bluuebunny/crossref_metadata_2025 huggingface dataset:
A puzzling case of antibiotics leading to increased virulence in Burkholderia thailandensis
Authors: Thapa Sudarshan Singh , Grove Anne
Published: April 2020

Burkholderia thailandensis, a nonpathogenic strain, has been used as a model organism to study the mechanism of gene regulation and virulence in pathogenic strains (B. pseudomallei and B. mallei). B. pseudomallei and B. mallei are the causative agents of melioidosis and glanders respectively, fatal diseases, which occur in animals as well as humans. These bacteria in general are resistant to a large number of commonly used antibiotics, creating a hurdle in treatment of the diseases. One of the most commonly prescribed medicines for treatment of glanders and melioidosis is co‐trimoxazole, (a cocktail of trimethoprim and sulfamethoxazole). Surprisingly, exposure of Caenorhabditis elegans to B. thailandensis grown on a sub‐lethal dosage of trimethoprim led to an increase in death of C. elegans by ~ 40%. This observed increase in virulence is in part due to an increase in production of malleilactone, as trimethoprim leads to ~50‐fold increase in malA expression and ~10‐fold increase of malR (an activator of malA). We have previously shown that genes involved in production of various secondary metabolites including malleilactone are under direct/indirect regulation of the global regulator MftR (Major facilitator transport Regulator), but MftR is not a direct target of trimethoprim. ChiP data show that malR is directly regulated by MftR, suggesting that some compound accumulated under the effect of trimethoprim could act as ligand for MftR altering its binding to malR. Our data indicate that under effect of trimethoprim, the purine salvage pathway is disturbed indicated by decrease in purine catabolism (urate level decreases by ~ 0.6‐fold) while xanthine (a ligand for MftR) level increases by ~2‐fold. Allopurinol, an Xdh inhibitor, leads to similar upregulation of malA and malR as trimethoprim, supporting the notion that xanthine accumulation is sufficient to cause upregulation of the virulence genes. Thus, our results indicate that xanthine accumulated under the effect of trimethoprim alters binding of MftR to malR leading to increased expression of malA making B. thailandensis more virulent. Further, ΔmftR strain showed ~ 46% increase in death of C. elegans compared to wild type supporting the theme of MftR‐mediated regulation of the mal gene cluster. Our data shows that sublethal dosage of trimethoprim has unintended consequence of increasing virulence in Burkholderia via interrupting purine salvage pathway.
Getting the message: expansion of a bacterial three‐hybrid assay to mRNA‐protein interactions
Authors: Berry Katherine E. , LeBlanc Hannah , Nguyen Linh
Published: April 2020

Non‐coding small RNAs (sRNAs) contribute to bacterial biofilm formation and virulence, but there is much we do not understand about their molecular mechanisms. Hfq is the best‐studied bacterial RNA chaperone protein and has become the paradigmatic example for how RNA‐binding proteins facilitate mRNA regulation by sRNAs. Here we present progress on establishing and broadening the capabilities of a recently developed bacterial three‐hybrid (B3H) assay for studying interactions between Hfq and its RNA targets. While Hfq interactions with both mRNAs and sRNAs are critical for its function, only sRNA‐Hfq interactions have been accessible in the B3H assay to date. In order to expand the assay to mRNA‐Hfq interactions, we first demonstrated the assay’s capacity to detect interactions between synthetic mRNA mimics and Escherichia coli Hfq. Furthermore, we have developed constructs that support detection of interactions between mRNA 5′UTRs and two E. coli RNA‐chaperone proteins: Hfq and ProQ. This development points to the potential of the B3H assay to detect a wider variety of RNA‐protein interactions than previously possible, and will allow us to dissect the molecular mechanisms of these interactions. Current efforts aim to understand the relationship between the translational status of an mRNA fragment and its ability to bind protein partners in the B3H assay. In addition, we are working to understand the relationship between the in vivo B3H data on RNA‐protein interactions and the binding energetics of these interactions, which are typically measured in vitro. The results of these efforts will provide a deeper understanding of the implications of B3H data for RNA‐protein binding and regulatory mechanisms. By expanding both the capabilities of this bacterial three‐hybrid assay and our understanding of the data it provides, we aim to broaden its utility in studying bacterial gene regulation and RNA‐protein interactions.

Support or Funding Information

This work was supported by National Institutes of Health [R15GM135878], Mount Holyoke College and the Henry Luce foundation.
KMT2D Haploinsufficiency in Kabuki Syndrome Impairs Differentiation of Neural Crest Cells
Authors: Lavery William J. , Kotliar Michael , Chang Ching F. , Brugmann Samantha A. , Barski Artem , Lindsley Andrew W.
Published: April 2020

Objective

Kabuki Syndrome (KS), caused by haploinsufficiency in lysine methyltransferase KMT2D, is a multi‐system disease characterized by distinct facial dysmorphisms, mild‐to‐moderate intellectual disability, renal and cardiac abnormalities, short stature and humoral immune deficiency. Many of the tissues in which these abnormalities manifest are derived from or developmentally modulated by the neural crest cell (NCC) lineage. This study’s objective was to determine which features of the KS phenotype can be directly linked to genetic and epigenetic dysregulation in KMT2D haploinsufficient NCCs.

Methods

Peripheral blood mononuclear cells from a human donor haploinsufficient for KMT2D (KS patient, KMT2D+/−) or wild type (WT) control were used to generate induced pluripotent stem (iPS) cells. WT and KS NCCs were generated from WT and KS iPS cells, respectively. Whole transcriptome RNA sequencing (RNAseq) was performed on iPS cells (WT and KS) and NCCs (WT and KS) to characterize differential gene expression. Chromatin immunoprecipitation sequencing (ChIPseq) of histone 3 lysine 4 trimethylation (H3K4Me3) and histone 3 lysine 27 acetylation (H3K27Ac) was performed on NCCs (WT and KS) to characterize differential histone modifications at gene regulatory elements. Analysis of RNAseq and ChIPseq data was performed using BioWardrobe platform.

Results

Gene set enrichment analysis of transcriptome data indicates that, compared to WT NCCs, KMT2D haploinsufficient NCCs have ectopic expression of several genes critical to normal NCC organization, morphogenesis and tissue differentiation, including NCC transcriptional repressor Msh homeobox 2 (MSX2). Analysis also identified several genes with decreased expression in KMT2D+/− NCCs, including transcription factor AP‐2 alpha (TFAP2A), which serves as a master regulator of NCC lineage progression. ChIPseq data indicate differential enrichment of H3K4Me3 and H3K27Ac histone modifications at the TFAP2A locus in WT compared to KS NCCs, which suggests that TFAP2A may have a direct role in the impaired differentiation of NCCs in KS. Further studies to explore the roles of TFAP2A and other genes identified by these data are underway.

Conclusion

These results show that KMT2D haploinsufficiency impairs differentiation of the NCC lineage cells through epigenetic and transcriptomic dysregulation, thus supporting the classification of KS as a neurocristopathy.

Support or Funding Information

NIH T32 Training Grant 5T32AI060515‐14 (WJL), R35 DE027557 (SB), Center for Pediatric Genomics Cincinnati Research Foundation (AWL).
Withdrawn: A Controllable Protein Translation Strategy for Exploring Cellular Signal Transduction Based on Reading through Premature Termination Codons
Authors: Cheng Xiyao , Huang Yongqi , Su Zhengding
Published: April 2020

Withdrawal: Xiyao Cheng, Yongqi Huang, and Zhengding Su. A Controllable Protein Translation Strategy for Exploring Cellular Signal Transduction Based on Reading through Premature Termination Codons. The FASEB Journal. 34:s1. doi: 10.1096/fasebj.2020.34.s1.09195.

The above abstract, published online on April 19, 2020, in Wiley Online Library (wileyonlinelibrary.com), has been withdrawn by agreement between the authors, FASEB, and Wiley Periodicals, Inc. The abstract has been withdrawn at the request of the authors.
Characterizing the role of an α1,6‐mannanase needed for Neurospora crassa cell wall biogenesis
Authors: Patel Pavan , Free Stephen
Published: April 2020

Cell wall components such as glucans, chitins and mannans are needed for fungal virulence. The dfg‐5 gene encodes a GH‐76 α‐ 1,6 mannanase that is needed to cleave the α‐1,6‐mannose backbone of fungal N‐linked oligosaccharide‐associated galactomannans. The processed galactomannan is then used as substrate by GH‐72 lichenin transferases, that cross‐link it into the cell wall. However the mechanism of how DFG‐5 processes the galactomannan structure remains unclear. Here, we characterize the DFG‐5 protein. A pair of adjacent aspartic acid (D) or glutamic acid (E) residues are normally found at the active sites of GH‐76 family glycosylhydrolases. We used site directed mutagenesis to mutate 7 conserved aspartate and glutamate residues in DFG‐5 protein. Complementation analysis demonstrates that D76, D116, D117, and E130 mutations renders DFG‐5 inactive. 3D Modeling of DFG5 protein strongly suggests that D116 and D117 define the active site based on homology with other characterized glycosylhydrolases. Modeling of DFG‐5 mannanase shows it has a second groove containing D76 and E130. To further characterize DFG‐5, we generated a HIS6 tagged version of the protein and demonstrated that it complemented a dfg‐5 gene deletion. A number of glycoproteins co‐purify with the HIS6‐tagged DFG‐5 and these co‐purifying glycoproteins require galactomannan for DFG‐5 binding. Three known cell wall glycoproteins co‐purify with DFG‐5 mannanse.

Support or Funding Information

Funding for the study is provided by the UB Foundation.
When the Efferent Becomes Afferent ‐ the Cellular Response of Sympathetic Neurons to Sensory Stimuli
Authors: Davis Harvey , Musa Ayesha , Li Dan , Herring Neil , Paterson David
Published: April 2020

Responding to the so called “inflammatory soup” has often been regarded as the sole domain of sensory afferent neurons. We tested the hypothesis that efferent sympathetic neurons can also directly respond to traditionally afferent stimuli. We used single cell RNA‐sequencing (scRNAseq) to explore transcript expression of membrane G protein and ion channel coupled receptors in the sympathetic neuronal population within the stellate ganglia of 5–6‐week‐old Wistar rats. Agonist responses were then evaluated using Fura‐2 AM calcium imaging and perforated patch clamp recordings in isolated cultured neurons.

ScRNAseq identified transcript expression of transient receptor potential (TRP) channels (TRPV2>TRM7>TRPC1>>TRM4). Certain TRP channel activators (10 mM camphor n=18, 10 mM cannabidiol n=16, and 100 mM 2‐APB n=15) caused large baseline calcium influxes, which were blocked by the TRP channel inhibitor ruthenium red (RuR, 10 mM; Camphor n=7; Cannabidiol n=24; 2‐APB n=11). An accompanying depolarization of the resting membrane potential and spontaneous firing was also observed, which was reduced by RuR. Other TRP channel activators (100 mM menthol n=15, 1 mM capsaicin n=16, 500 mM AITC n=9, 1 mM vanillin n=15, 300 nM JT010 n=15, 100 mM cavracol n=13, 10 mM icillin n=10, 50 mM naltriben n=15, pH=6.5 n=16 and 100 ng/ml IGF‐1 n=12) failed to produce significant responses.

ScRNAseq also identified transcript expression of receptors for inflammatory mediators, for example histamine (HRH3) and prostaglandin E2 (PTGER2 and PTGER3). Stimulation of these receptors (100 mM histamine n=11 and 1 mM prostaglandin E2 n=8) had no effect on baseline [Ca2+]I, but were able to reduce Ca2+ influx in response to KCl depolarization (50 mM) or nicotine (100 mM).

Overall, these data are important when interpreting stellate ganglia sympathetic neurons as purely efferent, with evidence that neuronal excitability can be directly influenced by sensory signaling pathways.

Support or Funding Information

This project was supported by the Wellcome trust and the British Heart Foundation
Comparing the Accuracy of Physiologic and Triangular Pseudoflow Waveforms for Estimating Wave Reflection Indices in Adults
Authors: Shenouda Ninette , Stock Joseph M. , Patik Jordan C. , Chirinos Julio A. , Edwards David G.
Published: April 2020

Central aortic pressure waveforms contain valuable prognostic information in addition to central systolic pressure. Using wave separation analysis, aortic pressure‐flow relations can be used to decompose aortic pressure waveforms into forward (Pf) and backward‐travelling (Pb) components. Pf and Pb are used to calculate reflection magnitude (RM=Pb/Pf) which is predictive of heart failure and all‐cause mortality. Aortic blood flow can be measured non‐invasively via ultrasonography or estimated using a triangular pseudoflow waveform; however, the latter can underestimate true flow and thus overestimate Pb and RM. Recently, a physiologic pseudoflow waveform has been developed that improves on the assumptions of triangulation and so may provide a more accurate estimation of wave reflection.

PURPOSE

The purpose of this study was to compare estimates of Pf, Pb, and RM derived from physiologic and triangular pseudoflow waveforms relative to measured flow. We hypothesized that the physiologic waveform would provide a more accurate estimate of wave reflection indices compared to the triangular waveform.

METHODS

In 75 adults (age 18–75 yrs), radial artery pressure waveforms were acquired with applanation tonometry and a generalized transfer function was used to synthesize central pressure waveforms. Aortic blood flow velocities were measured via echocardiography and estimated using physiologic (in‐house software) and triangular (SphygmoCor CvMS) pseudoflow waveforms. Pressure‐flow relations were established offline and wave separation analysis was performed. Accuracy of the physiologic and triangular‐derived estimates of Pf, Pb, and RM were assessed using 95% equivalence testing, paired t‐tests comparing the mean absolute biases, and Bland‐Altman plots.

RESULTS

Using a 10% equivalence zone as the maximum acceptable difference, the physiologic and triangular pseudoflow waveforms were both comparable to the measured flow waveform in deriving Pf. However, only the physiologic waveform yielded similar estimates of Pb and RM compared to the measured waveform. Further, the physiologic waveform yielded lower mean absolute biases compared to the triangular waveform for Pf (1.7 ± 2.1 vs. 2.8 ± 2.6 mmHg, p<0.001), Pb (1.0 ± 1.1 vs. 3.3 ± 2.7 mmHg, p<0.001), and RM (3 ± 3 vs. 14 ± 13%, p<0.001). Lastly, Bland‐Altman biases were smaller, and 95% limits of agreement were narrower, for all wave reflection indices derived from the physiologic waveform than from the triangular waveform.

CONCLUSION

These data suggest that the physiologic pseudoflow waveform provides a more accurate estimate of wave reflection indices compared to the triangular waveform, and thus may offer a superior alternative when true flow cannot be measured.
Use of Conditioned Media from Electrically‐Stimulated C2C12 Myotubes to Examine the Effect of Muscle Contractile Secretions on Breast Cancer Cell Proliferation
Authors: Kump David S. , Hughes Alvysha S. , Justice Brittany , Murphy Cangela
Published: April 2020

Background & Purpose

Evidence suggests that secretions following muscle contraction may have benefits regarding breast cancer. This study aimed to examine whether conditioned media from electrically‐stimulated C2C12 myotubes would inhibit proliferation of breast cancer cells in vitro.

Methods

Media collected from electrically‐stimulated C2C12 myotubes was mixed 1:1 with fresh media and applied to either T47D subluminal type breast cancer cells or a K14TRT triple‐negative breast cancer cell line for 24 hours. Breast cancer cell proliferation was measured using the MTT method. In addition, media from electrically‐stimulated and non‐stimulated C2C12 myotubes were subjected to mass spectrometry for comparison.

Results

Results provided an inconsistent picture regarding the approach. When applied at a 1:1 ratio, the conditioned media unexpectedly accelerated the proliferation of T47D cells by more than two‐fold. When applied to K14TRT cells, one experiment demonstrated no effect, while a second showed a robust inhibition of proliferation. The disparate results are likely due to the wide natural variance in proliferation in this cell line. Mass spectrometry analysis did not yield any potential differences in secreted molecules between electrically‐stimulated and non‐stimulated media.

Conclusions

The present results suggest that further work is needed to explore the utility of media from electrically‐stimulated myotubes as a model for muscle contraction‐induced secretions and their effect on breast cancer. Future work will include a dose‐response analysis and the use of ELISA to identify potential molecules.

Support or Funding Information

This study was partially supported by a WSSU Grant from the Office of Science Initiatives. Mass spectrometry was conducted by the Wake Forest Baptist Comprehensive Cancer Center Proteomics and Metabolomics Shared Resource, which is supported by the National Cancer Institute’s Cancer Center Support Grant award number P30CA012197
Transgenerational effects following chronic circadian disturbance given to mother rats in F1 generation rats
Authors: Choi Jeonghyun , Hong Yonggeun
Published: April 2020

It has not been documented whether circadian disturbance of mother may influence fetal neurodevelopment. Therefore, this study aims to find the root cause of congenital neuronal defects, which is explosively increasing today, in one of the environmental factors; circadian disturbance. Female rats were randomly divided into 3 groups; L/D group with 12/12 hr light and dark cycle, LL group with 24 hr constant light, DD group with 24 hr constant dark. All subjected females were assigned to each light condition one week prior to pregnancy. Then, they were mated with males, and F1 offsprings were obtained at three weeks after the mating. The future generations were exposed exactly same light conditions. The animals in each group were randomly divided into control and melatonin‐treated subgroups. All animals were engaged in physiological, histological, and neurobehavioral analyses. LL and DD groups showed several abnormal features and decreased cecum/colon length (p<.05). Also, LL and DD groups showed lower activation of intracranial δ‐wave than L/D group showed. However, in LL and DD groups treated with melatonin improved intestinal integrity and activated intracranial δ‐wave. Axonal sprouting was evaluated by Golgi‐cox staining for identifying the abnormal neurodevelopment in all groups. The LL and DD groups showed reduction of axonal sprouting than L/D group. However, axonal branching in neuron increased in LL and DD groups treated with melatonin. In addition, neurobehavioral tests including open field test, novel objective recognition test, social interaction test, spontaneous alternation Y‐maze test, and elevated plus maze test were performed. LL and DD groups showed abnormal neurobehaviors (p<.05), whereas, melatonin treatment resulted in the normalized neurobehaviors (p<.05). In conclusion, F1 generation born from mothers exposed to chronic circadian disturbance showed abnormal neurodevelopment and neurobehaviors. Melatonin was considered as a therapeutic agent for congenital neuronal deficit.

Support or Funding Information

Fundings: National Research Foundation (NRF‐2017R1A2A2A01067169, NRF‐2019R1A6A3A01091422), Korea
Elevated serum melatonin under constant darkness enhances neural repair in spinal cord injury through regulation of circadian clock proteins expression
Authors: Hong Yunkyung , Jin Yunho , Park Kanghui , Hong Yonggeun
Published: April 2020

We investigated the effects of environmental lighting conditions regulating endogenous melatonin production on neural repair, following experimental spinal cord injury (SCI). Rats were divided into three groups randomly: the SCI + L/D (12/12‐h light/dark), SCI + LL (24‐h constant light), and SCI + DD (24‐h constant dark) groups. Controlled light/dark cycle was pre‐applied 2 weeks before induction of spinal cord injury. There was a significant increase in motor recovery as well as body weight from postoperative day (POD) 7 under constant darkness. However, spontaneous elevation of endogenous melatonin in cerebrospinal fluid was seen at POD 3 in all of the SCI rats, which was enhanced in SCI + DD group. Augmented melatonin concentration under constant dark condition resulted in facilitation of neuronal differentiation as well as inhibition of primary cell death. In the rostrocaudal region, elevated endogenous melatonin concentration promoted neural remodeling in acute phase including oligodendrogenesis, excitatory synaptic formation, and axonal outgrowth. The changes were mediated via NAS‐TrkB‐AKT/ERK signal transduction co‐regulated by the circadian clock mechanism, leading to rapid motor recovery. In contrast, exposure to constant light exacerbated the inflammatory responses and neuroglial loss. These results suggest that light/dark control in the acute phase might be a considerable environmental factor for a favorable prognosis after SCI.

Support or Funding Information

National Research Foundation (NRF‐2013R1A2A2A01067169, NRF‐2015R1C1A2A01055691, NRF‐2017R1D1A1B03029565), Korea.
The implication of melatonin and Botulinum toxin type A in neural reconstruction and muscle repair after spinal cord injury
Authors: Jin Yunho , Hong Yonggeun
Published: April 2020

Spinal cord is responsible for neural transmission of nerve impulses between brain and the whole body. Spinal cord injury (SCI) is caused mostly by trauma leading to motor and sensory deficits, and autonomic dysfunctions. This injury involves primary and secondary mechanisms; primary SCI is the mechanical damage to the spinal cord while secondary SCI means further prolonged insults initiated by the original damage. The therapeutic aims for SCI usually focus on the suppression of secondary SCI by means of therapeutic drugs of surgery. In this study, melatonin and Botulinum toxin type A (BTA) was exploited to accelerate the recovery of SCI. First, contribution of melatonin, which is known to have neuroprotective roles, toward the recovery from SCI through regulation of autophagy and endoplasmic reticulum (ER) stress was examined. Following SCI, muscle spasticity occurs interrupting normal functional movements. BTA was administrated to alleviate muscle spasticity for the restoration of normal movements. Melatonin was proved to protect cells from excessive autophagic cell death, and to activate ER stress response leading to the restoration of cellular homeostasis. Furthermore, dendritic branches were preserved by melatonin. Muscle degradation was suppressed following BTA injection. Furthermore, BTA injection accelerated the recovery of muscle contraction‐regulatory proteins and removal of damaged cells, and restored presynaptic homeostasis via autophagy activation. In conclusion, melatonin may result in the recovery of SCI via the regulation of autophagy and ER stress response. And, BTA was proved to draw functional recovery by alleviating muscle spasticity after SCI. In other words, melatonin or BTA could be suggested as therapeutic candidates for the recovery of SCI.

Support or Funding Information

National Research Foundation (NRF‐2017R1A2A2A01067169), Korea.
Role of Jumonji‐C Histone Demethylase in the Development of Hepatocellular Carcinoma
Authors: Jeong Do-Won , Chun Yang-Sook
Published: April 2020

Reprogramming in lipid metabolism has received increasing recognition as a hallmark of cancer cells. In cancer cells, large demands of lipids were required to meet excessive synthesis of membranes, energy production and activation of intracellular signaling pathways during cell proliferation. To meet these demands, cancer cells express increased levels of lipogenic enzymes responsible for FAs synthesis, which is a target of crucial transcriptional regulator, sterol regulatory element binding protein 1c (SREBP1c). SREBP1c is a well‐known master transcription factor for de novo lipogenesis and is found frequently increased in many human cancers. However, the molecular mechanisms involved in the regulation of SREBP1c remain incompletely understood. In this study, we uncover the role of jumonji‐C histone demethylase (JHDM) as a repressor of SREBP1c, thereby preventing lipogenesis and proliferation of hepatocellular carcinoma (HCC) cells. JHDM directly interacts with SREBP1c at basic helix‐loop‐helix domain of SREBP1c and plant homeodomain of JHDM, promoting proteasomal degradation of SREBP1c protein. In addition, knockdown of JHDM stimulates mRNA levels of lipogenic enzymes and induces lipogenesis in HCC cells. However, JHDM‐loss‐induced lipogenesis is recovered by SREBP1c knockdown, implying that SREBP1c is crucial for the effects of JHDM toward lipogenesis. In contrast, ectopic expression of JHDM shows protective effects on oleic acid‐induced lipid accumulation in HCC cells. We also found that JHDM knockdown using si‐RNAs promotes mRNA expressions of cell cycle regulators, accompanied with enhanced cell growth, colony formation and spheroid formation. However, the effects of JHDM knockdown which promote HCC proliferation are reversed by knockdown of SREBP1c. In accordance with these data, JHDM overexpression represses cell growth in HCC cells, indicating that JHDM acts as a tumor suppressor by inhibiting SREBP1c‐mediated lipogenesis and cell cycle regulation. Overall, our findings suggest that JHDM serves as a molecular bridge between lipid metabolism and cancer development through regulation of SREBP1c and thus JHDM could be a therapeutic target in defense against HCC.

Support or Funding Information

This work was supported by grants from the National Research Foundation of Korea (2016R1A2B4013377, 2018R1A2B6007241, 2019R1A2C2083886).
Spatiotemporal Organization of Metabolic Networks in Living Cells
Authors: Kennedy Erin Laurel , Jeon Miji , Chauhan Krishna , Augustine Farhan , An Songon , Kyoung Minjoung
Published: April 2020

Cellular metabolic networks are dynamic. Organization between membrane‐bound organelles and cytoplasmic metabolic pathways are crucial to cell survival and function. This is the case for the glucose metabolism, where cytoplasmic enzymes have been observed to form spatially organized enzyme assemblies. We have quantified the sub‐cellular organization of these glucose metabolic assemblies with the mitochondria and studied a mitochondrial‐associated population of assemblies that are functionally linked to the mitochondrial metabolism. This research was completed with a home‐built lattice light sheet microscope and other biophysical and biochemical methods, which provided crucial spatiotemporal insight into the orchestration of metabolic functions that provides a starting point to map 4D functional metabolic networks.

Support or Funding Information

NIH Research Project Grant R01 GM134086, NIH Research Project Grant R01 GM125981, Research Grant R03 CA219609, Student support provided by NIH Chemistry Biology Interface Program (T32 GM066706), Student support provided by an NIGMS Initiative for Maximizing Student Development Grant (2 R25‐GM55036)
Metformin attenuates lead‐induced inflammatory and apoptotic lung injury through modulation of P53 and TNF‐α pathways in rats
Authors: Attafi Ibraheem M. , Al-Bakheet Saleh A. , Maayah Zaid H. , Ansari Mushtaq A. , Ahmad Sheikh E. , AL-Alallah Ibrahim A. , Alanazi Fawaz E. , Korashy Hesham M.
Published: April 2020

Exposure to heavy metals, such as Lead (Pb) is a matter of growing concern and potential risk factor for developing inflammatory lung diseases. The aim of this study was to a) investigate the protective effect of metformin against Pb‐induced lung injury in rat and b) investigate the underlying mechanisms specifically the role of oxidative stress, inflammation and apoptosis. For this purpose, twenty‐four adult male Wistar albino rats weighing about 230 gm were randomly distributed into six study groups of four rats each. The first group received normal saline (0.5 ml/kg/day) for 6 consecutive days and served as a a Control. The second group received metformin (500 mg/kg) for 6 consecutive days. The third and fourth groups received a single dose of normal saline (0.5 ml/kg/day) for 3 consecutive days followed by lead nitrate (Pb) (25 and 50 mg/kg, i.p.) for 3 consecutive days, respectively. The fifth and sixth groups received metformin (500 mg/kg) for 3 consecutive days followed by Pb 25 and 50 mg/kg, i.p. plus metformin 500 mg/kg for additional 3 consecutive days, respectively. One day after the last dose, animals were a and blood samples were collected via cardiac puncture for determination the levels of Pb. Rats lung tissues were excised and processed for histopathological examination, determination the levels of mRNA and protein expression of target genes, Analysis of apoptosis and TNF‐α production were determined by flow cytometer. Our results showed that exposure to Pb caused induction of inflammatory cells infiltration, cell apoptosis, modulation of genes involving in inflammation and oxidation, induction of p53 and TNF‐α were demonstrated in Pb‐induced lung injury. Administration of metformin 500 mg/kg for 6 days resulted in a protective effect on the Pb‐induced inflammatory lung injury through induction of the expression levels of NQO1, SOD, HO‐1, and p53 genes. In addition, metformin treatment significantly reduced the production of TNF‐α and expression of IL‐6 while induced the expression of IL‐10 in Pb‐treated groups. In conclusion, these observations suggest that metformin processes anti‐inflammatory, antiapoptotic and the induction of antioxidants properties against Pb‐induced lung injuries.
The cytosolic formate dehydrogenase FdsABG from Cupriavidus necator. Crystal structure of the FdsBG fragment which is responsible for the oxidative half‐reaction.
Authors: Blaha Gregor , Young Tynan , Niks Dimitri , Hakopian Sheron , Tam Timothy K. , Yu Xuejun , Hille Russ
Published: April 2020

Soluble formate dehydrogenase FdsABG, is a member of the NADH dehydrogenase superfamily, which is one of the most broadly distributed family of enzymes. Unlike most members of this superfamily, formate dehydrogenase FdsABG is a cytosolic enzyme. Physiologically, the FdsABG from Cupriavidus necator oxidizes formate to CO2 by reducing NAD+ to NADH. In the process, reducing equivalents are transferred from formate to the molybdenum center of FdsA, from where they are passed through a chain of iron‐sulfur clusters to the FMN of FdsB and on to NAD+. These two steps of accepting reducing equivalents from the formate and passing them on to NAD+ are considered the reductive and oxidative half‐reactions of the formate dehydrogenase reaction. Because FdsB‐ and FdsG‐like domains are found in all NADH dehydrogenases and can perform the oxidative half reaction independent of FdsA, we embarked on a crystallographic study of FdsBG of C. necator formate dehydrogenase, with the aim of putting the oxidative half reaction of the enzyme in a structural context.

Support or Funding Information

United States Department of Energy Grant DE‐SC0010666 (to R. H.)
Autophagy and Heat Shock Protein 70 Expression During Acute Heat Stress in Isosmotic and Hyperosmotic Conditions in Peripheral Blood Mononuclear Cells from Young Adults: Preliminary Data
Authors: Cote Melissa D. , McCormick James J. , Rutherford Maura M. , King Kelli E. , Meade Robert D. , Notley Sean R. , Akerman Ashley P. , Schmidt Madison D. , Kenny Glen P.
Published: April 2020

Heat‐stress induced dehydration is associated with elevations in extracellular osmolality. To counteract the resultant osmotic stress, cells employ several stress response systems including the heat shock protein (HSP) and autophagy pathways. HSPs refold damaged proteins, whereas autophagy acts to degrade damaged cellular components, thereby promoting cellular survival. Accumulating evidence indicates a coordinated response between the HSP and autophagy systems, whereby both systems are activated under conditions of heat stress. However, it remains unknown whether elevated extracellular osmolality augments the HSP and autophagic response to whole‐body heat stress in humans. Thus, the purpose of this study was to examine changes in HSP (HSP70) and autophagy resulting from elevated intravascular osmolality during acute heat exposure. We hypothesized that both autophagy and HSP70 protein expression would be greater under hyperosmotic stress compared to isosmotic conditions. On separate days (separated by ~1 week), six young men (mean [SD]; 27 [5] years) received a 90‐min intravenous infusion with either isosmotic (ISO; 0.9% NaCl; 0.200 mL·kg−1·min−1) or hyperosmotic (HYP; 3.0% NaCl; 0.125 mL·kg−1·min−1) saline, followed by passive whole‐body heating to ~0.8°C above baseline esopha‐‐‐geal temperature using a water perfused suit. Serum osmolality was determined via freeze‐point depression from blood samples at baseline and at the end of passive heating. Peripheral blood mononuclear harvested at these time‐points were analyzed via Western blotting to assess changes in HSP70 protein and proteins indicative of autophagic activity including microtubule associated protein 1 light chain 3 beta‐II (LC3‐II) and p62/sequesterome‐1 (p62). All values were normalized to β‐actin and reported as fold changes relative to baseline. Data are presented as means and standard deviations (SD) and were compared between‐conditions at the end of passive heating using paired (2‐tailed) t‐tests (α=0.05). At the end of heating, serum osmolality was higher during HYP when compared to ISO (298 [SD 3] vs. 291 [SD 3]; p<0.001). During passive heating, LC3‐II was increased (2.20 [SD 2.12] vs. 1.62 [SD 1.47]; p=0.04) and p62 protein was reduced (0.47 [SD 0.33] vs. 1.20 [SD 0.56]; p=0.01) in HYP compared to ISO, indicating greater autophagic activation during HYP. Despite this, HSP70 protein was lower under HYP conditions following heating than ISO (0.72 [SD 0.24] vs. 1.20 [SD 0.19]; p=0.04). Contrary to our hypothesis, these preliminary findings indicate greater reliance on the autophagic response rather than simultaneous activation of HSP70 during acute heat exposure with super‐imposed hyperosmotic stress.

Support or Funding Information

National Science and Engineering Research Council of Canada
The Influence of Salt Loading on the Kidney Injury Biomarker Neutrophil Gelatinase‐Associated Lipocalin in Healthy Young Adults
Authors: Barnett Alex M. , Babcock Matthew C. , Watso Joseph C. , Migdal Kamila U. , Farqhuar William B. , Robinson Austin T.
Published: April 2020

Purpose

The kidneys play a critical role in blood pressure (BP) regulation. In hypertensive patients and in rodent models, long‐term high dietary sodium (Na+) causes a reduction in kidney function (reduced glomerular filtration rate; GFR). Rodent studies demonstrate that high dietary Na+ increases levels of novel kidney injury biomarkers, independent of changes in BP and GFR. However, it is unclear if short‐term high Na+ increases kidney injury biomarkers in healthy humans. Therefore, we tested the hypothesis that short‐term high Na+ intake increases urinary excretion of the novel kidney injury marker Neutrophil Gelatinase‐Associated Lipocalin (NGAL) in healthy, young adults.

Methods

Twenty participants (12 M/8 F; age: 24±4 years; BMI: 23.1±0.6 kg/m2; BP: 112 ±10/64 ±9 mmHg, mean ± SD) participated in a double‐blind, placebo‐controlled, randomized, crossover study. For 10‐days, participants were asked to consume a 2,300 mg/day Na+ diet and supplement with salt (3,900 mg/day of Na+) or placebo (dextrose) capsules. The conditions were separated by at least two weeks and female participants were tested during the placebo phase of their oral contraceptive cycle. Participants collected their urine for the final 24 hours of each supplementation period to assess urine flow rate, and Na+ excretion to verify diet adherence. Brachial BP was measured after at least five minutes of rest. We measured urine and serum creatinine (kinetic modification via the Jaffe reaction) to estimate GFR via creatinine clearance. We used enzyme‐linked immunosorbent assays (Toronto Bioscience) to measure urinary NGAL. Statistical analysis included paired t‐tests and Wilcoxon matched‐pairs signed rank tests.

Results

Compared to placebo, Na+ supplementation increased urinary Na+ excretion (139.9±68.4 vs. 282.5±69.8 mmol/24 hours, p<0.01) but there was no difference in mean arterial BP (77±7 vs. 77±6 mmHg, p=0.64). Serum creatinine concentrations (0.78±0.12 vs. 0.77±0.09 mg/dL, p=0.55) were not different between conditions. However, compared to placebo, Na+ supplementation increased creatinine clearance (110.5±32.9 vs. 145.0±24.9 mL/min, p<0.01) and increased urinary NGAL excretion indexed to urine flow rate (16.9±21.9 vs. 28.4±36.1 ng/min, p=0.02).

Discussion

Our preliminary data suggests that salt loading does not alter serum creatinine concentration but increases creatinine clearance and urinary excretion of the kidney injury marker NGAL in healthy young adults.

Support or Funding Information

Funding

Supported by NIH grants R01HL128388 and K01HL147998, and ACSM Foundation Doctoral Student Research Grant 17‐00521
In vitro and in vivo characterisation of structure‐based nucleosome binding peptides
Authors: Teles Kaian Amorim , Fernandes Vinicius , Torres Isabel , Leite Manuela , Lobbia Vincenzo , Honorato Rodrigo , Lopes-de-Oliveira Paulo , Van Ingen Hugo , Grisolia Cesar , Treptow Werner , Santos Guilherme Martins
Published: April 2020

The modulation of chromatin is orchestrated by Nucleosome Binding Proteins (NBPs). The nucleosome surface has been hypothesized as a therapeutic target due to its capacity to modulate chromatin architecture. Herein, we evaluated if Nucleosome Binding Peptides (NBPeps) would be able to occupy the nucleosome surface directly, thereby modulating chromatin status and influencing phenotypic outcomes. To understand how the nucleosome structure is affected by NBPeps, we performed biochemical assays indicating that distinct NBPeps present differential actions on the nucleosome structure despite binding to similar target regions on the nucleosome. Cellbased assays and animal models demonstrated that the NBPeps penetrate the cell and have specific effects on cell physiology and phenotypic outcome, suggesting that NBPeps might have important therapeutic implications.

Support or Funding Information

This work has been supported by CNPq for KT,VF and IT fellowships, Fap‐DF grant number 04/2017 for GS, iNEXT grant number 6756, funded by the Horizon 2020 programme of the European Union, for HI.
Diosmetin and its glycoside, diosmin, improve atopic dermatitis in DNCB‐induced hairless mice.
Authors: Kim Su-Nam , Park Sang-A , Bong Sim-Kyu , Lee Jinwoo , Park No-June , Yang Min Hye , Kim Yong Kee
Published: April 2020

Naturally derived diosmetin and its glycoside diosmin are known to be effective in treating inflammatory disease. The aim of this study was to investigate the anti‐atopic dermatitis effect on diosmetin and diosmin 2,4‐dinitrochlorobenzen (DNCB)‐induced atopic dermatitis (AD) models. Oral administration of diometin or diosmin inhibited the development of DNCB‐induced AD‐like lesions in hairless mice by inhibiting transdermal moisture loss (TEWL) and improving skin hydration. Total serum immunoglobulin E (IgE) and interleukin‐4 (IL‐4) levels were also reduced in AD‐like lesion hairless mice skin samples treated with diosmetin or diosmin. In vitro, however, only diosmetin reduced the level of IL‐4 mRNA in RBL‐2H3 cells. Diosmin considered to be effective in the treatment of AD and skin inflammatory diseases by being converted into diosmetin in the body by pharmacokinetic metabolism. Thus oral administration of diosmetin and diosmin might be useful for the treatment of AD and cutaneous inflammatory diseases.

Support or Funding Information

This research was supported by the Bio & Medical Technology Development Program of the National Research Foundation (NRF)& funded by the Korean government (MSIT) (Nos. NRF‐2019M3A9I3080263, NRF‐2019M3A9I3080265, and NRF‐2019M3A9I3080266).
Mitochondria Are Involved in Ferroptosis Induced by Cardiac Ischemia‐Reperfusion in Rats
Authors: Chapa Xavier , Tyurina Yulia , Ayala-Arroyo Esteban , Rodriguez-Graciani Keishla Marie , Jang Sehwan , Bayir Hulya , Kagan Valerian , Javadov Sabzali
Published: April 2020

Cardiac ischemia/reperfusion injury (IRI) activates several cell death mechanisms, the priority of which depends on the severity and duration of ischemia. Recently, ferroptosis, an iron dependent mechanism of regulated cell death, was discovered to participate in cardiac IRI; however, mechanisms underlying ferroptotic signaling in the heart remain unknown. Mitochondria as the main source of ROS and ATP play a central role in cell death, especially in high energy consuming organs such as the heart. Here, we investigated the role of mitochondria in IRI‐induced ferroptosis in the rat heart. Isolated and Langendorff‐mode perfused rat hearts underwent 25‐min of global ischemia followed by 60‐min reperfusion in the presence and absence of the anti‐ferroptotic agents, ferrostatin‐1 (Fer‐1, 1 μM) and liproxstatin‐1 (Lip‐1, 0.2 μM). The compounds were present in the perfusion media 10 min prior to ischemia and throughout the entire period of reperfusion. Cardiac hemodynamic parameters including the left ventricular development pressure (LVDP), heart rate, and rates of contractility/relaxation were monitored continuously whereas LDH activity in the cardiac effluent was determined at reperfusion. Respiration rates, ROS production, and swelling were analyzed in mitochondria isolated from the heart at the end of reperfusion. Results showed that hearts treated with Fer‐1 and Lip‐1 had significantly higher post‐ischemic recovery of cardiac function than the control group (LVDP: 85% and 82%, respectively vs. 34%, p<0.05 for both). LDH release was reduced by ~50 % in the hearts treated with Fer‐1 and Lip‐1. A significant increase in the state 3 respiration rate and respiratory control index for mitochondrial ETC complex I was found in Fer‐1‐ or Lip‐1‐treated hearts. Likewise, Fer‐1 and Lip‐1 significantly reduced IRI‐induced basal and Ca2+‐induced mitochondrial ROS production. We observed no difference in Ca2+‐induced mitochondrial swelling between control and treated groups. IR‐induced ferroptotic phospholipid signaling was attenuated in the presence of Fer‐1 or Lip‐1. In conclusion, our study demonstrates that ferroptosis affect cardiac/mitochondrial function during IRI suggesting that mitochondria can be engaged in ferroptotic signaling in the heart.

Support or Funding Information

This study was supported by the NIH NIGMS (Grants SC1GM128210 and R25GM061838‐18).
Psychometric Properties of the Chinese Version of the Pieper-Zulkowski Pressure Ulcer Knowledge Test in Nursing Practice: A Cross-sectional Survey
Authors: Nie Wenbo , Tang Jing , Zulkowski Karen , Wang Lisheng , Zan Tao
Published: April 2020

OBJECTIVE

To translate the Pieper-Zulkowski Pressure Ulcer Knowledge Test into Chinese and analyze the internal consistency of the adapted questionnaire.

METHODS

The Pieper-Zulkowski Pressure Ulcer Knowledge Test was translated into Chinese, and the internal consistency and content validity of the translated test were assessed. Further, the authors conducted a cross-sectional survey using the test among 476 RNs in six hospitals in four cities of China.

MAIN RESULTS

The Cronbach α was .93 for all items and .83, .82, and .84 for the prevention, staging, and wound description subscales, respectively. The validity of content was acceptable (content validity index = 0.83-1.00). The average correct scores were as follows: total, 69.37%; prevention, 73.38%; staging, 69.61%; and wound description, 64.87%. Nurses with more advanced professional titles or wound care certifications scored significantly higher than other test takers. Participants who had conducted internet research about pressure injuries or read the pressure injury guidelines also had significantly higher scores than those who had not.

CONCLUSIONS

The translated instrument can effectively measure Chinese nurses’ knowledge about pressure injuries.
Who Wants To Be a Nurse? Understanding Emirati Female Students’ Knowledge and Attitudes About Nursing as a Career
Authors: Maymoun Nazneen , Sohail M. Sadiq
Published: April 2020

AIM

The aim of this study was to increase understanding of female high school students’ intentions to pursue a nursing career. The objective is to explore theories on attitudinal research and propose a research model that examines the relationship between attitude, knowledge, and impact on intention to pursue a nursing career.

BACKGROUND

There is a paucity of research on understanding the reasons for the shortage of local nurses in the United Arab Emirates.

METHOD

Data were collected from 157 female high school students. The research model was empirically tested using standard procedures.

RESULTS

Findings support some of the proposed direct and interactional effects. Attitude is a predictor of intention of high school students to pursue a nursing profession. The interaction effect of attitude and knowledge significantly predicts intention.

CONCLUSION

The study calls for further investigation on the effects of attitude and knowledge on students’ intention to pursue a nursing career and bring about changes in thinking.
Direction and Severity of Root Compression Affects the Clinical Outcome After Decompression?
Authors: Ko Sangbong , Lee Hohyoung , Chae Seungbum , Choi Wonkee , Lee Sungho
Published: April 2020

Study Design:

This was a retrospective cohort study.

Objective:

To evaluate the correlation between preoperative radiologic degree and the postoperative prognosis in lumbar foraminal stenosis (LFS) patients who required decompression.

Summary of Background Data:

There have been only a few studies for postoperative prognosis according to the degree of preoperative stenosis in patients requiring surgery.

Materials and Methods:

The shape and severity of LFS was classified according to Lee et al’s classification. Radiating leg pain was assessed preoperatively and at the last follow-up. Oswestry Disability Index (ODI), Roland Morris Disability Questionnaire (RMDQ), SF-36 Physical Component Score (PCS), and SF-36 Mental Component Score (MCS) were surveyed at the last follow-up. Mean follow-up was 6.3 years.

Results:

Among 130 patients, 34 cases were classified as Grade 1T (transverse), 31 cases as Grade 1V (vertical), 32 cases as Grade 2, and 33 cases as Grade 3. The mean age was 65.0 years. Radiating leg pain (Visual Analogue Scale) preoperatively and lastly was 6.8 and 2.3, respectively in Grade 1T; 7.4 and 3.3, respectively in Grade 1V; 7.7 and 3.0, respectively in Grade 2; and 7.4 and 2.9, respectively in Grade 3. ODI and RMDQ at the last follow-up were 13.9 and 6.3, respectively in Grade 1T; 23.1 and 11.2, respectively in Grade 1V; 18.8 and 12.3, respectively in Grade 2; 21.6 and 11.7, respectively in Grade 3. Grade 1T had significantly lower value than Grade 1V, 2 and 3 in ODI and RMDQ, respectively. SF-36 PCS and MCS at the last follow-up were 62.2 and 70.5, respectively in Grade 1T; 33.7 and 40.3, respectively in Grade 1V; 42.1 and 48.2, respectively in Grade 2; 37.0 and 48.5, respectively in Grade 3. Grade 1T had significantly higher value than grade 1V, 2 and 3 in SF-36 PCS and MCS, respectively.

Conclusions:

Decompression for LFS was effective for reducing leg radiating pain. Functional outcome and quality of life was better in mild transverse type implying the role of dorsal root ganglion and dynamic stenosis.