Dynamic Anchoring of PKA Is Essential during Oocyte Maturation

SummaryIn the final stages of ovarian follicular development, the mouse oocyte remains arrested in the first meiotic prophase, and cAMP-stimulated PKA plays an essential role in this arrest. After the LH surge, a decrease in cAMP and PKA activity in the oocyte initiates an irreversible maturation process that culminates in a second arrest at metaphase II prior to fertilization [1]. A-kinase anchoring proteins (AKAPs) mediate the intracellular localization of PKA and control the specificity and kinetics of substrate phosphorylation [2]. Several AKAPs have been identified in oocytes including one at 140 kDa [3, 4] that we now identify as a product of the Akap1 gene. We show that PKA interaction with AKAPs is essential for two sequential steps in the maturation process: the initial maintenance of meiotic arrest and the subsequent irreversible progression to the polar body extruded stage. A peptide inhibitor (HT31) that disrupts AKAP/PKA interactions stimulates oocyte maturation in the continued presence of high cAMP. However, during the early minutes of maturation, type II PKA moves from cytoplasmic sites to the mitochondria, where it associates with AKAP1, and this is shown to be essential for maturation to continue irreversibly

Impact of a Multimodal Antimicrobial Stewardship Program on Pseudomonas aeruginosa Susceptibility and Antimicrobial Use in the Intensive Care Unit Setting

Objective. To study the impact of our multimodal antibiotic stewardship program on Pseudomonas aeruginosa susceptibility and antibiotic use in the intensive care unit (ICU) setting. Methods. Our stewardship program employed the key tenants of published antimicrobial stewardship guidelines. These included prospective audits with intervention and feedback, formulary restriction with preauthorization, educational conferences, guidelines for use, antimicrobial cycling, and de-escalation of therapy. ICU antibiotic use was measured and expressed as defined daily doses (DDD) per 1,000 patient-days. Results. Certain temporal relationships between antibiotic use and ICU resistance patterns appeared to be affected by our antibiotic stewardship program. In particular, the ICU use of intravenous ciprofloxacin and ceftazidime declined from 148 and 62.5 DDD/1,000 patient-days to 40.0 and 24.5, respectively, during 2004 to 2007. An increase in the use of these agents and resistance to these agents was witnessed during 2008–2010. Despite variability in antibiotic usage from the stewardship efforts, we were overall unable to show statistical relationships with P. aeruginosa resistance rate. Conclusion. Antibiotic resistance in the ICU setting is complex. Multimodal stewardship efforts attempt to prevent resistance, but such programs clearly have their limits

Six Year Refractive Change among White Children and Young Adults: Evidence for Significant Increase in Myopia among White UK Children

OBJECTIVE:To determine six-year spherical refractive error change among white children and young adults in the UK and evaluate differences in refractive profiles between contemporary Australian children and historical UK data. DESIGN:Population-based prospective study. PARTICIPANTS:The Northern Ireland Childhood Errors of Refraction (NICER) study Phase 1 examined 1068 children in two cohorts aged 6-7 years and 12-13 years. Prospective data for six-year follow-up (Phase 3) are available for 212 12-13 year olds and 226 18-20 year olds in each cohort respectively. METHODS:Cycloplegic refractive error was determined using binocular open-field autorefraction (Shin-Nippon NVision-K 5001, cyclopentolate 1%). Participants were defined by spherical equivalent refraction (SER) as myopic SER ≤-0.50D, emmetropic -0.50D<SER<+2.00 or hyperopic SER≥+2.00D. MAIN OUTCOME MEASURES:Proportion and incidence of myopia. RESULTS:The proportion of myopes significantly increased between 6-7 years (1.9%) and 12-13 years (14.6%) (p<0.001) but not between 12-13 and 18-20 years (16.4% to 18.6%, p = 0.51). The estimated annual incidence of myopia was 2.2% and 0.7% for the younger and older cohorts respectively. There were significantly more myopic children in the UK at age 12-13 years in the NICER study (16.4%) than reported in Australia (4.4%) (p<0.001). However by 17 years the proportion of myopia neared equivalence in the two populations (NICER 18.6%, Australia 17.7%, p = 0.75). The proportion of myopic children aged 12-13 years in the present study (2006-2008) was 16.4%, significantly greater than that reported for children aged 10-16 years in the 1960's (7.2%, p = 0.01). The proportion of hyperopes in the younger NICER cohort decreased significantly over the six year period (from 21.7% to 14.2%, p = 0.04). Hyperopes with SER ≥+3.50D in both NICER age cohorts demonstrated persistent hyperopia. CONCLUSIONS:The incidence and proportion of myopia are relatively low in this contemporary white UK population in comparison to other worldwide studies. The proportion of myopes in the UK has more than doubled over the last 50 years in children aged between 10-16 years and children are becoming myopic at a younger age. Differences between the proportion of myopes in the UK and in Australia apparent at 12-13 years were eliminated by 17 years of age

A genome-wide association study of early menopause and the combined impact of identified variants

Early menopause (EM) affects up to 10% of the female population, reducing reproductive lifespan considerably. Currently, it constitutes the leading cause of infertility in the western world, affecting mainly those women who postpone their first pregnancy beyond the age of 30 years. The genetic aetiology of EM is largely unknown in the majority of cases. We have undertaken a meta-analysis of genome-wide association studies (GWASs) in 3493 EM cases and 13 598 controls from 10 independent studies. No novel genetic variants were discovered, but the 17 variants previously associated with normal age at natural menopause as a quantitative trait (QT) were also associated with EM and primary ovarian insufficiency (POI). Thus, EM has a genetic aetiology which overlaps variation in normal age at menopause and is at least partly explained by the additive effects of the same polygenic variants. The combined effect of the common variants captured by the single nucleotide polymorphism arrays was estimated to account for ∼30% of the variance in EM. The association between the combined 17 variants and the risk of EM was greater than the best validated non-genetic risk factor, smokin

New genetic loci link adipose and insulin biology to body fat distribution.

Body fat distribution is a heritable trait and a well-established predictor of adverse metabolic outcomes, independent of overall adiposity. To increase our understanding of the genetic basis of body fat distribution and its molecular links to cardiometabolic traits, here we conduct genome-wide association meta-analyses of traits related to waist and hip circumferences in up to 224,459 individuals. We identify 49 loci (33 new) associated with waist-to-hip ratio adjusted for body mass index (BMI), and an additional 19 loci newly associated with related waist and hip circumference measures (P < 5 × 10(-8)). In total, 20 of the 49 waist-to-hip ratio adjusted for BMI loci show significant sexual dimorphism, 19 of which display a stronger effect in women. The identified loci were enriched for genes expressed in adipose tissue and for putative regulatory elements in adipocytes. Pathway analyses implicated adipogenesis, angiogenesis, transcriptional regulation and insulin resistance as processes affecting fat distribution, providing insight into potential pathophysiological mechanisms

Regulatory T Cell Responses in Participants with Type 1 Diabetes after a Single Dose of Interleukin-2: A Non-Randomised, Open Label, Adaptive Dose-Finding Trial

BACKGROUND: Interleukin-2 (IL-2) has an essential role in the expansion and function of CD4+ regulatory T cells (Tregs). Tregs reduce tissue damage by limiting the immune response following infection and regulate autoreactive CD4+ effector T cells (Teffs) to prevent autoimmune diseases, such as type 1 diabetes (T1D). Genetic susceptibility to T1D causes alterations in the IL-2 pathway, a finding that supports Tregs as a cellular therapeutic target. Aldesleukin (Proleukin; recombinant human IL-2), which is administered at high doses to activate the immune system in cancer immunotherapy, is now being repositioned to treat inflammatory and autoimmune disorders at lower doses by targeting Tregs. METHODS AND FINDINGS: To define the aldesleukin dose response for Tregs and to find doses that increase Tregs physiologically for treatment of T1D, a statistical and systematic approach was taken by analysing the pharmacokinetics and pharmacodynamics of single doses of subcutaneous aldesleukin in the Adaptive Study of IL-2 Dose on Regulatory T Cells in Type 1 Diabetes (DILT1D), a single centre, non-randomised, open label, adaptive dose-finding trial with 40 adult participants with recently diagnosed T1D. The primary endpoint was the maximum percentage increase in Tregs (defined as CD3+CD4+CD25highCD127low) from the baseline frequency in each participant measured over the 7 d following treatment. There was an initial learning phase with five pairs of participants, each pair receiving one of five pre-assigned single doses from 0.04 × 106 to 1.5 × 106 IU/m2, in order to model the dose-response curve. Results from each participant were then incorporated into interim statistical modelling to target the two doses most likely to induce 10% and 20% increases in Treg frequencies. Primary analysis of the evaluable population (n = 39) found that the optimal doses of aldesleukin to induce 10% and 20% increases in Tregs were 0.101 × 106 IU/m2 (standard error [SE] = 0.078, 95% CI = -0.052, 0.254) and 0.497 × 106 IU/m2 (SE = 0.092, 95% CI = 0.316, 0.678), respectively. On analysis of secondary outcomes, using a highly sensitive IL-2 assay, the observed plasma concentrations of the drug at 90 min exceeded the hypothetical Treg-specific therapeutic window determined in vitro (0.015-0.24 IU/ml), even at the lowest doses (0.040 × 106 and 0.045 × 106 IU/m2) administered. A rapid decrease in Treg frequency in the circulation was observed at 90 min and at day 1, which was dose dependent (mean decrease 11.6%, SE = 2.3%, range 10.0%-48.2%, n = 37), rebounding at day 2 and increasing to frequencies above baseline over 7 d. Teffs, natural killer cells, and eosinophils also responded, with their frequencies rapidly and dose-dependently decreased in the blood, then returning to, or exceeding, pretreatment levels. Furthermore, there was a dose-dependent down modulation of one of the two signalling subunits of the IL-2 receptor, the β chain (CD122) (mean decrease = 58.0%, SE = 2.8%, range 9.8%-85.5%, n = 33), on Tregs and a reduction in their sensitivity to aldesleukin at 90 min and day 1 and 2 post-treatment. Due to blood volume requirements as well as ethical and practical considerations, the study was limited to adults and to analysis of peripheral blood only. CONCLUSIONS: The DILT1D trial results, most notably the early altered trafficking and desensitisation of Tregs induced by a single ultra-low dose of aldesleukin that resolves within 2-3 d, inform the design of the next trial to determine a repeat dosing regimen aimed at establishing a steady-state Treg frequency increase of 20%-50%, with the eventual goal of preventing T1D. TRIAL REGISTRATION: ISRCTN Registry ISRCTN27852285; ClinicalTrials.gov NCT01827735.This is the final version of the article. It first appeared from the Public Library of Science via http://dx.doi.org/10.1371/journal.pmed.100213

Genetic Testing to Inform Epilepsy Treatment Management From an International Study of Clinical Practice

IMPORTANCE: It is currently unknown how often and in which ways a genetic diagnosis given to a patient with epilepsy is associated with clinical management and outcomes. OBJECTIVE: To evaluate how genetic diagnoses in patients with epilepsy are associated with clinical management and outcomes. DESIGN, SETTING, AND PARTICIPANTS: This was a retrospective cross-sectional study of patients referred for multigene panel testing between March 18, 2016, and August 3, 2020, with outcomes reported between May and November 2020. The study setting included a commercial genetic testing laboratory and multicenter clinical practices. Patients with epilepsy, regardless of sociodemographic features, who received a pathogenic/likely pathogenic (P/LP) variant were included in the study. Case report forms were completed by all health care professionals. EXPOSURES: Genetic test results. MAIN OUTCOMES AND MEASURES: Clinical management changes after a genetic diagnosis (ie, 1 P/LP variant in autosomal dominant and X-linked diseases; 2 P/LP variants in autosomal recessive diseases) and subsequent patient outcomes as reported by health care professionals on case report forms. RESULTS: Among 418 patients, median (IQR) age at the time of testing was 4 (1-10) years, with an age range of 0 to 52 years, and 53.8% (n = 225) were female individuals. The mean (SD) time from a genetic test order to case report form completion was 595 (368) days (range, 27-1673 days). A genetic diagnosis was associated with changes in clinical management for 208 patients (49.8%) and usually (81.7% of the time) within 3 months of receiving the result. The most common clinical management changes were the addition of a new medication (78 [21.7%]), the initiation of medication (51 [14.2%]), the referral of a patient to a specialist (48 [13.4%]), vigilance for subclinical or extraneurological disease features (46 [12.8%]), and the cessation of a medication (42 [11.7%]). Among 167 patients with follow-up clinical information available (mean [SD] time, 584 [365] days), 125 (74.9%) reported positive outcomes, 108 (64.7%) reported reduction or elimination of seizures, 37 (22.2%) had decreases in the severity of other clinical signs, and 11 (6.6%) had reduced medication adverse effects. A few patients reported worsening of outcomes, including a decline in their condition (20 [12.0%]), increased seizure frequency (6 [3.6%]), and adverse medication effects (3 [1.8%]). No clinical management changes were reported for 178 patients (42.6%). CONCLUSIONS AND RELEVANCE: Results of this cross-sectional study suggest that genetic testing of individuals with epilepsy may be materially associated with clinical decision-making and improved patient outcomes