiency of FuGENE HD© lipofection and AMAXA© nucleofection on rat Schwann cells (SC).

Methods: The ischiadic and median nerves of 6-8 week old Lewis rats were cultured in modified melanocyte-growth medium. SCs were genetically transfected with green fluorescent protein (GFP) as reporter gene using FuGENE HD© lipofection and AMAXA© nucleofection.

Efficacy of various durations of in vitro predegeneration on the cell count and purity of rat Schwann-cell cultures.

The efficacy of Schwann-cell cultivation can be enhanced by in vitro predegeneration of the harvested cells compared to immediate culture. The aim of this study was to improve Schwann-cell culture efficacy by comparing three different durations of predegeneration. The sciatic and median nerves of 6-8-week-old Lewis rats were harvested and subjected to either 2-day, 7-day, or 14-day predegeneration in Dulbecco's Modified Eagle's Medium supplemented with 10% fetal calf serum and 1% Penicillin/Streptomycin.

Lipid metabolism in peroxisomes in relation to human disease.

Peroxisomes were long believed to play only a minor role in cellular metabolism but it is now clear that they catalyze a number of important functions. The importance of peroxisomes in humans is stressed by the existence of a group of genetic diseases in man in which one or more peroxisomal functions are impaired. Most of the functions known to take place in peroxisomes have to do with lipids.

Control analysis of biliary lipid secretion.

Unlabelled: Biliary lipid secretion is a complex process involving a multitude of metabolic pathways. It has always been assumed that bile salt secretion (BSec) fully controls this process. Recently we have demonstrated, that mdr2 P-glycoprotein (P-gp) is an important controlling step as well.

Signals on proteins, intracellular targeting and inborn errors of organellar metabolism.

Newly synthesized polypeptides contain signals that direct them to the appropriate intracellular organelles and the organelles contain receptors that recognize the signals. Protein synthesis occurs either on free ribosomes or on ribosomes bound to the endoplasmic reticulum. The proteins synthesized on bound ribosomes are co-translationally translocated into the lumen of the endoplasmic reticulum and contain or acquire targeting information for retention in the endoplasmic reticulum or for sorting to lysosomes and other compartments of the secretory and endocytic pathways.

Clinical and biochemical characteristics of peroxisomal disorders: an update.

Peroxisomal disorders represent a recently recognized group of inherited diseases in man, now comprising 14 different disorders. If discussion is restricted to those peroxisomal disorders in which there is neurological involvement (thereby excluding hyperoxaluria and acatalasaemia), results over the least few years have shown that analysis of very-long-chain fatty acids (VLCFAs) is a highly reliable initial test to establish whether or not one is dealing with a peroxisomal disorder. Rhizomelic chondrodysplasia punctata, its recently identified variant form and glutaryl-CoA oxidase deficiency will show no abnormalities and must be identified by other means.