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Along with the nasal epithelium, the lung epithelium is a portal of entry for sudden acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and many other respiratory viruses. In the case of SARS-CoV-2, the virus surface spike proteins bind to the ACE-2 receptor to facilitate entry into respiratory epithelium. Alveolar type 2 (AT2) cells are committed respiratory progenitor cells responsible for the integrity and regeneration of the respiratory epithelium and production of respiratory surfactant proteins. They express high levels of surface ACE-2 and thus are a leading target for primary infection by the SARS-CoV-2. This study describes a method to directly differentiate TERT-immortalized human cord blood-derived multilineage progenitor cells (MLPC) to AT2-like cells for the purpose of generating an in vitro cellular platform for viral studies. Differentiation was confirmed with the acquisition of AT2 and absence of alveolar type 1 (AT1) specific markers by confocal microscopy. Expression of the ACE-2 receptor was confirmed by immunofluorescence antibody staining, qRT-PCR, and binding of biotinylated SARS-CoV-2 spike (S and S1) proteins. The binding of biotinylated spike proteins was specifically blocked by unlabeled spike proteins and neutralizing antibodies. Additionally, it was demonstrated that the spike protein was internalized after binding to the surface membrane of the cells. We defined the culture conditions that enabled AT2-like cells to be repeatedly passaged and cryopreserved without further differentiation to AT1. Our method provides a stable and renewable source of AT2 cells for respiratory viral binding, blocking and uptake studies., Graphical Abstract AT2-like cells differentiated from TERT-immortalized human MLPC expressed ACE-2, bound SARS-CoV-2 spike proteins and differentiated between RBD-specific and non-RBD-specific neutralizing antibodies. This long-lived cell line could provide a reliable tool to study spike protein binding and inhibition by antibodies or other potential therapeutics designed to inhibit infection by the SARS-CoV-2 virus.Image, graphical abstract

The goal of the current study is to assess whether the scores of Brief Rating of Aggression by Children and Adolescents (BRACHA) at the emergency room (ER) can predict the aggressive incidents at pediatric psychiatric inpatient units. The study aims to identify predictors for two outcome measurements: 1) hazard rates for the first aggressive incident and 2) numbers of days between admission and the first aggressive incident, using the Cox regression model and Poisson regression model, respectively. The clinical records of a total of 5,610 adolescents admitted into the pediatric psychiatric inpatient units of Cincinnati Children's Hospital Medical Center were extracted for the analysis. The aggressive incident was defined as a score0 from any category of the Overt Aggression Scale (OAS) and the high-aggressive incident was defined as a score ≥ 2 from any category of the OAS. The results indicate that the BRACHA score was not associated with high-aggressive incidents (hazard ratio: 0.98, p = 0.7543). Similarly, the BRACHA scores was only associated with the number of days from admission to the first aggressive incident (Poisson regression coefficient: 0.24, p 0.0001) but not the number of days from admission to the first high-aggressive incident (Poisson regression coefficient: 0.03, p = 0.3994). Furthermore, the second peak of first aggressive incidents during the hospitalization highlights the importance of interventions at the end of the inpatient treatment course. To summarize, BRACHA scores based on initial assessments at the ER could correlate with first aggressive incidents, but not the first high-aggressive incidents.

Background aims Adoptive transfer of suppressive CD4+CD25+ thymic regulatory T cells (tTregs) can control auto- and alloimmune responses but typically requires in vitro expansion to reach the target cell number for efficacy. Although the adoptive transfer of expanded tTregs purified from umbilical cord blood ameliorates graft-versus-host disease in patients receiving hematopoietic stem cell transplantation for lymphohematopoietic malignancy, individual Treg products of 100 × 106 cells/kg are manufactured over an extended 19-day time period using a process that yields variable products and is both laborious and costly. These limitations could be overcome with the availability of ‘off the shelf’ Treg. Results Previously, the authors reported a repetitive restimulation expansion protocol that maintains Treg phenotype (CD4+25++127-Foxp3+), potentially providing hundreds to thousands of patient infusions. However, repetitive stimulation of effector T cells induces a well-defined program of exhaustion that leads to reduced T-cell survival and function. Unexpectedly, the authors found that multiply stimulated human tTregs do not develop an exhaustion signature and instead maintain their Treg gene expression pattern. The authors also found that tTregs expanded with one or two rounds of stimulation and tTregs expanded with three or five rounds of stimulation preferentially express distinct subsets of a group of five transcription factors that lock in Treg Foxp3expression, Treg stability and suppressor function. Multiply restimulated Tregs also had increased transcripts characteristic of T follicular regulatory cells, a Treg subset. Discussion These data demonstrate that repetitively expanded human tTregs have a Treg-locking transcription factor with stable FoxP3 and without the classical T-cell exhaustion gene expression profile—desirable properties that support the possibility of off-the-shelf Treg therapeutics.

Mammalian target of rapamycin (mTOR) inhibitors are clinically effective at treating some complex lymphatic malformations (LM). The mTOR inhibitor rapamycin blocks the phosphoinositide 3-kinase (PI3K) pathway, which is commonly mutated in this condition. Although rapamycin is effective at controlling symptoms of LM, treatment courses are long, not all LMs respond to treatment, and many patients relapse after treatment has stopped. Concurrent rat sarcoma virus (RAS) pathway abnormalities have been identified in LM, which may limit the effectiveness of rapamycin. Protein tyrosine phosphatase-2 (SHP2) controls the RAS pathway upstream, and SHP2 inhibitors are being investigated for treatment of various tumors. The objective of this study was to determine the impact of SHP2 inhibition in combination with rapamycin on LM growth in vitro. Using primary patient cells isolated from a surgically resected LM, we found that combination treatment with rapamycin and the SHP2 inhibitor SHP099 caused a synergistic reduction in cell growth, migration and lymphangiogenesis. These results suggest that combination treatment targeting the PI3K and RAS signaling pathways may result in effective treatment of LMs of the head and neck.

The goal of this study was to evaluate a novel risk stratification scheme to categorize patients on the basis of risk to either an operating room or device laboratory with rescue strategy.Lead extraction can be complicated by lethal issues such as vascular and cardiac rupture. Currently, the optimal site for lead extraction has not been well established.A risk stratification scheme was developed from previously available risk factors for major complications. Patients were prospectively risk stratified between October 2013 and January 2016. High-risk procedures were performed in the operating room with ready surgical services; intermediate-risk procedures were performed in the device laboratory.In total, 349 leads were removed from 187 patients (age 61.0 ± 17.2 years; 66.3% men) over 27 months. Seventy-two patients (38.5%) were categorized as high risk. Median implant duration of the oldest lead per patient was 11.2 years (interquartile range: 7.9 to 14.9 years) in the operating room group versus 2.6 years (interquartile range: 1.6 to 4.9 years) in the device laboratory group (p 0.001). Clinical success in the operating room (95.8%) and device laboratory (99.1%) groups was similar (p = 0.16). A higher incidence of major complications occurred in the high-risk group (operating room group: 6.9%; device laboratory: 0.0%; p = 0.007). In-hospital mortality (operating room group: 8.3%; device laboratory: 2.6%; p = 0.09) and long-term (2-year) survival (operating room: 70.8%; device laboratory: 84.4%; p = 0.07) rates were similar.Use of a novel risk stratification scheme in guiding the selection of operating room versus device laboratory for lead extraction is feasible, safe, and efficacious. Intermediate-risk procedures can be performed safely in the device laboratory with rescue strategy, without excess surgical resource utilization.

Intradetrusor onabotulinumtoxinA injection is an effective advanced treatment for overactive bladder. While the effective dosages have been well studied, very little data exist on treatment efficacy using differing injection techniques. The objective of this study was to determine whether the efficacy of a reduced injection technique of 5 injection sites was noninferior to the standard technique of 20 injection sites of intradetrusor onabotulinumtoxinA for treatment of overactive bladder.In this randomized noninferiority trial, men and women seeking treatment of overactive bladder with intradetrusor onabotulinumtoxinA injections were randomized to receive 100 units administered via either a reduced injection technique of 5 injection sites or a standard injection technique of 20 injection sites. Subjects completed a series of standardized questionnaires at baseline and at 4-12 weeks postprocedure to determine symptom severity and treatment efficacy. The primary outcome was treatment efficacy as determined by Overactive Bladder Questionnaire-Short Form with a noninferiority margin of 15 points. Secondary outcomes were incidence of urinary tract infection and urinary retention requiring catheterization.Data from 77 subjects were available for analysis with 39 in the control arm (20 injections) and 38 in the study arm (5 injections). There was a significant improvement in both arms from baseline to follow-up in Overactive Bladder Questionnaire-Short Form and International Consultation on Incontinence Questionnaire scores (p 0.001). Overall treatment success was 68% with no statistically significant difference between arms. A significant difference between arms was found on the Overactive Bladder Questionnaire-Short Form quality of life survey favoring the control arm (confidence interval [CI]: 0.36-20.5, p = 0.04). However, there were no significant differences between arms in the remaining validated questionnaires. The study arm did not demonstrate noninferiority to the control arm. Subjects in the study arm were significantly more likely to express a willingness to undergo the procedure again (odds ratio = 3.8, 95% CI: 1.42-10.67, p = 0.004). Adverse events did not differ between arms.A reduced injection technique for administration of intradetrusor onabotulinumtoxinA demonstrates similar efficacy to the standard injection technique but did not demonstrate noninferiority. Subjects preferred the reduced injection technique over the standard technique. A reduced injection technique is a safe and effective alternative to the standard technique.

Summary Methods to deliver gene editing agents in vivo as ribonucleoproteins could offer safety advantages over nucleic acid delivery approaches. We report the development and application of engineered DNA-free virus-like particles (eVLPs) that efficiently package and deliver base editor or Cas9 ribonucleoproteins. By engineering VLPs to overcome cargo packaging, release, and localization bottlenecks, we developed fourth-generation eVLPs that mediate efficient base editing in several primary mouse and human cell types. Using different glycoproteins in eVLPs alters their cellular tropism. Single injections of eVLPs into mice support therapeutic levels of base editing in multiple tissues, reducing serum Pcsk9 levels 78% following 63% liver editing, and partially restoring visual function in a mouse model of genetic blindness. In vitro and in vivo off-target editing from eVLPs was virtually undetected, an improvement over AAV or plasmid delivery. These results establish eVLPs as promising vehicles for therapeutic macromolecule delivery that combine key advantages of both viral and nonviral delivery., Graphical abstract, Highlights • Engineered virus-like particles (eVLPs) overcome three bottlenecks to protein delivery • DNA-free eVLPs efficiently deliver gene editing proteins with minimal off-target editing • Base editor eVLPs reduced serum Pcsk9 levels 78% following 63% liver editing in mice • Base editor eVLPs improved visual function in a mouse model of genetic blindness, Engineered, DNA-free virus-like particles efficiently deliver gene editing proteins, have minimal off-target effects, can be applied in vivo to deliver base editors to multiple organs, and are used to improve visual function in a mouse model of genetic blindness.

Classic implants for operative intervention of mid-shaft clavicle fractures in adolescents can become symptomatic, requiring removal. This study compares 1) biomechanical properties and 2) clinical outcomes in adolescents of mid-shaft clavicle fracture fixation with bioabsorbable versus metal implants.Six synthetic clavicles with mid-shaft fractures underwent bioabsorbable plating. A testing frame applied 10 non-destructive torsion and 10 axial compression cycles, followed by cantilever bending to failure. Stiffness was calculated; maximum failure load and failure mode were recorded. Results were compared to previous data for locked metal constructs. Retrospective review of surgically treated clavicle fractures over three years included functional and radiographic outcomes.Bioabsorbable plates had lower torsional stiffness (P 0.001) and maximum cantilever load (P 0.0001) than locked metal plates. There was no significant difference in compression stiffness (P = 0.2) or cantilever bending stiffness (P = 0.4). Primary failure of metal plates was screw pull-out compared to plate bending in bioabsorbable constructs. Seven patients with bioabsorbable implants were included. All patients with bioabsorbable constructs achieved radiographic union, but 71% lost reduction. Despite fracture angulation, all achieved normal shoulder function after one year determined by QuickDASH evaluation. None required a second surgery.Bioabsorbable implants had lower torsional stiffness and cantilever failure load, but comparable compression stiffness to metal implants. Bioabsorbable implants failed via gradual bending versus the catastrophic failure seen in metal implants. The clinical review allows understanding of the sequelae of this lower failure load of bioabsorbable plates where their use allowed in fracture displacement, yet achievement of ultimate radiographic union and acceptable functional outcomes.

Highlights: • The holopelagic amphipod Paraphronima is shown to contain hidden genetic diversity. • Diversity appears biogeographically restricted to regional scales. • A new species is described using a morpho-molecular approach. • Compound eye characters are shown to be useful in distinguishing species. • The inclusion of compound eyes is recommended in taxonomic studies of Hyperiidea. Holopelagic animals were long assumed to have widespread geographic distributions due to the failure to recognize hydrographic species' barriers in the open ocean. As molecular genetic tools are more commonly used to study the ocean's inhabitants, diversity is found to be substantially higher than when inferred from morphological taxonomies alone. Here, we investigate the morphological and genetic diversity of hyperiid amphipods within the genus Paraphronima, currently comprising two supposedly cosmopolitan species. By combining phylogenetic analyses and four species delimitation methods (GMYC, mPTP, bPTP, ABGD), we reveal substantial species-level genetic variation. Instead of two species inhabiting multiple ocean basins, the biogeography of Paraphronima species appears to be limited to more regional scales. Moreover, there is morphological evidence to corroborate the observed genetic diversity. By using an integrative morpho-molecular approach, a third species from the Gulf of California, Paraphronima robisoni sp. nov., is described. Interestingly, the morphological characters that best distinguish the species within the genus are characters of the compound eyes, which have rarely been used for taxonomy despite being the most obvious and varied features of hyperiids. Our results warrant further investigation of presumably cosmopolitan holopelagic amphipods, while we recommend the inclusion of eye morphology in future taxonomic studies.

Summary While prime editing enables precise sequence changes in DNA, cellular determinants of prime editing remain poorly understood. Using pooled CRISPRi screens, we discovered that DNA mismatch repair (MMR) impedes prime editing and promotes undesired indel byproducts. We developed PE4 and PE5 prime editing systems in which transient expression of an engineered MMR-inhibiting protein enhances the efficiency of substitution, small insertion, and small deletion prime edits by an average 7.7-fold and 2.0-fold compared to PE2 and PE3 systems, respectively, while improving edit/indel ratios by 3.4-fold in MMR-proficient cell types. Strategic installation of silent mutations near the intended edit can enhance prime editing outcomes by evading MMR. Prime editor protein optimization resulted in a PEmax architecture that enhances editing efficacy by 2.8-fold on average in HeLa cells. These findings enrich our understanding of prime editing and establish prime editing systems that show substantial improvement across 191 edits in seven mammalian cell types., Graphical abstract, Highlights • Pooled CRISPRi screens reveal that MMR inhibits prime editing efficiency and precision • PE4 and PE5 enhance editing outcomes through co-expression of dominant negative MLH1 • Programming additional silent mutations can enhance prime editing by evading MMR • PEmax editor improves prime editing efficacy in synergy with PE4, PE5, and epegRNAs, PE4 and PE5 are efficient and precise prime editing systems developed by leveraging insights into the way DNA repair pathways impact genome editing outcomes

Blockers of the renin-angiotensin system are effective in the treatment of experimental and clinical diabetic nephropathy. An approach different from blocking the formation or action of angiotensin II (1-8) that could also be effective involves fostering its degradation. Angiotensin-converting enzyme 2 (ACE2) is a monocarboxypeptidase that cleaves angiotensin II (1-8) to form angiotensin (1-7). Therefore, we examined the renal effects of murine recombinant ACE2 in mice with streptozotocin-induced diabetic nephropathy as well as that of amplification of circulating ACE2 using minicircle DNA delivery prior to induction of experimental diabetes. This delivery resulted in a long-term sustained and profound increase in serum ACE2 activity and enhanced ability to metabolize an acute angiotensin II (1-8) load. In mice with streptozotocin-induced diabetes pretreated with minicircle ACE2, ACE2 protein in plasma increased markedly and this was associated with a more than 100-fold increase in serum ACE2 activity. However, minicircle ACE2 did not result in changes in urinary ACE2 activity as compared to untreated diabetic mice. In both diabetic groups, glomerular filtration rate increased significantly and to the same extent as compared to non-diabetic controls. Albuminuria, glomerular mesangial expansion, glomerular cellularity, and glomerular size were all increased to a similar extent in minicircle ACE2-treated and untreated diabetic mice, as compared to non-diabetic controls. Recombinant mouse ACE2 given for 4 weeks by intraperitoneal daily injections in mice with streptozotocin-induced diabetic nephropathy also failed to improve albuminuria or kidney pathology. Thus, a profound augmentation of ACE2 confined to the circulation failed to ameliorate the glomerular lesions and hyperfiltration characteristic of early diabetic nephropathy. These findings emphasize the importance of targeting the kidney rather than the circulatory renin angiotensin system to combat diabetic nephropathy.

Background Anxiety is common following a traumatic brain injury (TBI), but who is most at risk, and to what extent, is not well understood. Methods Longitudinal data from a randomly-selected community sample (Wave 1: 7397, Wave 2: 6621 and Wave 3: 6042) comprising three adult cohorts (young: 20–24 years of age, middle-aged: 40–44, older: 60–64), were analysed. The association between TBI history, anxiety and comorbid depression was assessed, controlling for age, sex, marital/employment status, medical conditions, recent life events, alcohol consumption, social support and physical activity. Results Thirteen percent of the sample had sustained a TBI by Wave 3, 35% of whom had sustained multiple TBIs. Cross-sectional analyses revealed that clinically-significant anxiety was more common in people who had sustained a TBI. Longitudinal analyses demonstrated an increased risk of anxiety post-TBI, even after controlling for potential demographic, health and psychosocial confounds. Anxiety was more common than depression, although 10% of those with a TBI experienced comorbid anxiety/depression. Limitations TBIs were not medically confirmed and anxiety and depression were only assessed every four years by self-report, rather than clinical interview. Sample attrition resulted in the retention of healthier individuals at each wave. Conclusions TBIs are associated with a lifelong increased risk of experiencing clinically-significant anxiety, highlighting the chronic nature of TBI sequelae. Positive lifestyle changes (e.g., increasing physical activity, reducing alcohol consumption) may decrease the risk of anxiety problems in the early years after a TBI. Comorbid anxiety and depression was common, indicating that both should be monitored and treated.

To describe national trends in cystectomy at the time of urinary diversion for benign indications. Multiple practice patterns exist regarding the necessity for concomitant cystectomy with urinary diversion for benign end-stage lower urinary tract dysfunction. Beyond single-institution reports, limited data are available to describe how concurrent cystectomy is employed on a national level.A representative sample of patients undergoing urinary diversion for benign indications with or without concurrent cystectomy was identified from the Healthcare Cost and Utilization Project Nationwide Inpatient Sample from 1998 to 2011. Using multivariate logistic regression models, we identified hospital- and patient-level characteristics associated with concomitant cystectomy with urinary diversion.There was an increase in the proportion of concomitant cystectomy at the time of urinary diversion from 20% to 35% (P .001) between 1998 and 2011. The increase in simultaneous cystectomy over time occurred at teaching hospitals (vs community hospitals), in older patients, in male patients, in the Medicare population (vs private insurance and Medicaid), and in those with certain diagnoses.There has been an overall increase in the use of cystectomy at the time of urinary diversion for benign indications on a national level, although the indications driving this clinical decision appear inconsistent.

Summary Transparent zooplankton and nekton are often nearly invisible when viewed under ambient light in the pelagic zone [1–3]. However, in this environment, where the light field is directional (and thus likely to cause reflections), and under the bioluminescent searchlights of potential predators, animals may be revealed by reflections from their body surface [4–7]. We investigated the cuticle surfaces of seven species of hyperiids (Crustacea; Amphipoda) using scanning electron microscopy and found two undocumented features that may reduce reflectance. We found that the legs of Cystisoma spp. (n = 5) are covered with an ordered array of nanoprotuberances 200 ± 20 nm SD in height that function optically as a gradient refractive index material [6, 8, 9]. Additionally, we observed that Cystisoma and six other species of hyperiids are covered with a monolayer of homogenous spheres (diameters ranging from 52 ± 7 nm SD on Cystisoma spp. to 320 ± 15 nm SD on Phronima spp.). Optical modeling using effective medium theory and transfer matrix methods demonstrated that both the nanoprotuberances and the monolayers reduce reflectance by as much as 100-fold, depending on the wavelength and angle of the incident light and the thickness of the gradient layer. Even though we only consider surface reflectance and not internal light scattering, our study demonstrates that these nanoprotuberances and spheres can improve crypsis in a featureless habitat where the smallest reflection can render an animal vulnerable to visual predation.

Recessive dystrophic epidermolysis bullosa (RDEB) is a disease caused by mutations in the COL7A1 gene that result in absent or dysfunctional type VII collagen protein production. Clinically, RDEB manifests as early and severe chronic cutaneous blistering, damage to internal epithelium, an increased risk for squamous cell carcinoma, and an overall reduced life expectancy. Recent localized and systemic treatments have shown promise for lessening the disease severity in RDEB, but the concept of ex vivo therapy would allow a patient's own cells to be engineered to express functional type VII collagen. Here, we review gene delivery and editing platforms and their application toward the development of next-generation treatments designed to correct the causative genetic defects of RDEB.

The Flipped Classroom design has arguably revolutionized teaching across the globe. In this article we report the first comprehensive meta-analysis of its effects on student performance, relative to traditional teaching models, across disciplines and education level. We included 198 studies comprising 33,678 students in our meta-analysis. There were 174 studies conducted at the tertiary level, 21 at secondary, and three at primary school level. Overall, the flipped classroom had a moderate positive effect (g = .50) on student performance. The flipped classroom was beneficial regardless of discipline, with effect sizes ranging from weak (i.e., for IT, g = 0.30; k = 14) to strong (i.e., for humanities, g = 0.98; k = 34). Testing of other moderators relating to group sizes, in-class activities, pre-class testing, and the sophistication of a flipped classroom design provide evidence to suggest that the primary contributing factor to the flipped classroom effect is the opportunity it provides for structured, active learning and problem-solving.

Background: Ambroxol (ABX) has been suggested as an augmentative pharmacological agent for neuronopathic Gaucher disease (nGD). This study assessed the long-term safety and efficacy of combined therapy with high-dose ABX and enzyme replacement therapy (ERT) in nGD. Methods: Four patients with nGD were enrolled. ABX ERT therapy was administered for 4·5 years. Ambroxol was initiated at a dose of 1·5 mg/kg/day, and the dose was escalated up to 27 mg/kg/day. The changes in glucocerebrosidase (GBA) activity, biochemical, safety, and neurocognitive findings were assessed. Findings: Enhanced residual GBA activity was observed in all patients, as evidenced in both in vitro and in vivo studies. Mean seizure frequencies markedly decreased from the baseline, and the neurocognitive function was maintained. Lyso-Gb1, a biomarker for the severity and progression of GD, gradually decreased in all patients. High-dose ABX was well-tolerated with no severe adverse events. Interpretation: Long-term treatment with high-dose ABX ERT was safe and shows promise for arresting the progression of the neurological manifestations in GD. Clinical Trial Number: The study protocol was registered at the Clinical Research Information Service, Korea Centers for Disease Control and Prevention, Ministry of Health and Welfare (Republic of Korea) (no. KCT0003218). Funding Statement: This research was supported in part by the Bio & Medical Technology Development Program of the National Research Foundation (NRF) funded by the Korean government (NRF- 2015K1A4A3046807, NRF-2016M3A9B4915706 and NRF-2018M3A9H1078335) and by ISU ABXIS, Gyeonggi-do, Korea. Declaration of Interests: "None declared." Ethics Approval Statement: The study protocol was approved by the Institutional Review Board and the Ministry of Food and Drug Safety, Korea (no. 30449).

Objective To compare the efficacy and safety in a contemporary cohort of women who were offered either a pubovaginal sling (PVS) or a synthetic midurethral sling (MUS) after the US Food and Drug Administration notification and made an informed decision on procedure option. Methods A total of 201 women were given the option between a PVS and an MUS. Prior anti-incontinence surgery and concomitant surgery other than hysterectomy were not allowed. Minimal follow-up was 12 months. Patients were prospectively followed with validated quality of life questionnaires. Cure, voiding complaints, and complications were compared between the groups. Results Ninety-one women (45%) underwent PVS and 110 underwent MUS (55%). Median follow-up was 13.8 months. There was no difference in baseline characteristics between the groups except for the prevalence of urge incontinence. Subjective improvement in questionnaire scores was significant for both groups. Cure rate was accomplished in 75.8% of the PVS group patients compared with 80.9% of the MUS group patients (hazard ratio, 1.35; 95% confidence interval, 0.69-2.7; P = .38). Complications and voiding difficulty were similar between the groups. Conclusion In this contemporary cohort of women considered suitable candidates for either a PVS or an MUS, both offer comparable efficacy and complication rates. PVS may be safely offered to patients who would otherwise be good candidates for MUS if they are concerned with the implantation of mesh.

Summary The mesopelagic habitat is a vast space that lacks physical landmarks and is structured by depth, light penetration, and horizontal currents. Solar illumination is visible in the upper 1,000 m of the ocean, becoming dimmer and spectrally filtered with depth—generating a nearly monochromatic blue light field [1]. The struggle to perceive dim downwelling light and bioluminescent sources and the need to remain unseen generate contrasting selective pressures on the eyes of mesopelagic inhabitants [2]. Hyperiid amphipods are cosmopolitan members of the mesopelagic fauna with at least ten different eye configurations across the family—ranging from absent eyes in deep-living species to four enlarged eyes in mesopelagic individuals [3–7]. The hyperiid amphipod Paraphronima gracilis has a pair of bi-lobed apposition compound eyes, each with a large upward-looking portion and a small lateral-looking portion. The most unusual feature of the P. gracilis eye is that its upward-looking portion is resolved into a discontinuous retina with 12 distinct groups, each serving one transverse row of continuously spaced facets. On the basis of eye morphology, we estimated spatial acuity (2.5° ± 0.11°, SEM; n = 25) and optical sensitivity (30 ± 3.4 μm 2 ⋅ sr, SEM; n = 25). Microspectrophotometry showed that spectral sensitivity of the eye peaked at 516 nm (±3.9 nm, SEM; n = 6), significantly offset from the peak of downwelling irradiance in the mesopelagic realm (480 nm). Modeling of spatial summation within the linear retinal groups showed that it boosts sensitivity with less cost to spatial acuity than more typical configurations.

a b s t r a c t Background: Depression is one of the most frequently reported psychological problems following TBI, however prevalence estimates vary widely. Methodological and sampling differences may explain some of this variability, but it is not known to what extent. Methods: Data from 99 studies examining the prevalence of clinically diagnosed depression (MDD/ dysthymia) and self-reports of depression (clinically significant cases or depression scale scores) fol- lowing adult, non-penetrating TBI were analysed, taking into consideration diagnostic criteria, measure, post-injury interval, and injury severity. Results: Overall, 27% of people were diagnosed with MDD/dysthymia following TBI and 38% reported clini- cally significant levels of depression when assessed with self-report scales. Estimates of MDD/dysthymia varied according to diagnostic criteria (ICD-10: 14%; DSM-IV: 25%; DSM-III: 47%) and injury severity (mild: 16%; severe: 30%). When self-report measures were used, the prevalence of clinically significant cases of depression differed between scales (HADS: 32%; CES-D: 48%) method of administration (phone: 26%; mail 46%), post-injury interval (range: 33-42%), and injury severity (mild: 64%; severe: 39%). Conclusion: Depression is very common after TBI and has the potential to impact on recovery and quality of life. However, the diagnostic criteria, measure, time post-injury and injury severity, all impact on prevalence rates and must therefore be considered for benchmarking purposes. Crown Copyright © 2014 Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).

Spontaneous reversion of disease-causing mutations has been observed in some genetic disorders. In our clinical observations of severe generalized recessive dystrophic epidermolysis bullosa (RDEB), a currently incurable blistering genodermatosis caused by loss-of-function mutations in COL7A1 that results in a deficit of type VII collagen (C7), we have observed patches of healthy-appearing skin on some individuals. When biopsied, this skin revealed somatic mosaicism resulting from the self-correction of C7 deficiency. We believe this source of cells could represent an opportunity for translational “natural” gene therapy. We show that revertant RDEB keratinocytes expressing functional C7 can be reprogrammed into induced pluripotent stem cells (iPSCs) and that self-corrected RDEB iPSCs can be induced to differentiate into either epidermal or hematopoietic cell populations. Our results give proof in principle that an inexhaustible supply of functional patient-specific revertant cells can be obtained—potentially relevant to local wound therapy and systemic hematopoietic cell transplantation. This technology may also avoid some of the major limitations of other cell therapy strategies, e.g., immune rejection and insertional mutagenesis, which are associated with viral- and nonviral- mediated gene therapy. We believe this approach should be the starting point for autologous cellular therapies using “natural” gene therapy in RDEB and other diseases.

Identifying the cause of an out-of-hospital cardiac arrest can be challenging. In a patient with chest pain, documented ventricular tachycardia, a structurally normal heart, and non-obstructive coronary artery disease, what else should be considered? A 48 year old man with Graves' Disease suffered

The purpose of this article was to review the epidemiology, pathophysiology, and treatment options for stress urinary incontinence in the obese female patient and draw conclusions regarding the optimal treatment of this condition in this unique patient population. Obesity results in increased intra-abdominal pressure and this leads to weakening of the pelvic floor innervation and musculature. Weight loss through lifestyle modification and bariatric surgery improves stress urinary incontinence. Success of stress urinary incontinence surgery in obese women is similar to nonobese patients. Obese women should not be excluded from potentially curative surgery based on their body mass index (BMI) alone.

The development of research into the history of tree growth and inferred summer temperature changes in Yamalia spanning the last 2000 years is reviewed. One focus is the evolving production of tree-ring width (TRW) and tree-ring maximum-latewood density (MXD) larch (Larix sibirica) chronologies, incorporating different applications of Regional Curve Standardisation (RCS). Another focus is the comparison of independent data representing past tree growth in adjacent Yamalia areas: Yamal and Polar Urals, and the examination of the evidence for common growth behaviour at different timescales. The sample data we use are far more numerous and cover a longer time-span at Yamal compared to the Polar Urals, but Yamal has only TRW, while there are both TRW and MXD for the Polar Urals. We use more data (sub-fossil and from living trees) than in previous dendroclimatic studies in this region. We develop a new TRW chronology for Yamal, more than 2000 years long and running up to 2005. For the Polar Urals we develop new TRW and MXD chronologies that show good agreement at short (

Recessive dystrophic epidermolysis bullosa (RDEB) is characterized by a functional deficit of type VII collagen protein due to gene defects in the type VII collagen gene (COL7A1). Gene augmentation therapies are promising, but run the risk of insertional mutagenesis. To abrogate this risk, we explored the possibility of using engineered transcription activator-like effector nucleases (TALEN) for precise genome editing. We report the ability of TALEN to induce site-specific double-stranded DNA breaks (DSBs) leading to homology-directed repair (HDR) from an exogenous donor template. This process resulted in COL7A1 gene mutation correction in primary fibroblasts that were subsequently reprogrammed into inducible pluripotent stem cells and showed normal protein expression and deposition in a teratoma-based skin model in vivo. Deep sequencing-based genome-wide screening established a safety profile showing on-target activity and three off-target (OT) loci that, importantly, were at least 10 kb from a coding sequence. This study provides proof-of-concept for TALEN-mediated in situ correction of an endogenous patient-specific gene mutation and used an unbiased screen for comprehensive TALEN target mapping that will cooperatively facilitate translational application.

A B S T R A C T Objective: To examine middle ear volume in patients with aural atresia and investigate the role of middle ear volume as an adjunct measure in determining surgical candidacy. Methods: We performed a retrospective review of children with aural atresia in a tertiary academic pediatric otolaryngology practice. High resolution multiplanar CT scans of the temporal bones were analyzed for middle ear volume and staged according to existing clinical grading scales. Atretic ears were compared to the nonatretic ears of the same patient as well as to ears of a control population. Results: The average age of patients at the time CT was performed was 4.7 years (range

Recent warming in the Antarctic has led to increased production of icebergs; however, the ecological effects of icebergs on pelagic communities within the Southern Ocean have not been well-studied. We used a 10 m 2 MOCNESS to collect macrozooplankton and micronekton in the upper 300 m of the water column near free-drifting icebergs in the Atlantic sector of the Southern Ocean during three seasons: December 2005 (late spring), June 2008 (late fall) and March-April 2009 (late summer). Communities were dominated in all three seasons by Antarctic krill ( Euphausia superba ) and salps ( Salpa thompsoni ), which collectively comprised 60-95% of the community wet biomass in most cases. During our spring and summer cruises, mean biomass was elevated by 3.1-4.3x at a distance of 0.37 km from large icebergs vs . 9.26 km away. These differences were not statistically significant, and no trend in biomass with distance was apparent in samples from fall 2008, when total biomass was an order of magnitude lower. Biomass levels near icebergs during Dec 2005 and Mar-Apr 2009 were comparable to values reported from marginal ice zones, suggesting that waters around icebergs support macrozooplankton and micronekton communities comparable in magnitude to those in some of the most productive areas of the Southern Ocean. Sample variance also was significantly higher within 1.85 km of icebergs during Dec 2005 and Mar-Apr 2009, reflecting increased patchiness on scales sampled by the MOCNESS (20–40×10 3 m 3 filtered per sample). This pattern was not significant during Jun 2008. Large predatory medusae were observed within 1.85 km of icebergs and in Iceberg Alley, an area through which icebergs pass frequently, but were virtually absent in areas remote from icebergs. Small euphausiids showed an inverse distribution, with low densities in areas populated by large medusae. A shift in community composition from a near-iceberg assemblage dominated by herbivores to a carnivore-dominated community in Iceberg Alley may reflect a transition from bottom-up to top-down control with increasing distance and time. Body sizes of dominant species varied seasonally but did not show consistent trends with distance from icebergs. Concentrations of photosynthetic pigments in the guts of E. superba and S. thompsoni corresponded broadly to patterns in surface chlorophyll a concentrations and were comparable to maximum gut pigment concentrations measured in animals collected from highly productive marginal ice zones. Our results suggest that the macrozooplankton and micronekton assemblages near free-drifting icebergs can be quantitatively and qualitatively different from those in surrounding, iceberg-free waters, perhaps due to both bottom-up and top-down processes as well as physical forcing by the passage of a large object through the upper ocean.

A small ROV was used to collect plankton, make video surveys and take hydrographic measurements in close proximity to six free-drifting, Antarctic icebergs. The icebergs studied ranged in size from 32 km in length. Large icebergs have a greater scale of influence than do smaller ones and iceberg-mediated differences in the hydrographic characteristics of their surrounding water depend on the scale sampled. Irrespective of size, temperature generally decreased in close proximity to an iceberg while salinity increased. Chlorophyll a was often lower in the surface waters near the iceberg, relative to the surface waters further away. Tabular icebergs typically had 3 distinct underwater features: shelf, side and bottom. Ablation pockets were a common feature of subsurface ice. The ice itself is a dynamic and seemingly harsh environment with relatively few macrofauna living on it. Those that do inhabit the ice face are either highly specialized or highly mobile. Species composition of zooplankton within 40 m of an iceberg did not change relative to distance. However, biomass was generally greater within 5 m of an iceberg than it was 15 to 40 m distant.

Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disease characterized by mutations to the α-L-iduronidase (IDUA) gene resulting in inactivation of the IDUA enzyme. The loss of IDUA protein results in the progressive accumulation of glycosaminoglycans within the lysosomes resulting in severe, multi-organ system pathology. Gene replacement strategies have relied on the use of viral or nonviral gene delivery systems. Drawbacks to these include laborious production procedures, poor efficacy due to plasmid-borne gene silencing, and the risk of insertional mutagenesis. This report demonstrates the efficacy of a nonintegrating, minicircle (MC) DNA vector that is resistant to epigenetic gene silencing in vivo. To achieve sustained expression of the immunogenic IDUA protein we investigated the use of a tissue-specific promoter in conjunction with microRNA target sequences. The inclusion of microRNA target sequences resulted in a slight improvement in long-term expression compared to their absence. However, immune modulation by costimulatory blockade was required and permitted for IDUA expression in MPS I mice that resulted in the biochemical correction of pathology in all of the organs analyzed. MC gene delivery combined with costimulatory pathway blockade maximizes safety, efficacy, and sustained gene expression and is a new approach in the treatment of lysosomal storage disease.

Lead macrodislodgement is an undesirable complication after implantation of cardiac implantable electronic devices (CIED). The mechanisms of macrodislodgement, including Twiddler, Reel and Ratchet types, have been previously described in case reports and case series. The prevalence of each type has

Nascent, full length, immunoglobulin (Ig) heavy (H)-chains are post-translationally associated with H-chain-binding protein (BiP or GRP78) in the endoplasmic reticulum (ER). The first constant (C) domain, CH1 of a C gene (Cmu, Cgamma, Calpha), is important for this interaction. The contact is released upon BiP replacement by conventional Ig light (L)-chain (kappa or lambda). Incomplete or mutated H-chains with removed variable (VH) and/or C(H)1 domain, as found in H-chain disease (HCD), can preclude stable BiP interaction. Progression in development after the preB cell stage is dependent on surface expression of IgM when association of a micro H-chain with a L-chain overcomes the retention by BiP. We show that IgM lacking the BiP-binding domain is displayed on the cell surface and elicits a signal that allows developmental progression even without the presence of L-chain. The results are reminiscent of single chain Ig secretion in camelids where developmental processes leading to the generation of fully functional H-chain-only antibodies are not understood. Furthermore, in the mouse the largest secondary lymphoid organ, the spleen, is not required for H-chain-only Ig expression and the CD5 survival signal may be obsolete for cells expressing truncated IgM.

This paper describes the development and first results of the ''Community Integrated Assessment System'' (CIAS), a unique multi-institutional modular and flexible integrated assessment system for modelling climate change. Key to this development is the supporting software infrastructure, SoftIAM. Through it, CIAS is distributed between the communities of institutions which has each contributed modules to the CIAS system. At the heart of SoftIAM is the Bespoke Framework Generator (BFG) which enables flexibility in the assembly and composition of individual modules from a pool to form coupled models within CIAS, and flexibility in their deployment onto the available software and hardware resources. Such flexibility greatly enhances modellers' ability to re-configure the CIAS coupled models to answer different questions, thus tracking evolving policy needs. It also allows rigorous testing of the robustness of IA modelling results to the use of different component modules representing the same processes (for example, the economy). Such processes are often modelled in very different ways, using different paradigms, at the participating institutions. An illustrative application to the study of the relationship between the economy and the earth's climate system is provided.

The rigid cell wall peptidoglycan (murein) is a single giant macromolecule whose shape determines the shape of the bacterial cell. Insight into morphogenetic mechanism(s) responsible for determining the shape of the murein sacculus itself has begun to emerge only in recent years. The discovery that MfreB and Mbl are cytoskeletal actin homologues that form helical structures extending from pole to pole in rod-shaped cells has opened an exciting new field of microbial cell biology. MreB (in Gram-negative rods) and Mbl (in Gram-positive species) are essential for murein synthesis along the lateral wall and hence, the rod shape of the cell. Known members of the morphogenetic system include MreB (or Mbl), MreC, MreD and PBP2, but Rod A and murein biosynthetic enzymes involved in peptidoglycan precursor synthesis and assembly are likely to be recruited to the same multimolecular apparatus. However, the actual role of MreB in assembly of the morphogenetic complex is still not clear and little is known about regulatory mechanisms controlling the switch from lateral murein elongation to septa1 murein synthesis at the time of cell division.

H + -form Nafion 212 films were chemically degraded via Fenton's reagent and characteristic polymer chain motions were analyzed using broadband dielectric spectroscopy. The β -relaxation peak maximum frequency ( f max ) shifts to higher temperatures upon degradation, reflecting slower chain motions. Perhaps this is due to an upward shift in average molecular weight caused by low molecular weight fragments being leached out of the membrane during the degradation experiment. Permittivity data for degraded and undegraded materials were fitted to the Havriliak–Negami equation. The quantities extracted from these fits were the relaxation time ( τ ) and parameters that reflect the breadth and asymmetry of the distribution of τ . These parameters were also used in the Vogel–Fulcher–Tammann–Hesse (VFTH) equation which was well-fitted to relaxation time versus temperature data. The increase in the Vogel temperature with degradation reflects more restricted chains. The distribution of relaxation time curve shifts to longer times, broadens, and is transformed from being bi- to tri-modal which reflects an increase in microstructural heterogeneity after degradation. It is concluded that dielectric spectroscopy can be a powerful tool in assessing the chemical degradation of Nafion membranes in the fuel cell environment.

The 2A-like sequences from members of the picornavirus family were utilized to construct a tricistronic vector bearing the human iduronidase (IDUA) gene along with the firefly luciferase and DsRed2 reporter genes. The 2A-like sequences mediate a cotranslational cleavage event resulting in the release of each individual protein product. Efficient cleavage was observed and all three proteins were functional in vitro and in vivo, allowing for supratherapeutic IDUA enzyme levels and the coexpression of luciferase and DsRed2 expression, which enabled us to track gene expression.

Previous studies have identified Sleeping Beauty transposons as efficient vectors for nonviral gene delivery in mammalian cells. However, studies demonstrating the usefulness of transposons as gene delivery vehicles into adult stem cells are lacking. Multipotent adult progenitor cells (MAPC) are nonhematopoietic stem cells with the capacity to form most, if not all, cell types of the body and as such hold great therapeutic potential. The whole-body biodistribution and persistence of MAPC are unknown, and such data would help direct clinical applications. We have nucleofected murine MAPC with two plasmid-based Sleeping Beauty transposons encoding the red fluorescent protein (DsRed2) and firefly luciferase. Transgenic euploid MAPC clones maintained their characteristic multilineage differentiation potential in vitro. DsRed2 and luciferase expression allowed for MAPC detection in vivo and in tissue sections. To confirm that transgenesis occurred by transposition into the genome of MAPC, we mapped Sleeping Beauty transposon integration sites in two MAPC clones using splinkerette PCR. This novel dual-reporter imaging approach based on the transgenesis of MAPC with Sleeping Beauty transposons sheds light on the homing patterns of MAPC and paves the way for quantification of MAPC engraftment in real time in vivo. ispartof: Molecular Therapy vol:12 issue:1 pages:42-48 ispartof: location:United States status: published

This paper is concerned with dendroclimatic research aimed at representing the history of very large-scale temperature changes. It describes recent analyses of the data from a widespread network of tree-ring chronologies, made up of ring width and densitometric measurement data spanning three to six centuries. The network was built over many years from trees selected to maximise their sensitivity to changing temperature. This strategy was adopted so that temperature reconstructions might be achieved at both regional and very large spatial scales. The focus here is on the use of one growth parameter: maximum latewood density (MXD). The detailed nature of the temperature sensitivity of MXD across the whole network has been explored and the dominant common influence of mean April–September temperature on MXD variability is demonstrated. Different approaches to reconstructing past temperature for this season include the production of detailed year-by-year gridded maps and wider regional integrations in the form of subcontinental and quasi-hemispheric-scale histories of temperature variability spanning some six centuries. These ‘hemispheric’ summer series can be compared with other reconstructions of temperature changes for the Northern Hemisphere over the last millennium. The tree-ring-based temperature reconstructions show the clear cooling effect of large explosive volcanic eruptions. They also exhibit greater century-timescale variability than is apparent in the other hemispheric series and suggest that the late 15th and the 16th centuries were cooler than indicated by some other data. However, in many tree-ring chronologies, we do not observe the expected rate of ring density increases that would be compatible with observed late 20th century warming. This changing climate sensitivity may be the result of other environmental factors that have, since the 1950s, increasingly acted to reduce tree-ring density below the level expected on the basis of summer temperature changes. This prevents us from claiming unprecedented hemispheric warming during recent decades on the basis of these tree-ring density data alone. Here we show very preliminary results of an investigation of the links between recent changes in MXD and ozone (the latter assumed to be associated with the incidence of UV radiation at the ground). D 2003 Elsevier B.V. All rights reserved.

The availability of the draft sequence of the human genome has created a pressing need to assign functions to each of the 35,000 or so genes that it defines. One useful approach for this purpose is to use model organisms for both bioinformatic and functional comparisons. We have developed a complementation system, based on the model eukaryote Saccharomyces cerevisiae, to clone human cDNAs that can functionally complement yeast essential genes. The system employs two regulatable promoters. One promoter, tetO (determining doxycycline-repressible expression), is used to control essential S. cerevisiae genes. The other, pMET3 (which is switched off in the presence of methionine), is employed to regulate the expression of mammalian cDNAs in yeast. We have demonstrated that this system is effective for both individual cDNA clones and for cDNA libraries, permitting the direct selection of functionally complementing clones. Three human cDNA libraries have been constructed and screened for clones that can complement specific essential yeast genes whose expression is switched off by the addition of doxycycline to the culture medium. The validity of each complementation was checked by showing that the yeast cells stop their growth in the presence of doxycycline and methionine, which represses the expression of the yeast and mammalian coding sequence, respectively. Using this system, we have screened 25 tetO replacement strains and succeeded in isolating human cDNAs complementing six essential yeast genes. In this way, we have uncovered a novel human ubiquitin-conjugating enzyme, have isolated a human cDNA clone that may function as a signal peptidase and have demonstrated that the functional segment of the human Psmd12 proteosome sub-unit contains a PINT domain.

To ensure the fidelity of DNA replication, cells activate a stress-response pathway when DNA replication is perturbed. This pathway regulates not only progress through the cell cycle but also transcription, apoptosis, DNA repair/recombination and DNA replication itself. Mounting evidence has suggested that this pathway is important for the maintenance of genomic integrity. Here, we discuss recent findings about how this pathway is activated by replication stress and how it regulates the DNA-replication machinery to alleviate the stress.

Adoptive T-cell transfer of T-cells with transgenic high avidity T-cell receptors (TCR) holds a lot of promise as therapeutic approach, however comprising certain challenges. Endogenous and transferred TCR-chains compete for surface expression and may pair inappropriately, potentially leading to autoimmunity. Designer nucleases like transcription activator-like effector nucleases (TALEN) and RNA-guided nucleases of the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated (Cas) system introduce specific DNA double strand breaks (DSB) at their target sites. This ability can be exploited for targeted genome editing. DSB repair through non-homologous end joining (NHEJ) can result in permanent gene knockout. Alternatively, if a homology-containing donor template is provided, transgenes can be inserted into the specific target locus by homology directed repair (HDR). In order to disrupt endogenous TCR expression, we assembled seven TALEN monomers and two guide RNAs targeting the constant regions of the TCR α-chain (TRAC) and the TCR β-chain (TRBC1 and TRBC2), respectively. Here we show the specific DSB induction by TALEN and CRISPR-Cas nucleases at their target sites, examined in K562 and T-cells using Cel1-assay and deep sequencing. Compared to nucleofection of TALEN-expressing plasmids, electroporation of mRNA considerably improved TCR knockout efficiency in primary T-cells. Using TRAC-TALEN and TRBC-TALEN mRNA, surface CD3 expression was successfully eliminated in about 75% and 60% of T-cells, respectively. The nuclease specificity was examined by identifying integrase-deficient lentiviral vector (IDLV)-marked DSB using LAM-PCR and deep sequencing. Although clustered IDLV integration sites were detected at the TALEN and CRISPR target sites, we did not detect any off-target activity, indicating high specificity. Additionally, K562 cells were nucleofected with TALEN-expressing plasmids and a donor template containing a GFP expression cassette flanked by 800bp homologous sequences to the TRAC locus. Thereby we were able to verify targeted gene addition through HDR of TALEN-mediated DSB in about 10% of treated cells. TALEN and CRISPR that show high efficiency and specificity to their target sequence will be used for generating T-cells with high avidity TCR.

Apres une discussion sur l'ecologie du betail, l'alimentation humaine, la composition du groupe domestique et l'allocation alimentaire des pasteurs Il Chamus de la region du lac Baringo au centre-ouest du Kenya, l'A. utilise les donnees d'Hodder pour evaluer une explication alternative des symboles et de la culture materielle de cette population. L'etude montre l'importance du lait et des produits laitiers comme ressource alimentaire. Le nombre de gourdes decorees et le degre de l'emploi de symbole sur les contenants laitiers dependent du nombre d'enfants et de co-epouses, et de la taille du troupeau dans un foyer. Les femmes produisent et se servent de moins de gourdes decorees avec l'âge

Objective To document a cumulative experience with removal of the Telectronics 330–801 Accufix atrial lead after it was withdrawn from the market. Design We reviewed our results with 96 at tempted lead extraction procedures through Dec. 1, 1995. Material and Methods All patients underwent preoperative assessment, including fluoroscopic screening of the lead. Patients had blood typing and screening done before the extraction procedure, and extractions were performed with cardiac surgical intervention available. Pacing dependence was assessed to determine the need for temporary pacing. Results The mean patient age was 65.7 ± 1.6 years, and the mean duration of lead implantation was 31.0 ± 1.4 months. Of the 96 lead extractions attempted, 94 were successful. Of the 96 cases, the retention wires were normal in 29, fractured without protrusion in 34, and fractured with protrusion in 33. Fifty-three leads were removed with use of simple traction only, and seven leads were removed by using a locking stylet and telescoping sheaths. The Cook workstation was used in the removal of 32 leads through the femoral vein; in 1 of these 32 patients, the retention wire was removed before lead extraction. The retention wire was removed but the lead was left intact in one patient. One lead was removed during an operative maze procedure. Associated complications were minimal. Conclusion Although lead extraction is a poten­tially fatal procedure, the Telectronics 330–801 Accu­fix atrial lead can be extracted safely with minimal morbidity when performed by experienced personnel.