bluuebunny/crossref_metadata_2025_split huggingface dataset:
Association of NRF2 and NQO1 Polymorphisms and Susceptibility to Emphysema
Authors: Marzec Jacqui M , Hertzberg Andrew , Yoon Hyoung‐Kyu , Kleeberger Steve 
Published: June 2021

RationaleEmphysema is defined as airway enlargement due to loss of alveolar structure and diminished elasticity, and is mainly caused by cigarette smoking and air pollution. NRF2 activates antioxidants such as NQO1 in response to oxidative stress, and deficiency increases susceptibility to cigarette smoke‐induced emphysema in mice. The objective of this study was to determine whether functional polymorphisms in NRF2 and NQO1 associate with emphysema susceptibility.MethodsEmphysema cases and controls were recruited for study. Regions of the NRF2 promoter and NQO1 promoter were amplified from genomic DNA and sequenced.ResultsIndividuals homozygous for the NRF2 rs35652124 G allele were at increased risk for development of emphysema compared to those with wildtype alleles [OR 2.514; CI: 1.173, 5.389]. Individuals homozygous for the NQO1 rs689455 C allele were at decreased risk for emphysema compared to those with wildtype alleles [OR 0.299; CI: 0.124, 0.722].ConclusionsIn this translational investigation, we found that the NRF2 rs35652124 and the NQO1 rs689455 mutations confer differential risk of emphysema. Previous investigations demonstrated similar risk effects of NRF2 and NQO1 promoter SNPs for susceptibility to acute lung injury. These investigations suggest that NRF2 and NQO1 are pivotal determinants of susceptibility to oxidative stress‐induced lung diseases.
Purinergic receptors stimulate Calcium transport in pancreatic duct cells
Authors: Novak Ivana , Hansen Mette Rud , Krabbe Simon 
Published: June 2021

Normal pancreatic juice contains <1 mM concentrations of Ca2+, but little is known about Ca2+ homeostasis, which may be an important function of duct epithelium. Our aim was to investigate whether duct cells from human and rat pancreas express Ca2+‐transporting and Ca2+‐binding proteins and whether purinergic receptors regulate Ca2+ transport, as is known for anion transport. Expression analysis revealed that pancreatic ducts of rat and human duct cell line CFPAC‐1 (also PANC‐1 and Capan‐1) express the Na+/Ca2+ exchanger (NCX), as well as the Ca2+‐sensing receptor and the Ca2+‐binding protein calbindin 9kD. These proteins localize to the luminal and lateral membranes of pancreatic duct epithelia. Ca2+ imaging experiments showed that activation of purinergic P2X and P2Y receptors stimulated NCX in rat and human duct cells. Application of ATP to CFPAC‐1 monolayers stimulated Ca2+ transport from the luminal to the basolateral side. Taken together, these results show that pancreatic ducts express a number of Ca2+‐handling proteins and we suggest that these together with purinergic receptors may play a role in regulation of pancreatic Ca2+ transport.Support: PhD scholarship, University of Copenhagen and DNSRC 272‐05‐0420
Regulation of V‐ATPase, NBCe1‐A and NBCe1‐B in hippocampus
Authors: Martin Henno , Oehlke Oliver , Roussa Eleni 
Published: June 2021

The electrogenic sodium‐bicarbonate‐cotransporter (NBCe1) is a key player in regulation of intracellular pH in several cell types. In the CNS however, modes of intracellular pH regulation reveal considerable differences, compared to other tissues, because neuronal activity is accompanied by rapid pH shifts. Here we have investigated expression and regulation of V‐ATPase, and N‐terminal variants of NBCe1 in mouse brain.V‐ATPase, NBCe1‐A and NBCE1‐B expression was detected in hippocampus, cerebral cortex, olfactory bulb and cerebellum. However, NBCe1 N‐terminal variants revealed distinct cellular distribution in mouse brain and showed differential activity‐dependent regulation in vitro. After induction of extracellular acidosis or alkalosis gene expression and protein expression of V‐ATPase, NBCe1‐A and NBCe1‐B in hippocampal cultures were not changed. However, acute extracellular acidosis resulted to V‐ATPase and NBCe1‐A redistribution from the cytosol towards the plasma membrane.The results suggest redistribution of acid‐base transporters as main regulation mechanismus during acid‐base disturbances. Moreover, NBCe1 may be involved in neuronal modulation, and pH regulation during neuronal activity.
Mineralcorticoid Receptor (MR) Antagonism Attenuates Glomerular Filtration Barrier Remodeling in the Transgenic Ren2 Rat
Authors: Whaley‐Connell Adam , Habibi Javad , Wei Yongzhong , Gutweiler Alex , Jellison Jessica , Wiedmeyer Charles E , Ferrario Carlos M , Sowers James R 
Published: June 2021

Renin‐angiotensin‐aldosterone system (RAAS) activation is associated with glomerular filtration barrier/podocyte injury & accompanying proteinuria. To explore potential MR mediated renal effects, Ren2 transgenic rats, which manifest increased tissue RAAS activity with elevated serum aldosterone levels, were used. Six week old Ren2 and Sprague‐Dawley rats were treated with a low dose of the MR antagonist spironolactone (SP)(0.24 mg/day) or vehicle for 21 days. Ren2 rats exhibited albuminuria, loss of podocyte‐specific proteins and podocyte foot‐process effacement on ultrastructural analysis, along with reduced expression of mediators of insulin metabolic signaling. Ren2 kidneys also manifested increased NADPH oxidase/ROS and redox‐sensitive Rho kinase activation in conjunction with enhanced renal tissue levels of MR, angiotensin type 1 receptor, angiotensin II and angiotensin‐(1‐12). SP treatment reduced albuminuria and tissue RAAS expression and improved podocyte structural and protein integrity along with improvements in insulin metabolic signaling. MR blockade directly provides protection in this model of renal RAAS activation, in part, through reductions in redox‐sensitive ROK impairment of insulin metabolic signaling and glomerular remodeling.Supported by NIH R01 HL73101‐01A1 (JRS) & HL‐51952 (CMF), VA Merit (JRS), VA VISN 15 (AWC), & Missouri Kidney Program (AWC).
Microvascular Loss and Progression of Renal Injury in Renovascular Disease: Renoprotective Effects of VEGF
Authors: Chade Alejandro R. , Iliescu Radu 
Published: June 2021

Chronic renal artery stenosis (RAS) induces renal functional impairment by promoting microvascular (MV) endothelial dysfunction, damage, and loss, which contributes to the progression of renal injury. Vascular endothelial growth factor (VEGF), crucial for preservation of microvasculature, promotes endothelial repair and modulates renal fibrosis.We hypothesized that administration of VEGF would preserve the renal MV architecture and function during the evolution of RAS.RAS was induced in 12 pigs. VEGF‐165 was intra‐renally infused at the onset of the stenosis (RAS+VEGF, 0.05 ug/kg) in 6 pigs monitored for 6 weeks, and compared to normal controls (n=6). Additional RAS+VEGF pigs underwent revascularization (RV) after 6 weeks, VEGF repeated (RAS+VEGF+RV), and observed for 4 more weeks (n=3). Normal and RAS+RV animals (n=3) served as time‐controls.Single‐kidney hemodynamics and function were assessed in vivo using ultra‐fast CT after 6 and 10 weeks. Renal MV architecture was investigated ex‐vivo using 3D micro‐CT and CD31 immunostaining, and morphology evaluated in cross‐sections stained with trichrome.The degree of RAS and increase in blood pressure were similar in RAS and RAS+VEGF animals. However, decreased RBF, perfusion, and GFR in RAS were normalized in RAS+VEGF and accompanied by augmented renal capillary density and decreased fibrosis. Furthermore, RV normalized GFR and blood pressure only in RAS+VEGF+RV pigs.A single intra‐renal infusion of VEGF improved renal function and decreased fibrosis of the stenotic kidney. This study shows the feasibility of a novel targeted therapeutic approach and supports renoprotective effects of VEGF during the evolution of RAS by preserving renal function not only at the early stage of the disease, but also by improving the responses to revascularization.
Critical Role of αB Crystalline on Stretch‐Induced FAK Activation in Cardiomyocytes
Authors: Pereira Michelle B. M. , Santos Aline M , Franchini Kleber G. 
Published: June 2021

AimFocal adhesion kinase (FAK) is activated and plays a role in cardiomyocyte hypertrophy induced by mechanical stress. FAK activation may depend on cooperation with protein partners. We investigated whether interaction with alpha‐B crystalline (αB Crys) regulates FAK activation by mechanical stress in cardiomyocytes.MethodsαB Crys and FAK expression and phosphorylation at Tyr397 (pFAK) were quantified in neonatal rat ventricular myocytes (NRVMs). FAK/αB Crys interaction was investigated by co‐immunoprecipitation and pulldown assays. αB Crys depletion by RNAi was used to examine its effect on FAK activation.ResultsCyclic stretch (10%, 2 hs) increased pFAK (2‐fold) and enhanced αB Crys expression (4‐fold) and phosphorylation at Ser59 (3‐fold). Stretch also induced FAK/αB Crys association. Pulldown assays with recombinant FAK domains indicated that the interaction with αB Crys is mediated by FAK C‐terminal domain. FAK immunostaining was coincident with myofilaments in non‐stretched while in stretched cells it was predominantly located in the nuclei. αB Crys was predominantly located at the nuclei of stretched NRVMs, coincident with FAK staining. Depletion of αB Crys markedly reduced FAK activation and re‐location to the nucleus.ConclusionThis study demonstrates that αB Crys associates and regulates FAK activation in cardiomyocytes in response to mechanical stress.
Case report: Anatomic variant of forearm arteries and its clinical correlation
Authors: Kumka Myroslava 
Published: June 2021

MethodsTraditional macro‐micro dissections of 30 upper limbs from 15 embalmed cadavers of both sexes and of different ages were performed.Summary of results: An atypical course and branching pattern of forearm vessels was found unilaterally in one male cadaver. The ulnar artery originated as one of the two terminal branches of the brachial artery and was significantly smaller than commonly seen. It descended superficially, covered only by the skin and the anterbrachial fascia. The branching pattern of the radial artery was also atypical. A common interosseous artery which typically arises from the ulnar artery, in this case emerged as a short branch of the radial artery distal to the radial tuberosity. While passing toward the interosseous membrane, the common interosseous artery was separated from the median and anterior interosseous nerves by the ulnar head of the pronator teres muscle and branched out into the anterior and posterior interosseous and ulnar recurrent arteries that supply the major part of the anterior forearm region.ConclusionThis anatomical variant of forearm arteries should be considered in diagnosis and treatment since it could complicate radiographic, surgical, and orthopaedic procedures in the forearm, such as radial artery screening for grafting purposes, ligation of a lacerated radial or ulnar artery, and assessment of forearm vessels by color Doppler ultrasonography.
Presence of Markers of Femoroacetabular Impingement in the Asymptomatic Population
Authors: Raynor Charys Megan , Bryant Dianne , Spouge Alison , Birmingham Trevor B , Willits Kevin 
Published: June 2021

PurposeTo determine: 1) the prevalence of femoroacetabular impingement (FAI) in asymptomatic individuals, and 2) whether there is an association between the presence of FAI and age and degenerative disease. Methods: 88 volunteers were verified as asymptomatic through physical examination and then underwent an MRI of the hip. Different criteria values were used to classify the presence or absence of cam (alpha angle of 45, 50 and 55 degrees) and pincer (centre‐edge (CE) angle of 35 and 40 degrees) impingement.ResultsThe prevalence of FAI ranged from 23.9 to 67.0% depending on the classification criteria applied. There was no significant association between FAI and age. Prevalence of FAI, identified using high criteria values (55 degree alpha angle and 40 degree CE angle), predicts the presence of acetabular cartilage lesions. Prevalence of FAI, identified using lower criteria values, was not a significant predictor of degenerative disease.ConclusionFAI is prevalent in the asymptomatic population. Further research is necessary to verify the validity and reliability of criteria used to diagnose impingement and to determine the validity of the diagnosis. Support: AANA and SIRF.Grant Funding SourceInternal
Enzymology of DNA Repair in the radiation‐resistant extremophile Deinococcus radiodurans
Authors: Cao Zheng , Shadrick William R , Julin Douglas 
Published: June 2021

The bacterium Deinococcus radiodurans exhibits extraordinary resistance to the effects of ionizing radiation, and other genotoxic agents. We are using genetic and biochemical approaches to define the enzymatic pathways by which D. radiodurans repairs DNA damage.D. radiodurans genome loci DR1902 (RecD) and DR1572 (the putative helD gene) encode DNA helicases. Cells with a helD deletion are more sensitive than wild‐type to hydrogen peroxide and methyl‐methanesulfonate, but they are not sensitive to gamma or UV irradiation. Both helicases have a protein domain that is homologous to other superfamily 1 helicases, and an additional N‐terminal domain. The N‐terminal domain of HelD is essential for DNA unwinding by this enzyme. We are attempting to purify the helicase domain of RecD as defined by limited proteolysis, and to examine the kinetics of DNA unwinding by the domain and the full‐length RecD protein. Genome locus DR1126 encodes RecJ, a single‐strand‐specific exonuclease in the RecF pathway for DNA recombination and repair in E. coli. We have deleted the gene from the D. radiodurans genome by standard homologous recombination techniques. We are unable to delete the gene from each of the ~10 chromosomal copies in the cell, suggesting that recJ may be an essential gene. Cells carrying the partial deletion are more sensitive to UV irradiation than wild‐type, suggesting an important role for RecJ in D. radiodurans.
Experimental model of the interplay between TBL1‐mediated activation and Siah‐1‐induced poly‐ubiquitination of β‐catenin
Authors: Dimitrova Yoana N. , Chazin Walter J. 
Published: June 2021

β‐catenin is a critical factor in the Wnt signaling pathway and its function is maintained by balancing activation and degradation signals. TBL1 has been identified as a transcriptional co‐activator of β‐catenin. Remarkably, TBL1 has also been associated with an SCFTBL1‐like E3 ubiquitin ligase, initiated by Siah‐1 upon DNA damage, while Siah‐1 itself functions as a potent E3 ligase. Our studies focus on understanding the complex interplay between TBL1‐mediated activation and Siah‐1‐induced poly‐ubiquitination of β‐catenin.Protocols for recombinant production of the full‐length TBL1 and Siah‐1 have been developed and the proteins were used in in vitro ubiquitination assays to probe the poly‐ubiquitination of β‐catenin. Our results have shown that Siah‐1 is able to efficiently poly‐ubiquitinate β‐catenin as a simple E3 ligase. Interestingly, the addition of TBL1 is seen to inhibit β‐catenin ubiquitination. In order to understand how TBL1 and Siah‐1 compete for β‐catenin, we have mapped the binding domains in Siah‐1‐β ‐catenin and TBL1‐β‐catenin complexes and characterized the strength of their interactions. Combining our biochemical and biophysical data with computational approaches, we have designed a model of the molecular events regulating the functional state ofβ‐catenin. Support: NIH 5R01GM075156‐04.
Two human holocarboxylase synthetase proteins display similar kinetic properties
Authors: Ingaramo Maria , Beckett Dorothy 
Published: June 2021

Human Holocarboxylase Synthetase (HCS) catalyzes transfer of the cofactor biotin to biotin‐dependent carboxylases. Consequently, this enzyme is of fundamental importance for fatty acid synthesis, gluconeogenesis, and amino acid catabolism. In addition, in agreement with its role in transcription regulation, it has been proposed that HCS biotinylates histones. There are two major forms of HCS present in human cells that differ by 58 amino acids at the N‐terminus. In order to determine if these residues have any effect on HCS catalytic activity, both forms were recombinantly expressed, purified, and subjected to biochemical characterization.The oligomeric state and kinetic behavior of full length HCS and 58‐HCS were characterized. Equilibrium sedimentation experiments indicate that both HCS forms are monomers even in the presence of the intermediate biotinoyl‐5′‐adenylate. The steady state kinetic analysis as a function of biotin, ATP or biotin accepting substrate concentrations indicate that full length HCS and 58HCS behave similarly. Therefore the additional N‐terminal amino acids do not affect catalysis and are not involved in biotin acceptor fragment recognition. Furthermore, the results indicate that the distinct roles of HCS in transcription regulation and metabolism reflect properties other than the fundamental kinetic characteristics of the two enzyme forms.
A bifunctional enzyme of gluthathione‐dependent formaldehyde dehydrogenase and S‐nitrosogluthathione reductase from Antrodia camphorata
Authors: Wen Lisa , Huang Chih‐Yu , Ken Chuian‐Fu , Lin Chi‐Tsai 
Published: June 2021

Glutathione‐dependent formaldehyde dehydrogenase (GFD) plays important roles in formaldehyde detoxification and antioxidation. A gene encoding GFD from A. camphorate was identified based on sequence homology. The deduced amino acid sequence of 378 residues is conserved among the reported GFDs. To characterize the Ac‐GFD, the coding region was subcloned into a vector pET‐20b(+) and transformed into E. coli. The recombinant GFD was expressed and purified by Ni2+‐nitrilotriacetic acid Sepharose. This purified enzyme showed a single band on a 10 % SDS‐PAGE. The enzyme retained 50% GFD activity after heating at 50 o C for 5 min. The enzyme is bifunctional. In addition to the GFD activity, it also functions as an effective S‐nitrosoglutathione reductase (GSNOR) presumably to safeguard against nitrosative stress. The Km values for S‐hydroxymethylglutathione and S‐ nitrosoglutathione were 1.197 mM and 0.276 mM, respectively.Supported by the National Science Council of the Republic of China under grant NSC 96‐2313‐B‐019‐001 to C‐T. L.
18beta‐glycyrrhetinic acid prevents free fatty acid‐induced lipotoxicity by inhibiting ER stress and oxidative stress
Authors: Wu Xudong , Zha Weibin , Sun Lixin , Pecora Beth S , Cao Risheng , Gurley Emily C , Studer Elaine J , Hylemon Phillip B , Pandak William M , Zhang Luyong , Zhou Huiping 
Published: June 2021

The endoplasmic reticulum (ER) stress signal pathway has been linked to free fatty acid (FFA)‐induced inflammatory response and disease progression of nonalcoholic fatty liver disease in clinic. We have previously shown that 18?β‐glycyrrhetinic acid (GA) prevents FFA‐induced hepatic lipotoxicity by inhibiting cathepsin B expression and activity. But the underlying cellular mechanism remains unclear. This study determined if GA prevents FFA‐induced lipotoxicity by regulating ER stress signaling pathways.MethodsMouse macrophages, primary rat hepatocytes and Kupffer cells were used in this study. Cells were treated with palmitic acid (PA) and oleic acid (OA) in the presence or absence of GA. Activation of the UPR was determined by real time RT‐PCR and Western Blot analysis. The apoptosis was detected by fluorescence microscopy using Annexin V‐FITC/propidium iodide. Reactive oxygen species (ROS) was measured using dichlorofluorescein diacetate (DCFH‐DA).ResultsPA dose‐dependently induced UPR activation, ROS formation, and apoptosis both in macrophages and hepatocytes, while OA had no effect. GA not only inhibited PA‐induced UPR activation, but also inhibited PA‐induced ROS formation. PA‐induced ROS formation and cathepsin B expression were inhibited in CHOP knockout macrophages and hepatocytes.ConclusionGA prevents FFA‐induced lipotoxicity by inhibiting ER stress and oxidative stress.
Membrane penetration of the FYVE domain is modulated by pH
Authors: He Ju , Vora Mohsin , Filonov Grigory S. , Musselman Catherine A. , Verkhusha Vladislav V. , Stahelin Robert V. , Kutateladze Tatiana G. 
Published: June 2021

The FYVE domain (Fab1, YOTB, Vac1, EEA1) binds phosphatidylinositol 3‐phosphate [PtdIns(3)P] in membranes of early endosomes and penetrates bilayers. Here, we investigate the membrane anchoring mechanism by NMR, liposome binding, monolayer surface tension, mutagenesis, and in‐vivo localization experiment. Our results show that the extent of the FYVE domain insertion into PtdIns(3)P‐enriched membranes is substantially enhanced by acidic conditions and contacts with anionic lipids. The EEA1 FYVE domain associates with POPC/POPE/PtdIns(3)P vesicles with a Kd of 49 nM at pH 6.0, however binding is ~~24 fold weaker at pH 8.0. The decrease of the binding affinity is primarily due to much faster dissociation of the FYVE domain from membranes in basic media. The membrane penetration is modulated by the tandem His residues in the RRHHCRXCG signature motif, mutations of which abolish the pH‐sensitivity. The increased membrane association in vitro and in vivo by the Hrs, RUFY1, Vps27p and WDFY1 FYVE domains in a low pH environment demonstrates that pH‐dependency is a general function of the FYVE protein family.Supported by NIH grants GM071424 and CA113472 (TGK).
Structure of an activated G protein‐coupled receptor kinase reveals its receptor‐docking domain
Authors: Tesmer John JG , Singh Puja , Huang Chih‐chin 
Published: June 2021

G protein‐coupled receptor (GPCR) kinases (GRKs) play a key role in the desensitization of GPCRs by phosphorylating the cytoplasmic loops and tails of activated receptors. How GRKs recognize and are in turn activated by these receptors is not understood. High resolution crystal structures have been determined for GRK1, GRK2, and GRK6, but in each of these structures the kinase domain assumes an open, inactive conformation, and key structural elements believed to be involved in the recognition of GPCRs are missing or poorly ordered. We have now determined the crystal structure of a GRK bound to a nucleotide substrate analog wherein the kinase domain assumes a closed, catalytically active conformation. In this structure, missing structural elements have coalesced to form a receptor‐docking domain that explains how receptor binding promotes kinase activation. Based on this structure we can model a GRK‐GPCR complex, leading us to propose that GRKs and heterotrimeric G proteins recognize activated receptors via similar mechanisms.Support for this research was provided by NIH grants HL071818 and HL086865.
The blood cobalamin levels were associated with liver injuries and tumour progression: a prognostic indicator in patients with hepatocellular carcinoma
Authors: Kuo Chang Sheng , Lin Ching Yih , Lu Chin Li , Wu Meng Ying , Syu Huang Rwei Fen 
Published: June 2021

The aims of the study were to investigate the potential factors to influence serum vitamin B12 levels which may serve as a prognostic indicator in patients with hepatocellular carcinoma (HCC). Ninety patients with HCC and 90 healthy controls provided a blood specimen for B12 analysis. Demographic and disease‐related variables were previously recorded. Compared with the controls, HCC patients had a significantly higher level of mean serum B12 (1355 ± 1365 vs. 717 ± 589 pg/mL, respectively, P = 0.0001). There were significant correlations between serum B12, liver injuries (GOT, GPT and ALT levels, P < 0.05), alpha‐fetal protein levels (r = 0.382, P = 0.001), and tumour size (r = 0.300, P = 0.004). As patients were followed up for at least 30 months or until death, HCC cases in the highest terile of serum B12 levels (mean survival 13.5 months) had a significantly worse survival than those in the second (mean survival 25.0 months) and lowest tertile groups (mean survival 30.7 months; P for trend = 0.001). Thus, the data suggest that elevated blood vitamin B12 levels of HCC patients were associated with liver injuries and tumour progression. High B12 levels predict poor survival in patients with HCC.The study was supported by the grants (DOH95, 96‐TD‐F‐113‐008) to RF Huang.