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Metagenomic next-generation sequencing (mNGS) of cerebrospinal fluid (CSF) is an agnostic method for broad-based diagnosis of central nervous system (CNS) infections. Here we analyzed the 7-year performance of clinical CSF mNGS testing of 4,828 samples from June 2016 to April 2023 performed by the University of California, San Francisco (UCSF) clinical microbiology laboratory. Overall, mNGS testing detected 797 organisms from 697 (14.4%) of 4,828 samples, consisting of 363 (45.5%) DNA viruses, 211 (26.4%) RNA viruses, 132 (16.6%) bacteria, 68 (8.5%) fungi and 23 (2.9%) parasites. We also extracted clinical and laboratory metadata from a subset of the samples (n = 1,164) from 1,053 UCSF patients. Among the 220 infectious diagnoses in this subset, 48 (21.8%) were identified by mNGS alone. The sensitivity, specificity and accuracy of mNGS testing for CNS infections were 63.1%, 99.6% and 92.9%, respectively. mNGS testing exhibited higher sensitivity (63.1%) than indirect serologic testing (28.8%) and direct detection testing from both CSF (45.9%) and non-CSF (15.0%) samples (P < 0.001 for all three comparisons). When only considering diagnoses made by CSF direct detection testing, the sensitivity of mNGS testing increased to 86%. These results justify the routine use of diagnostic mNGS testing for hospitalized patients with suspected CNS infection., Competing Interests: Competing interests A.P.W., B.O., T.B., B.M. and S.M. are employed by and own equity in Delve Bio. C.Y.C. is a co-founder of Delve Bio and on the scientific advisory board for Delve Bio, Flightpath Biosciences, Biomeme, Mammoth Biosciences, BiomeSense and Poppy Health. He is also an inventor on US patent 11380421, ‘Pathogen detection using next-generation sequencing’, under which algorithms for taxonomic classification, filtering and pathogen detection are used by SURPI+ software. C.Y.C. receives research support from Delve Bio and Abbott Laboratories, Inc. M.R.W. is a co-founder and on the scientific advisory board for Delve Bio. The other authors declare no competing interests., (© 2024. The Author(s).)

The COVID-19 pandemic has created a global health crisis, with challenges arising from the ongoing evolution of the SARS-CoV-2 virus, the emergence of new strains, and the long-term effects of COVID-19. Aiming to overcome the development of viral resistance, our study here focused on developing broad-spectrum pan-coronavirus antiviral therapies by targeting host protein quality control mechanisms essential for viral replication. Screening an in-house compound library led to the discovery of three candidate compounds targeting cellular proteostasis. The three compounds are (1) the nucleotide analog cordycepin, (2) a benzothiozole analog, and (3) an acyldepsipeptide analog initially developed as part of a campaign to target the mitochondrial ClpP protease. These compounds demonstrated dose-dependent efficacy against multiple coronaviruses, including SARS-CoV-2, effectively inhibiting viral replication in vitro as well as in lung organoids. Notably, the compounds also showed efficacy against SARS-CoV-2 delta and omicron strains. As part of this work, we developed a BSL2-level cell-integrated SARS-CoV-2 replicon, which could serve as a valuable tool for high-throughput screening and studying intracellular viral replication. Our study should aid in the advancement of antiviral drug development efforts., (© 2024. The Author(s).)

Thin cell culture membranes in organ-on-a-chip (OOC) devices are used to model a wide range of thin tissues. While early and most current platforms use microporous or fibrous elastomeric or thermoplastic membranes, there is an emerging class of devices using extra-cellular matrix (ECM) protein-based membranes to improve their biological relevance. These ECM-based membranes present physiologically relevant properties, but they are difficult to integrate into OOC devices due to their relative fragility. Additionally, the specialized fabrication methods developed to date make comparison between methods difficult. This work presents the development and characterization of a method to produce ultrathin matrix-derived membranes (UMM) in OOC devices that requires only a preassembled thermoplastic device and a micropipette, decoupling the device and UMM fabrication processes. Control over the thickness and permeability of the UMM is demonstrated, along with integration of the UMM in a device enabling high-resolution on-chip microscopy. The reliability of the UMM fabrication method is leveraged to develop a medium-throughput well-plate format device with 32 independent UMM-integrated samples. Finally, proof-of-concept cell culture experiments are demonstrated. Due to its simplicity and controllability, the presented method has the potential to overcome technical barriers preventing wider adoption of physiologically relevant ECM-based membranes in OOC devices., (© 2024 The Authors. Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

Studying human fetal lungs can inform how developmental defects and disease states alter the function of the lungs. Here, we sequenced >150,000 single cells from 19 healthy human pseudoglandular fetal lung tissues ranging between gestational weeks 10-19. We capture dynamic developmental trajectories from progenitor cells that express abundant levels of the cystic fibrosis conductance transmembrane regulator (CFTR). These cells give rise to multiple specialized epithelial cell types. Combined with spatial transcriptomics, we show temporal regulation of key signalling pathways that may drive the temporal and spatial emergence of specialized epithelial cells including ciliated and pulmonary neuroendocrine cells. Finally, we show that human pluripotent stem cell-derived fetal lung models contain CFTR-expressing progenitor cells that capture similar lineage developmental trajectories as identified in the native tissue. Overall, this study provides a comprehensive single-cell atlas of the developing human lung, outlining the temporal and spatial complexities of cell lineage development and benchmarks fetal lung cultures from human pluripotent stem cell differentiations to similar developmental window., (© 2024. The Author(s).)

The lower respiratory tract is a hierarchical network of compliant tubular structures that are made from extracellular matrix proteins with a wall lined by an epithelium. While microfluidic airway-on-a-chip models incorporate the effects of shear and stretch on the epithelium, week-long air-liquid-interface culture at physiological shear stresses, the circular cross-section, and compliance of native airway walls have yet to be recapitulated. To overcome these limitations, a collagen tube-based airway model is presented. The lumen is lined with a confluent epithelium during two-week continuous perfusion with warm, humid air while presenting culture medium from the outside and compensating for evaporation. The model recapitulates human small airways in extracellular matrix composition and mechanical microenvironment, allowing for the first time dynamic studies of elastocapillary phenomena associated with regular breathing and mechanical ventilation, as well as their impacts on the epithelium. A case study reveales increasing damage to the epithelium during repetitive collapse and reopening cycles as opposed to overdistension, suggesting expiratory flow resistance to reduce atelectasis. The model is expected to promote systematic comparisons between different clinically used ventilation strategies and, more broadly, to enhance human organ-on-a-chip platforms for a variety of tubular tissues., (© 2024 The Authors. Small published by Wiley‐VCH GmbH.)

Nowadays there are multiple types of contraceptive methods, from reversible to permanent, for those choosing to delay pregnancy. Misconceptions about contraception and infertility are a key factor for discontinuation or the uptake of family planning methods. Regaining fertility (the ability to conceive) after contraceptive discontinuation is therefore pivotal. Technical studies to date have evaluated return to fertility by assessing pregnancy as an outcome, with variable results, or return to ovulation as a surrogate measure by assessing hormone levels (such as progesterone, LH, FSH) with or without transvaginal ultrasound. In general, relying on time to pregnancy as an indicator of return to fertility following contraceptive method discontinuation can be problematic due to variable factors independent of contraceptive effects on fertility, hormone clearance, and fertility recovery. Since the ability to conceive after contraceptive method discontinuation is a critical factor influencing product uptake, it is important to have robust biomarkers that easily and accurately predict the timing of fertility return following contraception and isolate that recovery from extrinsic and circumstantial factors. The main aim of this review is to summarize the current approaches, existing knowledge, and gaps in methods of evaluating return-to-fertility as well as to provide insights into the potential of new biomarkers to more accurately predict fertility restoration after contraceptive discontinuation. Biomarker candidates proposed in this document include those associated with folliculogenesis, cumulus cell expansion, follicular rupture and ovulation, and endometrial transport and receptivity which have been selected and scored on predefined criteria meant to evaluate their probable viability for advancement. The review also describes limitations, regulatory requirements, and a potential path to clinically testing these selected biomarkers. It is important to understand fertility restoration after contraceptive method discontinuation to provide users and health providers with accurate evidence-based information. Predictive biomarkers, if easy and low-cost, have the potential to enable robust evaluation of RTF, and provide potential users the information they desire when selecting a contraceptive method. This could lead to expanded uptake and continuation of modern contraception and inform the development of new contraceptive methods to widen user's family planning choices., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Cordova-Gomez, Wong, Sims, Doncel and Dorflinger.)

Introducing or correcting disease-causing mutations through genome editing in human pluripotent stem cells (hPSCs) followed by tissue-specific differentiation provide sustainable models of multiorgan diseases, such as cystic fibrosis (CF). However, low editing efficiency resulting in extended cell culture periods and the use of specialised equipment for fluorescence activated cell sorting (FACS) make hPSC genome editing still challenging. We aimed to investigate whether a combination of cell cycle synchronisation, single-stranded oligodeoxyribonucleotides, transient selection, manual clonal isolation, and rapid screening can improve the generation of correctly modified hPSCs. Here, we introduced the most common CF mutation, ΔF508, into the CFTR gene, using TALENs into hPSCs, and corrected the W1282X mutation using CRISPR-Cas9, in human-induced PSCs. This relatively simple method achieved up to 10% efficiency without the need for FACS, generating heterozygous and homozygous gene edited hPSCs within 3-6 weeks in order to understand genetic determinants of disease and precision medicine.

Introduction: Poor or inconsistent adherence to daily oral pre-exposure prophylaxis (PrEP) has emerged as a key barrier to effective HIV prevention. The advent of potent long-acting (LA) antiretrovirals (ARVs) in conjunction with advances in controlled release technologies has enabled LA ARV drug delivery systems (DDS) capable of providing extended dosing intervals and overcome the challenge of suboptimal drug adherence with daily oral dosing., Areas Covered: This review discusses the current state of the LA PrEP field, recent advances, and emerging technologies, including ARV prodrug modifications and new DDS. Technological challenges, knowledge gaps, preclinical testing considerations, and future directions important in the context of clinical translation and implementation of LA HIV PrEP are discussed., Expert Opinion: The HIV prevention field is evolving faster than ever and the bar for developing next-generation LA HIV prevention options continues to rise. The requirements for viable LA PrEP products to be implemented in resource-limited settings are challenging, necessitating proactive consideration and product modifications during the design and testing of promising new candidates. If successfully translated, next-generation LA PrEP that are safe, affordable, highly effective, and accepted by both end-users and key stakeholders will offer significant potential to curb the HIV pandemic.

Human embryonic stem cells (ES) and induced pluripotent stem cells (iPSC) are powerful tools that have the potential to generate in vitro human lung epithelial cells. However, challenges in efficiency and reproducibility remain in utilizing the cells for therapy discovery platforms. Here, we optimize our previously published protocols to efficiently generate three developmental stages of the lung model (fetal lung epithelial progenitors, fLEP; immature airway epithelial spheroid, AES; air-liquid interface culture, ALI), and demonstrate its potential for cystic fibrosis (CF) drug discovery platforms. The stepwise approach directs differentiation from hPSC to definitive endoderm, anterior ventral foregut endoderm, and fetal lung progenitor cells. The article also describes the generation of immature airway epithelial spheroids in Matrigel with epithelial cells sorted by a magnetic-activated cell sorting system, and the generation of adult-like airway epithelia through air-liquid interface conditions. We demonstrate that this optimized procedure generates remarkably higher cystic fibrosis transmembrane conductance regulator (CFTR) expression and function than our previous method, and thus is uniquely suitable for CF research applications. © 2022 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: hESC/hiPSC differentiation to fetal lung progenitors Basic Protocol 2: Formation of airway epithelial spheroids Alternate Protocol 1: Cryopreservation of airway epithelial spheroids Basic Protocol 3: Differentiation and maturation in air-liquid interface culture Alternate Protocol 2: Differentiation and maturation of epithelial progenitors from airway epithelial spheroids in ALI culture., (© 2022 The Authors. Current Protocols published by Wiley Periodicals LLC.)

For those people with cystic fibrosis carrying rare CFTR mutations not responding to currently available therapies, there is an unmet need for relevant tissue models for therapy development. Here, we describe a new testing platform that employs patient-specific induced pluripotent stem cells (iPSCs) differentiated to lung progenitor cells that can be studied using a dynamic, high-throughput fluorescence-based assay of CFTR channel activity. Our proof-of-concept studies support the potential use of this platform, together with a Canadian bioresource that contains iPSC lines and matched nasal cultures from people with rare mutations, to advance patient-oriented therapy development. Interventions identified in the high-throughput, stem cell-based model and validated in primary nasal cultures from the same person have the potential to be advanced as therapies., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

The discovery of the Cystic fibrosis (CF) gene in 1989 has paved the way for incredible progress in treating the disease such that the mean survival age of individuals living with CF is now ~58 years in Canada. Recent developments in gene targeting tools and new cell and animal models have re-ignited the search for a permanent genetic cure for all CF. In this review, we highlight some of the more recent gene therapy approaches as well as new models that will provide insight into personalized therapies for CF., (© 2021. The Author(s).)

The cystic fibrosis (CF) transmembrane conductance regulator (CFTR) protein is a cAMP-activated anion channel that is critical for regulating fluid and ion transport across the epithelium. This process is disrupted in CF epithelia, and patients harbouring CF-causing mutations experience reduced lung function as a result, associated with the increased rate of mortality. Much progress has been made in CF research leading to treatments that improve CFTR function, including small molecule modulators. However, clinical outcomes are not necessarily mutation-specific as individuals harboring the same genetic mutation may present with varying disease manifestations and responses to therapy. This suggests that the CFTR protein may have alternative functions that remain under-appreciated and yet can impact disease. In this mini review, we highlight some notable research implicating an important role of CFTR protein during early lung development and how mutant CFTR proteins may impact CF airway disease pathogenesis. We also discuss recent novel cell and animal models that can now be used to identify a developmental cause of CF lung disease., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Huang, Quach, Lee, Dierolf, Moraes and Wong.)

Fetal lung underdevelopment, also known as pulmonary hypoplasia, is characterized by decreased lung growth and maturation. The most common birth defect found in babies with pulmonary hypoplasia is congenital diaphragmatic hernia (CDH). Despite research and clinical advances, babies with CDH still have high morbidity and mortality rates, which are directly related to the severity of lung underdevelopment. To date, there is no effective treatment that promotes fetal lung growth and maturation. Here, we describe a stem cell-based approach in rodents that enhances fetal lung development via the administration of extracellular vesicles (EVs) derived from amniotic fluid stem cells (AFSCs). Using fetal rodent models of pulmonary hypoplasia (primary epithelial cells, organoids, explants, and in vivo), we demonstrated that AFSC-EV administration promoted branching morphogenesis and alveolarization, rescued tissue homeostasis, and stimulated epithelial cell and fibroblast differentiation. We confirmed this regenerative ability in in vitro models of lung injury using human material, where human AFSC-EVs obtained following good manufacturing practices restored pulmonary epithelial homeostasis. Investigating EV mechanism of action, we found that AFSC-EV beneficial effects were exerted via the release of RNA cargo. MicroRNAs regulating the expression of genes involved in lung development, such as the miR17-92 cluster and its paralogs, were highly enriched in AFSC-EVs and were increased in AFSC-EV-treated primary lung epithelial cells compared to untreated cells. Our findings suggest that AFSC-EVs hold regenerative ability for underdeveloped fetal lungs, demonstrating potential for therapeutic application in patients with pulmonary hypoplasia., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Cell lineage conversion of fibroblasts to specialized cell types through transdifferentiation may provide a fast and alternative cell source for regenerative medicine. Here we show that transient transduction of fibroblasts with the four reprogramming factors (Oct4, Sox2, Klf4, and c-Myc) in addition to the early lung transcription factor Nkx2-1 (also known as Ttf1), followed by directed differentiation of the cells, can convert mouse embryonic and human adult dermal fibroblasts into induced lung-like epithelial cells (iLEC). These iLEC differentiate into multiple lung cell types in air liquid interface cultures, repopulate decellularized rat lung scaffolds, and form lung epithelia composed of Ciliated, Goblet, Basal, and Club cells after transplantation into immune-compromised mice. As proof-of-concept, differentiated human iLEC harboring the Cystic Fibrosis mutation dF508 demonstrated pharmacological rescue of CFTR function using the combination of lumacaftor and ivacaftor. Overall, this is a promising alternative approach for generation of patient-specific lung-like progenitors to study lung function, disease and future regeneration strategies.

The design of photocatalysts able to reduce CO 2 to value-added chemicals and fuels could enable a closed carbon circular economy. A common theme running through the design of photocatalysts for CO 2 reduction is the utilization of semiconductor materials with high-energy conduction bands able to generate highly reducing electrons. Far less explored in this respect are low-energy conduction band materials such as WO 3 . Specifically, we focus attention on the use of Pd nanocrystal decorated WO 3 nanowires as a heretofore-unexplored photocatalyst for the hydrogenation of CO 2 . Powder X-ray diffraction, thermogravimetric analysis, ultraviolet-visible-near infrared, and in situ X-ray photoelectron spectroscopy analytical techniques elucidate the hydrogen tungsten bronze, H y WO 3- x , as the catalytically active species formed via the H 2 spillover effect by Pd. The existence in H y WO 3- x of Brønsted acid hydroxyls OH, W(V) sites, and oxygen vacancies (V O ) facilitate CO 2 capture and reduction reactions. Under solar irradiation, CO 2 reduction attains CO production rates as high as 3.0 mmol g cat -1 hr -1 with a selectivity exceeding 99%. A combination of reaction kinetic studies and in situ diffuse reflectance infrared Fourier transform spectroscopy measurements provide a valuable insight into thermochemical compared to photochemical surface reaction pathways, considered responsible for the hydrogenation of CO 2 by Pd@H y WO 3- x .

Background: Therapies targeting certain CFTR mutants have been approved, yet variations in clinical response highlight the need for in-vitro and genetic tools that predict patient-specific clinical outcomes. Toward this goal, the CF Canada-Sick Kids Program in Individual CF Therapy (CFIT) is generating a "first of its kind", comprehensive resource containing patient-specific cell cultures and data from 100 CF individuals that will enable modeling of therapeutic responses., Methods: The CFIT program is generating: 1) nasal cells from drug naïve patients suitable for culture and the study of drug responses in vitro, 2) matched gene expression data obtained by sequencing the RNA from the primary nasal tissue, 3) whole genome sequencing of blood derived DNA from each of the 100 participants, 4) induced pluripotent stem cells (iPSCs) generated from each participant's blood sample, 5) CRISPR-edited isogenic control iPSC lines and 6) prospective clinical data from patients treated with CF modulators., Results: To date, we have recruited 57 of 100 individuals to CFIT, most of whom are homozygous for F508del (to assess in-vitro: in-vivo correlations with respect to ORKAMBI response) or heterozygous for F508del and a minimal function mutation. In addition, several donors are homozygous for rare nonsense and missense mutations. Nasal epithelial cell cultures and matched iPSC lines are available for many of these donors., Conclusions: This accessible resource will enable development of tools that predict individual outcomes to current and emerging modulators targeting F508del-CFTR and facilitate therapy discovery for rare CF causing mutations., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Age-related decreases in cortical thickness observed during adolescence may be related to fluctuations in sex and stress hormones. We examine this possibility by relating inter-regional variations in age-related cortical thinning (data from the Saguenay Youth Study) to inter-regional variations in expression levels of relevant genes (data from the Allen Human Brain Atlas); we focus on genes coding for glucocorticoid receptor (NR3C1), androgen receptor (AR), progesterone receptor (PGR), and estrogen receptors (ESR1 and ESR2). Across 34 cortical regions (Desikan-Killiany parcellation), age-related cortical thinning varied as a function of mRNA expression levels of NR3C1 in males (R2 = 0.46) and females (R2 = 0.30) and AR in males only (R2 = 0.25). Cortical thinning did not vary as a function of expression levels of PGR, ESR1, or ESR2 in either sex; this might be due to the observed low consistency of expression profiles of these 3 genes across donors. Inter-regional levels of the NR3C1 and AR expression interacted with each other vis-à-vis cortical thinning: age-related cortical thinning varied as a function of NR3C1 mRNA expression in brain regions with low (males: R2 = 0.64; females: R2 = 0.58) but not high (males: R2 = 0.0045; females: R2 = 0.15) levels of AR mRNA expression. These results suggest that glucocorticoid and androgen receptors contribute to cortical maturation during adolescence.

Hypertension is one of the major causes of morbidity and mortality. Worldwide, Hypertension is estimated to cause 7.5 million deaths, about 12.8% of the total of all deaths. This accounts for 57 million disability adjusted life years (DALYS) or 3.7% of total DALYS. This led WHO to set a target of 25% reduction in prevalence by 2025. To reach that, WHO has adopted non-conventional methods for the management of hypertension? Despite worldwide popularity of such non-conventional therapies, only small volume of evidence exists that supports its effectiveness. This review attempted to make a critical appraisal of the evidence, with the aim to (1) describe the therapeutic modalities frequently used, and (2) review the current level of evidence attributable to each modality. Databases from Cochrane Library, MEDLINE, PUBMED, and EMBASE were searched from 2005-2015. A total of 23 publications have been identified and selected. Out of these, 15 systematic reviews and/or meta-analysis of RCTs, 5 RCTs, 1 non-RCT, and 2 observational studies without control. Among those 23 publications, therapeutic modalities identified are: fish oil, qigong, yoga, coenzyme Q10, melatonin, meditation, vitamin D, vitamin C, monounsaturated fatty acids, dietary amino-acids, chiropractic, osteopathy, folate, inorganic nitrate, beetroot juice, beetroot bread, magnesium, and L-arginine. The followings were found to have weak or no evidence: fish oil, yoga, vitamin D, monounsaturated fatty acid, dietary amino-acids, and osteopathy. Those found to have significant reduction in blood pressure are: magnesium, qigong, melatonin, meditation, vitamin C, chiropractic, folate, inorganic nitrate, beetroot juice and L-arginine. Coenzyme Q10on the other hand, showed contradicting results were some studies found weak or no effect on blood pressure while others showed significant blood pressure reduction effect. By virtue of the research designs and methodologies, the evidence contributed from these studies is at level 1. Results from this review suggest that certain non-conventional therapies may be effective in treating hypertension and improving cardiac function and therefore considered as part of an evidence-based approach.

Income inequality is associated with poor health and social outcomes. Negative social comparisons and competition may involve the hypothalamic-pituitary-adrenal (HPA) and hypothalamic-pituitary-gonadal (HPG) axes in underlying some of these complex inter-relationships. Here we investigate brain maturation, indexed by age-related decreases in cortical thickness, in adolescents living in neighborhoods with differing levels of income inequality and household income. We examine whether inter-regional variations relate to those in glucocorticoid receptor (HPA) and androgen receptor (HPG) gene expression. For each sex, we used a median split of income inequality and household income (income-to-needs ratio) to create four subgroups. In female adolescents, the high-inequality low-income group displayed the greatest age-related decreases in cortical thickness. In this group, expression of glucocorticoid and androgen receptor genes explained the most variance in these age-related decreases in thickness across the cortex. We speculate that female adolescents living in high-inequality neighborhoods and low-income households may experience greater HPA and HPG activity, leading to steeper decreases in cortical thickness with age.

Pulmonary disease is the major cause of morbidity and mortality in patients with cystic fibrosis, a disease caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene. Heterogeneity in CFTR genotype-phenotype relationships in affected individuals plus the escalation of drug discovery targeting specific mutations highlights the need to develop robust in vitro platforms with which to stratify therapeutic options using relevant tissue. Toward this goal, we adapted a fluorescence plate reader assay of apical CFTR-mediated chloride conductance to enable profiling of a panel of modulators on primary nasal epithelial cultures derived from patients bearing different CFTR mutations. This platform faithfully recapitulated patient-specific responses previously observed in the "gold-standard" but relatively low-throughput Ussing chamber. Moreover, using this approach, we identified a novel strategy with which to augment the response to an approved drug in specific patients. In proof of concept studies, we also validated the use of this platform in measuring drug responses in lung cultures differentiated from cystic fibrosis iPS cells. Taken together, we show that this medium throughput assay of CFTR activity has the potential to stratify cystic fibrosis patient-specific responses to approved drugs and investigational compounds in vitro in primary and iPS cell-derived airway cultures., Competing Interests: COMPETING INTERESTS The authors declare that they have no competing interests.

Cystic fibrosis (CF) is a fatal recessive genetic disorder caused by a mutation in the gene encoding CF transmembrane conductance regulator (CFTR) protein. Alteration in CFTR leads to thick airway mucus and bacterial infection. Cell therapy has been proposed for CFTR restoration, but efficacy has been limited by low engraftment levels. In our previous studies, we have shown that using a pre-conditioning regimen in combination with optimization of cell number and time of delivery, we could obtain greater bone marrow cell (BMC) retention in the lung. Here, we found that optimized delivery of wild-type (WT) BMC contributed to apical CFTR expression in airway epithelium and restoration of select ceramide species and fatty acids in CFTR -/- mice. Importantly, WT BMC delivery delayed Pseudomonas aeruginosa lung infection and increased survival of CFTR -/- recipients. Only WT BMCs had a beneficial effect beyond 6 months, suggesting a dual mechanism of BMC benefit: a non-specific effect early after cell delivery, possibly due to the recruitment of macrophages and neutrophils, and a late beneficial effect dependent on long-term CFTR expression. Taken together, our results suggest that BMC can improve overall lung function and may have potential therapeutic benefit for the treatment of CF., (Copyright © 2017. Published by Elsevier Inc.)

This Mini-Review describes sex differences in 66 quantitative characteristics of the brain and body measured in a community-based sample of 1,024 adolescents 12-18 years of age, members of the Saguenay Youth Study. Using an extensive phenotyping protocol, we have obtained measures in a number of domains, including brain structure, cognition, mental health, substance use, body composition, metabolism, cardiovascular reactivity, and life style. For each measure, we provide estimates of effect size (Cohen's d) and sex-specific correlations with age (Pearson R). In total 59 of the 66 characteristics showed sex differences (at a nominal P < 0.05), with small (32), medium-sized (13), and large (11) effects. Some, but not all, of these sex differences increase during adolescence; this appears to be the case mostly for anatomical and physiological measures. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Heart failure is a multifactorial disease with poor prognosis. There are many hypotheses regarding the cause of heart failure. Leading among them are the hemodynamic and the neuro-hormonal hypotheses. Although the energy depletion hypothesis has been fairly recent, there is evidence suggesting that declining bioenergy plays a major role in heart failure. This review explored the myocardial energy depletion hypothesis from the role of micronutrients in correcting and alleviating symptoms of heart failure. Even though focus was on key nutrients such as coenzyme Q10, thiamine, riboflavin, L-carnitine, and taurine, emphasis was on the combined effect of multiple micronutrients as a whole. Search from databases from 2000 to 2015 produced four clinical studies using multiple micronutrients on heart failure. Evidence from the studies show that using high doses of multiple micronutrients may have positive effects on heart failure and simultaneously support the myocardial energy depletion hypothesis.

Silicon constitutes 28% of the earth's mass. Its high abundance, lack of toxicity and low cost coupled with its electrical and optical properties, make silicon unique among the semiconductors for converting sunlight into electricity. In the quest for semiconductors that can make chemicals and fuels from sunlight and carbon dioxide, unfortunately the best performers are invariably made from rare and expensive elements. Here we report the observation that hydride-terminated silicon nanocrystals with average diameter 3.5 nm, denoted ncSi:H, can function as a single component heterogeneous reducing agent for converting gaseous carbon dioxide selectively to carbon monoxide, at a rate of hundreds of μmol h(-1) g(-1). The large surface area, broadband visible to near infrared light harvesting and reducing power of SiH surface sites of ncSi:H, together play key roles in this conversion. Making use of the reducing power of nanostructured hydrides towards gaseous carbon dioxide is a conceptually distinct and commercially interesting strategy for making fuels directly from sunlight.

Minimally invasive transforaminal lumbar interbody fusion (MI-TLIF) has been scrutinized for having a complex learning curve. Careful assessment of MI-TLIF complications and critical analyses of prevention may aid a safe adoption of this technique. The current report focuses on the incidence of interbody cage extrusions following MI-TLIF in a series of 513 patients. The authors discuss their experience with graft extrusions and provide methods to minimize this complication. This study retrospectively reviewed 513 prospectively followed patients who underwent MI-TLIF over a 10 year period. The inclusion criteria consisted of all patients who underwent one to three level MI-TLIF, from whom the incidence of cage extrusion was analyzed. Cage extrusion was defined as an interbody graft migrating outside the cephalad and caudal vertebral body posterior margin. Cage extrusions were diagnosed by comparing the intraoperative radiographs to the postoperative radiographs. Patients with >10° coronal curves, significant sagittal malalignment, infection, and preoperative instrumentation failure were excluded. Of 513 patients undergoing MI-TLIF, five patients (0.97%) were diagnosed with cage migrations. The mean follow-up duration was 13.6 ± standard deviation of 8.8 months. Complications included asymptomatic cage migration alone (two patients) neurological decline (two patients) and epidural hematoma (one patient). On average, cage migrations cost a university hospital an additional $US17,217 for revision treatment. While the incidence of cage migrations is low (0.97%), it can lead to postoperative complications that require revision surgery and increased hospital costs. The risk for this significant complication can be minimized with proper technique and patient selection., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Heart failure is a progressive cardiovascular disorder and, in most cases, begins with atherosclerosis and ischemic heart disease. The prognosis of patients with heart failure is poor, even with improvement on the management of all forms of ischemic heart disease. There have been studies on heart failure using a single nutrient or a combination of multiple nutrients. Results are mixed. The aim of this study was to assess the influence of multiple micronutrient supplementation using the quality of life measure on patients with heart failure secondary to ischemic heart disease. This prospective case series followed 12 patients for a period between 3 to 8 months, using the Minnesota Living with Heart Failure Questionnaire (MLHFQ) as the sole outcome measure. The primary outcome was a score change over time between the start and endpoint of treatment. Change in MLHFQ mean total score was 27.08 ± 20.43 and mean symptoms score was 4.67 ± 3.34. Paired t-test showed a difference between baseline and endpoint of treatment (P < 0.001), which was statistically significant. A high dose of multiple micronutrients may have beneficial effects on cardiac function in patients with symptomatic heart failure. This study indicates the need for long-term controlled studies to test the efficacy and safety of this economic approach in managing heart failure.

Psoriasis is a chronic inflammatory skin disease characterized by thickened, silvery-scaled patches. There is currently no cure and treatments only attempt to reduce the severity of symptoms. This study reports the case of a 36-year-old female who presented to the clinic with severe psoriasis and had been treated with topical steroid cream for the past 14 years. After adherence to prescribed dietary changes for 6 months, including abundant intake of vegetables, minimal consumption of meat, and avoidance of junk food and sugar in food or drinks, as well as nutritional supplementation with Vitacor Plus, ProLysinC, VitaCforte and LysinC Drink mix, the patient experienced complete resolution of psoriatic patches on her body.

OBJECT Patients with symptomatic intradural-extramedullary (ID-EM) tumors may be successfully treated with resection of the lesion and decompression of associated neural structures. Studies of patients undergoing open resection of these tumors have reported high rates of gross-total resection (GTR) with minimal long-term neurological deficit. Case reports and small case series have suggested that these patients may be successfully treated with minimally invasive surgery (MIS). These studies have been limited by small patient populations. Moreover, there are no studies directly comparing perioperative outcomes between patients treated with open resection and MIS. The objective of this study was to compare perioperative outcomes in patients with ID-EM tumors treated using open resection or MIS. METHODS A retrospective review was performed using data collected from 45 consecutive patients treated by open resection or MIS for ID-EM spine tumors. These patients were treated over a 9-year period between April 2003 and October 2012 at Northwestern University and the University of Chicago. Statistical analysis was performed to compare perioperative outcomes between the two groups. RESULTS Of the 45 patients in the study, 27 were treated with the MIS approach and 18 were treated with the open approach. Operative time was similar between the two groups: 256.3 minutes in the MIS group versus 241.1 minutes in the open group (p = 0.55). Estimated blood loss was significantly lower in the MIS group (133.7 ml) compared with the open group (558.8 ml) (p < 0.01). A GTR was achieved in 94.4% of the open cases and 92.6% of the MIS cases (p = 0.81). The mean hospital stay was significantly shorter in the MIS group (3.9 days) compared with the open group (6.1 days) (p < 0.01). There was no significant difference between the complication rates (p = 0.32) and reoperation rates (p = 0.33) between the two groups. Multivariate analysis demonstrated an increased rate of complications in cervical spine tumors (OR 15, p = 0.05). CONCLUSIONS Thoracolumbar ID-EM tumors may be safely and effectively treated with either the open approach or an MIS approach, with an equivalent rate of GTR, perioperative complication rate, and operative time. Patients treated with an MIS approach may benefit from a decrease in operative blood loss and shorter hospital stays.

Objectives: We aimed to examine the impact of demographic and treatment factors on overall survival among spinal ependymoma patients., Methods: Using data from Surveillance, Epidemiology, and End Results (SEER) from 1990-2008, we evaluated subjects with histologically confirmed spinal cord ependymoma., Results: We identified 1,353 patients with spinal ependymoma (mean age 43.5 years). Among these, 26 had anaplastic ependymoma (AE), 374 had myxopapillary ependymoma (MPE), and 953 had other low-grade ependymoma (non-MPE). Median follow-up was 61 months. Overall survival at 5 years was 97% for MPE, 92% for low-grade non-MPE, and 73% for AE. Gross total resection (GTR) was achieved in 58% of subjects with MPE, 51% with low-grade non-MPE, and 50% with AE. Radiation occurred more often after subtotal resection (STR) than after GTR, and more often among those with anaplastic histology. Histology and extent of surgical resection were significant prognostic factors in multivariate analyses. Compared to MPE, subjects with low-grade non-MPE had a higher risk of mortality at 5 years (HR 2.35, P = 0.03), as did subjects with AE (HR 8.63, P < 0.01). Compared to GTR, STR was associated with an increased risk of mortality at 5 years (HR 2.2, P = 0.01), as was biopsy only (HR 2.05, P = 0.03) and no surgery (HR 4.97, P < 0.01). Among patients with either STR or GTR, adjuvant radiotherapy did not confer a survival benefit at 5 years (STR: HR 2.29, P = 0.07, and GTR: HR 2.2, P = 0.12)., Discussion: We found that lower grade histology and higher extent of surgical resection were significant prognostic factors for more favourable survival outcomes for spinal ependymoma patients.

Sore throat is a common presenting complaint in the outpatient setting. Most cases are nonbacterial in origin, but those that are bacterial are usually the result of group A β-hemolytic streptococcus. Guidelines exist to help physicians decide whether to treat with an antibiotic. Lemierre syndrome is a dangerous potential sequela of pharyngitis that results in septic thrombophlebitis of the internal jugular (IJ) vein. A high index of suspicion is needed to consider this diagnosis in the workup of pharyngitis and should be aggressively treated once identified. Consideration should be given to completing blood cultures and neck imaging because of clinical suspicion. The case study discussed here illustrates the presentation, evaluation, and treatment of Lemierre syndrome., (© Copyright 2015 by the American Board of Family Medicine.)

OBJECT Transforaminal lumbar interbody fusion (TLIF) has become one of the preferred procedures for circumferential fusion in the lumbar spine. Over the last decade, advances in surgical techniques have enabled surgeons to perform the TLIF procedure through a minimally invasive approach (MI-TLIF). There are a few studies reported in the medical literature in which perioperative complication rates of MI-TLIF were evaluated; here, the authors present the largest cohort series to date. They analyzed intraoperative and perioperative complications in 513 consecutive MI-TLIF-treated patients with lumbar degenerative disc disease. METHODS The authors performed a retrospective review of prospectively collected data on 513 consecutive patients treated over a 10-year period for lumbar degenerative disc disease using MI-TLIF. All patients undergoing either a first-time or revision 1- or 2-level MI-TLIF procedure were included in the study. Demographic, intraoperative, and perioperative data were collected and analyzed using bivariate analyses (Student t-test, analysis of variance, odds ratio, chi-square test) and multivariate analyses (logistic regression). RESULTS A total of 513 patients underwent an MI-TLIF procedure, and the perioperative complication rate was 15.6%. The incidence of durotomy was 5.1%, and the medical and surgical infection rates were 1.4% and 0.2%, respectively. A statistically significant increase in the infection rate was seen in revision MI-TLIF cases, and the same was found for the perioperative complication rate in multilevel MI-TLIF cases. Instrumentation failure occurred in 2.3% of the cases. After analysis, no statistically significant difference was seen in the rates of durotomy during revision and multilevel surgeries. There was no significant difference between the complication rates when stratified according to presenting diagnosis. CONCLUSIONS To the authors' knowledge, this is the largest study of perioperative complications in MI-TLIF in the literature. A total of 513 patients underwent MI-TLIF (perioperative complication rate 15.6%). The most common complication was a durotomy (5.1%), and there was only 1 surgical wound infection (0.2%). There were significantly more perioperative infections in revision MI-TLIF cases and more perioperative complications in multilevel MI-TLIF cases. The results of this study suggest that MI-TLIF has a similar or better perioperative complication profile than those documented in the literature for open-TLIF treatment of degenerative lumbar spine disease.

The contribution of bone marrow cells (BMC) in lung repair is controversial. We previously reported a subpopulation of BMC that express Clara cell secretory protein (CCSP). To determine the contribution of endogenous CCSP(+) BMC to airway regeneration, we performed bone marrow transplantation studies using the CCtk mouse, which expresses a thymidine kinase suicide gene under regulation of the CCSP promoter. Mice were transplanted with wild-type or CCtk BMC and treated with ganciclovir to eliminate CCSP(+) cells. After airway injury using naphthalene, mice depleted of CCSP(+) BMC had more inflammatory cells in lung and decreased levels of oxygen in arterial blood. They also had reduced expression of airway epithelial genes and less Clara cells compared to control mice that had intact CCSP(+) BMC and bone marrow derived CCSP(+) cells in the airways. After naphthalene injury, administration of CCSP reproduced the beneficial effect of CCSP(+) BMC by improving recovery of airway epithelium, reducing lung inflammation and increasing oxygen in arterial blood from mice depleted of CCSP(+) BMC. Our data demonstrate that ablation of CCSP(+) BMC delays airway recovery and suggests the beneficial effect of CCSP(+) BMC in lung recovery is in part due to production of CCSP itself.

Airway epithelial cells are of great interest for research on lung development, regeneration and disease modeling. This protocol describes how to generate cystic fibrosis (CF) transmembrane conductance regulator protein (CFTR)-expressing airway epithelial cells from human pluripotent stem cells (PSCs). The stepwise approach from PSC culture to differentiation into progenitors and then mature epithelia with apical CFTR activity is outlined. Human PSCs that were inefficient at endoderm differentiation using our previous lung differentiation protocol were able to generate substantial lung progenitor cell populations. Augmented CFTR activity can be observed in all cultures as early as at 35 d of differentiation, and full maturation of the cells in air-liquid interface cultures occurs in <5 weeks. This protocol can be used for drug discovery, tissue regeneration or disease modeling.

Invasive central nervous system aspergillosis is a rare form of fungal infection that presents most commonly in immunocompromised individuals. There have been multiple previous reports of aspergillus vertebral osteomyelitis and spinal epidural aspergillus abscess; however to our knowledge there are no reports of intramedullary aspergillus infection. We present a 19-year-old woman with active acute lymphoblastic leukemia who presented with several weeks of fevers and bilateral lower extremity weakness. She was found to have an intramedullary aspergillus abscess at T12-L1 resulting from adjacent vertebral osteomyelitis and underwent surgical debridement with ultra-sound guided aspiration and aggressive intravenous voriconazole therapy. To our knowledge this is the first reported case of spinal aspergillosis invading the intramedullary cavity. Though rare, this entity should be included in the differential for immunocompromised patients presenting with fevers and neurologic deficit. Early recognition with aggressive neurosurgical intervention and antifungal therapy may improve outcomes in future cases., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

In this study, evidence-based medicine is used to assess optimal surgical and medical management of patients with post-operative deep wound infection following spinal instrumentation. A computerized literature search of the PubMed database was performed. Twenty pertinent studies were identified. Studies were separated into publications addressing instrumentation retention versus removal and publications addressing antibiotic therapy regimen. The findings were classified based on level of evidence (I-III) and findings were summarized into evidentiary tables. No level I or II evidence was identified. With regards to surgical management, five studies support instrumentation retention in the setting of early deep infection. In contrast, for delayed infection, the evidence favors removal of instrumentation at the time of initial debridement. Surgeons should be aware that for deformity patients, even if solid fusion is observed, removal of instrumentation may be associated with significant loss of correction. A course of intravenous antibiotics followed by long-term oral suppressive therapy should be pursued if instrumentation is retained. A shorter treatment course may be appropriate if hardware is removed., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Objectives: To investigate whether associations of smoking with depression and anxiety are likely to be causal, using a Mendelian randomisation approach., Design: Mendelian randomisation meta-analyses using a genetic variant (rs16969968/rs1051730) as a proxy for smoking heaviness, and observational meta-analyses of the associations of smoking status and smoking heaviness with depression, anxiety and psychological distress., Participants: Current, former and never smokers of European ancestry aged ≥16 years from 25 studies in the Consortium for Causal Analysis Research in Tobacco and Alcohol (CARTA)., Primary Outcome Measures: Binary definitions of depression, anxiety and psychological distress assessed by clinical interview, symptom scales or self-reported recall of clinician diagnosis., Results: The analytic sample included up to 58 176 never smokers, 37 428 former smokers and 32 028 current smokers (total N=127 632). In observational analyses, current smokers had 1.85 times greater odds of depression (95% CI 1.65 to 2.07), 1.71 times greater odds of anxiety (95% CI 1.54 to 1.90) and 1.69 times greater odds of psychological distress (95% CI 1.56 to 1.83) than never smokers. Former smokers also had greater odds of depression, anxiety and psychological distress than never smokers. There was evidence for positive associations of smoking heaviness with depression, anxiety and psychological distress (ORs per cigarette per day: 1.03 (95% CI 1.02 to 1.04), 1.03 (95% CI 1.02 to 1.04) and 1.02 (95% CI 1.02 to 1.03) respectively). In Mendelian randomisation analyses, there was no strong evidence that the minor allele of rs16969968/rs1051730 was associated with depression (OR=1.00, 95% CI 0.95 to 1.05), anxiety (OR=1.02, 95% CI 0.97 to 1.07) or psychological distress (OR=1.02, 95% CI 0.98 to 1.06) in current smokers. Results were similar for former smokers., Conclusions: Findings from Mendelian randomisation analyses do not support a causal role of smoking heaviness in the development of depression and anxiety., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Background: Minimally invasive transforaminal lumbar interbody fusion (MI-TLIF) is commonly used for the treatment of degenerative lumbar spinal disorders. The rate of postoperative neurological deficits is traditionally low. New neurological postoperative complications may be underreported. We report our infrequent rate of MI-TLIF procedures complicated by postoperative weakness., Methods: A database of 340 patients was evaluated, all of whom underwent MI-TLIF procedures performed between January 2002 and June 2012 by the senior author. We identified four cases (1.2%) whose postoperative course was complicated with bilateral lower extremity weakness. We retrospectively reviewed their past medical history, operative time, estimated blood loss, length of hospital stay, changes in intraoperative neurophysiological monitoring, and pre- and postoperative neurological exams., Results: The average age of the four patients was 65.5 years(range: 62-75 years), average body mass index (BMI) was 25.1 (range: 24.1-26.6), and there were three females and one male. All patients had preoperative degenerative spondylolisthesis (either grade I or grade II). All patients were placed on a Wilson frame during surgery and underwent unilateral left-sided MI-TLIF. Three out of the four patients had a past medical history significant for abdominal or pelvic surgery and one patient had factor V Leiden deficiency syndrome., Conclusions: The rate of new neurological deficits following an MI-TLIF procedure is low, as documented in this study where the rate was 1.2%. Nonetheless, acknowledgement and open discussion of this serious complication is important for surgeon education. Of interest, the specific etiology or pathophysiology behind these complications remains relatively unknown (e.g. direct neural injury, traction injury, hypoperfusion, positioning complication, and others) despite there being some similarities between the patients and their perioperative courses.

The pituitary gland is a key structure in the hypothalamic-pituitary-gonadal (HPG) axis--it plays an important role in sexual maturation during puberty. Despite its small size, its volume can be quantified using magnetic resonance imaging (MRI). Here, we study a cohort of 962 typically developing adolescents from the Saguenay Youth Study and estimate pituitary volumes using a newly developed multi-atlas segmentation method known as the MAGeT Brain algorithm. We found that age and puberty stage (controlled for age) each predicts adjusted pituitary volumes (controlled for total brain volume) in both males and females. Controlling for the effects of age and puberty stage, total testosterone and estradiol levels also predict adjusted pituitary volumes in males and pre-menarche females, respectively. These findings demonstrate that the pituitary gland grows during adolescence, and its volume relates to circulating plasma-levels of sex steroids in both males and females., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Transforaminal lumbar interbody fusion (TLIF) is an important surgical option for the treatment of back pain and radiculopathy. The minimally invasive TLIF (MI-TLIF) technique is increasingly used to achieve neural element decompression, restoration of segmental alignment and lordosis, and bony fusion. This article reviews the surgical technique, outcomes, and complications in a series of 144 consecutive 1- and 2-level MI-TLIFs in comparison with an institutional control group of 54 open traditional TLIF procedures with a mean of 46 months' follow-up. The evidence base suggests that MI-TLIF can be performed safely with excellent long-term outcomes., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Posterior approaches for decompression in minimally invasive spine surgery are increasingly used for a wide range of pathology. Surgeons and patients must understand these risks in order to identify, manage, and ideally prevent complications. Technical intraoperative complications, recurrences and reoperations, infections, and medical complications associated with the surgery are considered for common posterior minimally invasive decompression procedures of the cervical and lumbar spine. Methods of possibly avoiding these complications are also discussed. This article then aggregates the relevant data to allow concise understanding of the complications associated with these procedures., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The lateral transpsoas approach to the lumbar spine has become an increasingly popular method to achieve fusion. Although this approach requires less tissue dissection, a smaller incision, decreased operative time, reduced blood loss and postoperative pain, and shorter hospital stay, it carries the potential for serious neurologic and visceral complications. This article reviews these complications in detail and proposes mechanisms for their avoidance., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Objective: Minimally invasive spine surgery (MISS) techniques have similar long-term outcomes compared to open surgery for patients undergoing 1- or 2-level discectomy, foraminotomy, or laminectomy. However, the rate of cerebrospinal fluid (CSF) leaks with both techniques has not been well established in the literature. This study sought to compare the rate and clinical impact of CSF leak in open lumbar foraminotomy, discectomy, or laminectomy with comparable MISS approaches., Methods: A total of 863 patients undergoing 1- or 2-level discectomy, foraminotomy, or laminectomy by either MISS an open technique were evaluated for CSF leaks. Variables assessed included operative time, blood loss, CSF leaks, hospital stay, days with lumbar drain, days of postoperative flat bed rest, and postoperative intervention. Statistical analyses include univariate analysis (Student t test, analysis of variance, odds ratio, χ(2)) and bivariate analysis (logistic regression)., Results: In the MISS group there were 15 CSF leaks (4.7%), and 49 CSF leaks (9.0%) in the open group. Eight patients in the open group required lumbar drainage, compared to zero patients in the MISS group. Twelve patients required reoperation for persistent CSF leak in the open group, compared to zero patients in the MISS group. Patients undergoing open spine surgery were 2 times more likely to have a CSF leak (odds ratio = 2.3, 95% confidence interval = 1.2 to 3.7, P = .01). Patients undergoing MISS had significantly lower reoperation rates for CSF leak repairs (open = 25% of open CSF leak cases, MISS = 0%, P < .01)., Conclusions: In this study, there was a statistically significant decreased rate of CSF leak between an MISS approach and an open surgical approach. Furthermore, CSF leaks in open surgery have a higher probability of requiring lumbar drainage or reoperation to repair the durotomy., (Copyright © 2014 Elsevier Inc. All rights reserved.)