Characterization of the Structural Requirements for the NADase Activity of Bacterial Toll/IL-1R domains in a Course-based Undergraduate Research Experience.

TLRs initiate innate immune signaling pathways via Toll/IL-1R (TIR) domains on their cytoplasmic tails. Various bacterial species also express TIR domain-containing proteins that contribute to bacterial evasion of the innate immune system. Bacterial TIR domains, along with the mammalian sterile α and TIR motif-containing protein 1 and TIRs from plants, also have been found to exhibit NADase activity. Initial X-ray crystallographic studies of the bacterial TIR from Acinetobacter baumannii provided insight into bacterial TIR structure but were unsuccessful in cocrystallization with the NAD+ ligand, leading to further questions about the TIR NAD binding site. In this study, we designed a Course-Based Undergraduate Research Experience (CURE) involving 16-20 students per year to identify amino acids crucial for NADase activity of A. baumannii TIR domain protein and the TIR from Escherichia coli (TIR domain-containing protein C). Students used structural data to identify amino acids that they hypothesized would play a role in TIR NADase activity, and created plasmids to express mutated TIRs through site-directed mutagenesis. Mutant TIRs were expressed, purified, and tested for NADase activity. The results from these studies provide evidence for a conformational change upon NAD binding, as was predicted by recent cryogenic electron microscopy and hydrogen-deuterium exchange mass spectrometry studies. Along with corroborating recent characterization of TIR NADases that could contribute to drug development for diseases associated with dysregulated TIR activity, this work also highlights the value of CURE-based projects for inclusion of a diverse group of students in authentic research experiences.

Balanced crystalloids (RInger's lactate) versus normal Saline in adults with diabetic Ketoacidosis in the Emergency Department (BRISK-ED): a pilot randomised controlled trial.

BACKGROUND: Current diabetic ketoacidosis (DKA) treatment guidelines recommend using normal saline (NS); however, NS may delay DKA resolution by causing more hyperchloremic metabolic acidosis compared with balanced crystalloids. This study's objective was to determine the feasibility of a future multicentred randomised controlled trial (RCT) comparing intravenous Ringer's lactate (RL) with NS in managing ED patients with DKA. METHODS: We conducted a parallel-arm, triple-blind, pilot RCT of adults (≥18 years) with DKA at a Canadian academic tertiary care ED. The primary feasibility outcome was recruitment rate (target ≥41.3% of eligible participants over the 1-year study period); the primary efficacy outcome was time elapsed from ED presentation to DKA resolution. The superiority margin for a clinically significant difference was chosen to be a 40% time reduction to DKA resolution. We also assessed the need to break allocation concealment and loss to follow-up. Patients with clinical suspicion for DKA were screened for inclusion and enrolled patients were randomised 1:1 to receive RL or NS. Patients, clinicians and outcome assessors were blinded to allocation. RESULTS: We enrolled 52 (25 RL, 27 NS) of 60 eligible patients (86.7%), exceeding our target recruitment rate. There were more patients in the NS group with type 1 diabetes, and more patients in the RL group had an admission co-diagnosis in addition to DKA. For the 44 participants with confirmed laboratory evidence of resolution, median (IQR) time to DKA resolution for RL versus NS was 15.7 (10.4-18.8) and 12.7 (7.9-19.2) hours, respectively. There were no cases where blinding was broken, and there was no loss to follow-up. CONCLUSIONS: This pilot trial demonstrated our protocol's feasibility by exceeding our target recruitment rate. Our results may be used to inform future multicentre trials to compare the safety and efficacy of RL and NS in managing DKA in the ED. TRIAL REGISTRATION NUMBER: NCT04926740.

Balanced crystalloids (RInger's lactate) versus normal Saline in adults with diabetic Ketoacidosis in the Emergency Department (BRISK-ED): a protocol for a pilot randomized controlled trial.

BACKGROUND: Current guidelines for diabetic ketoacidosis (DKA) recommend treatment with normal saline (NS). However, NS, with its high chloride concentrations, may worsen acidosis and contribute to a hyperchloremic metabolic acidosis. Alternatives to NS are balanced crystalloids (e.g. Ringer's Lactate [RL]) which have chloride concentrations similar to human plasma; therefore, treatment with balanced crystalloids may lead to faster DKA resolution. A recent systematic review and meta-analysis by Catahay et al. (2022) demonstrated the need for more blinded, high-quality trials comparing NS versus RL in the treatment of DKA. METHODS: We describe a protocol for BRISK-ED (Balanced crystalloids [RInger's lactate] versus normal Saline in adults with diabetic Ketoacidosis in the Emergency Department). Our study is a single-centre, triple-blind, pilot randomized controlled trial (RCT) of adults (≥ 18 years) with DKA presenting to an academic tertiary care ED in London, Canada. Patients with clinical suspicion for DKA will be screened and those found to not meet DKA criteria or have euglycemic DKA will be excluded. We will aim to recruit 52 patients with DKA and will randomize them 1:1 to receive intravenous RL or NS. The primary feasibility outcome will be recruitment rate, and the primary efficacy outcome will be time elapsed from ED presentation to DKA resolution. Secondary outcomes include time to insulin infusion discontinuation, intensive care unit admission, in-hospital death, and major adverse kidney events within 30 days, defined as a composite of: i) death, ii) new renal replacement therapy, or iii) final serum creatinine ≥ 200% baseline at the earliest of hospital discharge or 30 days after ED presentation. Patients, clinicians, and outcome assessors will be blinded to allocation group. We will follow an intention-to-treat analysis. Gehan-Wilcoxon, Mann-Whitney U, or chi-square tests will be used to compare groups as appropriate. DISCUSSION: The results of this pilot study will inform the design and feasibility of a full-scale, multicentre RCT to assess fluid choice in adult ED patients with DKA. If proven to demonstrate faster resolution of DKA, administration of balanced crystalloids may replace NS in diabetes treatment guidelines and improve patient and health systems outcomes. TRIAL REGISTRATION: ClinicalTrials.gov, Registration # NCT04926740; Registered June 15, 2021.

Telehealth Interventions Within the Scope of Occupational Therapy Practice: A Systematic Review.

IMPORTANCE: With the increasing use of telehealth, it is imperative to synthesize the existing evidence to examine the effectiveness of telehealth interventions and inform practitioners and reimbursement entities. OBJECTIVE: To examine the effectiveness of telehealth occupational therapy interventions across the lifespan, delivered either independently or as part of an interdisciplinary team. DATA SOURCES: MEDLINE, CINAHL, SAGE, PsycInfo, Cochrane Database of Systematic Reviews, OTseeker, and OT Search and hand searches of systematic reviews, relevant journals, and known occupational therapy telehealth research. STUDY SELECTION AND DATA COLLECTION: Following Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines, we screened and appraised articles that included an occupational therapy intervention delivered via telehealth and that were peer reviewed, written in English, published between 2009 and 2019, and Levels 1b to 3b evidence. FINDINGS: Twenty full-text articles (8 Level 1b, 9 Level 2b, and 3 Level 3b) met the inclusion criteria. Strong strength of evidence supports the use of telehealth approaches for occupational therapy interventions for people with neurological and pain conditions. Moderate strength of evidence supports the use of telehealth interventions to support education outcomes. Low strength of evidence was found for other outcomes for children with developmental disorders and additional conditions. CONCLUSIONS AND RELEVANCE: There is evidence to support that occupational therapy interventions delivered via telehealth are similarly effective as those delivered face-to-face, especially for neurological and pain conditions. What This Article Adds: The findings include an updated synthesis of telehealth occupational therapy interventions provided separately and with interdisciplinary health care teams. They expand occupational therapy's scope of practice to include interventions provided across the lifespan for rehabilitation and habilitation needs and include effectiveness by conditions.

DNA methylation patterns and gene expression from amygdala tissue of mature Brahman cows exposed to prenatal stress.

Prenatal stress can alter postnatal performance and temperament of cattle. These phenotypic effects may result from changes in gene expression caused by stress-induced epigenetic alterations. Specifically, shifts in gene expression caused by DNA methylation within the brain's amygdala can result in altered behavior because it regulates fear, stress response and aggression in mammals Thus, the objective of this experiment was to identify DNA methylation and gene expression differences in the amygdala tissue of 5-year-old prenatally stressed (PNS) Brahman cows compared to control cows. Pregnant Brahman cows (n = 48) were transported for 2-h periods at 60 ± 5, 80 ± 5, 100 ± 5, 120 ± 5, and 140 ± 5 days of gestation. A non-transported group (n = 48) were controls (Control). Amygdala tissue was harvested from 6 PNS and 8 Control cows at 5 years of age. Overall methylation of gene body regions, promoter regions, and cytosine-phosphate-guanine (CpG) islands were compared between the two groups. In total, 202 genes, 134 promoter regions, and 133 CpG islands exhibited differential methylation (FDR ≤ 0.15). Following comparison of gene expression in the amygdala between the PNS and Control cows, 2 differentially expressed genes were identified (FDR ≤ 0.15). The minimal differences observed could be the result of natural changes of DNA methylation and gene expression as an animal ages, or because this degree of transportation stress was not severe enough to cause lasting effects on the offspring. A younger age may be a more appropriate time to assess methylation and gene expression differences produced by prenatal stress.

Biorenewable triblock copolymers consisting of l-lactide and ε-caprolactone for removing organic pollutants from water: a lifecycle neutral solution.

BACKGROUND: Current methods of removing organic pollutants from water are becoming ineffective as the world population increases. In this study, a series of biorenewable triblock copolymers with hydrophobic poly(ε-caprolactone) block and hydrophilic poly(l-lactide) blocks were synthesized and tested as agents to remove environmental pollutants from an aqueous solution. The percent of pollutant removed and equilibrium inclusion constants were calculated for the polymers. These values were compared to previously known removal agents for their effectiveness. RESULTS: Triblock copolymer samples removed over 70% of the polycyclic aromatic hydrocarbon (PAH) phenanthrene from an aqueous solution, with selectivity for the adsorption of phenanthrene over other PAHs tested. The inclusion constant was 7.4 × 105 M-1 and adsorption capacity was 5.8 × 10-7 mol phenanthrene/g polymer. Rose Bengal was used to further probe the nature of interactions between the copolymers and a small molecule guest. Solid samples of the block-poly(l-lactide)-block-poly(ε-caprolactone)-block-poly(l-lactide) (PLLA-PCL-PLLA) systems were found to rapidly remove over 90% of Rose Bengal from aqueous solution, resulting in a complete disappearance of the characteristic pink color. Solutions of the copolymers in dichloromethane also removed Rose Bengal from water with a similar level of efficiency. Large inclusion constant values were obtained, ranging from 1.0 × 105 to 7.9 × 105 M-1, and the average adsorption capacity value of 6.2 × 10-7 mol/g polymer was determined. Aged polymer samples exhibited different adsorption characteristics and mechanistic theories for the removal of Rose Bengal were determined. CONCLUSION: The triblock copolymer consisting of l-lactide and ε-caprolactone was effective in removing various organic pollutants in aqueous environments. It is a biorenewable material which leads to minimal waste production during its lifecycle. These polymers were in general more effective in removing organic pollutants than commercially available pollution removal systems.