A puromycin selectable cell line for the enrichment of mouse embryonic stem cell-derived V3 interneurons

INTRODUCTION: Spinal V3 interneurons (INs) are a commissural, glutamatergic, propriospinal neuron population that holds great potential for understanding locomotion circuitry and local rewiring after spinal cord injury. Embryonic stem cells hold promise as a cell source. However, the inevitable heterogeneity resulting from differentiation protocols makes studying post-mitotic stem cell-derived neuron populations difficult because proliferative glia quickly overtake a culture. Previously, an induction protocol for V3 INs was established. However, because of the heterogeneous population resulting from the induction protocol, functional characterization of the induced cells was not possible. METHODS: A selectable murine transgenic embryonic stem cell (ESC) line (Sim1-Puro) was generated by recombineering. The expression of the puromycin resistance enzyme, puromycin N-acetyl-transferase (PAC), was knocked into the locus of a post-mitotic V3 IN marker (Sim1), allowing Sim1 gene regulatory elements to control PAC expression. The resulting cell line was characterized for Sim1 expression by in situ hybridization, for glutamatergic marker expression by immunocytochemistry and quantitative real time polymerase chain reaction (qRT-PCR), and for functional maturation by electrophysiology. RESULTS: Puromycin selection significantly enriched the population for V3 INs, allowing long-term characterization. The selected population expressed the neuronal marker β-III tubulin and the glutamatergic neuron marker VGluT2. The selected V3 INs also exhibited appropriate functional maturation, as assessed by electrophysiology, and remained glutamatergic for 2 weeks. CONCLUSION: The Sim1-Puro cell line provides a simple, high throughput method for generating large numbers of V3 INs from mouse ESCs for future in vitro and cell transplantation studies. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13287-015-0213-z) contains supplementary material, which is available to authorized users

Does social isolation mediate the association between hearing loss and cognition in adults? A systematic review and meta-analysis of longitudinal studies

Background: There has been extensive research on the relationship between hearing and cognitive impairment in older adults but little examination of the role of mediating factors. Social isolation is a potential mediator, occurring because of hearing loss, and contributing to accelerated cognitive decline. Previous systematic reviews on this topic area have not considered the temporal nature of hearing loss and cognitive impairment exclusively or examined potential mediators within a longitudinal study design. Methods: A systematic review was conducted. Electronic searches were performed in Web of Science, PubMed (Medline), Scopus, EMBASE, PsychInfo, and ProQuest (PsychArticles and ProQuest Dissertation and Theses) based on a search string of keywords relating to hearing loss, social isolation, and cognitive impairment/dementia in June 2023. Papers were critically appraised using the CASP checklists for cohort studies. Risk of bias in the selected studies was assessed using the Item Bank for Assessment of Risk of Bias and Precision for Observational Studies of Interventions or Exposures. Results: Eleven of the 15 included studies provide evidence of a dose-dependent association between hearing threshold (40 dB HL or greater) and later cognitive impairment or incident dementia. Only one study included social isolation as a mediator, which was found to not be a significant contributing factor. The meta-analysis of 5 studies pooled hazard ratio for cognitive impairment due to hearing loss is 1.11 (95% CI: 1.06 to 1.15, p < 0.001). The pooled hazard ratio for incident dementia due to hearing loss was HR 1.21 (95% CI: 1.11 to 1.31, p = 0.002). Conclusion: The analysis of included studies indicate that hearing threshold level affects later cognitive status or dementia diagnosis. There is not enough evidence to determine the role of social isolation as a mediator. Future epidemiology studies need to measure different elements of social isolation and ensure that hearing and cognition are measured at multiple time points

Optimizing preoperative antibiotics in patients with β-lactam allergies: A role for pharmacy

PURPOSE: Patients with a reported β-lactam allergy (BLA) are often given alternative perioperative antibiotic prophylaxis, increasing risk of surgical site infections (SSIs), acute kidney injury (AKI), and Clostridioides difficile infection (CDI). The purpose of this study was to implement and evaluate a pharmacist-led BLA clarification interview service in the preoperative setting. METHODS: A pharmacist performed BLA clarification telephone interviews before elective procedures from November 2018 to March 2019. On the basis of allergy history and a decision algorithm, first-line preoperative antibiotics, alternative antibiotics, or allergy testing referral was recommended. The pharmacist intervention (PI) group was compared to a standard of care (SOC) group who underwent surgery from November 2017 to March 2018. RESULTS: Eighty-seven patients were included, with 50 (57%) and 37 (43%) in the SOC and PI groups, respectively. The most common surgeries included orthopedic surgery in 41 patients (47%) and neurosurgery in 17 patients (20%). In the PI group, all BLA labels were updated after interview. Twenty-three patients were referred for allergy testing, 12 of the 23 (52%) completed BLA testing, and penicillin allergies were removed for 9 of the 12 patients. Overall, 28 of the 37 (76%) pharmacy antibiotic recommendations were accepted. Cefazolin use significantly increased from 28% to 65% after the intervention (P = 0.001). SSI occurred in 5 (10%) patients in the SOC group and no patients in the PI group (P = 0.051). All of these SSIs were associated with alternative antibiotics. Incidence of AKI and CDI was similar between the groups. No allergic reactions occurred in either group. CONCLUSION: Implementation of a pharmacy-driven BLA reconciliation significantly increased β-lactam preoperative use without negative safety outcomes

The Evolving Status of Fellowships and Mini-Fellowships in Diagnostic Radiology: A Survey of Program Directors and Chief Residents

RATIONALE AND OBJECTIVES: Recent changes in radiology fellowships include musculoskeletal radiology adopting a match system, interventional radiology transitioning away from diagnostic radiology to offer direct-entry programs, and a common fellowship application timeline created by the Society for Chairs of Academic Radiology Departments (SCARD). The concept of mini-fellowships has also emerged with the elimination of the oral American Board of Radiology examinations that had been administered in the final year of residency training prior to 2014. This paper seeks to assess the opinions of fellowship program directors, residency program directors, and chief residents regarding these recent changes. MATERIALS AND METHODS: This is a cross-sectional study using a web-based survey posed to fellowship program directors, residency program directors, and chief residents in 2020. Questions sought to explore current attitudes toward the following topics: (1) a common fellowship application timeline; (2) a common fellowship match; and (3) the status of mini-fellowships in diagnostic radiology. In addition, the number of fellowship positions for each subspecialty was estimated using subspecialty society directories, Accreditation Council for Graduate Medical Education (ACGME) data, and individual program websites. RESULTS: Deidentified responses were collected electronically and aggregated. The three respondent groups preferred a common fellowship application timeline at rates of 67% among fellowship program directors, 80% residency program directors, and 74% residents. A common match system across all subspecialties was preferred at rates of 50% fellowship program directors, 74% residency program directors, and 26% chief residents. There was widespread reported compliance with the SCARD fellowship timeline policy. Subspecialty programs using the match system reported interviewing greater numbers of applicants per position. Fellowship directors and chief residents reported that the most common duration of mini-fellowship experiences was 2 to 3 months. CONCLUSION: There is a division between chief residents and program directors regarding the preference for a common radiology match. Adopting a radiology-wide fellowship match would increase the number of interviews required. The SCARD fellowship timeline policy has been successful, and there is support across stakeholders regarding the common timeline. Mini-fellowships are highly variable in length and structure

Cdc42 Regulates Extracellular Matrix Remodeling in Three Dimensions*

Extracellular matrix (ECM) actively participates in normal cell regulation and in the process of tumor progression. The Rho GTPase Cdc42 has been shown to regulate cell-ECM interaction in conventional two-dimensional culture conditions by using dominant mutants of Cdc42 in immortalized cell lines that may introduce nonspecific effects. Here, we employ three-dimensional culture systems for conditional gene targeted primary mouse embryonic fibroblasts that better simulate the reciprocal and adaptive interactions between cells and surrounding matrix to define the role of Cdc42 signaling pathways in ECM organization. Cdc42 deficiency leads to a defect in global cell-matrix interactions reflected by a decrease in collagen gel contraction. The defect is associated with an altered cell-matrix interaction that is evident by morphologic changes and reduced focal adhesion complex formation. The matrix defect is also associated with a reduction in synthesis and activation of matrix metalloproteinase 9 (MMP9) and altered fibronectin deposition patterning. A Cdc42 mutant rescue experiment found that downstream of Cdc42, p21-activated kinase (PAK), but not Par6 or WASP, may be involved in regulating collagen gel contraction and fibronectin organization. Thus, in addition to the previously implicated roles in intracellular regulation of actin organization, proliferation, and vesicle trafficking, Cdc42 is essential in ECM remodeling in three dimensions

Cardiac Biomarkers and Risk of Atrial Fibrillation in Chronic Kidney Disease: The CRIC Study.

Background We tested associations of cardiac biomarkers of myocardial stretch, injury, inflammation, and fibrosis with the risk of incident atrial fibrillation (AF) in a prospective study of chronic kidney disease patients. Methods and Results The study sample was 3053 participants with chronic kidney disease in the multicenter CRIC (Chronic Renal Insufficiency Cohort) study who were not identified as having AF at baseline. Cardiac biomarkers, measured at baseline, were NT-proBNP (N-terminal pro-B-type natriuretic peptide), high-sensitivity troponin T, galectin-3, growth differentiation factor-15, and soluble ST-2. Incident AF ("AF event") was defined as a hospitalization for AF. During a median follow-up of 8 years, 279 (9%) participants developed a new AF event. In adjusted models, higher baseline log-transformed NT-proBNP (N-terminal pro-B-type natriuretic peptide) was associated with incident AF (adjusted hazard ratio [HR] per SD higher concentration: 2.11; 95% CI, 1.75, 2.55), as was log-high-sensitivity troponin T (HR 1.42; 95% CI, 1.20, 1.68). These associations showed a dose-response relationship in categorical analyses. Although log-soluble ST-2 was associated with AF risk in continuous models (HR per SD higher concentration 1.35; 95% CI, 1.16, 1.58), this association was not consistent in categorical analyses. Log-galectin-3 (HR 1.05; 95% CI, 0.91, 1.22) and log-growth differentiation factor-15 (HR 1.16; 95% CI, 0.96, 1.40) were not significantly associated with incident AF. Conclusions We found strong associations between higher NT-proBNP (N-terminal pro-B-type natriuretic peptide) and high-sensitivity troponin T concentrations, and the risk of incident AF in a large cohort of participants with chronic kidney disease. Increased atrial myocardial stretch and myocardial cell injury may be implicated in the high burden of AF in patients with chronic kidney disease

The role of hydropower in South Asia’s energy future

With rising energy demand in Asia, the high potential for hydropower development and the need for low-carbon energy development, hydropower would seem to have a significant role in South Asia’s energy future. However, the extent of hydropower development will depend on several risk factors, including the cost of alternative energy sources, the environmental sustainability of hydropower and social issues of equitable development. Using a risk-analysis framework, it is concluded that the future of hydropower will depend on how well policies and institutions manage the risks, facilitate efficient financial markets, and promote fair and friendly cross-border electricity trade

Unbiased Metagenomic Sequencing for Pediatric Meningitis in Bangladesh Reveals Neuroinvasive Chikungunya Virus Outbreak and Other Unrealized Pathogens.

The burden of meningitis in low-and-middle-income countries remains significant, but the infectious causes remain largely unknown, impeding institution of evidence-based treatment and prevention decisions. We conducted a validation and application study of unbiased metagenomic next-generation sequencing (mNGS) to elucidate etiologies of meningitis in Bangladesh. This RNA mNGS study was performed on cerebrospinal fluid (CSF) specimens from patients admitted in the largest pediatric hospital, a World Health Organization sentinel site, with known neurologic infections (n = 36), with idiopathic meningitis (n = 25), and with no infection (n = 30), and six environmental samples, collected between 2012 and 2018. We used the IDseq bioinformatics pipeline and machine learning to identify potentially pathogenic microbes, which we then confirmed orthogonally and followed up through phone/home visits. In samples with known etiology and without infections, there was 83% concordance between mNGS and conventional testing. In idiopathic cases, mNGS identified a potential bacterial or viral etiology in 40%. There were three instances of neuroinvasive Chikungunya virus (CHIKV), whose genomes were &gt;99% identical to each other and to a Bangladeshi strain only previously recognized to cause febrile illness in 2017. CHIKV-specific qPCR of all remaining stored CSF samples from children who presented with idiopathic meningitis in 2017 (n = 472) revealed 17 additional CHIKV meningitis cases, exposing an unrecognized meningitis outbreak. Orthogonal molecular confirmation, case-based clinical data, and patient follow-up substantiated the findings. Case-control CSF mNGS surveys can complement conventional diagnostic methods to identify etiologies of meningitis, conduct surveillance, and predict outbreaks. The improved patient- and population-level data can inform evidence-based policy decisions.IMPORTANCE Globally, there are an estimated 10.6 million cases of meningitis and 288,000 deaths every year, with the vast majority occurring in low- and middle-income countries. In addition, many survivors suffer from long-term neurological sequelae. Most laboratories assay only for common bacterial etiologies using culture and directed PCR, and the majority of meningitis cases lack microbiological diagnoses, impeding institution of evidence-based treatment and prevention strategies. We report here the results of a validation and application study of using unbiased metagenomic sequencing to determine etiologies of idiopathic (of unknown cause) cases. This included CSF from patients with known neurologic infections, with idiopathic meningitis, and without infection admitted in the largest children's hospital of Bangladesh and environmental samples. Using mNGS and machine learning, we identified and confirmed an etiology (viral or bacterial) in 40% of idiopathic cases. We detected three instances of Chikungunya virus (CHIKV) that were &gt;99% identical to each other and to a strain previously recognized to cause systemic illness only in 2017. CHIKV qPCR of all remaining stored 472 CSF samples from children who presented with idiopathic meningitis in 2017 at the same hospital uncovered an unrecognized CHIKV meningitis outbreak. CSF mNGS can complement conventional diagnostic methods to identify etiologies of meningitis, and the improved patient- and population-level data can inform better policy decisions