bluuebunny/crossref_metadata_2025 huggingface dataset:
Treatment Adherence Among Persons Receiving Concurrent Multidrug-Resistant Tuberculosis and HIV Treatment in KwaZulu-Natal, South Africa
Authors: Stephens Fay , Gandhi Neel R. , Brust James C.M. , Mlisana Koleka , Moodley Pravi , Allana Salim , Campbell Angie , Shah Sarita
Published: June 2019

Background:

Success in multidrug-resistant tuberculosis (MDR-TB) and HIV treatment requires high medication adherence despite high pill burdens, frequent adverse events, and long treatment duration, which may jeopardize adherence. We prospectively compared MDR-TB/HIV-coinfected persons to those with MDR-TB alone to determine the impact of concurrent treatment on adherence and outcomes.

Methods:

We assessed medication adherence monthly using 3-day recall, 30-day recall, and visual analog scale and examined adherence to monthly study visits (months 0–12). We determined the proportion of participants fully adherent (no reported missed doses) to MDR-TB vs. HIV treatment by each measure. We assessed the association of medication and clinic visit adherence with MDR-TB treatment success (cure or completion, 18–24 months) and HIV virologic suppression.

Results:

Among 200 patients with MDR-TB, 63% were women, median age was 33 years, 144 (72%) were HIV-infected, and 81% were receiving antiretroviral therapy (ART) at baseline. Adherence to medications (81%–98% fully adherent across all measures) and clinic visits (80% missed ≤1 visit) was high, irrespective of HIV status. Adherence to ART was significantly higher than to MDR-TB treatment by all self-reported measures (3-day recall: 92% vs. 84%, respectively; P = 0.003). In multivariable analysis, the adjusted risk ratio of unsuccessful MDR-TB treatment increased with every missed visit: 1.50, 2.25, and 3.37 for unsuccessful treatment, for 1, 2, and ≥3 missed visits.

Conclusions:

Adherence to ART was higher than to MDR-TB treatment among persons with MDR-TB/HIV coinfection. Missed clinic visits may be a simple measure for identifying patients at risk of unsuccessful MDR-TB treatment outcome.
Project Shikamana: Community Empowerment-Based Combination HIV Prevention Significantly Impacts HIV Incidence and Care Continuum Outcomes Among Female Sex Workers in Iringa, Tanzania
Authors: Kerrigan Deanna , Mbwambo Jessie , Likindikoki Samuel , Davis Wendy , Mantsios Andrea , Beckham S.Wilson , Leddy Anna , Shembilu Catherine , Mwampashi Ard , Aboud Said , Galai Noya
Published: June 2019

Objective:

To determine the impact of a community empowerment model of combination HIV prevention (Project Shikamana) among female sex workers (FSW) in Iringa, Tanzania.

Methods:

We conducted a 2-community randomized trial. Intervention elements included the following: (1) Community‐led drop‐in center and mobilization activities; (2) venue-based peer education, condom distribution, and HIV testing; (3) peer service navigation; (4) provider sensitivity trainings; and (5) SMS reminders. We used time-location sampling to enroll 496 FSW and conducted a survey and blood draws to screen for HIV and assess viral load at 0 and 18 months. We conducted an intent-to-treat analysis using logistic and Poisson regression and inverse probability weighting for primary outcomes.

Results:

The analysis included 171 HIV-positive and 216 HIV-negative FSW who completed baseline and 18-month study visits. Participants in the intervention were significantly less likely to become infected with HIV at 18-month follow-up (RR 0.38; P = 0.047), with an HIV incidence of 5.0% in the intervention vs. 10.4% control. Decreases in inconsistent condom use over time were significantly greater in the intervention (72.0%–43.6%) vs. control (68.8%–54.0%; RR 0.81, P = 0.042). At follow-up, we observed significant differences in behavioral HIV care continuum outcomes, and positive, but nonsignificant, increases in viral suppression (40.0%–50.6%) in the intervention vs. control (35.9%–47.4%). There was a strong association of between higher intervention exposure and HIV outcomes including viral suppression.

Conclusions:

Project Shikamana is the first trial of community empowerment-based combination prevention among FSW in Africa to show a significant reduction in HIV incidence warranting its broader implementation and evaluation.
Can Ex Vivo Magnetic Resonance Imaging of Rectal Cancer Specimens Improve the Mesorectal Lymph Node Yield for Pathological Examination?
Authors: Stijns Rutger , Philips Bart , Wauters Carla , de Wilt Johannes , Nagtegaal Iris , Scheenen Tom
Published: June 2019

Purpose

The aim of this study was to use 7 T ex vivo magnetic resonance imaging (MRI) scans to determine the size of lymph nodes (LNs) in total mesorectal excision (TME) specimens and to increase the pathological yield of LNs with MR-guided pathology.

Materials and Methods

Twenty-two fixated TME specimens containing adenocarcinoma were scanned on a 7 T preclinical MRI system with a T1-weighted 3-dimensional gradient echo sequence with frequency-selective lipid excitation (repetition time/echo time, 15/3 milliseconds; resolution, 0.293 mm3) and a water-excited 3-dimensional multigradient echo (repetition time, 30 milliseconds; computed echo time, 6.2 milliseconds; resolution, 0.293 mm3) pulse sequence.

The first series of 11 TME specimens (S1) revealed the number and size of LNs on both ex vivo MRI and histopathology. The second series of 11 TME specimens (S2) was used to perform MR-guided pathology. The number, size, and percentages of yielded LNs of S1 and S2 were compared.

Results

In all specimens (22/22), a median number of 34 LNs (interquartile range, 26–34) was revealed on ex vivo MRI compared with 14 LNs (interquartile range, 7.5–21.5) on histopathology (P= 0.003). Mean size of all LNs did not differ between the 2 series (ex vivo MRI: 2.4 vs 2.5 mm,P= 0.267; pathology: 3.6 vs 3.5 mm,P= 0.653). The median percentages of harvested LNs compared with nodes visible on ex vivo MRI per specimen for both series were not significantly different (40% vs 43%,P= 0.718). By using a size threshold of greater than 2 mm, the percentage improved to 71% (S1) and to 78% (S2,P= 0.895). The median number of harvested LNs per specimen did not increase by performing MR-guided pathology (S1, 14 LNs; S2, 20 LNs;P= 0.532).

Conclusions

Ex vivo MRI visualizes more LNs than (MR-guided) pathology is able to harvest. Current pathological examination was not further improved by MR guidance. The majority of LNs or LN-like structures visible on ex vivo MRI below 2 mm in size remain unexplained, which warrants a 3-dimensional approach for pathological reconstruction of specimens.
Molecular screening of the human parvoviruses B19 and bocavirus 1 in the study of congenital diseases as applied to symptomatic pregnant women and children
Authors: Salbetti Maria Belen , Pedranti Mauro Sebastian , Barbero Paula , Molisani Paula , Lazzari Martina , Olivera Nicolas , Isa Maria Beatriz , Bertoldi Ariel , Moreno Laura , Adamo Maria Pilar
Published: June 2019

Introduction. B19 virus (B19V) and bocavirus 1 (HBoV1) are human pathogenic parvoviruses that are prevalent worldwide and are responsible for a diverse and not yet fully established spectrum of clinical manifestations.

Objective. To screen B19V and HBoV1 in patients with clinical manifestations associated with acquisition of the infection during gestation.

Methods. A retrospective, observational study was performed that included serum samples from patients without a previous known aetiology. B19V and HBoV1 were determined by end-point PCR. Positive samples were genotyped.

Results. A total of 106 serum samples were analysed, 61 from pregnant women and 45 from neonates and paediatric patients. None were positive for HBoV1, while B19V was detected in 37/106 [34.9 %, 95 % confidence interval (CI): 26.5–44.4] of the samples studied. In the group of pregnant women, 28/61 (45.9 %, 95 % CI: 34.0–58.3) were B19V-positive, and 2 of them had foetal anaemia followed by hydrops and foetal death, 3 were associated with a history of recurrent pregnancy loss and there was 1 case of spontaneous abortion. B19V was also detected in cases of maternal febrile exanthema, polyhydramnios, oligohydramnios and foetal ascites. In the group of children, 9/45 (20.0 %, 95 % CI: 10.9–33.8) neonatal patients were B19V-positive, and this was associated with foetal hydrops, TORCH syndrome and cardiac alterations. The nucleotide sequences analysed confirmed the identity of B19V genotype 1.

Conclusions. We found no evidence to indicate the presence of HBoV1 in maternal blood or in the newborns/paediatric patients (hence providing no support for the supposed vertical transmission). On the other hand, the high frequency of B19V in the pathologies studied indicates the importance of molecular diagnosis in both the mother and the child. Future efforts should contribute to early detection and characterization of infections.
Mechanistic and compositional studies of the autophagy-inducing areca nut ingredient
Authors: Chiu Chang-Ta , Liu Shyun-Yeu , Yen Ching-Yu , Tsai Meng-Ting , Chang Huei-Cih , Liu Young-Chau , Lin Mei-Huei
Published: June 2019

Areca nut (AN) is a popular chewing carcinogen worldwide causing a variety of diseases such as oral and esophageal carcinomas. We previously found that the partially purified 30-100 kDa fraction of AN extract (ANE 30-100K) induces autophagy in oral carcinoma OECM-1 cells and some other different types of cells. Since autophagy is known to play important roles in tumor establishment and development, the underlying mechanisms of ANE 30-100K-induced autophagy (AIA) is worthy of further investigation. In this study, we further demonstrated that the cytotoxic concentration of ANE 30-100K induces some typical autophagy hallmarks in esophageal carcinoma (CE81T/VGH) cells in an Atg5-dependent manner. Furthermore, the endocytosis inhibitor (methyl-β-cyclodextrin) and two caveolin shRNAs, as well as two proteasome inhibitors (lactacystin and epoxomicin), were shown to attenuate ANE 30-100K-induced cytotoxicity and LC3-II accumulation significantly in OECM-1 and CE81T/VGH cells. Finally, we also analyzed the carbohydrate compositions of ANE 30-100K by phenol-sulfuric acid method and high performance anion exchange chromatography with pulse amperic detector. The results showed that ANE 30-100K contains about 67% carbohydrate and is composed of fucose (5.938%), arabinose (24.631%), glucosamine (8.066%), galactose (26.820%), glucose (21.388%), and mannose (13.157%). Collectively, these results suggest that caveolin-mediated endocytosis and proteasome are required for AIA and the major components of ANE 30-100K are carbohydrates. This study may have provided new knowledges of the action mechanisms and compositions of ANE 30-100K.
Kin structure and roost fidelity in greater noctule bats
Authors: Santos João D. , Meyer Christoph F.J. , Ibáñez Carlos , Popa-Lisseanu Ana G. , Juste Javier
Published: June 2019

Roost fidelity is an important aspect of mammalian biology. Studying the mechanisms underlying philopatry can help us understand a species’ energetic requirements, ecological constraints and social organisation. Temperate bat species notably exhibit a comparatively high degree of female philopatry, resulting in maternity colonies segregated at the mitochondrial level. We focus on the greater noctule,Nyctalus lasiopterus, to study this behaviour in more depth. We make use of microsatellite data for 11 markers across 84 individuals residing in Maria Luisa Park in Seville, Spain. At the time of sampling, this urban park boasted the highest number of registered bats of this species, among which three social groups were observed to segregate spatially. We studied the distribution of pairs of individuals across filial relationship categories and relatedness estimates relative to the social group of each individual. This analysis was complemented by information on roost-use frequency among a subset of genotyped bats. We found no significant relationship between roost use and genetic distance, but there was evidence of higher group-sharing with increased levels of relatedness. Mother-daughter pairs shared the same group more often than expected, as did pairs of individuals of relatedness above 0.36. We discuss the implications of these results in terms of the behavioural ecology of temperate bats and for conservation efforts aimed at preserving them.
Surpassing the physical Nyquist limit to produce super-resolution cryo-EM reconstructions
Authors: Feathers J. Ryan , Spoth Katherine A. , Fromme J. Christopher
Published: June 2019

The resolution of cryo-EM reconstructions is fundamentally limited by the Nyquist frequency, which is half the sampling frequency of the detector and depends upon the magnification used. In principle, super-resolution imaging should enable reconstructions to surpass the physical Nyquist limit by increasing sampling frequency, yet there are no reports of reconstructions that do so. Here we report the use of super-resolution imaging with the K3 direct electron detector to produce super-resolution single-particle cryo-EM reconstructions significantly surpassing the physical Nyquist limit. We also present a comparative analysis of a sample imaged at four different magnifications. This analysis demonstrates that lower magnifications can be beneficial, despite the loss of higher resolution signal, due to the increased particle numbers imaged. To highlight the potential utility of lower magnification data collection, we produced a 3.5 Å reconstruction of jack bean urease with particles from a single micrograph.

Abstract Figure
Tangential flow microfluidics for the capture and release of nanoparticles and extracellular vesicles on conventional and ultrathin membranes
Authors: Dehghani Mehdi , Lucas Kilean , Flax Jonathan , McGrath James , Gaborski Thomas
Published: June 2019

Membranes have been used extensively for the purification and separation of biological species. A persistent challenge is the purification of species from concentrated feed solutions such as extracellular vesicles (EVs) from biological fluids. We investigated a new method to isolate micro- and nano-scale species termed tangential flow for analyte capture (TFAC), which is an extension of traditional tangential flow filtration (TFF). Initially, EV purification from plasma on ultrathin nanomembranes was compared between both normal flow filtration (NFF) and TFF. NFF resulted in rapid formation of a protein cake which completely obscured any captured EVs and also prevented further transport across the membrane. On the other hand, TFF showed capture of CD63 positive EVs with minimal contamination. We explored the use of TFF to capture target species over membrane pores, wash and then release in a physical process that does not rely upon affinity or chemical interactions. This process of TFAC was studied with model particles on both ultrathin nanomembranes and conventional thickness membranes (polycarbonate track-etch). Successful capture and release of model particles was observed using both membranes. Ultrathin nanomembranes showed higher efficiency of capture and release with significantly lower pressures indicating that ultrathin nanomembranes are well-suited for TFAC of delicate nanoscale particles such as EVs.
Identifying Novel Roles for Peptidergic Signaling in Mice
Authors: Powers Kathryn G. , Ma Xin-Ming , Eipper Betty A. , Mains Richard E.
Published: June 2019

Despite accumulating evidence demonstrating the essential roles played by neuropeptides, it has proven challenging to use this information to develop therapeutic strategies. Peptidergic signaling can involve juxtacrine, paracrine, endocrine and neuronal signaling, making it difficult to define physiologically important pathways. One of the final steps in the biosynthesis of many neuropeptides requires a single enzyme, peptidylglycine α-amidating monooxygenase (PAM), and lack of amidation renders most of these peptides biologically inert. PAM, an ancient integral membrane enzyme that traverses the biosynthetic and endocytic pathways, also affects cytoskeletal organization and gene expression. While mice, zebrafish and flies lackingPam(PamKO/KO) are not viable, we reasoned that cell-type specific elimination ofPamexpression would generate mice that could be screened for physiologically important and tissue-specific deficits.PamcKO/cKOmice, with loxP sites flanking the 2 exons deleted in the globalPamKO/KOmouse, were indistinguishable from wildtype mice. EliminatingPamexpression in excitatory forebrain neurons reduced anxiety-like behavior, increased locomotor responsiveness to cocaine and improved thermoregulation in the cold. A number of amidated peptides play essential roles in each of these behaviors. Although atrial natriuretic peptide (ANP) is not amidated,Pamexpression in the atrium exceeds levels in any other tissue. EliminatingPamexpression in cardiomyocytes increased anxiety-like behavior and improved thermoregulation. Atrial and serum levels of ANP fell sharplyPamMyh6-cKO/cKOin mice and RNASeq analysis identified changes in gene expression in pathways related to cardiac function. Use of this screening platform should facilitate the development of new therapeutic approaches targeted to peptidergic pathways.

SIGNIFICANCE

Peptidergic signaling, which plays key roles in the many pathways that control thermoregulation, salt and water balance, metabolism, anxiety, pain perception and sexual reproduction, is essential for the maintenance of homeostasis. Despite the fact that peptides generally signal through G protein coupled receptors, it has proven difficult to use knowledge about peptide synthesis, storage and secretion to develop effective therapeutics. Our goal was to develop anin vivobioassay system that would reveal physiologically meaningful deficits associated with disturbed peptidergic signaling. We did so by developing a system in which an enzyme essential for the production of many bioactive peptides could be eliminated in a tissue-specific manner.
Phosphorylation of multiple proteins involved in ciliogenesis by Tau Tubulin kinase 2
Authors: Bernatik Ondrej , Pejskova Petra , Vyslouzil David , Hanakova Katerina , Zdrahal Zbynek , Cajanek Lukas
Published: June 2019

Primary cilia (PC) are organelles necessary for proper implementation of developmental and homeostasis processes. To initiate their assembly, coordinated actions of multiple proteins are needed. Tau tubulin kinase 2 (TTBK2) is a key player in the cilium assembly pathway, controlling final step of cilia initiation. The function of TTBK2 in ciliogenesisis is critically dependent on its kinase activity, however, precise mechanism of TTBK2 action is so far incompletely understood, due to very limited information about its relevant substrates. In this study we identify CEP83, CEP89, CCDC92, Rabin8 and DVL3 as substrates of TTBK2 kinase activity. Further, we characterise a set of phosphosites of the newly identified substrates and CEP164, induced by TTBK2in vitroandin vivo. Intriguingly, we further show that identified TTBK2 phosphosites and consensus sequence delineated from those are distinct from motifs previously assigned to TTBK2. Finally, we address functional relevance of selected phosphorylations of CEP164 and provide evidence that the examined TTBK2-induced phosphorylations of CEP164 are relevant for the process of cilia formation. In summary, our work provides important insight into substrates-TTBK2 kinase relationship and suggests that phosphorylation of substrates on multiple sites by TTBK2 is probably involved in the control of ciliogenesis in human cells.
Mapping Co-regulation Pathways among Ligand Binding sites in RyR1
Authors: Chirasani V. R , Popov K. I , Meissner G. , Dokholyan N. V
Published: June 2019

Ryanodine receptor 1 (RyR1) is an intracellular calcium (Ca2+) release channel required for skeletal muscle contraction. RyR1 is co-regulated by multiple activators – Ca2+, ATP and caffeine (CFF), yet the mechanism of co-regulation and the action synergy of these activators is unknown. Here, we report the detailed network of allosteric connections between the three ligand sites and the pore region in (i) Ca2+bound – closed, (ii) ATP/CFF bound – closed, (iii) Ca2+/ATP/CFF bound – closed and (iv) Ca2+/ATP/CFF bound – open RyR1 states. We find that two dominant paths mediate the communication between the Ca2+binding site and pore region in Ca2+-only state. ATP/CFF-only occupied – closed RyR1 has two additional paths with CFF-but not ATP-occupied path using part of the closed Ca2+-only pathway. In the presence of Ca2+, ATP and CFF, major differences between the open and closed states are identified with both using part of the paths of the closed Ca2+-only and ATP/CFF-only states. We find that the three activators Ca2+, ATP, and CFF propagate their effects to the pore region through a network of partially overlapping pathways. Such coordination of allosteric pathway underlies the molecular basis of synergy of channel regulation by multiple activators.

Statement of Significance

RyRs are a group of Ca2+channels that bind to several endogenous modulators and regulate Ca2+release through closed-to-open gating transition. Despite the high-resolution structural data available for RyR1, the allosteric mechanism of RyR1 gating remains elusive. In this study, we employed graph-theoretical approach to demonstrate the allosteric network of synergistic interaction among various activators in RyR1. To our knowledge, for the first time we were able to identify the co-regulation among ligand sites in RyR1 to regulate the closed-to-open gating transition. The explored allosteric coupling in RyR1 may assist in designing advanced therapeutics for several debilitating diseases. Our findings in this study will assist to design new strategies for controlled allosteric regulation of RyR1 functionality in future.
Tuberculosis treatment outcome: The case of women in Ethiopia and China, Ten-Years Retrospective Cohort study
Authors: Mirutse Gebremeskel , Fang Mingwang , kahsay Alemayehu Bayray , Ma Xiao
Published: June 2019

Every year tuberculosis kills above half million women all over the world. Nonetheless, the difference in the size of deaths among countries was not compared. Hence, this study is aimed to compare the death size of two countries. Socio demographic and clinical data of women treated for all form of tuberculosis in the past ten years 2007-2016 were collected from total of eight hospitals and six treatment centers of Tigray and Zigong respectively. Then, collected data were entered into SPSS version 21 then we estimated the magnitude of TB, level of treatment success and assessed factors associated with the unsuccessful TB outcome. In the past ten years, a total of 5603(41.5%) and 4527 (24.5%) tuberculosis cases were observed in Tigray and Zigong respectively. Of those with treatment outcome record a total of 2602(92%) in Tigray and 3916(96.7%) in Zigong were successfully treated. Total of 170 (6%) cases in Tigray and 36(0.8%) cases in Zigong were dead. In Tigray cases like retreatment (aOR, 0.29; 95% CI: 0.16-0.53) and multi drug resistant (aOR, 0.31; 95% CI: 0.003, 0.27) were less likely to show treatment success. But, HIV co-infected TB cases (aOR, 3.58; 95% CI: 2.47, 5.18) were more likely to show treatment success. In Zigong, women with MDR TB (Adjusted OR, 0.90; 95%CI: 0.24, 0.34) were less likely to show treatment success. On the other hand women in the age category of 15-49 (adjusted OR, 1.55; 95% CI: 1.08, 2.206) showed treatment success. Big number of tuberculosis cases and death were observed in Tigray comparing with Zigong. Hence, a relevant measure should be considered to improve treatment outcome of women in Tigray.
Large-scale, dynamin-like motions of the human guanylate binding protein 1 revealed by multi-resolution simulations
Authors: Barz Bogdan , Loschwitz Jennifer , Strodel Birgit
Published: June 2019

Guanylate binding proteins (GBPs) belong to the dynamin-related superfamily and exhibit various functions in the fight against infections. The functions of the human guanylate binding protein 1 (hGBP1) are tightly coupled to GTP hydrolysis and dimerization. Despite known crystal structures of the hGBP1 monomer and GTPase domain dimer, little is known about the dynamics of hGBP1. To gain a mechanistic understanding of hGBP1, we performed sub-millisecond multi-resolution molecular dynamics simulations of both the hGBP1 monomer and dimer. We found that hGBP1 is a highly flexible protein that undergoes a hinge motion similar to the movements observed for other dynamin-like proteins. Another large-scale motion was observed for the C-terminal helixα13, providing a molecular view for theα13–α13 distances previously reported for the hGBP1 dimer. Most of the loops of the GTPase domain were found to be flexible, disclosing why GTP binding is needed for hGBP1 dimerization to occur.

Author summary

Gunaylate binding proteins are key fighters against microbial and viral pathogens. In the human body there are seven types of such proteins, among which is the guanylate binding protein 1 (hGBP1). This protein is able to perform its function only once it is activated by binding and converting guanosinetriphosphat (GTP) to guanosinediphosphat and guanosinemonophosphat via hydrolysis. In concert with the conversion of GTP the dimerization of hGBP1 occurs, which can further interact with the lipid membrane of the pathogen and disrupt it. While the crystal structure of the protein is known, the activation and dimerization steps are not well understood at molecular level as studying them experimentally is difficult. An alternative approach is given by molecular simulations, allowing us to elucidate the protein dynamics closely connected to these steps. From our simulations applied to both the hGBP1 monomer and dimer we identified large-scale motions taking place in hGBP1 that had not been reported before. We discuss the relevance of these motions in terms of their biological function, such as possible membrane damage caused by one of the motions or locking the protein in the dimer state.
Probing the Subcellular Distribution of Phosphatidylinositol Reveals a Surprising Lack at the Plasma Membrane
Authors: Zewe James P. , Miller April , Sangappa Sahana , Wills Rachel C. , Goulden Brady D. , Hammond Gerald R. V.
Published: June 2019

The polyphosphoinositides (PPIn) are central regulatory lipids that direct membrane function in eukaryotic cells. Understanding how their synthesis is regulated is crucial to revealing these lipids’ role in health and disease. PPIn are derived from the major structural lipid, phosphatidylinositol (PI). However, although the distribution of most PPIn have been characterized, the subcellular localization of PI available for PPIn synthesis is not known. Here, we have used several orthogonal approaches to map the subcellular distribution of PI, including localizing exogenous fluorescent PI, as well as detecting lipid conversion products of endogenous PI after acute chemogenetic activation of PI-specific phospholipase and 4-kinase. We report that PI is broadly distributed throughout intracellular membrane compartments. However, there is a surprising lack of PI in the plasma membrane compared to the PPIn. These experiments implicate regulation of PI supply to the plasma membrane, as opposed to regulation of PPIn-kinases, as crucial to the control of PPIn synthesis and function at the PM.

Zewe et al develop approaches to map the subcellular distribution of the major phospholipid, phosphatidylinositol (PI), revealing that the lipid is present in most membranes except for plasma membrane, where it is mainly found as PI4P and PI(4,5)P2.
Depletion of Ric-8B leads to reduced mTORC2 activity
Authors: Nagai Maíra H. , Gutiyama Luciana M. , Xavier Victor P. S. , Machado Cleiton F. , Reis Alice H. , Donnard Elisa R. , Galante Pedro A. F. , Abreu Jose G. , Festuccia William T. , Malnic Bettina
Published: June 2019

mTOR, a serine/threonine protein kinase that is involved in a series of critical cellular processes, can be found in two functionally distinct complexes, mTORC1 and mTORC2. In contrast to mTORC1, little is known about the mechanisms that regulate mTORC2. Here we show that mTORC2 activity is reduced in mice with a hypomorphic mutation of the Ric-8B gene. Ric-8B is a highly conserved protein that acts as a non-canonical guanine nucleotide exchange factor (GEF) for heterotrimeric Gαs/olf type subunits. We found that Ric-8B hypomorph embryos are smaller than their wild type littermates, fail to close the neural tube in the cephalic region and die during mid-embryogenesis. Comparative transcriptome analysis revealed that signaling pathways involving GPCRs and G proteins are dysregulated in the Ric-8B mutant embryos. Interestingly, this analysis also revealed an unexpected impairment of the mTOR signaling pathway.

Phosphorylation of Akt at Ser 473 is downregulated in the Ric-8B mutant embryos, indicating a decreased activity of mTORC2. In contrast, phosphorylation of S6, a downstream target of mTORC1, is unaltered. Knockdown of the endogenous Ric-8B gene in HEK293T cells leads to reduced phosphorylation levels of Akt at Ser 473, but not of S6, further supporting the selective involvement of Ric-8B in mTORC2 activity. Our results reveal a crucial role for Ric-8B in development and provide novel insights into the signals that regulate mTORC2 activity.

Author Summary

Gene inactivation in mice can be used to identify genes that are involved in important biological processes and that may contribute to disease. By using this approach, we found that the Ric-8B gene is essential for embryogenesis and for the normal development of the nervous system. Ric-8B mutant mouse embryos are smaller than their wild type littermates and show neural tube defects at the cranial region. This approach also allowed us to identify the biological pathways that are involved in the observed phenotypes, the G protein and mTORC2 signaling pathways. mTORC2 plays particular important roles also in the adult brain, and has been implicated in neurological disorders. Ric-8B is highly conserved in mammals, including humans. Our mutant mice provide a model to study the complex molecular and cellular processes underlying the interplay between Ric-8B and mTORC2 in neuronal function.
Trickle infection and immunity toTrichuris muris
Authors: Glover Maya , Colombo Stefano A.P. , Thornton David J. , Grencis Richard K.
Published: June 2019

The majority of experiments investigating the immune response to gastrointestinal helminth infection use a single bolus infection. However,in situindividuals are repeatedly infected with low doses. Therefore, to model natural infection, mice were repeatedly infected (trickle infection) with low doses ofTrichuris muris. Trickle infection resulted in the slow acquisition of immunity reflected by a gradual increase in worm burden followed by a partial expulsion. Flow cytometry revealed that the CD4+ T cell response shifted from Th1 dominated to Th2 dominated, which coincided with an increase in Type 2 cytokines. The development of resistance following trickle infection was associated with increased worm expulsion effector mechanisms including goblet cell hyperplasia, Muc5ac production and increased epithelial cell turn over. Depletion of CD4+ T cells reversed resistance confirming their importance in protective immunity following trickle infection. In contrast, depletion of group 2 innate lymphoid cells did not alter protective immunity.T. muristrickle infection resulted in a dysbiotic mircrobiota which began to recover alpha diversity following the development of resistance.

These data support trickle infection as a robust and informative model for analysis of immunity to chronic intestinal helminth infection more akin to that observed under natural infection conditions and confirms the importance of CD4+ T cell adaptive immunity in host protection.

Author Summary

Infection with parasitic worms (helminths) is a considerable cause of morbidity in humans. Understanding how we respond to infection is crucial to developing novel therapies. Laboratory models of helminth infection have been a valuable tool in understanding fundamental immune responses to infection. However, typically an individual mouse will be infected with a large, single-dose of the parasite. This is in contrast to the natural scenario in which individuals will receive frequent low level exposures. What is unknown is how repeated infection alters the development of immunity to infection. We have developed a laboratory model to tackle this question. We infected mice with the model helminthTrichuris murison a weekly basis and assessed a range of responses in comparison with a more traditional infection system. We found striking differences in the dynamics of the infection, the host immune response, and in changes to host gut microbial populations. Our study shows how resistance to helminth infection can develop over time in response to repeat infection, and provides a model system that better reflects human immunity to this parasite.
Defining the Subcellular Distribution and Metabolic Channeling of Phosphatidylinositol
Authors: Pemberton Joshua G. , Kim Yeun Ju , Sengupta Nivedita , Eisenreichova Andrea , Toth Daniel J. , Boura Evzen , Balla Tamas
Published: June 2019

Phosphatidylinositol (PtdIns) is an essential structural component of eukaryotic membranes that also serves as the common precursor for polyphosphoinositide (PPIn) lipids. Despite the recognized importance of PPIn species for signal transduction and membrane homeostasis, there is still a limited understanding of how the dynamic regulation of PtdIns synthesis and transport contributes to the turnover of PPIn pools. To address these shortcomings, we capitalized on the substrate selectivity of a bacterial enzyme, PtdIns-specific PLC, to establish a molecular toolbox for investigations of PtdIns distribution and availability within intact cells. In addition to its presence within the ER, our results reveal low steady-state levels of PtdIns within the plasma membrane (PM) and endosomes as well as a relative enrichment of PtdIns within the cytosolic leaflets of the Golgi complex, peroxisomes, and outer mitochondrial membranes. Kinetic studies also demonstrate the requirement for sustained PtdIns supply from the ER for the maintenance of monophosphorylated PPIn species within the PM, Golgi complex, and endosomal compartments.

Pemberton et al. characterize a molecular toolbox for the visualization and manipulation of phosphatidylinositol (PtdIns) within intact cells. Results using these approaches define the steady-state distribution of PtdIns across subcellular membrane compartments as well as provide new insights into the relationship between PtdIns availability and polyphosphoinositide turnover.
Abbreviated assessment of psychopathology in patients with suspected seizure disorders
Authors: Malpas Charles B , Wang Albert D , Leong Michelle , Johnstone Benjamin , Rayner Genevieve , Kalincik Tomas , Kwan Patrick , O’Brien Terence J , Velakoulis Dennis
Published: June 2019

Purpose

Psychopathology is common in patients undergoing investigation for seizure-related disorders. Psychometric examination using self-report instruments, such as the SCL-90-R, can assist diagnosis. The SCL-90-R, however, is a lengthy instrument and might not be tolerated by all patients. We assessed several abbreviated forms of the SCL-90-R in patients undergoing video encephalographic monitoring (VEM).

Method

687 patients completed the SCL-90-R and scores were computed for the full SCL-90-R and five abbreviated forms. Correlations and mean differences were computed between different forms. Classification accuracy was assessed via receiver operating characteristic (ROC) curves, and measurements models were examined using confirmatory factor analysis (CFA).

Results

All abbreviated forms were strongly correlated with the SCL-90-R for general psychopathology (r= .93 – .99), depression (r= .89 – .95), anxiety (r= .97 – .98), psychosis (r= .95 – .99), and obsessive-compulsive symptoms (r= .97). Classification performance was similar across forms for depression and anxiety, with high negative predictive values (.90 – .94) and lower positive predictive values (.34 – .38). Classification performance for psychotic and obsessive-compulsive disorders was poor. Differences were observed between the full SCL-90-R and its abbreviated forms across most domains (d= 0.00 – 0.65). The published measurement model was most strongly validated for the SCL-27, SCL-14, and the SCL-K-9.

Conclusions

These five SCL-90-R abbreviated forms show high convergent validity with the full version. In patients undergoing investigation for seizure-related disorders, the BSI or BSI-18 are most appropriate where screening for both depression and anxiety is required. The SCL-K-9 is appropriate when only a single measure of global psychological distress is required. None of the instruments were able to detect psychotic or obsessive-compulsive symptoms with great accuracy. Caution should be exercised when making direct comparisons across the different forms.
Binding of single-mutant epidermal growth factor (EGF) ligands alter the stability of the EGF receptor dimer and promote growth signaling
Authors: Pascarelli Stefano , Merzhakupova Dalmira , Uechi Gen-Ichiro , Laurino Paola
Published: June 2019

The Epidermal Growth Factor Receptor (EGFR) is a membrane-anchored tyrosine kinase that is able to selectively respond to multiple extra-cellular stimuli. Previous studies have indicated that the modularity of this system is affected by ligand-induced differences in the stability of the dimerized receptor in a process known as “Biased signaling”. However, this hypothesis has not been explored using single-mutant ligands thus far. Herein, we developed a new approach to identify residues responsible for functional divergence combining the conservation and co-evolution information of ortholog and paralog genes encoding the epidermal growth factor (EGF) ligand. Then, we mutated these residues and assessed the mutants’ effects on the receptor by employing a combination of molecular dynamics (MD) and biochemical techniques. Although the EGF mutants had comparable binding affinities to the wild type ligand for EGFR, the EGF mutants induced a different phosphorylation and cell growth pattern in multiple cell lines. The MD simulations of the EGF mutants show a long-range effect on the receptor dimer interface. For the first time in this study, a single mutation in EGF is shown to be enough to alter the activation of the pathway at the cellular level. These results also support the theory of biased signaling in the tyrosine kinase receptor system and demonstrate a promising new way to study ligand-receptor interactions.
Mutations of the YidC Insertase alleviate stress from σM-dependent membrane protein overproduction in Bacillus subtilis
Authors: Zhao Heng , Sachla Ankita J. , Helmann John D.
Published: June 2019

In Bacillus subtilis, the extracytoplasmic function σ factor σM regulates cell wall synthesis and is critical for intrinsic resistance to cell wall targeting antibiotics. The anti-σ factors YhdL and YhdK form a complex that restricts the basal activity of σM, and the absence of YhdL leads to runaway expression of the σM regulon and cell death. Here, we report that this lethality can be suppressed by gain-of-function mutations in spoIIIJ, which encodes the major YidC membrane protein insertase in B. subtilis. B. subtilis PY79 SpoIIIJ contains a single amino acid substitution in the substrate-binding channel (Q140K), and this allele suppresses the lethality of high SigM. Analysis of a library of YidC variants reveals that increased charge (+2 or +3) in the substrate-binding channel can compensate for high expression of the σM regulon. Derepression of the σM regulon induces secretion stress, oxidative stress and DNA damage responses, all of which can be alleviated by the YidCQ140K substitution. We further show that the fitness defect caused by high σM activity is exacerbated in the absence of SecDF protein translocase or σM-dependent induction of the Spx oxidative stress regulon. Conversely, cell growth is improved by mutation of specific σM-dependent promoters controlling operons encoding integral membrane proteins. Collectively, these results reveal how the σM regulon has evolved to up-regulate membrane-localized complexes involved in cell wall synthesis, and to simultaneously counter the resulting stresses imposed by regulon induction.

Author Summary

Bacteria frequently produce antibiotics that inhibit the growth of competitors, and many naturally occurring antibiotics target cell wall synthesis. In Bacillus subtilis, the alternative σ factor σM is induced by cell wall antibiotics, and upregulates genes for peptidoglycan and cell envelope synthesis. However, dysregulation of the σM regulon, resulting from loss of the YhdL anti-σM protein, is lethal. We here identify charge variants of the SpoIIIJ(YidC) membrane protein insertase that suppress the lethal effects of high σM activity. Further analyses reveal that induction of the σM regulon leads to high level expression of membrane proteins that trigger envelope stress, and this stress is countered by specific genes in the σM regulon.
Wavefront sensing at X-ray free-electron lasers
Authors: Seaberg Matthew , Cojocaru Ruxandra , Berujon Sebastien , Ziegler Eric , Jaggi Andreas , Krempasky Juraj , Seiboth Frank , Aquila Andrew , Liu Yanwei , Sakdinawat Anne , Lee Hae Ja , Flechsig Uwe , Patthey Luc , Koch Frieder , Seniutinas Gediminas , David Christian , Zhu Diling , Mikeš Ladislav , Makita Mikako , Koyama Takahisa , Mancuso Adrian P. , Chapman Henry N. , Vagovič Patrik
Published: June 2019

Here a direct comparison is made between various X-ray wavefront sensing methods with application to optics alignment and focus characterization at X-ray free-electron lasers (XFELs). Focus optimization at XFEL beamlines presents unique challenges due to high peak powers as well as beam pointing instability, meaning that techniques capable of single-shot measurement and that probe the wavefront at an out-of-focus location are desirable. The techniques chosen for the comparison include single-phase-grating Talbot interferometry (shearing interferometry), dual-grating Talbot interferometry (moiré deflectometry) and speckle tracking. All three methods were implemented during a single beam time at the Linac Coherent Light Source, at the X-ray Pump Probe beamline, in order to make a direct comparison. Each method was used to characterize the wavefront resulting from a stack of beryllium compound refractive lenses followed by a corrective phase plate. In addition, difference wavefront measurements with and without the phase plate agreed with its design to within λ/20, which enabled a direct quantitative comparison between methods. Finally, a path toward automated alignment at XFEL beamlines using a wavefront sensor to close the loop is presented.
Polarization control with an X-ray phase retarder for high-time-resolution pump–probe experiments at SACLA
Authors: Kubota Y. , Suzuki M. , Katayama T. , Yamamoto K. , Tono K. , Inubushi Y. , Seki T. , Takanashi K. , Wadati H. , Yabashi M.
Published: June 2019

Control of the polarization of an X-ray free-electron laser (XFEL) has been performed using an X-ray phase retarder (XPR) in combination with an arrival timing diagnostic on BL3 of the SPring-8 Angstrom Compact free-electron LAser (SACLA). To combine with the timing diagnostic, a pink beam was incident on the XPR crystal and then monochromated in the vicinity of samples. A high degree of circular polarization of ∼97% was obtained experimentally at 11.567 keV, which agreed with calculations based on the dynamical theory of X-ray diffraction. This system enables pump–probe experiments to be operated using circular polarization with a time resolution of 40 fs to investigate ultrafast magnetic phenomena.
Flow separation control of NACA-2412 airfoil with bio-inspired nose
Authors: Raj Mohamed Mohamed Arif , Guven Ugur , Yadav Rajesh
Published: June 2019

Purpose

The purpose of this paper is to achieve an optimum flow separation control over the airfoil using passive flow control method by introducing bio-inspired nose near the leading edge of the NACA 2412 airfoil.

Design/methodology/approach

Two distinguished methods have been implemented on the leading edge of the airfoil: forward facing step, which induces multiple accelerations at low angle of attack, and cavity/backward facing step, which creates recirculating region (axial vortices) at high angle of attack.

Findings

The porpoise airfoil (optimum bio-inspired nose airfoil) delays the flow separation and improves the aerodynamic efficiency by increasing the lift and decreasing the parasitic drag. The maximum increase in aerodynamic efficiency is 22.4 per cent, with an average increase of 8.6 per cent at all angles of attack.

Research limitations/implications

The computational analysis has been done for NACA 2412 airfoil at low subsonic speed.

Practical implications

This design improves the aerodynamic performance and increases structural strength of the aircraft wing compared to other conventional high-lift devices and flow-control devices.

Originality/value

Different bio-inspired nose designs which are inspired by the cetacean species have been analysed for NACA 2412 airfoil, and optimum nose design (porpoise airfoil) has been found.
When in Rome, do as the Romans do
Authors: Memon Mumtaz Ali , Mirza Muhammad Zeeshan , Lim Bibiana , Umrani Waheed Ali , Hassan Mohamed Ayyub , Cham Tat Huei , Shahzad Khurram
Published: June 2019

Purpose

International students are considered as a huge and potentially lucrative segment for various products of the tourism industry in Malaysia. One of the tourism products is the local food they consume. The purpose of this paper is to investigate key factors that contribute to international students’ intention to consume Malay food. Drawing on the theory of planned behavior (TPB), the present study examines the impact of attitude, subject norm and perceived behavioral control (PBC) on international students’ intention to consume Malay food.

Design/methodology/approach

A quantitative approach using self-administered questionnaire was adopted. A total of 163 samples were collected from international students studying in five different Malaysian higher education institutions. A partial least squares structural equation modeling technique was used to test the hypothesized model.

Findings

The findings highlight that attitude, subject norms and PBC significantly motivate international students to consume local food. The subject norm toward consuming local food was revealed as the strongest predictor of international students’ intention to consume Malay food.

Originality/value

Although this is one of the early studies that uses the TPB in assessing the factors that influence international students’ intention to consume local food, this paper is driven by and aimed for practical consideration and thus making a significant contribution to practice. Specifically, the findings of the present study provide important implications for tourism industry practitioners to plan and implement initiatives that may promote local food among international students studying in Malaysian institutions.