bluuebunny/crossref_metadata_2025 huggingface dataset:
Beyond tokenism in quality management policy and programming: moving from participation to meaningful involvement of people with HIV in New York State
Authors: Coren Freda , Brown Margaret K , Ikeda Daniel J , Tietz Daniel , Steinbock Clemens , Baim-Lance Abigail , Agins Bruce D
Published: January 2021

Background

Consumer involvement in health-care policy and quality management (QM) programming is a key element in making health systems people-centered. Involvement of health-care consumers in these areas, however, remains underdeveloped and under-prioritized. When consumer involvement is actively realized, few mechanisms for assessing its impact have been developed. The New York State Department of Health (NYSDOH) embraces consumer involvement of people with HIV in QM as a guiding principle, informed by early HIV/AIDS advocacy and a framework of people-centered quality care.

Method

HIV consumer involvement is implemented statewide and informs all quality of care programming as a standard for QM in health-care organizations, implemented through four key several initiatives: (i) a statewide HIV Consumer Quality Advisory Committee; (ii) leadership and QM trainings for consumers; (iii) specific tools and activities to engage consumers in QM activities at state, regional and health-care facility levels and (iv) formal organizational assessments of consumer involvement in health-care facility QM programs.

Results

We review the literature on this topic and place the methods used by the NYSDOH within a theoretical framework for consumer involvement.

Conclusion

We present a model that offers a paradigm for practical implementation of routine consumer involvement in QM programs that can be replicated in other health-care settings, both disease-specific and general, reflecting the priority of active participation of consumers in QM activities at all levels of the health system.
Current knowledge of Krüppel-like factor 5 and vascular remodeling: providing insights for therapeutic strategies
Authors: Xie Ziyan , Chen Junye , Wang Chenyu , Zhang Jiahao , Wu Yanxiang , Yan Xiaowei
Published: January 2021

Vascular remodeling is a pathological basis of various disorders. Therefore, it is necessary to understand the occurrence, prevention, and treatment of vascular remodeling. Krüppel-like factor 5 (KLF5) has been identified as a significant factor in cardiovascular diseases during the last two decades. This review provides a mechanism network of function and regulation of KLF5 in vascular remodeling based on newly published data and gives a summary of its potential therapeutic applications. KLF5 modulates numerous biological processes, which play essential parts in the development of vascular remodeling, such as cell proliferation, phenotype switch, extracellular matrix deposition, inflammation, and angiogenesis by altering downstream genes and signaling pathways. Considering its essential functions, KLF5 could be developed as a potent therapeutic target in vascular disorders.
Assessing the accuracy of cosmological parameters estimated from velocity – density comparisons via simulations
Authors: Hollinger Amber M , Hudson Michael J
Published: January 2021

A promising method for measuring the cosmological parameter combination fσ8 is to compare observed peculiar velocities with peculiar velocities predicted from a galaxy density field using perturbation theory. We use N-body simulations and semi-analytical galaxy formation models to quantify the accuracy and precision of this method. Specifically, we examine a number of technical aspects, including the optimal smoothing length applied to the density field, the use of dark matter haloes or galaxies as tracers of the density field, the effect of noise in the halo mass estimates or in the stellar-to-halo mass relation, and the effect of finite survey volumes. We find that for a Gaussian smoothing of $4 \mbox{$\, h^{-1}$}$ Mpc, the method has only small systematic biases at the level of 5 per cent. Cosmic variance affects current measurements at the 5 per cent level due to the volume of current redshift data sets.
Reopening Interventional Pain Practices during the Early Phase of the COVID-19 Global Pandemic
Authors: Huynh Lisa , Chang Richard G , Chhatre Akhil , Sayeed Yusef , MacVicar John , McCormick Zachary L , Duszynski Belinda , Smith Clark
Published: January 2021

Objective

Examine how interventional pain physicians navigated the early phase of reopening practices during the coronavirus disease 2019 (COVID-19) pandemic.

Methods

In June/July 2020, Spine Intervention Society members were queried about practice demographics, perception of COVID-19 prevalence, financial impact, and implementation of new tools and procedures when re-opening practices.

Results

Of the 2,295 members approached, 195 (8%) completed the survey. A majority (71%) reported using risk stratification tools and changing scheduling patterns. Nearly 70% performed initial assessments via telehealth and 87% for follow-up encounters. More than 80% performed symptom/temperature checks upon in-person clinic/facility entrance, and 63% screened patients via phone. Most (58%) did not test patients for COVID-19 for office visits, while 38% tested only if symptomatic. For epidural injections, intra-articular injections, and radiofrequency neurotomy procedures, 43% reported not testing patients, while 36% tested patients only if symptomatic. Most (70%) required patients to wear a mask upon entering the clinic/facility. For nonprocedure encounters, respondents used surgical masks (85%), gloves (35%), face shields/goggles (24%), N95 respirators (15%), and gowns (6%). Some (66%) discussed unique COVID-19 risks/complications and 26% provided written information. Most did not make changes to steroid dosage (67%) or peri-procedural anticoagulation management (97%). The vast majority (81%) estimated that COVID-19 will have a moderate-severe financial impact on their practice.

Conclusions

COVID-19 has dramatically affected interventional pain practices with regard to telehealth, in-clinic precautions, screening/testing protocols, and patient counseling. Practice patterns will continue to evolve as we learn more about the disease and improve methods to provide safe and effective care.
Using 3-Dimensional Motion Analysis to Optimize Treatment Planning for a Patient With Dropfoot: Case Report
Authors: Feng Jing , Bompiani Erin , Coats Scott , Patrick Do K , Aiona Michael
Published: January 2021

Objective

This study demonstrated the use of computerized motion analysis to assist in evidence-based clinical decision-making.

Case Description

A 15-year-old girl who had right hemiparesis after a stroke was referred for 3-dimensional computerized motion analysis to determine the effect of 3 devices intended to control her dropfoot and to assist in developing a treatment plan. Four conditions were tested and compared: barefoot, lateral support ankle brace, functional electrical stimulation (FES) device, and dropfoot cuff.

Results

Kinematics showed the right ankle had significant dropfoot during swing phase (32.7 degrees of plantarflexion at terminal swing) in barefoot. The lateral support ankle brace, FES device, and dropfoot cuff reduced terminal swing plantarflexion to 27.2 degrees, 17.6 degrees, and 15.3 degrees, respectively, though ankle kinematics remained abnormal because of inadequate dorsiflexion. Improvements in gait variable score with FES (−8.2 degrees) or dropfoot cuff (−8.7 degrees) were significantly more than that with the lateral support brace (−2.2 degrees), and the difference in gait variable score between FES and dropfoot cuff was insignificant. Compared with the barefoot condition, the lateral support brace condition did not show a clinically significant difference in gait profile score; however, the gait profile scores of both FES and dropfoot cuff conditions showed clinically significant improvement (−1.7 degrees and −2.1 degrees, respectively).

Conclusion

Objective data delineated subtle changes among 3 devices and led to the recommendation to discontinue the lateral support ankle brace, continue using her night ankle-foot orthosis and FES device, with the dropfoot cuff as a backup when she feels leg fatigue or skin irritation, and consider serial casting or surgical calf lengthening.

Impact

Computerized motion analysis provides quantitative evaluation of subtle differences in the effect of braces with different designs, which are hard for the human eye to discern. The objective data inform and validate treatment decision-making. The recommendations were made as a result of evidence-based practice.
THE UNIFIED INTERACTION MODEL: SIMULATIONS OF TL DOSE RESPONSE AND EXPERIMENTAL VERIFICATION OF INTENDED DOSE RESPONSE LINEARITY BY POSTIRRADIATION PHOTON EXCITATION
Authors: Ginzburg Dimitry , Oster Leonid , Biderman Shlomo , Reshes Galina , Eliyahu Ilan
Published: January 2021

The Unified Interaction Model (UNIM) simulates the linear/supralinear dose response of the glow peaks of LiF:Mg,Ti (TLD-100) and other thermoluminescent materials and the dependence of the supralinearity on photon/electron energy. The UNIM is based on the radiation action of spatially correlated trapping centres (TCs) and luminescent centres (LCs), which results in localised (geminate) electron/hole recombination by quantum mechanical tunnelling. The linear dose response is mainly attributed to geminate recombination. UNIM simulations of the dose response of glow peak 5 in LiF:Mg,Ti following 500 and 8 keV electron/photon irradiation are discussed. In addition, simulations of postirradiation photo-excitation that redistribute the electrons and holes in the various TCs and LCs are demonstrated to extend the region of linear dose response and reduce the supralinearity. Experimental verification of dose–response linearity for levels of dose ≤30 Gy following both 3.65 and 5 eV photon excitation is presented.
Dynamics of above- and belowground responses of silver birch saplings and soil gases to soil freezing and waterlogging during dormancy
Authors: Repo Tapani , Domisch Timo , Roitto Marja , Kilpeläinen Jouni , Wang Ai-Fang , Piirainen Sirpa , Heiskanen Juha , Makita Naoki , Lehto Tarja , Sutinen Sirkka
Published: January 2021

Winter precipitation and soil freeze–thaw events have been predicted to increase in boreal regions with climate change. This may expose tree roots to waterlogging (WL) and soil freezing (Fr) more than in the current climate and therefore affect tree growth and survival. Using a whole-tree approach, we studied the responses of silver birch (Betula pendula Roth.) saplings, growing in mineral soil, to 6-week Fr and WL in factorial combinations during dormancy, with accompanying changes in soil gas concentrations.

Physiological activation (dark-acclimated chlorophyll fluorescence and chlorophyll content index) and growth of leaves and shoot elongation and stem diameter growth started earlier in Fr than NoFr (soil not frozen). The starch content of leaves was temporarily higher in Fr than NoFr in the latter part of the growing season. Short and long root production and longevity decreased, and mortality increased by soil Fr, while there were no significant effects of WL. Increased fine root damage was followed by increased compensatory root growth. At the beginning of the growing season, stem sap flow increased fastest in Fr + WL, with some delay in both NoWL (without WL) treatments. At the end of the follow-up growing season, the hydraulic conductance and impedance loss factor of roots were higher in Fr than in NoFr, but there were no differences in above- and belowground biomasses. The concentration of soil carbon dioxide increased and methane decreased by soil Fr at the end of dormancy. At the beginning of the growing season, the concentration of nitrous oxide was higher in WL than in NoWL and higher in Fr than in NoFr. In general, soil Fr had more consistent effects on soil greenhouse gas concentrations than WL. To conclude, winter-time WL alone is not as harmful for roots as WL during the growing season.
Pollination promotes ABA synthesis but not sexual reproduction in the apomictic speciesZanthoxylum bungeanumMaxim.
Authors: Fei Xitong , Shi Qianqian , Lei Yu , Wang Shujie , Qi Yichen , Hu Haichao , Wei Anzhi
Published: January 2021

Apomixis is a form of reproduction that does not involve the fertilization of female gametes by male gametes but instead involves the production of offspring directly from the female parent. The offspring of apomixis are genetically identical to the female parent and inherit its traits. Therefore, apomixis has great potential for application to agricultural genetic breeding. However, it remains unclear whether apomictic species require pollination, and the impacts of pollination on such species are poorly understood. We investigated the effects of pollination on the apomictic species Zanthoxylum bungeanum Maxim. by analyzing its fertilization process, assembling its transcriptome, and measuring hormone concentrations, fruit setting rate and gene expression levels. Transcriptome sequencing of pollinated and unpollinated fruits resulted in a total of 69,131 PacBio reads. Of these, 7102 genes were up-regulated and 6491 genes were down-regulated. Analysis of the differentially expressed genes (DEGs) and construction of a weighted gene co-expression network showed that many DEGs were involved in plant hormone signal transduction, suggesting that hormonal signaling during development differs between pollinated and unpollinated fruit. The germination rate of Z. bungeanum pollen in vitro was only 11%, and pollen could not germinate in the embryo sac to complete fertilization. Although pollination did not enable Z. bungeanum to complete the sexual reproduction process, it significantly increased abscisic acid (ABA) concentration and fruit setting rate. Spraying 100 μg l−1 ABA also significantly increased the fruit setting rate. Therefore, ABA appears to be a key factor in the regulation of fruit setting in apomictic Z. bungeanum. Based on these results, we suggest that some male plants be cultivated in Z. bungeanum plantations or exogenous ABA be sprayed to increase the likelihood of pollination and thereby increase the fruit setting rate.
Overexpression of Solanum tuberosum Respiratory Burst Oxidase Homolog A (StRbohA) Promotes Potato Tolerance to Phytophthora infestans
Authors: Soliman Atta , Adam Lorne R. , Rehal Pawanpuneet K. , Daayf Fouad
Published: January 2021

Reactive oxygen species (ROSs) represent one of the first lines of plants’ biochemical defense against pathogens. Plants’ respiratory burst oxidase homologs (RBOHs) produce ROSs as byproducts in several cellular compartments. In potato tubers, Solanum tuberosum respiratory burst oxidase homolog (StRBOHs) are involved in suberization and healing of wounded tissues. StRbohA has been tested in the model plant Arabidopsis thaliana, which led to enhanced plant defense against the soilborne pathogen Verticillium dahliae. Here, we showed that overexpressing StRbohA in potato plants increases plant tolerance to the oomycete Phytophthora infestans, the causal agent of late blight disease. Transgenic potato plants expressing StRbohA showed reduced disease symptoms (necrosis) compared with the wild type. In parallel, the expression of pathogenesis-related genes (PRs); RBOHs; antioxidation-related genes CPRX1, PRX2, APRX1, CAT1, and CAT2; and genes involved in the biosynthesis pathways of jasmonic and salicylic acids (ICS, PAL1, PAL2, LOX1, LOX2, and LOX3) exhibited significant increases in transgenic plants in response to infection. After higher expression of RBOHs, ROSs accumulated more in inoculation sites of the transgenic plants. ROSs act as signals that activate gene expression in the salicylic acid (SA) biosynthesis pathway, leading to the accumulation of SA and triggering SA-based defense mechanisms. SA-responsive PRs showed higher expression in the transgenic plants, which resulted in the restriction of pathogen growth in plant tissues. These results demonstrate the effective role of StRbohA in increasing potato defense against P. infestans.
Prediction of individuals at high-risk of chronic kidney disease during treatment with lithium for bipolar disorder
Authors: Hayes Joseph F , Osborn David PJ , Francis Emma , Ambler Gareth , Tomlinson Laurie A , Boman Magnus , Wong Ian , Geddes John , Dalman Christina , Lewis Glyn
Published: January 2021

Background

Lithium is the most effective treatment in bipolar disorder. Its use is limited by concerns about risk of chronic kidney disease (CKD). We aimed to develop a model to predict risk of CKD following lithium treatment initiation, by identifying individuals with a high-risk trajectory of renal function.

Methods

We used United Kingdom Clinical Practice Research Datalink (CPRD) electronic heath records (EHRs) from 2000-2018. CPRD Aurum for prediction model development and CPRD Gold for external validation. We used elastic net to generate a prediction model from potential features. We performed discrimination and calibration assessments in an external validation data set.

We included all patients aged ≥16 with bipolar disorder prescribed lithium. To be included patients had to have ≥1 year of follow-up before lithium initiation, ≥3 estimated glomerular filtration rate (eGFR) measures after lithium initiation (to be able to determine a trajectory) and a normal (≥60 mL/min/1.73m2) eGFR at lithium initiation (baseline). In the Aurum development cohort 1609 fulfilled these criteria. The Gold external validation cohort included 934 patients.

We included 44 potential baseline features in the prediction model, including sociodemographic, mental and physical heath and drug treatment characteristics. We compared a full model with the 3-variable five-year kidney failure risk equation (KFRE) and a 3-variable elastic net model.

We used group-based trajectory modelling to identify latent trajectory groups for eGFR. We were interested in the group with deteriorating renal function (the high-risk group).

Findings

The high-risk group included 191 (11.87%) of the Aurum cohort and 137 (14.67%) of the Gold cohort, of these 168 (87.96%) and 117 (85.40%) respectively developed CKD 3a or more severe during follow-up. The model, developed in Aurum, had a ROC area of 0.879 (95%CI 0.853-0.904) in the Gold external validation data set. At the empirical optimal cut-point defined in the development dataset, the model had a sensitivity of 0. 91 (95%CI 0.84-0.97) and a specificity of 0.74 (95% CI 0.67-0.82). However, a 3-variable elastic net model (including only age, sex and baseline eGFR) performed similarly well (ROC area 0.888; 95%CI 0.864-0.912), as did the KFRE (ROC area 0.870; 95%CI 0.841-0.898).

Conclusions

Individuals at high-risk of a poor trajectory of renal function can be identified before initiation of lithium treatment by a simple equation including age, sex and baseline eGFR. We did not identify strong predicters of renal impairment specific to lithium treated patients.
Agonistic CD40 antibody therapy induces tertiary lymphoid structures but impairs the response to immune checkpoint blockade in glioma
Authors: van Hooren Luuk , Vaccaro Alessandra , Ramachandran Mohanraj , Vazaios Konstantinos , Libard Sylwia , van de Walle Tiarne , Georganaki Maria , Huang Hua , Pietilä Ilkka , Lau Joey , Ulvmar Maria H. , Karlsson Mikael C.I. , Zetterling Maria , Mangsbo Sara M. , Jakola Asgeir S. , Bontell Thomas Olsson , Smits Anja , Essand Magnus , Dimberg Anna
Published: January 2021

Gliomas are brain tumors characterized by immunosuppression. Immunostimulatory agonistic CD40 antibodies (αCD40) are in clinical development for solid tumors but are yet to be evaluated for glioma. Here, systemic delivery of αCD40 led to cytotoxic T cell dysfunction and impaired the response to immune checkpoint inhibitors in preclinical glioma models. This was associated with an accumulation of suppressive CD11b+B cells. However, αCD40 also induced tertiary lymphoid structures (TLS). In human glioma, TLS correlated with increased T cell infiltration indicating enhanced immune responses. Our work unveils the pleiotropic effects of αCD40 therapy in glioma, which is of high clinical relevance.
RASSF1C oncogene elicits amoeboid invasion, cancer stemness and invasive EVs via a novel SRC/Rho axis
Authors: Tognoli Maria Laura , Vlahov Nikola , Steenbeek Sander , Grawenda Anna M. , Eyres Michael , Cano-Rodriguez David , Scrace Simon , Kartsonaki Christiana , von Kriegsheim Alex , Willms Eduard , Wood Matthew J. , Rots Marianne G. , van Rheenen Jacco , O’Neill Eric , Pankova Daniela
Published: January 2021

Cell plasticity is a crucial hallmark leading to cancer metastasis. Upregulation of Rho/ROCK pathway drives actomyosin contractility, protrusive forces and contributes to the occurrence of highly invasive amoeboid cells in tumors. Cancer stem cells are similarly associated with metastasis, but how these populations arise in tumors is not fully understood. Here we show that the novel oncogene RASSF1C drives mesenchymal to amoeboid transition and stem cell attributes in breast cancer cells. Mechanistically, RASSF1C activates Rho/ROCK via SRC mediated RhoGDI inhibition, resulting in generation of actomyosin contractility. Moreover, we demonstrate that amoeboid cells display the cancer stem cell markers CD133, ALDH1 and the pluripotent marker Nanog; are accompanied by higher invasive potentialin vitroandin vivo;and employ extracellular vesicles to transfer the invasive phenotype to target cells and tissue. Importantly, the underlying RASSF1C driven biological processes concur to explain clinical data: namely, methylation of the RASSF1C promoter correlates with better survival in early stage breast cancer patients. Therefore, we propose the use ofRASSF1gene promoter methylation status as a biomarker for patient stratification.
Increased sensitivity to strong perturbations in a whole-brain model of LSD
Authors: Jobst Beatrice M. , Atasoy Selen , Ponce-Alvarez Adrián , Sanjuán Ana , Roseman Leor , Kaelen Mendel , Carhat-Harris Robin , Kringelbach Morten L. , Deco Gustavo
Published: January 2021

Lysergic acid diethylamide (LSD) is a potent psychedelic drug, which has seen a revival in clinical and pharmacological research within recent years. Human neuroimaging studies have shown fundamental changes in brain-wide functional connectivity and an expansion of dynamical brain states, thus raising the question about a mechanistic explanation of the dynamics underlying these alterations. Here, we applied a novel perturbational approach based on a whole-brain computational model, which opens up the possibility to externally perturb different brain regions in silico and investigate differences in dynamical stability of different brain states, i.e. the dynamical response of a certain brain region to an external perturbation. After adjusting the whole-brain model parameters to reflect the dynamics of functional magnetic resonance imaging (fMRI) BOLD signals recorded under the influence of LSD or placebo, perturbations of different brain areas were simulated by either promoting or disrupting synchronization in the regarding brain region. After perturbation offset, we quantified the recovery characteristics of the brain area to its basal dynamical state with the Perturbational Integration Latency Index (PILI) and used this measure to distinguish between the two brain states. We found significant changes in dynamical complexity with consistently higher PILI values after LSD intake on a global level, which indicates a shift of the brain’s global working point further away from a stable equilibrium as compared to normal conditions. On a local level, we found that the largest differences were measured within the limbic network, the visual network and the default mode network. Additionally, we found a higher variability of PILI values across different brain regions after LSD intake, indicating higher response diversity under LSD after an external perturbation. Our results provide important new insights into the brain-wide dynamical changes underlying the psychedelic state - here provoked by LSD intake - and underline possible future clinical applications of psychedelic drugs in particular psychiatric disorders.

Highlights

Novel offline perturbational method applied on functional magnetic resonance imaging (fMRI) data under the effect of lysergic acid diethylamide (LSD)

Shift of brain’s global working point to more complex dynamics after LSD intake

Consistently longer recovery time after model perturbation under LSD influence

Strongest effects in resting state networks relevant for psychedelic experience

Higher response diversity across brain regions under LSD influence after an external in silico perturbation
COVID-19 virtual patient cohort reveals immune mechanisms driving disease outcomes
Authors: Jenner Adrianne L. , Aogo Rosemary A. , Alfonso Sofia , Crowe Vivienne , Smith Amanda P. , Morel Penelope A. , Davis Courtney L. , Smith Amber M. , Craig Morgan
Published: January 2021

To understand the diversity of immune responses to SARS-CoV-2 and distinguish features that predispose individuals to severe COVID-19, we developed a mechanistic, within-host mathematical model and virtual patient cohort. Our results indicate that virtual patients with low production rates of infected cell derived IFN subsequently experienced highly inflammatory disease phenotypes, compared to those with early and robust IFN responses. In thesein silicopatients, the maximum concentration of IL-6 was also a major predictor of CD8+T cell depletion. Our analyses predicted that individuals with severe COVID-19 also have accelerated monocyte-to-macrophage differentiation that was mediated by increased IL-6 and reduced type I IFN signalling. Together, these findings identify biomarkers driving the development of severe COVID-19 and support early interventions aimed at reducing inflammation.

Author summary

Understanding of how the immune system responds to SARS-CoV-2 infections is critical for improving diagnostic and treatment approaches. Identifying which immune mechanisms lead to divergent outcomes can be clinically difficult, and experimental models and longitudinal data are only beginning to emerge. In response, we developed a mechanistic, mathematical and computational model of the immunopathology of COVID-19 calibrated to and validated against a broad set of experimental and clinical immunological data. To study the drivers of severe COVID-19, we used our model to expand a cohort of virtual patients, each with realistic disease dynamics. Our results identify key processes that regulate the immune response to SARS-CoV-2 infection in virtual patients and suggest viable therapeutic targets, underlining the importance of a rational approach to studying novel pathogens using intra-host models.
Quantification of fast molecular adhesion by fluorescence footprinting
Authors: Yasunaga Adam B. , Li Isaac T.S.
Published: January 2021

Rolling adhesion is a unique process in which the adhesion events are short-lived and operate under highly non-equilibrium conditions. These characteristics pose a challenge in molecular force quantification, wherein situmeasurement of such forces cannot be achieved with most molecular force sensors that probe near equilibrium. In this report, we demonstrated a quantitative adhesion footprint assay combining DNA-based non-equilibrium force probes and modelling to measure the molecular force involved in fast rolling adhesion. We were able to directly profile the ensemble molecular force distribution during rolling adhesion with a dynamic range between 0 – 18 pN. Our results showed that the shear stress driving bead rolling motility directly controls the molecular tension on the probe-conjugated adhesion complex. Furthermore, the shear stress can steer the dissociation bias of components within the molecular force probe complex, favouring either DNA probe dissociation or receptor-ligand dissociation.
Kinetic and structural mechanism for DNA unwinding by a non-hexameric helicase
Authors: Carney Sean P. , Ma Wen , Whitley Kevin D. , Jia Haifeng , Lohman Timothy M. , Luthey-Schulten Zaida , Chemla Yann R.
Published: January 2021

UvrD, a model for non-hexameric Superfamily 1 helicases, utilizes ATP hydrolysis to translocate stepwise along single-stranded DNA and unwind the duplex. To dissect the mechanism underlying DNA unwinding, we use optical tweezers to measure directly the stepping behavior of UvrD as it processes a DNA hairpin and show that UvrD exhibits a variable step size averaging ~3 base pairs. Analyzing stepping kinetics across ATP reveals the type and number of catalytic events that occur with different step sizes. These single-molecule data reveal a mechanism in which UvrD moves one base pair at a time but sequesters the nascent single strands, releasing them non-uniformly after a variable number of catalytic cycles. Molecular dynamics simulations point to a structural basis for this behavior, identifying the protein-DNA interactions responsible for strand sequestration. Based on structural and sequence alignment data, we propose that this stepping mechanism may be conserved among other non-hexameric helicases.
Genomic and phenotypic characterization of experimentally selected resistantLeishmania donovanireveals a role for dynamin-1 like protein in the mechanism of resistance to a novel anti-leishmanial compound
Authors: Hefnawy Aya , Negreira Gabriel , Jara Marlene , Cotton James A. , Maes Ilse , D’ Haenens Erika , Imamura Hideo , Cuypers Bart , Monsieurs Pieter , Mouchtoglou Christina , De Winter Hans , Berriman Matt , Sanders Mandy , Martin Julio , de Muylder Geraldine , Dujardin Jean-Claude , Sterckx Yann G.-J. , Domagalska Malgorzata Anna
Published: January 2021

The implementation of prospective drug resistance (DR) studies in the R&D pipelines is a common practice for many infectious diseases, but not for Neglected Tropical Diseases. Here, we explored and demonstrated the importance of this approach, using as paradigmsLeishmania donovani, the etiological agent of Visceral Leishmaniasis (VL), and TCMDC-143345, a promising compound of the GSK ‘Leishbox’ to treat VL. We experimentally selected resistance to TCMDC-143345 in vitro and characterized resistant parasites at genomic and phenotypic levels. We found that it took more time to develop resistance to TCMDC-143345 than to other drugs in clinical use and that there was no cross resistance to these drugs, suggesting a new and unique mechanism. By whole genome sequencing, we found two mutations in the gene encoding theL. donovanidynamin-1-like protein (LdoDLP1) that were fixed at highest drug pressure. Through phylogenetic analysis, we identified LdoDLP1 as a family member of the dynamin-related proteins, a group of proteins that impacts the shapes of biological membranes by mediating fusion and fission events, with a putative role in mitochondrial fission. We found thatL. donovanilines genetically engineered to harbor the two identified LdoDLP1 mutations were resistant to TCMDC-143345 and displayed altered mitochondrial properties. By homology modeling, we showed how the two LdoDLP1 mutations may influence protein structure and function. Taken together, our data reveal a clear involvement of LdoDLP1 in the adaptation/resistance ofL. donovanito TCMDC-143345.

Importance

Humans and their pathogens are continuously locked in a molecular arms race during which the eventual emergence of pathogen drug resistance (DR) seems inevitable. For neglected tropical diseases (NTDs), DR is generally studied retrospectively, once it has already been established in clinical settings. We previously recommended to keep one step ahead in the host-pathogen arms race and implement prospective DR studies in the R&D pipeline, a common practice for many infectious diseases, but not for NTDs. Here, usingLeishmania donovani, the etiological agent of Visceral Leishmaniasis (VL), and TCMDC-143345, a promising compound of the GSK ‘Leishbox’ to treat VL, as paradigms, we experimentally selected resistance to the compound and proceeded to genomic and phenotypic characterization of DR parasites. The results gathered in the present study suggest a new DR mechanism involving theL. donovanidynamin-1 like protein (LdoDLP1) and demonstrate the practical relevance of prospective DR studies.
Dynamics of antibodies to SARS-CoV-2 in convalescent plasma donors
Authors: Steenhuis Maurice , van Mierlo Gerard , Derksen Ninotska I.L. , Ooijevaar-de Heer Pleuni , Kruithof Simone , Loeff Floris L. , Berkhout Lea C. , Linty Federica , Reusken Chantal , Reimerink Johan , Hogema Boris , Zaaijer Hans , van de Watering Leo , Swaneveld Francis , van Gils Marit J. , Bosch Berend Jan , van Ham Marieke , Brinke Anja ten , Vidarsson Gestur , van der Schoot Ellen C. , Rispens Theo
Published: January 2021

The novel SARS-CoV-2 virus emerged in late 2019 and has caused a global health and economic crisis. The characterization of the human antibody response to SARS-CoV-2 infection is vital for serosurveillance purposes as well for treatment options such as transfusion with convalescent plasma or immunoglobin products derived from convalescent plasma. In this study, we measured antibody responses in 844 longitudinal samples from 151 RT-PCR positive SARS-CoV-2 convalescent adults during the first 34 weeks after onset of symptoms. All donors were seropositive at the first sampling moment and only one donor seroreverted during follow-up analysis. Anti-RBD IgG and anti-nucleocapsid IgG levels slowly declined with median half-life’s of 62 and 59 days during 2-5 months after symptom onset, respectively. The rate of decline of antibody levels diminished during extended follow-up. In addition, the magnitude of the IgG response correlated with neutralization capacity measured in a classic plaque reduction assay as well in our in-house developed competition assay. The result of this study gives valuable insight into the longitudinal response of antibodies to SARS-CoV-2.
No association between the SARS-CoV-2 variants and mortality rates in the Eastern Mediterranean Region
Authors: Omais Saad , Kharroubi Samer , Zaraket Hassan
Published: January 2021

As the novel coronavirus SARS-CoV-2 continues to spread in all countries, there is a growing interest in monitoring and understanding the impact of emerging strains on virus transmission and disease severity. Here, we analyzed SARS-CoV-2 genomic sequences reported in the Eastern Mediterranean Region (EMR) countries, as of 1 January 2021. The majority (∼75%) of these sequences originated from three out of 22 EMR countries, and 65.8% of all sequences belonged to GISAID clades GR, GH, G and GV. A delay ranging between 30-150 days from sample collection to sequence submission was observed across all countries, limiting the utility of such data in informing public health policies. We identified ten common non-synonymous mutations represented among SARS-CoV-2 in the EMR and several country-specific ones. Two substitutions, spike_D614G and NSP12_P323L, were predominantly concurrent in most countries. While the single incidence of NSP12_P323L was positively correlated with higher case fatality rates in EMR, no such association was established for the double (spike_D614G and NSP12_P323L) concurrent variant across the region. Our study identified critical data gaps in EMR highlighting the importance of enhancing surveillance and sequencing capacities in the region.
Neurophysiological and brain structural markers of cognitive frailty differs from Alzheimer’s disease
Authors: Kocagoncu Ece , Nesbitt David , Emery Tina , Hughes Laura , Henson Richard N. ,   Cam-CAN, Rowe James B.
Published: January 2021

With increasing life span, there is growing importance of understanding the mechanisms of successful cognitive ageing. In contrast, cognitive frailty has been proposed to be a precursor to Alzheimer’s disease. Here we test the hypothesis that cognitively frail adults represent a branch of healthy ageing, distinct from latent dementia. We used electro-magnetoencephalography and magnetic resonance imaging to investigate the structural and neurophysiological features of cognitive frailty in relation to healthy aging, and clinical presentations of mild cognitive impairment and Alzheimer’s disease. Cognitive performance of the cognitively frail group was similar to those with mild cognitive impairment. We used a novel cross-modal oddball task to induce mismatch responses to unexpected stimuli. Both controls and cognitively frail showed stronger mismatch responses and larger temporal grey matter volume, compared to people with mild cognitive impairment and Alzheimer’s disease. Our results suggest that cognitively frail represents a spectrum of normal ageing rather than incipient or undiagnosed Alzheimer’s disease. Lower cognitive reserve, hearing impairment and medical comorbidity might contribute to the aetiology of cognitive impairment.
Hierarchical Regulation of Autophagy During Adipocyte Differentiation
Authors: Ahmed Mahmoud , Lai Trang Huyen , Pham Trang Minh , Zada Sahib , Elashkar Omar , Hwang Jin Seok , Kim Deok Ryong
Published: January 2021

We previously showed that some adipogenic transcription factors such as CEBPB and PPARG directly and indirectly regulate autophagy gene expression in adipogenesis. The order and the effect of these events are undetermined. In this study, we modeled the gene expression, DNA-binding of transcriptional regulators, and histone modifications during adipocyte differentiation and evaluated the effect of the regulators on gene expression in terms of direction and magnitude. Then, we identified the overlap of the transcription factors and co-factors binding sites and targets. Finally, we built a chromatin states model based on the histone marks and studied their relation with the factors’ binding. Adipogenic factors differentially regulated autophagy genes as part of the differentiation program. Co-regulators associated with specific transcription factors and preceded them to the regulatory regions. Transcription factors differed in the binding time and location, and their effect on expression was either localized or long-lasting. Adipogenic factors disproportionately targeted genes coding for autophagy-specific transcription factors. To sum, a hierarchical arrangement between adipogenic transcription factors and co-factors drives the regulation of autophagy during adipocyte differentiation.
Estimating the rate of plasmid transfer with an adapted Luria–Delbrück fluctuation analysis
Authors: Kosterlitz Olivia , Tirado Adamaris Muñiz , Wate Claire , Elg Clint , Bozic Ivana , Top Eva M. , Kerr Benjamin
Published: January 2021

To increase our basic understanding of the ecology and evolution of conjugative plasmids, we need a reliable estimate of their rate of transfer between bacterial cells. However, accurate estimates of plasmid transfer have remained elusive due to biological and experimental complexity. Current methods to measure transfer rate can be confounded by many factors. A notable example involves plasmid transfer between different strains or species where the rate that one type of cell donates the plasmid is not equal to the rate at which the other cell type donates. Asymmetry in these rates has the potential to bias or constrain current transfer estimates, thereby limiting our capabilities for estimating transfer in microbial communities. Inspired by the classic fluctuation analysis of Luria and Delbrück, we develop a novel approach, the Luria-Delbrück method (‘LDM’), for estimating plasmid transfer rate. Our new approach embraces the stochasticity of conjugation departing from the current deterministic population dynamic methods. In addition, the LDM overcomes obstacles of traditional methods by not being affected by different growth and transfer rates for each population within the assay. Using stochastic simulations and experiments, we show that the LDM has high accuracy and precision for estimation of transfer rates compared to the most widely used methods, which can produce estimates that differ from the LDM estimate by orders of magnitude.

Significance Statement

Conjugative plasmids play significant roles in the ecology and evolution of microbial communities. Notably, antibiotic resistance genes are often encoded on conjugative plasmids. Thus, conjugation—the transfer of a plasmid copy from one cell to another—is a common way for antibiotic resistance to spread between important clinical pathogens. For both public health modeling and a basic understanding of microbial population biology, accurate estimates of this fundamental rate are of great consequence. We show that widely used methods can lead to biased estimates, deviating from true values by several orders of magnitude. Therefore, we developed a new approach, inspired by the classic fluctuation analysis of Luria and Delbrück, for accurately assessing the rate of plasmid conjugation under a variety of conditions.
Role of spatial patterning of N-protein interactions in SARS-CoV-2 genome packaging
Authors: Seim Ian , Roden Christine A. , Gladfelter Amy S.
Published: January 2021

Viruses must efficiently and specifically package their genomes while excluding cellular nucleic acids and viral sub-genomic fragments. Some viruses use specific packaging signals, which are conserved sequence/structure motifs present only in the full-length genome. Recent work has shown that viral proteins important for packaging can undergo liquid-liquid phase separation (LLPS), where one or two viral nucleic acid binding proteins condense with the genome. The compositional simplicity of viral components lends itself well to theoretical modeling compared to more complex cellular organelles. Viral LLPS can be limited to one or two viral proteins and a single genome that is enriched in LLPS-promoting features. In our previous study, we observed that LLPS-promoting sequences of SARS-CoV-2 are located at the 5ʹ and 3ʹ ends of the genome, whereas the middle of the genome is predicted to consist mostly of solubilizing elements. Is this arrangement sufficient to drive single genome packaging, genome compaction, and genome cyclization? We addressed these questions using a coarse-grained polymer model, LASSI, to study the LLPS of nucleocapsid protein with RNA sequences that either promote LLPS or solubilization. With respect to genome cyclization, we find the most optimal arrangement restricts LLPS-promoting elements to the 5ʹ and 3ʹ ends of the genome, consistent with the native spatial patterning. Genome compaction is enhanced by clustered LLPS-promoting binding sites, while single genome packaging is most efficient when binding sites are distributed throughout the genome. These results suggest that many and variably positioned LLPS-promoting signals can support packaging in the absence of a singular packaging signal which argues against necessity of such a feature. We hypothesize that this model should be generalizable to multiple viruses as well as cellular organelles like paraspeckles, which enrich specific, long RNA sequences in a defined arrangement.

Statement of significance

The COVID-19 pandemic has motivated research of the basic mechanisms of coronavirus replication. A major challenge faced by viruses such as SARS-CoV-2 is the selective packaging of a large genome in a relatively small capsid while excluding host and sub-genomic nucleic acids. Genomic RNA of SARS-CoV-2 can condense with the Nucleocapsid (N-protein), a structural protein component critical for packaging of many viruses. Notably, certain regions of the genomic RNA drive condensation of N-protein while other regions solubilize it. Here, we explore how the spatial patterning of these opposing elements promotes single genome compaction, packaging, and cyclization. This model informs futurein silicoexperiments addressing spatial patterning of genomic features that are experimentally intractable because of the length of the genome.
Resolving the molecular fingerprint of the distal carboxy tail in modulating CaV1 calcium dependent inactivation
Authors: Sang Lingjie , Vieira Daiana C. O. , Yue David T. , Ben-Johny Manu , Dick Ivy E.
Published: January 2021

Ca2+/calmodulin-dependent inactivation (CDI) of CaVchannels is a critical regulatory process required for tuning the kinetics of Ca2+entry for different cell types and physiologic responses. Calmodulin (CaM) resides on the IQ domain of the CaVcarboxy-tail, such that Ca2+binding initiates a reduction in channel open probability, manifesting as CDI. This regulatory process exerts a significant impact on Ca2+entry and is tailored by alternative splicing. CaV1.3 and CaV1.4 feature a long-carboxy-tail splice variant that modulates CDI through a competitive mechanism. In these channels, the distal-carboxy-tail (DCT) harbors an inhibitor of CDI (ICDI) module that competitively displaces CaM from the IQ domain, thereby diminishing CDI. While this overall mechanism is now well-described, the detailed interaction loci for ICDI binding to the IQ domain is yet to be elucidated. Here, we perform alanine-scanning mutagenesis of the IQ and ICDI domains and evaluate the contribution of neighboring regions. We identify multiple critical residues within the IQ domain, ICDI and the nearby A region of the channel, which are required for high affinity IQ/ICDI binding. Importantly, disruption of this interaction commensurately diminishes ICDI function, as seen by the re-emergence of CDI in mutant channels. Furthermore, analysis of the homologous ICDI region of CaV1.2 reveals a selective effect of this channel region on CaV1.3 channels, implicating a cross-channel modulatory scheme in cells expressing both channel subtypes. In all, these findings provide new insights into a molecular rheostat that fine tunes Ca2+entry and supports normal neuronal and cardiac function.
Self-organized stem cell-derived human lung buds with proximo-distal patterning and novel targets of SARS-CoV-2
Authors: Rosado-Olivieri E.A. , Razooky B. , Hoffmann H.-H. , De Santis R. , Rice C.M. , Brivanlou A.H
Published: January 2021

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the global COVID-19 pandemic and the lack of therapeutics hinders pandemic control1–2. Although lung disease is the primary clinical outcome in COVID-19 patients1–3, how SARS-CoV-2 induces tissue pathology in the lung remains elusive. Here we describe a high-throughput platform to generate tens of thousands of self-organizing, nearly identical, and genetically matched human lung buds derived from human pluripotent stem cells (hPSCs) cultured on micropatterned substrates. Strikingly,in vitro-derived human lung buds resemble fetal human lung tissue and displayin vivo-like proximo-distal coordination of alveolar and airway tissue differentiation whose 3D epithelial self-organization is directed by the levels of KGF. Single-cell transcriptomics unveiled the cellular identities of airway and alveolar tissue and the differentiation of WNThicycling alveolar stem cells, a human-specific lung cell type4. These synthetic human lung buds are susceptible to infection by SARS-CoV-2 and endemic coronaviruses and can be used to track cell type-dependent susceptibilities to infection, intercellular transmission and cytopathology in airway and alveolar tissue in individual lung buds. Interestingly, we detected an increased susceptibility to infection in alveolar cells and identified cycling alveolar stem cells as targets of SARS-CoV-2. We used this platform to test neutralizing antibodies isolated from convalescent plasma that efficiently blocked SARS-CoV-2 infection and intercellular transmission. Our platform offers unlimited, rapid and scalable access to disease-relevant lung tissue that recapitulate key hallmarks of human lung development and can be used to track SARS-CoV-2 infection and identify candidate therapeutics for COVID-19.