Significance of Premature Stop Codons in \u3cem\u3eenv\u3c/em\u3e of Simian Immunodeficiency Virus

The location of the translational termination codon for the transmembrane protein (TMP) varies in three infectious molecular clones of simian immunodeficiency virus from macaques (SIVmac). The SIVmac251 and SIVmac142 infectious clones have premature stop signals that differ in location by one codon; transfection of these DNAs into human HUT-78 cells yielded virus with a truncated TMP (28 to 30 kilodaltons [kDa]). The SIVmac239 infectious clone does not have a premature stop codon in its TMP-coding region. Transfection of HUT-78 cells with this clone initially yielded virus with a full-length TMP (41 kDa). At 20 to 30 days posttransfection, SIVmac239 virus with a 41-kDa TMP gradually disappeared coincident with the emergence of a virus with a 28-kDa TMP. Virus production dramatically increased in parallel with the emergence of a virus with a 28-kDa TMP. Sequence analysis of viral DNAs from these cultures showed that premature stop codons arising by point mutation were responsible for the change in size of the TMP with time. A similar selective pressure for truncated forms of TMP was observed when the SIVmac239 clone was transfected into human peripheral blood lymphocytes (PBL). In contrast, no such selective pressure was observed in macaque PBL. When the SIVmac239 clone was transfected into macaque PBL and the resultant virus was serially passaged in macaque PBL, the virus replicated very well and maintained a 41-kDa TMP for 80 days in culture. Macaque monkeys were infected with SIVmac239 having a 28-kDa TMP; virus subsequently recovered from T4-enriched lymphocytes of peripheral blood showed only the 41-kDa form of TMP. These results indicate that the natural form of TMP in SIVmac is the full-length 41-kDa TMP, just as in human immunodeficiency virus type 1. Viruses with truncated forms of TMP appear to result from mutation and selection during propagation in unnatural human cells

Prospective comparison of microbial culture and polymerase chain reaction in the diagnosis of corneal ulcer.

PURPOSE: To compare polymerase chain reaction (PCR) to microbial culture for the detection and identification of bacterial and fungal pathogens in microbial keratitis. DESIGN: Prospective cohort study. METHODS: A total of 108 consecutive corneal ulcers were cultured and analyzed by PCR using pan-bacterial and pan-fungal primers. PCR products were cloned, sequenced, and compared to culture results using standard bioinformatics tools. RESULTS: Of the 108 samples, 56 were culture-positive, 25 for bacteria and 31 for fungi; 52 were culture-negative. After eliminating false-positive PCR products, 94 of 108 were positive by PCR, 37 for bacteria and 57 for fungi. Nineteen of 25 bacterial culture-positive samples were positive by PCR, and 29 of 31 samples culture-positive for fungi were positive by PCR. The majority of sequenced PCR products matched the positive culture results. Of the 52 culture-negative samples, 46 (88%) yielded pathogen deoxyribonucleic acid (DNA) PCR products, 18 bacterial and 28 fungal. These represented a variety of species, including at least three novel previously uncultured microbes. CONCLUSIONS: PCR detects microbial DNA in the majority of bacterial and fungal corneal ulcers, and identifies potentially pathogenic organisms in a high proportion of culture-negative cases. Yield and concordance with culture are higher for fungal than bacterial ulcers. Practical use of the technique is limited by artefactual amplification of nonpathogenic organisms. PCR may be used as an adjunct to culture to identify potential pathogens in microbial keratitis

Molecularly Self-Assembled Nucleic Acid Nanoparticles for Targeted In Vivo siRNA Delivery

Nanoparticles are employed for delivering therapeutics into cells1,2. However, size, shape, surface chemistry and the presentation of targeting ligands on the surface of nanoparticles can affect circulation half-life and biodistribution, cell specific internalization, excretion, toxicity, and efficacy3-7. A variety of materials have been explored for delivering small interfering RNAs (siRNAs) - a therapeutic agent that suppresses the expression of targeted genes8,9. However, conventional delivery nanoparticles such as liposomes and polymeric systems are heterogeneous in size, composition and surface chemistry, and this can lead to suboptimal performance, lack of tissue specificity and potential toxicity10-12. Here, we show that self-assembled DNA tetrahedral nanoparticles with a well-defined size can deliver siRNAs into cells and silence target genes in tumours. Monodisperse nanoparticles are prepared through the self-assembly of complementary DNA strands. Because the DNA strands are easily programmable, the size of the nanoparticles and the spatial orientation and density of cancer targeting ligands (such as peptides and folate) on the nanoparticle surface can be precisely controlled. We show that at least three folate molecules per nanoparticle is required for optimal delivery of the siRNAs into cells and, gene silencing occurs only when the ligands are in the appropriate spatial orientation. In vivo, these nanoparticles showed a longer blood circulation time (t1/2 ∼ 24.2 min) than the parent siRNA (t1/2 ∼ 6 min)

Estimated event-free survival benefits with dapagliflozin in HF with mildly reduced or preserved ejection fraction

Background: Recent guidelines support consideration of sodium glucose co-transporter(SGLT)-2 inhibitors in the long-term management of heart failure (HF) with mildly reduced or preserved ejection fraction. Patients and clinicians may be interested in the expected lifetime benefits of SGLT-2 inhibitors. Objectives: To estimate event-free survival gains from long-term use of dapagliflozin in patients with HF with mildly reduced or preserved ejection fraction overall and in clinically relevant subgroups. Methods: In this pre-specified analysis of DELIVER, we applied validated nonparametric age-based methods to extrapolate potential gains in survival free from the primary endpoint (cardiovascular death or worsening HF event) from long-term use of dapagliflozin. Eligible participants had symptomatic HF, LVEF>40%, elevated natriuretic peptide levels, and structural heart disease. For every year between age 55 and 85, we estimated event-free survival using age at randomization rather than time from randomization as the time horizon. Residual lifespan free from primary endpoint was estimated based on area under the survival curve in each arm. Results: Among 6,263 participants, mean survival free from the primary endpoint for a 65-year-old participant was 12.1 (95%CI 11.0 to 13.2) years with dapagliflozin and 9.7 (95%CI 8.8 to 10.7) years with placebo, representing a 2.3 (95%CI 0.9 to 3.8) year event-free survival gain(P=0.002). Treatment gains in survival free from the primary endpoint ranged from 2.0 (95%CI -0.6 to 4.6) years in a 55-year-old to 1.2 (95%CI -0.1 to 2.4) years in a 75-year-old. Mean event-free survival was greater with dapagliflozin than with placebo across all 14 subgroups. Conclusions: Treatment with dapagliflozin is projected to extend event-free survival by up to 2-2.5 years among middle age and older individuals with HF with mildly reduced, preserved, or improved ejection fraction

Dapagliflozin in patients recently hospitalized with heart failure and mildly reduced or preserved ejection fraction

Background: Patients recently hospitalized for heart failure (HF) are at high risk for rehospitalization and death. Objectives: The purpose of this study was to investigate clinical outcomes and response to dapagliflozin in patients with HF with mildly reduced or preserved left ventricular ejection fraction (LVEF) who were enrolled during or following hospitalization. Methods: The DELIVER (Dapagliflozin Evaluation to Improve the LIVES of Patients With PReserved Ejection Fraction Heart Failure) trial randomized patients with HF and LVEF >40% to dapagliflozin or placebo. DELIVER permitted randomization during or shortly after hospitalization for HF in clinically stable patients off intravenous HF therapies. This prespecified analysis investigated whether recent HF hospitalization modified risk of clinical events or response to dapagliflozin. The primary outcome was worsening HF event or cardiovascular death. Results: Of 6,263 patients in DELIVER, 654 (10.4%) were randomized during HF hospitalization or within 30 days of discharge. Recent HF hospitalization was associated with greater risk of the primary outcome after multivariable adjustment (HR: 1.88; 95% CI: 1.60-2.21; P < 0.001). Dapagliflozin reduced the primary outcome by 22% in recently hospitalized patients (HR: 0.78; 95% CI: 0.60-1.03) and 18% in patients without recent hospitalization (HR: 0.82; 95% CI: 0.72-0.94; Pinteraction = 0.71). Rates of adverse events, including volume depletion, diabetic ketoacidosis, or renal events, were similar with dapagliflozin and placebo in recently hospitalized patients. Conclusions: Dapagliflozin safely reduced risk of worsening HF or cardiovascular death similarly in patients with and without history of recent HF hospitalization. Starting dapagliflozin during or shortly after HF hospitalization in patients with mildly reduced or preserved LVEF appears safe and effective. (Dapagliflozin Evaluation to Improve the LIVEs of Patients With PReserved Ejection Fraction Heart Failure [DELIVER]; NCT03619213)

Efficacy and safety of dapagliflozin according to frailty in patients with heart failure: a prespecified analysis of the DELIVER trial

Background: Frailty is increasing in prevalence and because frail patients are often perceived to have a less favorable benefit/risk profile, they may be less likely to receive new pharmacological treatments. We investigated the efficacy and tolerability of dapagliflozin according to frailty status in patients with heart failure and mildly reduced and preserved ejection fraction randomized in DELIVER. Methods: Frailty was measured using the Rockwood cumulative deficit approach. The primary endpoint was time to a first worsening heart failure event or cardiovascular death. Results: Of the 6263 patients randomized, a Frailty Index (FI) was calculable in 6258. In total, 2,354 (37.6%) patients had class 1 frailty (FI <0.210, i.e., not frail), 2,413 (38.6%) were in class 2 (FI 0.211-0.310, i.e., more frail), and 1,491 (23.8%) had class 3 frailty (FI >0.311, i.e., most frail). Greater frailty was associated with a higher rate of the primary endpoint (per 100 person years): FI class 1, 6.3 (95% CI 5.7-7.1); class 2, 8.3 (7.5-9.1); and class 3, 13.4 (12.1-14.7), P<0.001. The effect of dapagliflozin (as a hazard ratio) on the primary endpoint from FI class 1 to 3 was: 0.85 (95% CI, 0.68-1.06); 0.89 (0.74-1.08); and 0.74 (0.61-0.91), respectively (Pinteraction=0.40). Although frailer patients had worse KCCQ scores at baseline, the improvement with dapagliflozin was greater than in less frail patients: placebo-corrected improvement in KCCQ-OSS at 4 months FI class 1, 0.3 (95% CI -0.9 to 1.4); class 2, 1.5 (0.3-2.7); and class 3, 3.4 (1.7-5.1) [Pinteraction=0.021]. Adverse reactions and treatment discontinuation, while more frequent in frailer patients, were not more common with dapagliflozin than placebo, irrespective of frailty class. Conclusions: In DELIVER, frailty was common and associated with worse outcomes. The benefit of dapagliflozin was consistent across the range of frailty studied. The improvement in health-related quality of life with dapagliflozin occurred early and was greater in patients with greater frailty

Dapagliflozin across the range of ejection fraction in patients with heart failure: a patient-level, pooled meta-analysis of DAPA-HF and DELIVER

Whether the sodium–glucose cotransporter 2 inhibitor dapagliflozin reduces the risk of a range of morbidity and mortality outcomes in patients with heart failure regardless of ejection fraction is unknown. A patient-level pooled meta-analysis of two trials testing dapagliflozin in participants with heart failure and different ranges of left ventricular ejection fraction (≤40% and >40%) was pre-specified to examine the effect of treatment on endpoints that neither trial, individually, was powered for and to test the consistency of the effect of dapagliflozin across the range of ejection fractions. The pre-specified endpoints were: death from cardiovascular causes; death from any cause; total hospital admissions for heart failure; and the composite of death from cardiovascular causes, myocardial infarction or stroke (major adverse cardiovascular events (MACEs)). A total of 11,007 participants with a mean ejection fraction of 44% (s.d. 14%) were included. Dapagliflozin reduced the risk of death from cardiovascular causes (hazard ratio (HR) 0.86, 95% confidence interval (CI) 0.76–0.97; P = 0.01), death from any cause (HR 0.90, 95% CI 0.82–0.99; P = 0.03), total hospital admissions for heart failure (rate ratio 0.71, 95% CI 0.65–0.78; P < 0.001) and MACEs (HR 0.90, 95% CI 0.81–1.00; P = 0.045). There was no evidence that the effect of dapagliflozin differed by ejection fraction. In a patient-level pooled meta-analysis covering the full range of ejection fractions in patients with heart failure, dapagliflozin reduced the risk of death from cardiovascular causes and hospital admissions for heart failure (PROSPERO: CRD42022346524)

Global, regional, and national age-sex-specific mortality for 282 causes of death in 195 countries and territories, 1980-2017 : a systematic analysis for the Global Burden of Disease Study 2017

Background Global development goals increasingly rely on country-specific estimates for benchmarking a nation's progress. To meet this need, the Global Burden of Diseases, Injuries, and Risk Factors Study (GBD) 2016 estimated global, regional, national, and, for selected locations, subnational cause-specific mortality beginning in the year 1980. Here we report an update to that study, making use of newly available data and improved methods. GBD 2017 provides a comprehensive assessment of cause-specific mortality for 282 causes in 195 countries and territories from 1980 to 2017. Methods The causes of death database is composed of vital registration (VR), verbal autopsy (VA), registry, survey, police, and surveillance data. GBD 2017 added ten VA studies, 127 country-years of VR data, 502 cancer-registry country-years, and an additional surveillance country-year. Expansions of the GBD cause of death hierarchy resulted in 18 additional causes estimated for GBD 2017. Newly available data led to subnational estimates for five additional countries Ethiopia, Iran, New Zealand, Norway, and Russia. Deaths assigned International Classification of Diseases (ICD) codes for non-specific, implausible, or intermediate causes of death were reassigned to underlying causes by redistribution algorithms that were incorporated into uncertainty estimation. We used statistical modelling tools developed for GBD, including the Cause of Death Ensemble model (CODErn), to generate cause fractions and cause specific death rates for each location, year, age, and sex. Instead of using UN estimates as in previous versions, GBD 2017 independently estimated population size and fertility rate for all locations. Years of life lost (YLLs) were then calculated as the sum of each death multiplied by the standard life expectancy at each age. All rates reported here are age-standardised. Findings At the broadest grouping of causes of death (Level 1), non-communicable diseases (NC Ds) comprised the greatest fraction of deaths, contributing to 73.4% (95% uncertainty interval [UI] 72.5-74.1) of total deaths in 2017, while communicable, maternal, neonatal, and nutritional (CMNN) causes accounted for 186% (17.9-19.6), and injuries 8.0% (7.7-8.2). Total numbers of deaths from NCD causes increased from 2007 to 2017 by 22.7% (21.5-23.9), representing an additional 7.61 million (7. 20-8.01) deaths estimated in 2017 versus 2007. The death rate from NCDs decreased globally by 7.9% (7.08.8). The number of deaths for CMNN causes decreased by 222% (20.0-24.0) and the death rate by 31.8% (30.1-33.3). Total deaths from injuries increased by 2.3% (0-5-4-0) between 2007 and 2017, and the death rate from injuries decreased by 13.7% (12.2-15.1) to 57.9 deaths (55.9-59.2) per 100 000 in 2017. Deaths from substance use disorders also increased, rising from 284 000 deaths (268 000-289 000) globally in 2007 to 352 000 (334 000-363 000) in 2017. Between 2007 and 2017, total deaths from conflict and terrorism increased by 118.0% (88.8-148.6). A greater reduction in total deaths and death rates was observed for some CMNN causes among children younger than 5 years than for older adults, such as a 36.4% (32.2-40.6) reduction in deaths from lower respiratory infections for children younger than 5 years compared with a 33.6% (31.2-36.1) increase in adults older than 70 years. Globally, the number of deaths was greater for men than for women at most ages in 2017, except at ages older than 85 years. Trends in global YLLs reflect an epidemiological transition, with decreases in total YLLs from enteric infections, respirator}, infections and tuberculosis, and maternal and neonatal disorders between 1990 and 2017; these were generally greater in magnitude at the lowest levels of the Socio-demographic Index (SDI). At the same time, there were large increases in YLLs from neoplasms and cardiovascular diseases. YLL rates decreased across the five leading Level 2 causes in all SDI quintiles. The leading causes of YLLs in 1990 neonatal disorders, lower respiratory infections, and diarrhoeal diseases were ranked second, fourth, and fifth, in 2017. Meanwhile, estimated YLLs increased for ischaemic heart disease (ranked first in 2017) and stroke (ranked third), even though YLL rates decreased. Population growth contributed to increased total deaths across the 20 leading Level 2 causes of mortality between 2007 and 2017. Decreases in the cause-specific mortality rate reduced the effect of population growth for all but three causes: substance use disorders, neurological disorders, and skin and subcutaneous diseases. Interpretation Improvements in global health have been unevenly distributed among populations. Deaths due to injuries, substance use disorders, armed conflict and terrorism, neoplasms, and cardiovascular disease are expanding threats to global health. For causes of death such as lower respiratory and enteric infections, more rapid progress occurred for children than for the oldest adults, and there is continuing disparity in mortality rates by sex across age groups. Reductions in the death rate of some common diseases are themselves slowing or have ceased, primarily for NCDs, and the death rate for selected causes has increased in the past decade. Copyright (C) 2018 The Author(s). Published by Elsevier Ltd.Peer reviewe