bluuebunny/crossref_metadata_2025_split huggingface dataset:
Search for New KCa3.1‐Targeting Small Molecules and Monoclonal Antibodies
Authors: Shim Hesung , Nguyen Hai , Cui Yanjun , Colussi Paul , Bednenko Janna , Wulff Heike 
Published: April 2020

The intermediate‐conductance Ca2+‐activated K+ channel KCa3.1 (also known as SK4 or the Gárdos channel) is encoded by the KCNN4 gene and plays an important role in the activation of T‐ and B‐cells, mast cells, macrophages and microglia by regulating membrane potential, cellular volume and calcium signaling. KCa3.1 is further involved in the proliferation of dedifferentiated vascular smooth muscle cells and fibroblast. Accordingly, KCa3.1 inhibitors are therapeutically relevant as immunosuppressants and for the treatment of a wide range of fibroproliferative disorders. Using the recently solved cryo‐EM structures of the full length KCa3.1 channel in the closed and two open states we virtually screened a small library of clinically used drugs with two software packages (Glide and AutoDock Vina). The top 50 scoring compounds for the triarylmethane binding site in the inner pore varied significantly between the two programs and the three states. We are currently in the process of functionally confirming the more interesting hits using a thallium flux assay and whole‐cell patch‐clamp.In parallel we are also searching for KCa3.1 blocking monoclonal antibodies, since antibodies constitute attractive alternatives to small molecules because of their exquisite specificity and long half‐lives. Human KCa3.1 was recombinantly expressed in Tetrahymena thermophila, a unicellular eukaryote that has shown a remarkable ability to produce correctly folded human ion channels at high yield. The recombinant KCa3.1 was purified and adhered to magnetic beads that were subsequently used to pan two human phage display libraries. Library screening identified 34 unique anti‐KCa3.1 antibodies, out of which two were found to functionally block KCa3.1 current in whole‐cell patch‐clamp experiments. The selected clone, TTG‐252, inhibits human KCa3.1 current with an IC50 of 1.8 nM while showing no effect on human KCa1.1, KCa2.2, Kv11.1 (hERG) or Nav1.5. The antibody does not cross‐react to mouse or rat KCa3.1.Support or Funding InformationFunded by R21NS101876.
The effect of heat stress on motor preparation processing
Authors: Shibasaki Manabu , Ishikura Keiko , Sawai Norie , Hirai Minami , Kobayashi Fumino , Nakata Hiroki 
Published: April 2020

An excessively increased internal temperature is known to reduce exercise/sports performance. To clarify the effect of heat stress on motor preparation processing using electroencephalography (EEG), subjects performed two types of voluntary isometric exercise before (normothermia) and during heat stress on different days. Fifteen volunteers performed contingent negative variation (CNV) trials in Experiment 1, and fourteen volunteers performed movement‐related cortical potential (MRCP) trials in Experiment 2, while eleven subjects performed both trials. Whole body heating via a water‐perfused suit increased the esophageal temperature by 1.52 ± 0.14 °C in Experiment 1 and 1.40 ± 0.10 °C in Experiment 2. The mean amplitudes of the middle‐ and late‐CNV at Cz and Pz during heat stress were significantly reduced relative to those in normothermic trials (middle CNV, −3.3 ± 2.8 to −0.5 ± 3.2 μV at Cz, −3.8 ± 2.8 to −1.6 ± 3.0 μV at Pz; late CNV, −5.9 ± 3.5 to −3.1 ± 3.1 μV at Cz, −6.6 ± 2.9 to −3.7 ± 2.8 μV at Pz, P< 0.05 for each). However, in Experiment 2, there were no significant differences between normothermia and heat stress regarding the mean amplitudes of MRCPs including the Bereitschaftspotential (BP) and negative slope (NS′). These results suggest that mild hyperthermia influences cognitive processing being reflected by CNV with stimulus‐triggered movement, rather than motor‐related processing reflected by MRCPs related to self‐initiated movement.Support or Funding InformationJapan Society for the Promotion of Science 18H03166
Elucidating the Effects of Distal Charges on the Reduction Potential of the Rieske Protein
Authors: Lopez Rachel , Hunsicker-Wang Laura 
Published: April 2020

The Rieske protein plays a key role in both cellular respiration and photosynthesis, transferring electrons through the respective cytochrome bc1 and b6f complexes of each process. It is characterized by a [2Fe‐2S] cluster and is unique in that one iron is ligated by nitrogens from two solvent‐exposed histidine residues while the other is ligated by the sulfurs of two cysteines. In the electron transport chain (ETC), the Rieske protein oxidizes ubiquinol and reduces cytochrome c, transferring electrons between complexes II and IV.Previous work has demonstrated how hydrogen bonds, solvent accessibility of the ligands, and the presence of nearby charged residues can impact the reduction potential of the protein. However, the full extent of this third element, how far from the cluster these charged residues can reside and still impact the reduction potential, has yet to be explored. Two types of mutants have been created to probe the effects of these distal charges. First, mutations removing positively charged residues were predicted to decrease reduction potential. Likewise, mutations removing negatively charged residues were expected to increase reduction potential. Each mutation either resides a unique distance from the cluster or produces a different change in overall charge, demonstrating the impact of each factor both individually and additively. Lastly, the pKa values of the histidine ligands were also expected to be inversely related to reduction potential.Using pH‐dependent UV‐Visible spectrophotometry, the pKa values of these mutants were determined and correlated to these relationships. While the overall trend of pKa values increased as expected for the negative‐charge mutants, confounding elements also appeared, such as a third pKa sigmoid. Furthermore, E146R, a positive‐charge mutant, also demonstrated increased pKa values, opposite to the hypothesized trend. New mutants that implement a neutral to positive substitutions have been developed to probe the effect of making the protein more positive.Support or Funding InformationMurchison Summer Undergraduate Research Fellowship, Welch Foundation
Get Out of Your Shell: Protein Extraction from Carcinus maenas
Authors: Kelly Lauren , Myracle Angela D. , Grant Jennifer E. 
Published: April 2020

The European green crab, a species that is native to Europe and northern Africa, is an invasive species that has overpopulated the coast of Maine. These crabs end up in fisherman’s nets but presently harbor no economic value. Yet, crab shells in general contain some of the highest protein content among shelled organisms. If the proteins and peptides extracted from Carcinus maenas can be extracted from shells inexpensively, it is possible that the European green crab shell could be transformed from a waste to an economic product. The purpose of this research is to examine the peptide and protein content from the European green crab (Carcinus maenas), using SDS‐PAGE electrophoresis.Protein extraction from shells is a non‐trivial task. The major component of crab shell is calcium carbonate, deposited within a protein/chitin matrix. Challenges to protein purification abound, particularly because calcium is prone to precipitating out when changes to pH or ionic strength are employed to enrich the proteins. This work expands upon our efforts to optimize extraction conditions. We chose to focus on 24‐hour extractions using hydrochloric acid; conditions ranging from 1 M to 0.25 M HCl were tested for efficacy. Protein precipitation using ammonium sulfate was then performed in order to fractionate proteins. Here we present data on the success of this ammonium sulfate‐based approach which resulted in isolation of a solution that was roughly 0.3 microgram/microliter protein.This research may identify novel bioactive peptides; little work has been done on uses for the European green crab and this approach may generate value‐added economic ideas.Support or Funding InformationThis work was supported by a University of Wisconsin‐Stout Student Dissemination Grant to LK.
Paracrine Secretion of Cardiac‐Derived Erythropoietin is Required for Cardiogenesis
Authors: Allwood Melissa A. , Platt Mathew J. , Huber Jason S. , Jacyniak Kathy , Edgett Brittany A. , Klein Jordynn M. , Marrow Jade P. , Alshamali Razan , Romanova Nadya , Brunt Keith R. , Vickaryous Matthew K. , Simpson Jeremy A. 
Published: April 2020

Erythropoietin (EPO) is widely recognized as the principle regulator of erythropoiesis, however, extra‐erythropoietic functions have been identified including cytoprotection, cardiac inotropy, cellular proliferation, and embryonic development. Whole body deletion of either EPO or the EPO receptor is embryonic lethal with impaired cardiogenesis leading to ventricular hypoplasia. While multiple extra‐renal tissue and cell types produce EPO, whether the heart is a direct source remains unclear. Human recombinant EPO increases myocardial contractility and confers cytoprotection against cardiac injury, which suggests a role for EPO signalling in the heart. Our objectives were to (1) confirm whether the heart produces EPO and (2) determine if there is a role for paracrine EPO signalling during cardiogenesis. We generated constitutive, cardiomyocyte‐specific EPO knockout mice driven by the Mlc2v promoter (EPOfl/fl:Mlc2v‐cre+/−; EPOΔ/Δ‐CM). We confirmed that the heart is a source of EPO expression with a distinct circadian rhythm in adult hearts and increased expression during embryonic development. During cardiogenesis, cardiac EPO expression was reduced, but not eliminated, in EPOΔ/Δ‐CM hearts with decreased cardiac cell proliferation. This suggests EPO signalling is partially compensated by an alternate cardiac cell type during cardiac development. In adult EPOΔ/Δ‐CM mice, global cardiac mass was preserved while cardiomyocyte cross‐sectional area was increased. Taken together, cellular cardiomyocyte hypertrophy in the absence of gross organ hypertrophy, and the observed reduction in cardiac cell proliferation during cardiogenesis, points towards a reduction in the overall number of cardiomyocytes in EPOΔ/Δ‐CM mice. Collectively, these data identify the first physiological roles of extra‐renal EPO by confirming that the heart is a source of EPO and that paracrine expression is required for cardiogenesis. Further, cardiac EPO expression is a complex interplay of multiple cell types where loss of cardiomyocyte production results in compensation from other cardiac cell lineages.Support or Funding InformationThis work was funded in part by the Canadian Institutes of Health Research (JAS), the Natural Sciences and Engineering Research Council of Canada (KRB, MKV, and JAS), the Canadian Glycomics Network (JAS), and the Heart and Stroke Foundation of Canada (KRB and JAS). JAS is also a new investigator with the Heart and Stroke Foundation of Ontario.
Gestational High‐Altitude Hypoxia and Metabolomic Reprogramming in Pulmonary Arteries from Fetal Sheep
Authors: Leslie Eric , Jones Breanna , Alvarez Rafael , Dasgupta Chiranjib , Fiehn Oliver , Bosviel Remy , Newman John W. , La Frano Michael , Zhang Lubo , Wilson Sean M. 
Published: April 2020

Gestational high‐altitude hypoxia increases the risk of myriad diseases in human infants including pulmonary hypertension of the newborn. Fetal sheep lungs are susceptible to long‐term intrauterine hypoxia, with the lung structure and function exhibiting characteristics associated with the development of pulmonary hypertension. Previous studies showed thickening of the pulmonary arterial medial wall, dysregulation of arterial reactivity, and reduced right ventricular output in fetal lambs following exposure to high altitude long‐term hypoxia. To further explore the mechanisms associated with hypoxia‐induced aberrations in the fetal sheep lung, we examined the premise that metabolomic and proteomic changes are associated with phenotypic transformation caused by intrauterine hypoxia. To address this hypothesis, we performed metabolomic and proteomic analysis on pulmonary arteries isolated from near term fetal lambs that were exposed to intrauterine high‐altitude hypoxia (3801 m) for the latter 110+ days of pregnancy or those gestated near sea level. The data demonstrated gestational hypoxia caused a glycolytic shift, reduced fatty acid oxidation, and elevated pentose phosphate metabolism along with increased histone expression, illustrative of increases in nucleotide synthesis. Increased nucleotide synthesis in response to gestational hypoxia is suggestive of a shift towards a proliferative and synthetic phenotype, providing insight into the mechanisms related to gestational hypoxia‐mediated pulmonary remodeling and hypertension. These studies begin to shed light on the mechanistic underpinnings associated with the phenotypic transformations to the pulmonary vasculature that occur with intrauterine hypoxia.Support or Funding InformationSupported in part by NIH grants HD083132 (LZ), 1U24DK097154 (OF ) through a pilot project grant to SMW and HD098477 (SMW). Additional support was provided by USDA Intramural projects #2032‐51530‐022‐00D and #2032‐51530‐025‐00D (J.W.N.). The USDA is an equal opportunity provider, and employer.
Embryonic and adult‐derived resident renal macrophages have different distribution and are maintained by distinctive mechanisms at steady state and during Inflammation
Authors: Shen Xiao (Gregory) , Zhu Qian , Cao Yangyang , Shi Peng 
Published: April 2020

The mononuclear phagocytes, including macrophages, dendritic cells and monocytes, are present in the kidney and are greatly involved in the pathogenesis of most kidney diseases. However, it is difficult to discriminate them from each other in the kidney, partly due to the fact that the markers generally used to discriminate them in other organs, such as CD11c, MHC class II and CX3CR1, are highly expressed by many of them. Here, we developed a strategy of multicolor flow cytometry to clearly differentiate them and their subtypes. Their identities were further confirmed by fate mapping, function analyses and lineage‐specific depletion. Using this strategy, we were able to show their inflammatory kinetics and activation alteration in a murine model of renal ischemia‐reperfusion injury. Moreover, by lineage tracing, we found that in adult mice, there were about 25% renal macrophages derived from embryo which are self‐maintaining, while other macrophages were actively replenished from the bone marrow. The embryonic and adult‐derived macrophages were distinctive in transcriptome and in distributions, as the embryonic macrophages highly accumulated in the medulla. Renal epithelial cells highly express CX3CL1 and the CX3CL1‐CX3CR1 axis was required for the differentiation of monoctes to the adult‐derived macrophages but not for the recruitment of monocytes from blood into the kidney. Thus, our discriminating strategy can be widely exploited to analyze mononuclear phagocytes in the kidney and has unveiled the specificity of turnover of renal macrophages.Support or Funding InformationNational Natural Science Foundation of China (Grant# 81670378, 81700365 and 31771266)
Constructing Combinatorial Synthases Using Acyl Carrier Protein Chimeras
Authors: Armstrong Claire , Kokona Bashkim , Charkoudian Louise K. 
Published: April 2020

Synthases encoded by biosynthetic gene clusters in microorganisms are capable of producing structurally complex molecules of diverse function. The ability to mix‐and‐match enzymes from different synthases to create “unnatural natural products” could enable the environmentally‐responsible construction of new pharmaceutically relevant molecules. Acyl carrier proteins (ACPs) are central hubs within biosynthetic pathways as they mediate the transport of molecular building blocks and intermediates to enzymatic partners that extend and tailor the growing natural product. Past combinatorial biosynthesis efforts have failed because of incompatible ACP‐enzyme interactions. To overcome this obstacle, chimeric ACPs capable of cross‐talk between different biosynthetic pathways were designed to serve as a lynchpin in functional hybrid synthases. Chimeric ACPs were constructed by combining features of the actinorhodin polyketide synthase ACP (ACT ACP) and of E. coli fatty acid synthase ACP (AcpP). Given the premise that AcpP interacts with its cognate ketosynthase, FabF, whereas ACT ACP does not, the ability of chimeric ACPs to form a mechanistically‐relevant crosslink with FabF was evaluated. A colorimetric assay was used to assess ACP interactions and revealed the role of Helix II of AcpP in driving ACP‐FabF binding. Results suggest that the residues guiding ACP‐FabF compatibility may reside in Helix II. Future directions will include using site directed mutagenesis to investigate the specific residues responsible for ACP‐FabF recognition.Support or Funding InformationNational Institutes of Health (R15GM120704 to L.K.C)National Science Foundation CAREER Award (CHE‐1652424 to L.K.C.)Cottrell Scholars Award (24350 to L.K.C.)
Identification and structure‐guided enzyme engineering of an extremophilic protein phosphatase
Authors: Chen Tiantian , Vander Kooi Carl D. , Anwar Humza , Sharma Savita , Murphy Robert D. , Gentry Matthew S. , Vander Kooi Craig W. 
Published: April 2020

Glucan phosphatases are members of the dual‐specificity phosphatase (DSP) family of cysteine‐dependent phosphatases. In plants, glucan phosphatases regulate starch metabolism while in animals they regulate glycogen metabolism. All known glucan phosphatase family members possess a DSP with a highly conserved active site. There is significant interest in developing glucan phosphatases for utilization in industrial processes involving starch and other polyglucans. We identified a putative phosphatase from the thermophilic bacterium Thermogutta terrifontis (Tt‐phos). Tt‐phos was expressed, purified and found to have a high thermal stability and tyrosine phosphatase activity. Determination of the crystal structure of Tt‐phos demonstrated that it adopts a glucan phosphatase‐like fold and provided key details of its active site architecture. Structure‐guided design was utilized to engineer the enzyme to produce a novel chimeric enzyme that has robust glucan phosphatase activity at elevated temperature. This study establishes the structural mechanism of Tt‐phos and provides key insights into the relationship of glucan phosphatase to the broader phosphatase family. Further, these data demonstrate a successful application of protein engineering for industrial applications of glucan phosphatases.Support or Funding InformationThis work was funded by the NSF (CHE CLP:1808304). Data was collected at the Advanced Photon Source (APS) SER‐CAT.
Artificial intelligence-enabled rapid diagnosis of COVID-19 patients
Authors: Mei Xueyan , Lee Hao-Chih , Diao Kai-yue , Huang Mingqian , Lin Bin , Liu Chenyu , Xie Zongyu , Ma Yixuan , Robson Philip M. , Chung Michael , Bernheim Adam , Mani Venkatesh , Calcagno Claudia , Li Kunwei , Li Shaolin , Shan Hong , Lv Jian , Zhao Tongtong , Xia Junli , Long Qihua , Steinberger Sharon , Jacobi Adam , Deyer Timothy , Luksza Marta , Liu Fang , Little Brent P. , Fayad Zahi A. , Yang Yang 
Published: April 2020

AbstractFor diagnosis of COVID-19, a SARS-CoV-2 virus-specific reverse transcriptase polymerase chain reaction (RT-PCR) test is routinely used. However, this test can take up to two days to complete, serial testing may be required to rule out the possibility of false negative results, and there is currently a shortage of RT-PCR test kits, underscoring the urgent need for alternative methods for rapid and accurate diagnosis of COVID-19 patients. Chest computed tomography (CT) is a valuable component in the evaluation of patients with suspected SARS-CoV-2 infection. Nevertheless, CT alone may have limited negative predictive value for ruling out SARS-CoV-2 infection, as some patients may have normal radiologic findings at early stages of the disease. In this study, we used artificial intelligence (AI) algorithms to integrate chest CT findings with clinical symptoms, exposure history, and laboratory testing to rapidly diagnose COVID-19 positive patients. Among a total of 905 patients tested by real-time RT-PCR assay and next-generation sequencing RT-PCR, 419 (46.3%) tested positive for SARSCoV-2. In a test set of 279 patients, the AI system achieved an AUC of 0.92 and had equal sensitivity as compared to a senior thoracic radiologist. The AI system also improved the detection of RT-PCR positive COVID-19 patients who presented with normal CT scans, correctly identifying 17 of 25 (68%) patients, whereas radiologists classified all of these patients as COVID-19 negative. When CT scans and associated clinical history are available, the proposed AI system can help to rapidly diagnose COVID-19 patients.
Risk Analysis and Prediction for COVID19 Demographics in Low Resource Settings using a Python Desktop App and Excel Models
Authors: Kyagulanyi Ali , Muhanguzi Joel Tibabwetiza , Oscar Dembe 
Published: April 2020

AbstractWhile the novel covid19 disease caused by sar-cov-2 corona virus has proved a serious threat to mankind it being a pandemic, the rate at which technology in low resource income countries like Uganda has been used to predict the spread and impact of the disease in their economies has not been strongly employed. This paper presents a an excel model and desktop application software developed using open source python programming tools for carrying out risk analysis and prediction of demographics for covid19 disease. Prediction results for both models clearly stated using epidemiological curve, these results can vary based on the force of infection which varies based on government measures and actions. With a certain degree of certainty of the potential impact of the disease on low resource countries, it will foster proper planning and strategical methods to properly manage the pandemic.
Microbiome evaluation revealed salivary dysbiosis in addicts of betel nut preparations
Authors: Saleem Faizan , Mujtaba Ghulam , Kori Junaid Ahmed , Hassan Arshad , Azim M. Kamran 
Published: April 2020

AbstractBetel nut addiction is recognized as the causative agent of oral microbiome dysbiosis and other systematic disorders. A number of betel nut preparations containing ingredients such as slaked lime, catechu extract and tobacco are being commonly used particularly in South Asia. The underlying variations in the oral microbiome due to usage of betel nut preparations are poorly understood. We evaluated salivary microbiome in response to chewing of betel nut preparation(s). In order to assess the microbiome dynamics, metagenomic analysis of 16S rRNA gene (V3-V4 hypervariable region) from salivary bacteria in chewers of betel nut preparation (n = 16) and non-chewers (n = 55) was carried out by Greengenes and SILVA ribosomal sequence databases. It was observed that Gutka chewers demonstrated lower alpha diversity and number of bacterial genera than the non-chewers. Taxonomic assignment on phylum level revealed Firmicutes (p-value = 0.042 at 95% confidence interval) to be significantly more abundant in Gutka chewers in comparison with non-chewers. Beta diversity analysis at genus level by weighted unifrac distance matrices unveiled both groups to be divergent from each other. On the genus level, Veillonella (p-value = 0.015), Streptococcus (p-value = 0.026), Leptotrichia (p-value = 0.022) and Serratia (p-value = 0.022) species appeared to be significantly more abundant in Gutka chewers in comparison to non-chewers. The present study suggests salivary dysbiosis in response to gutka chewing and concludes that gutka chewers possess higher abundance of acidogenic and aciduric bacteria. This study contributes additional information regarding oral microbiome variations with response to gutka consumption.