bluuebunny/crossref_metadata_2025_split huggingface dataset:
Extracellular Superoxide Dismutase (ecSOD) Modulates TNFα Signaling in Human Lung Endothelial Cells
Authors: Miller Michael , Choi Hyehun , Lamb Fred 
Published: April 2020

Tumor necrosis factor‐α (TNFα) signaling requires superoxide (O2−•) to be produced extracellularly by NADPH oxidase, but it remains unclear how this redox signal is transduced into activation of intracellular signaling pathways. We previously demonstrated in vascular smooth muscle cells (VSMCs) that TNFα signaling is dependent upon the extracellular isoform of superoxide dismutase (ecSOD). In cultured VSMCs siRNA knockdown of ecSOD decreased downstream MAPK phosphorylation and NFκB activation. In contrast, integrin receptor activation was enhanced, inducing higher levels of focal adhesion kinase (FAK) phosphorylation. These findings hint at a role for ecSOD in cytokine signaling that is independent from an anti‐oxidant influence. Studies in mice have shown that conditional knockout of ecSOD induces a rapid and lethal ARDS phenotype. Due to the critical nature of TNFα and reactive oxygen in the development of acute respiratory distress syndrome (ARDS) we sought to determine the signaling role of extracellular O2−• and ecSOD in endothelial cells (ECs). We employed cultured human lung microvascular endothelial cells (HLMVEC) to assess the impact of ecSOD knockdown (siRNA) on TNFα signaling. ecSOD expression was reduced ~ 50% by a 72hr exposure to siRNA. In siRNA treated HLMVECs, phosphorylation of extracellular signal‐regulated kinase (ERK) following a 15 min exposure to TNFα was significantly increased (siControl vs siSOD3 – 0.82 vs 1.93 mean signal intensity; p = 0.016). This short‐term exposure to TNFα also induced a significant reduction in the abundance of ecSOD detected by Western blot (61.6% signal intensity after TNF treatment). This suggests either rapid TNFα‐induced degradation of ecSOD, or dissociation of the protein from the cell surface, where ecSOD is known to bind to either heparin sulfate or Fibulin 5, a component of the extracellular matrix that also binds to integrins. A transendothelial resistance (TEER) assay was employed to assess the role of ecSOD in formation of a tight EC monolayer. HLMVECs treated with ecSOD siRNA were significantly impaired in their ability to develop TEER compared to cells treated with scrambled siRNA. Together, these data suggest that ecSOD plays a critical role in normal endothelial cell function and in signaling in response to TNFα, including a role in regulation of endothelial monolayer integrity. Ongoing work seeks to define the contribution of ecSOD‐dependent signaling to EC inflammation in ARDS.Support or Funding InformationThis work was supported by R01HL128386 and 5T32HD060554‐10
Antioxidant Activity of Graviola Extracts Extracted with Different Solvents
Authors: Kang Myunghwa , Kim Dohee , Seo Hyungsuk , Lee Hajong , Choo Yeanhan , Kim Doungjin 
Published: April 2020

The gravureola (Annona muricata L.Graviola) has long been a traditional "natural remedy" for tropical Amazonians and is called a "God's gift." Graviola is attracting attention in medicine and pharmacy because it contains a lot of antioxidants, such as tannins, saponins, flavonoids and anthraquinones, which are highly researched. Therefore, this study was performed to extract gravolae from different solvents and to compare the antioxidant activity of gravureola extract.The extraction solvent used in this study was prepared by drying extracted with heat mentle by adding water, 30% ethanol and 80% ethanol. Antioxidant effect of each extract was measured by measuring total phenolic acid contents and flavonoids contents which measured DPPH radical scavenging activity, ABTs scavenging activity, SOD‐liked activity, reducing power and antioxidant content. As a result, the DPPH radical scavenging activity showed the highest activity at 20 minutes for the 30% ethanol extract and at 20 minutes for the 80% ethanol extract.ABTs scavenging activity showed similar effects in all extracts. The SOD‐liked activity was very high in 80% ethanol extract compared to other extracts. Reducing power was 80% ethanol> 30% ethanol> water. Total phenolic acid and flavonoids contents were 80% ethanol> 30% ethanol> water and contained less than 0.2mg / g. As a result, the antioxidant effect of graviola was the highest in 80% ethanol extract, suggesting the possibility of development as a functional material.
Regulation of Human Organic Cation Transporter 3 (hOCT3) by the Dual‐Specifity Tyrosine‐(Y)‐Phosphorylation Regulated Kinase (DYRK1A)
Authors: Ciarimboli Giuliano , Stapel Mira , Barz Vivien , Pavenstädt Hermann , Rinschen Markus M. 
Published: April 2020

hOCT3, a transmembrane protein with broad tissue distribution, is involved in the low‐affinity, high‐capacity transport of monoamine substrates across the cell membrane. In mice OCT3 is important for cerebral monoaminergic neurotransmission and for the effects of monoamines on behavior. The molecular regulation of hOCT3, however, is unknown.In this study, we have identified the dual‐specificity tyrosine‐(Y)‐phosphorylation regulated kinase 1A (DYRK1A) as a key interaction partner of hOCT3 using a membrane‐based Split‐Ubiquitin‐Yeast‐Two‐Hybrid technique, and pull‐down experiments in hOCT3 and DYRK1A transfected human embryonic kidney (HEK) cells. In these cells, DYRK1A overexpression resulted in hOCT3 phosphorylation at the serines 3 and 537 as determined by phosphoproteomic analysis, and in a significant inhibition of the transport of 4‐4‐dimethylamino‐styryl‐N‐methylpyridinium (ASP+), a fluorescent hOCT3 substrate. DYRK1A overexpression reduced the maximum transport velocity (Vmax) of hOCT3, without changing its affinity for ASP+. The hOCT3 transporter function was rescued by inhibition of DYRK1A with 100 nM harmine. In the seminoma cell line Tcam‐2, which endogenously expresses hOCT3 and DYRK1A, harmine treatment resulted in 1) a significant increased hOCT3 expression in the plasma membrane, as determined by biotinylation experiments; 2) an increase of ASP+ uptake; and 3) a decrease of hOCT3/DYRK1A co‐localization as determined by confocal microscopy. Moreover, in Tcam2 cells, phosphatase activation by 4 h incubation with 10 μM ceramide induced a significant increase of ASP+ uptake.Taken together, these data show that hOCT3 is regulated by DYRK1A via interaction, phosphorylation and change of intracellular distribution. The gene coding for the DYRK1A is localized in the Down syndrome (DS) critical region of chromosome 21 and is a strong candidate gene for DS‐associated mental retardation. Since hOCT3 functions as neurotransmitter reuptake transporter in the brain, we speculate that its inhibition by DYRK1A results in an impaired reuptake of neurotransmitters, and therefore an impoverishment of brain neurotransmitters content. This mechanism may be one of the molecular causes of mental retardation and susceptibility to Alzheimer diseases in DS patients.Support or Funding InformationDeutsche Forschungsgemeinschaft CI 107/11.1
A new molecular mechanism of evasion of apoptosis revealed by novel C‐kinase activity reporter
Authors: Kajimoto Taketoshi , Caliman Alisha D. , Tobias Irene S. , Okada Taro , Pilo Caila A. , Van An-Angela N. , McCammon J. Andrew , Nakamura Shun-ichi , Newton Alexandra C. 
Published: April 2020

It is well known that cells, especially cancer cells, have an ability of evasion of apoptosis by cellular stress like nutrient starvation. And the balance between apoptosis signal and apoptosis‐resistant signal will determine the fate of cells, dead or alive. Here we found new cell signaling system that plays a role in applying the brakes to cell death that in case cancer cells avoid apoptosis by cellular stress like starvation using newly developed biosensor and in silico docking simulation technique. The new cell signaling system is that second messenger, sphingosine 1‐phosphate (S1P), directly activates a key cell signaling protein, atypical protein kinase C (aPKC).First, we found that the inhibition of aPKC induces apoptosis of cancer cell lines. Next, for making clear the molecular mechanism of the aPKC‐induced apoptosis resistance, we generated a genetically encoded reporter with the same modular structure as the original C kinase activity reporter (CKAR) but with a unique substrate sequence that allows specific visualization of atypical PKC activity in cells. Using the atypical PKC‐specific CKAR (aCKAR) we found that intracellular S1P induces the activation of atypical PKC in an S1P receptor‐independent manner. Biochemical studies revealed that S1P directly binds to the kinase domain of atypical PKC isozymes, relieving autoinhibitory constraints to activate the enzyme. In silico docking studies were used to identify potential binding sites for aPKC, one of which was validated by biochemical and aCKAR imaging techniques.Now we got new insights about the player of evasion of apoptosis in cancer at the molecular level, and it has potential for development of new molecular‐targeted agents to release brakes against cell death.Support or Funding InformationThis study was supported by NIH to A.C.N., JSPS KAKENHI, Kobe University Grant for Japan‐US Collaboration, Nakatani Foundation Grant for Technology Development Research to T.K., NIH, HHMI, NBCR, NSF to J.A.M., JSPS KAKENHI to S.N. and T.O. A.D.C. was supported in part by the UCSD Graduate Training Program in Cellular and Molecular Pharmacology.
Effects of Various Hypoxia vs Normoxia on Metabolic and Skeletal Muscle Oxygenation Function during Graded Exercise in Healthy Males
Authors: Seo Jongbeom , Jung Won-Sang , Park Hun-Young , Lim Kiwon 
Published: April 2020

PurposeThe purpose of this study was to investigate the effect of various hypoxia vs. normoxia on metabolic and skeletal muscle oxygenation function during grated exercise and exercise performance.MethodsEleven healthy male participants (21.5 ± 2.3 years) performed graded exercise test (GXT) using cycle ergometer at sea‐level (760 mmHg) and under various hypobaric hypoxia (1,500 m simulated altitude; 634 mmHg, 3,000 m simulated altitude; 528 mmHg and 4,500 m simulated altitude; 433 mmHg) in a random order. GXT starts at 300 kg·m·min−1 (50 watts) and increases by 150 kg·m·min−1 (25 watts) every 2 minutes until exhaustion. The pedal frequency was set at 60 rpm. Metabolic parameters (peripheral capillary oxygen saturation; SPO2, heart rate; HR, minute ventilation; VE, oxygen consumption; VO2, carbon dioxide excretion; VCO2, respiratory exchange ratio; RER, oxygen pulse, and blood lactate) and skeletal muscle oxygen profiles (oxygenated hemoglobin and myoglobin; OxyHb, deoxygenated hemoglobin and myoglobin; DeoxyHb, and tissue oxygen saturation; StO2) were measured every 1 minute during the GXT, and it also evaluated at exercise load of 50, 75, 100, 125, and 150 watts on ergometers under various hypoxia vs. normoxia. Exercise performance was evaluated by maximal oxygen consumption, peak power, and exercise time obtained through GXT.ResultsIn metabolic parameters, HR, VE, VCO2, RER, and blood lactate showed a significant increase in hypoxia compared with normoxia. Also, the increase was more pronounced as the hypoxia became more severe. However, SPO2, VO2, and O2pulse presented a significant decrease in hypoxia compared with normoxia. Also, the decrease was more pronounced as the hypoxia became more severe. In skeletal muscle oxygen profiles, OxyHb showed a significant decrease in 3,000 m and 4,500 m simulated altitude compared with normoxia. In exercise performance, VO2max and peak power were significantly decreased in 3,000 m and 4,500 m simulated altitude compared with normoxia, and exercise time decreased significantly with increasing simulated altitude.ConclusionThese showed a decrease in exercise performance due to a decrease in metabolic parameters and skeletal muscle oxygenation in hypoxia vs. normoxia, and more pronounced as the hypoxia became more severe.Support or Funding Information
Beneficial Effect of TMAO in Spontaneously Hypertensive Heart Failure Rats is Associated with Diuretic and Hypotensive Actions.
Authors: Ufnal Marcin , Konop Marek , Gawrys-Kopczynska Marta , Maksymiuk Klaudia , Sozanski Krzysztof , Derzsi Ladislav 
Published: April 2020

There is an ongoing debate weather trimethylamine‐oxide (TMAO), a molecule present in seafood and a derivate of microbiota metabolism, is beneficial or harmful for the circulatory system. Numerous biophysical studies show that TMAO protects proteins exposed to osmotic and hydrostatic stress (HS). We hypothesized that TMAO may benefit the circulatory system by protecting cardiac proteins exposed to HS produced by the contracting heart.Male, 6‐week‐old, Sprague‐Dawley (SD, n=40) and Spontaneously‐Hypertensive‐Heart‐Failure (SHHF, n=18) rats were randomly divided into either control (Water) or TMAO treatment. After 50 weeks, half of SD were given Isoprenaline (s.c.) to increase arterial blood pressure (BP) and produce a mild, Takotsubo‐like cardiomyopathy (ISO). In vitro, cardiac lactic dehydrogenase (LDH) with or without TMAO was exposed to HS (changes in hydrostatic pressure similar, and higher than those present in the heart) for up to 24hrs and was evaluated using fluorescence correlation spectroscopy.After 58 weeks of the treatment survival was 100% in SD‐Water, SD‐TMAO, ISO‐TMAO and 90% in ISO‐WATER. In SHHF‐Water survival was 66% vs 100% in SHHF‐TMAO (p=0.065). In general, TMAO‐treated rats showed higher natriuresis and diuresis. In comparison to SHHF‐Water, SHHF‐TMAO showed significantly lower plasma NT‐proBNP, lower diastolic BP, lower liver angiotensinogen expression and a trend towards lower cardiac fibrosis. In vitro, exposure of LDH to HS with or without TMAO did not affect the protein structure. TMAO does not exert negative effects. In contrast, TMAO reduces mortality in HF rats that is associated with natriuretic, diuretic and hypotensive effects. HS produced by the contracting heart is neutral for cardiac LDH protein structure.Support or Funding InformationSupported by National Science Centre, Poland grant no 2018/31/B/NZ5/00038.
Novel insights into the fucose metabolism – challenging the old dogma
Authors: Sosicka Paulina , Ng Bobby G. , Wong Maurice , Xia Zhi-Jie , Scott David , Lebrilla Carlito B. , Freeze Hudson H. 
Published: April 2020

GDP‐Fucose used for glycan fucosylation is synthesized de novo from either glucose (Glc) or mannose (Man) or utilized directly from salvaged/exogenous fucose (Fuc). Meticulous radioisotope dilution studies performed in the 1970’s only in HeLa cells using 0.3 μM Fuc showed that the de novo pathway provides >90% of GDP‐Fuc and salvage <10%.Employing different Fuc‐specific lectins, we showed that at low concentrations (<20 μM), the entry of Fuc into cells relies on transport via GLUT1 (SLC2A1), but above this, most Fuc enters via amiloride‐sensitive macropinocytosis. Inhibitors of both clathrin‐ and caveolin‐dependent endocytosis had no effect on uptake. These findings were further confirmed using 3H Fuc in four cell lines.We used differentially labelled 13C Glc, Man and Fuc to study contributions of each pathway to Fuc in N‐glycans in six cell lines. Very low concentrations (0.5–30 μM) of exogenous Fuc progressively inhibit the de novo pathway, and completely by ~50 μM. Surprisingly, GDP‐Fuc derived from Glc is much more sensitive to this inhibition than that derived from Man, even though both contribute to GDP‐Man synthesis. When cultured in 30–50 μM Fuc, cells not only inhibit the de novo pathway, but also prefer exogenous over salvaged Fuc. Cells seem to distinguish multiple pools of GDP‐Fuc based on their origin.Using LC‐MS/MS and eight different cell lines, we studied the ongoing biosynthesis of >150 fucosylated N‐glycans containing 1–4 Fuc residues by comparing the proportion of 13C Fuc incorporation at various concentrations for 24h. Over the labeling period, cells had to choose between GDP‐Fuc synthesized de novo or from exogenous fucose. N‐glycans with a single Fuc show a proportional increase in 13C‐labeled species with increasing concentration of label. To our surprise, glycans with 2, 3, and 4 Fuc each required progressively higher exogenous 13C Fuc to become fully labeled.Based on LC‐MS/MS and lectin staining, 1–6αFuc labels more efficiently with exogenous Fuc than 1–2α, 1–3α and 1–4αFuc, suggesting differential access to exogenous sugar that depends partially on Fuc linkage. Km differences between fucosyltransferases cannot explain preferential use of GDP‐Fuc synthesized from exogenous monosaccharide over de novo produced substrate. Rather, it suggests the existence of separate, ill‐defined pools of GDP‐Fuc. Altogether it raises the question whether salvaged fucose equivalent to exogenous Fuc? Their behavior clearly differs from de novo synthesized GDP‐Fuc.Support or Funding InformationThe Rocket Fund and R01DK099551.
“A Whole New Perspective on How the Body Fits Together” – An Evaluation of a Cadaver Lab Experience for High School and Undergraduate Students
Authors: Cale Andrew , Byram Jessica , Schmalz Naomi 
Published: April 2020

BackgroundSince 2014, the Center for Anatomy and Physiology Education at Indiana University School of Medicine – Indianapolis has hosted interactive human cadaver lab tours for local high school and undergraduate students. During these graduate student‐led tours, touring students observe human anatomy on prosections and with isolated organs. Similar educational outreach initiatives have been shown to benefit adolescents by improving health literacy and encouraging interests in healthcare careers.AimTo evaluate students’ perceptions of the anatomy lab tours and their impact on students’ interests in healthcare careers.MethodsBetween April and December 2018, touring students (n=261) were invited to complete pre‐ and post‐tour surveys evaluating their past experiences with cadaver labs, tour expectations, career interests, and additional thoughts and feelings regarding the tour. Pre‐surveys were paper‐based and completed in‐person, while post‐surveys were administered online via REDCap and completed within one week of the tour. Responses were aggregated into pre‐ and post‐tour groups and analyzed using thematic analysis. This study was granted exempt status from the IRB at Indiana University (#1802755251).ResultsOf the 261 students who completed pre‐tour surveys, 204 (78%) completed the post‐tour survey. Before the tour, students anticipated learning about human anatomy (52%) and expected to only be able to see (and not feel) a cadaver (66%). Most students expressed excitement (80%) or nervousness (45%). A few students viewed the lab tour as an opportunity to test if they could see themselves in a healthcare career (4%). After the tour, most students indicated that the tour either met (77%) or exceeded (22%) their expectations. Students found the lab tour to be educational and interesting. They were surprised by the opportunity to interact with the cadaver (23%) and by the pre‐dissected appearance of the cadaver (26%). Numerous students (41%) also expressed an increased interest in healthcare careers after the lab tour.ConclusionOverall, students perceived the lab tours as engaging, educational experiences that not only improved their anatomical knowledge, but also reinforced or increased their interest in healthcare careers. Academic institutions interested in positively impacting their local students can implement a cadaver lab tour program by sharing access to their in‐house human cadaver lab and recruiting instructors to share their time and anatomy expertise.
The efficacy of Augmented Reality in improving the learning outcome
Authors: Bakhtiari Shahram 
Published: April 2020

Augmented Reality is gaining traction as an alternative and effective learning tool in studying human anatomy in colleges and universities. This technology provides an immersive approach to learn anatomical structure using HoloLens with an extensive range of maneuverability of Hologram, dissection mode, information boxes and most amazingly walking into the Hologram and visualizing the human body inside out.Human anatomy, as one of the introductory courses in health sciences is usually found challenging for the students and the capability of any learning tools to provide simplification and three‐dimensional visualization, in addition to eliciting excitement with advanced technology will be promising to improve the students’ learning experience.The inclusion of the Augmented Reality into the human anatomy course was able to improve the students’ final exam grade 18.9% and reduce the failure rate by 10%. The student’s satisfaction survey also showed 87.6% of the students believed that the Augmented Reality helped them to reach the objectives of the anatomy course.Support or Funding InformationWest Coast University
Agonism of adiponectin Receptor Prevents 7‐ketocholesterol‐induced Endothelial Inflammation through Suppressing Endoplasmic Reticulum and Oxidative Stresses
Authors: Zhang Yang , Chen Jiajie , Boini Krishna , Li Xiang 
Published: April 2020

Endoplasmic reticulum (ER) stress is involved in endothelial inflammation and injury by various cardiovascular risk factors including oxysterols. AdipoRon is a selective small‐molecule agonist of adiponectin receptor, which has been shown to mimics many beneficial effects of adiponectin on the cardiovascular system including endothelial cells. However, the molecular mechanisms that mediate adipoRon‐induced endothelial protection are not well understood. Here, we studied whether adipoRon treatment interferes with the induction of ER stress to protect the endothelial cells from oxysterol‐induced inflammation and injury in vitro and to inhibit vascular inflammation from acute hypercholesterolemia in vivo. In cultured murine endothelial cells, adipoRon prevented 7‐ketocholesterol‐induced increases in endothelial cell death, inflammatory gene expression, monocyte adhesion, and Nlrp3 inflammasome activation. In parallel, adipoRon inhibited 7‐ketocholesterol‐induced ER stress and reactive oxygen species (ROS) production. In mice with acute hypercholesterolemia, adipoRon pre‐treatment could inhibit ROS production and ER stress in the coronary arterial wall without alterations in the plasma cholesterol concentration. Moreover, adipoRon reduced expression of ICAM1 in the coronary arterial intima and decreased numbers of CD45+ immune cells that adhere to or accumulate around the coronary arteries. Together, our results indicate that the cardiovascular protective action of adipoRon is associated with its inhibitory effects on ROS production and ER stress and consequent inflammatory responses in endothelial cells.Support or Funding InformationNational Institutes of Health (HL122769 and HL122937)