bluuebunny/crossref_metadata_2025_split huggingface dataset:
Protective Role Of Antioxidant Supplementation In Tobacco Smoke Toxicity At The Blood‐Brain Barrier
Authors: Naik Pooja Shankar , Smith Quentin , Cucullo Luca 
Published: June 2021

Tobacco smoke (TS) is enriched in highly reactive oxidative species (e.g., hydrogen peroxide, peroxynitrate) which can damage the blood‐brain barrier (BBB) and trigger inflammatory events leading to the pathogenesis and progression of various neurological disorders. Physiological levels of antioxidants vitamin C and E are significantly reduced in smokers in fate of oxidative stress caused by TS exposure. We hypothesize that antioxidants can reduce the harmful effects of ROS‐dependent TS toxicity at the BBB.We exposed various in vitro BBB models to TS levels comparable to that found in chronic heavy smokers. Antioxidants supplementation of vitamin C and/or E at physiological concentration reduced TS‐induced vascular inflammatory response: release of proinflammatory cytokines from BBB endothelial cells and monocytes, release and activation of matrix metalloproteinases (MMPs), increased expression of vascular adhesion molecules. Furthermore, antioxidants reduced TS‐driven loss of BBB integrity and viability. Intravascular increase of nitric oxide (NO) by TS was also offset by antioxidants.Our results indicate that antioxidants can avert TS‐induced vascular pro‐inflammatory stimuli in vitro thus protecting the BBB. However, while antioxidants can reduce vascular oxidative stress, it might not be effective to counteract other teratogenic and pathophysiological stimuli induced by smoking.This work was supported by NIH/NIDA R01‐DA029121‐01A1 and Alternative Research Development Foundation (ARDF) to Dr. Luca Cucullo.
Behavior and memory evaluation in Wistar rats with hypothalamus arcuate nucleus lesion
Authors: Guimarães Ernesto da Silveira Goulart , de Caires Luiz Carlos , Musso Camila Manso , Vasconcellos Rebecca , Assis Diego , Andreazzi Ana Eliza , Mourão-Júnior Carlos Alberto , Garcia Raúl Marcel González 
Published: June 2021

The administration of L‐monosodium glutamate (MSG) in neonatal rats causes lesions in the arcuate nucleus of the hypothalamus leading to obesity development. Because of the many connections between these regions with important neuronal regulating centers, the aim of this study was to evaluate the behavioral and memory patterns of these animals.At the first 5 days of life, male Wistar rats received subcutaneous doses of MSG. At 90 days of life the animals was submit to the open field arena test regarding episodes of rearing, freezing, number of traversed squares and the time spent in the arena's center. We evaluated the animals’ memory using the object recognition test regarding the working memory (WM) and the long‐term memory (LTM). The Lee Index (LI) was calculated and the perigonadal fat pad was weighed to check the obesity and hypertrophy of adipocytes. The MSG rats showed a significant increase in the episodes of rearing, freezing and decrease in the number of squares traversed and in the time spent in the arena's center. In the LTM test MSG animals showed a reduced discrimination index, the WM test showed no significant difference. The MSG group had an increased LI and perigonadal fat pad mass.MSG obese rats showed a behavior that indicates a stress and anxiety profile, which may be due to the arcuate nucleus lesion. The impaired LTM can be explained by the some neural connections between the hypothalamus and hippocampus.Financial Support: Capes, CNPq, Fapemig and UFJF
Klotho Enhances Glucose‐induced Insulin Secretion by Upregulating Plasma Membrane Retention of TRPV2
Authors: Lin Yi , Sun Zhongjie 
Published: June 2021

BackgroundKlotho is a recently discovered anti‐aging gene. Our recent studies indicated that Klotho gene is expressed in mouse pancreatic islets and in an insulinoma cell line (MIN6 β cells) as evidenced by expression of Klotho mRNA and protein. The purpose of this study was to investigate if Klotho is involved in the regulation of insulin secretion in β cells.ResultsOverexpression of mouse Klotho protein significantly increased plasma membrane retention of TRPV2 (transient receptor potential V2), calcium entry, the glucose‐induced increase in intracellular calcium, and the glucose‐induced insulin secretion in MIN6 β cells. On the other hand, knockdown of Klotho by siRNA significantly decreased plasma membrane retention of TRPV2 and attenuated glucose‐induced calcium entry and insulin secretion. Tranilast, a selective inhibitor of TRPV2, abolished the promoting effects of overexpression of Klotho on glucose‐induced calcium entry and insulin secretion in MIN6 cells. Thus, TRPV2 lies in the downstream of Klotho in the regulation of glucose‐induced insulin secretion.ConclusionThe present study demonstrated, for the first time, that Klotho may increase glucose‐induced insulin secretion by up‐regulating membrane retention of TRVP2 and thus glucose‐induced calcium response. The findings reveal a previously unidentified role of Klotho in the regulation of insulin secretion in MIN6 β cells.
How does Parkinson disease severity influence kinetics and kinematics during a rapid force production task?
Authors: Willis Ryan , Dibble Leland E. , Rockwell W. Tyler , Marcus Robin L. , LaStayo Paul C. , Foreman K. Bo 
Published: June 2021

The cardinal signs of Parkinson disease (PD) include bradykinesia, hypokinesia, tremor, and rigidity. These symptoms worsen as the disease progresses, often resulting in postural instability. However, little attention has been given to the effects of disease severity on the kinetics and kinematics of individual lower extremity joints during rapid force production tasks necessary for balance responses. Therefore, we examined lower extremity joint moments and average angular velocity of 14 subjects with PD who were segregated into 2 groups (less severe [n=7, mean age=62.29±6.73] and more severe [n=7, mean age=73.00±6.19]) based on UPDRS motor subsection scores while ON medication (less severe=7.14±2.19, more severe=17.43±6.75). Subjects completed 4–5 counter movement jumps (CMJ) 1 hour after taking their medication. Kinetic and kinematic variables were recorded for the explosive phase of the CMJ. Subject's first 3 successful trials were averaged and examined using the Mann‐Whitney U Test (p≤0.05). Individuals with less severe involvement move significantly faster; however, their ability to produce torque around the hip, knee, and ankle is not significantly different compared to more severely affected individuals.Grant Funding Source: NIH Grant #: 1 R15 HD056478‐01
A Novel Anti‐apoptotic Role of DMP1 in Normal and Pathological Conditions
Authors: Rangiani Afsaneh , Sun Yao , Nowar Nour , Sahfiey Alex , Yuan Baozhi , Qin Chunlin , Kuro-o Makoto , Feng Jian Q 
Published: June 2021

IntroductionDeletion or mutations in Dmp1, expressed in both hard and soft tissues, leads to an increase of FGF23 and hypophosphatemic rickets. Deletion of FGF23 or Klotho (co‐receptor of FGF23) leads to a chronic kidney diseases like phenotype.PurposeTo test the hypothesis that DMP1 plays a protective role in renal osteodystrophy caused by deletion of Klotho.MethodsDmp1 KO, Klotho KO, Dmp1‐Klotho double‐KO (dKO) mice were generated, sacrificed and compared to WT at 6 wk; x‐ray, uCT, SEM, serum biochemistry, histology, TRAP and TUNEL methods were used for phenotype analyses.ResultsdKO mice displayed an identical serum biochemical phenotype (high Pi/FGF23, low PTH) and aging as those in single Klotho KO, with unexpected massive ectopic calcification in the dental pulp/root canal, aorta and kidney, plus a substantial increase in apoptosis (P<0.001, compared to single Klotho KO). These pathological changes are completely different from either Dmp1 KO (low in apoptosis with normal life span and no ectopic ossification) or Klotho KO (a moderate increase in apoptosis and ectopic ossification in soft tissue).ConclusionDMP1 has a possible protective role in conditions such as the renal osteodystrophy. In addition, DMP1 appears to have a novel protective role for odontoblasts, osteocytes and endothelial cells of aorta and kidney in the pro‐apoptotic conditions such as the renal osteodystrophy.Grant Funding Source: NIH grants to JQF for DEDE018486015209
Changes in semicircular canal morphology in response to selective breeding for high voluntary wheel running
Authors: Schutz Heidi , Jamniczky Heather A. , Hallgrímsson Benedikt , Garland Theodore 
Published: June 2021

Variation in semicircular canal (SCC) radius of curvature (R) and shape show correlations with locomotor agility among species of mammals. We used laboratory mice from 4 replicate lines bred for high voluntary wheel running (HR) and 4 non‐selected control lines (C) to examine responses in the SCC to 8 weeks of access to wheels and 21 generations of selection. Mouse crania were μCT scanned at 21 μm resolution and linear measurements, 3D landmarks, and centroid sizes of the SCC were taken. Linear measures were used to calculate R for all three canals and their mean, whereas landmarks were used to generate multivariate descriptors of 3D canal shape.HR mice weighed less than C and wheel access reduced body mass for both groups. ANCOVA showed that body mass was a significant positive predictor of R. However, when controlling for body mass we found no differences in R between HR and C, no effect of wheel access, and no interaction.SCC shape was also significantly correlated with body mass; controlling for body mass or centroid size, there were significant differences in SCC shape between HR and C mice. These results demonstrate that semicircular canal morphology is responsive to selection for locomotor activity and that canal shape may be more sensitive than canal size.Funding for this project provided by a U.C. Riverside Chancellor's Postdoctoral Fellowship to HS, Canadian Foundation for Innovation to BH and IOS‐1121273 to TG.
An active site mutant allele of human Pol ε is a mutator in vitro and in vivo
Authors: Agbor Anderson Ayuk , Pursell Zachary F 
Published: June 2021

DNA replication in eukaryotes is carried out primarily by three DNA polymerases (Pols): Pols α, δ and ε. While yeast Pol ε has been shown to function on the leading strand during DNA replication, the precise functions of human Pol ε in vivo remain relatively poorly understood.Wild type pol ε from both yeast and humans is a highly accurate enzyme. Active site mutant alleles of yeast Pol ε were previously shown to be mutators with unique error signatures both in vitro and in vivo. We found that human Exo− M630G Pol ε is a 50‐fold mutator over the wild type enzyme in vitro, with a strong preference for T•dT and T•dG errors.We introduced Exo− M630G Pol ε into human mismatch repair‐deficient HCT116 colorectal cancer cells. Mutations in this cell line arise primarily from errors made during replication. Cells transfected with Exo− M630G Pol ε exhibited an increase in HPRT1 mutant frequency over cells transfected with control vector. The results indicate that transfected Exo− M630G Pol ε is involved in replication, is a mutator during replication in vivo, and is at least partially dominant over the endogenous Exo+ Pol ε. Importantly, Exo− M630G Pol ε mutant lacking the C‐terminal tail, which is known to be essential in yeast, showed no increase in HPRT1 mutant frequency.With the human Exo‐ M630G Pol ε mutator allele, we now have the ability to probe Pol ε function during human DNA replication and repair of damaged DNA.NIH ES016780, NIH RR020152
Cell specific processing and release of the pro‐hormone candidate tumor suppressor, Ecrg4, from the epithelial cell surface
Authors: Dang Xitong , Podvin Sonia , Coimbra Raul , Eliceiri Brian , Baird Andrew 
Published: June 2021

ObjectiveTo study the processing of human Ecrg4 in epithelial cells.MethodsPlasmids encoding human Ecrg4, Ecrg4 mutants and Ecrg4‐derived peptides fused with green fluorescent protein were transiently expressed in the prostate cancer (PC‐3) or human embryonic kidney (HEK) epithelial cells. The expression of Ecrg4 was analyzed by immunoblotting, immunohistochemistry and fluorescence and confocal microscopy. Cell surface‐bound Ecrg4 was detected by cell surface fractionation.ResultsEcrg4 can be detected in the conditioned media of PC‐3 but not HEK cells 30 hours after transient transfection of pcDNA3‐hEcrg4. In both HEK and PC3 cells however, a “secreted” 14 kDa Ecrg4 product was found tethered to the outer cell surface and could not be released by washing cells with neutral (PBS), high salt (2M NaCl), acidic (Glycine, pH 2.8) or basic (Na2CO3, pH 11) buffers. Fusion of leader sequences with GFP established that Ecrg4 retention was mediated by amino acids 1–15 of the 30 amino acid leader sequence while Ecrg4(16–30) promoted retention. Mutagenesis studies further established that processing occurred at predicted furin and thrombin consensus sites at the cell surface.ConclusionEcrg4 is cell membrane protein that can be shed into media by cell stimulation. This surface bound ECRG4 can be further processed at the furin site to release the ligand‐like peptides in conditioned media.
Complex Peptide Biosensors for Detection of Intracellular Kinase Biomarkers
Authors: Morris May C , Kurzawa Laetitia , Van Ngoc , Pellerano Morgan 
Published: June 2021

Cyclin‐dependant kinases play a central role in coordinating cell cycle progression, and in sustaining proliferation of cancer cells, thereby constituting attractive pharmacological targets. We have developed a family of environmentally‐sensitive fluorescent peptide biosensors to probe the relative abundance and activities of CDK/cyclin kinases. CDKSENS and CDKACT sensors allow to probe CDK/Cyclin kinases in a sensitive and specific fashion in vitro. Complexation of these peptide biosensors with cell‐penetrating peptide carriers allows to form stable nanoparticles that can probe these kinases in living cells in a non‐invasive fashion (Kurzawa et al. PloS One 2011). This technology can be applied to monitor subtle alterations between healthy and cancer cell lines, to probe kinase levels and activities in tumour biopsies, and to monitor response to therapeutics in vitro and in vivo.Source of Funding: Grants to MCM “Chercheuse d'Avenir” Région Languedoc‐Roussillon and INCA
Nutlin‐3 activates MEK1/2‐ERK1/2 pathway via p53‐induced ROS accumulation
Authors: Lee Sun-Young , Ko Kyoung-Won , Choe Yun-Jeong , Kim Ho-Shik 
Published: June 2021

Tumor suppressor p53 activates growth arrest or apoptosis in response to genotoxic stress. Along with its growth suppressive effect, p53 was also known to stimulate MAPK pathway, of which mechanisms still remains to elusive. In human osteosarcoma U2OS cells, nutlin‐3, a small molecular antagonist of MDM2, induced phosphorylation of MEK1/2 and ERK1/2 accompanied by p53 activation, which was completely inhibited by p53 siRNA. Nutlin‐ 3 induced accumulation of ROS and subsequently, TEMPO, a ROS scavenger, prevented the phosphorylation of MEK1/2 and ERK1/2. Moreover, pifithrin‐μwhich blocks mitochondrial localization of p53 prevented ROS accumulation and MEK1/2‐ ERK1/2 phosphorylation as well. Knockdown of MEK1/2 and ERK1/2 activity using chemical inhibitors or small interfering RNA enhanced cell death induced by nutlin‐3. Collectively, it can be concluded that nutlin‐3 activates MEK1/2‐ERK1/2 via ROS generated by mitochondrial localization of p53 activation and nutlin‐3‐induced ERK1/2 activation may play a role in protecting U20S cells from p53‐dependent apoptosis, constituting negative feedback loop of p53.This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2011‐ 0027115).
Construction of non‐visual arrestins with enhanced specificity for individual G protein‐coupled receptors
Authors: Gimenez Luis E , Gurevich Vsevolod V 
Published: June 2021

We identified elements that drive arrestin‐3 (Arr3) binding to G protein coupled receptors. Based on the identification of residues that determine arrestin‐1 and ‐2 selectivity for cognate receptors and the analysis of the evolution in the arrestin family, we mutated “receptor discriminator” residues in Arr3. The recruitment of these mutants to different receptors was assessed using BRET‐based assays in cells. D260K and Q262P showed enhanced preference for M2 muscarinic (M2R), D1 (D1R), and D2 (D2R) dopamine receptors, over β2‐adrenergic receptors (β2AR). The specificity was further enhanced by combining D260K and Q262P. Conversely, Y239T enhanced Arr3 binding to β2AR, while reducing the binding to M2R, D1R, and D2R. Mutation Q257Y reduced arrestin recruitment to D2R. The Y239T+Q257Y combination virtually eliminated binding to β2AR, reduced D2R binding, and enhanced Arr3 recruitment to D1R. Thus, Y239T+Q257Y had opposite effects on Arr3 binding to D1R and D2R. Thus, manipulation of key residues on the receptor‐binding surface modifies receptor preference, allowing for the construction of non‐visual arrestins specific for particular GPCR subtypes. These findings pave the way to the construction of signaling‐biased arrestins targeting the receptor of choice for research or therapeutic purposes.NIH grants GM081756, GM077561 and EY011500 (VVG).
Non‐visual arrestins function as simple scaffolds assembling the MKK4–JNK3a2 signaling complex
Authors: Zhan Xuanzhi , Kaoud Tamer S , Dalby Kevin N , Gurevich Vsevolod V 
Published: June 2021

Arrestins are a small family of proteins with four mammalian members that play key roles in the regulation of multiple GPCR‐dependent and ‐independent signaling pathways. Although arrestins were reported to serve as scaffolds for MAP kinase cascades, promoting the activation of JNK3, ERK1/2, and p38, the molecular mechanisms involved were not elucidated and even the direct binding of arrestins with MAP kinases were never demonstrated. Here using purified proteins we show that both non‐visual arrestins directly bind JNK3α2 and its upstream activator MKK4, and that the affinity of arrestin‐3 for these kinases is higher than that of arrestin‐2. Reconstitution of the MKK4‐JNK3α2 signaling module from pure proteins in the presence of different arrestin‐3 concentrations showed that arrestin‐3 acts as a “true” scaffold, facilitating JNK3α2 phosphorylation by bringing the two kinases together. Both JNK3α2 phosphorylation by MKK4 and JNK3α2 activity towards its substrate ATF2 increase at low and then decrease at high arrestin‐3 levels, yielding bell‐shaped concentration dependence expected with true scaffolds that do not activate the upstream kinase or its substrate. Thus, direct binding of both kinases and true scaffolding is the molecular mechanism of arrestin‐3 action on the MKK4‐JNK3α2 signaling module.NIH grants GM081756, GM077561, EY011500 (VVG), GM059802 and Welch Foundation (F‐1390) (KND).
Structural and functional studies of plant glucan phosphatases
Authors: Husodo Satrio , Gentry Matthew S. 
Published: June 2021

Glucan phosphatases (GPs) dephosphorylate starch to regulate degradation. Historically, GPs have been defined as proteins containing a dual specificity phosphatase (DSP) domain and a carbohydrate binding module (CBM). To date there are two published GPs: Starch‐Excess‐4 (SEX4) and Like‐SEX4‐1 (LSF1). Our lab recently determined the structure of SEX4. These data identified a previously unknown domain in the C‐terminus of GPs, the so‐called CT‐domain. The discovery of this domain allowed us to identify a third member of the GP family, LSF2. LSF2 contains both a DSP and CT domain, but lacks a CBM. We cloned LSF2, purified recombinant protein, and demonstrated that it has glucan phosphatase activity even though it lacks a CBM. Furthermore, SEX4 and LSF2 possess different specificities: SEX4 dephosphorylates starch preferentially at the C6 position of glucose, while LSF2 acts on the C3 position. The function of LSF1 is still unknown as it lacks phosphatase activity thus far. We are performing crystallization screenings on LSF1 and LSF2 to obtain their structures, and we are using molecular modeling of LSF1 and LSF2 to determine key residues involved in glucan binding. In addition, we aligned the known orthologs of each gene and mapped the degree of conservation on the structures to infer critical regions for GP activity. Cumulatively, these results allow us to determine the molecular mechanism of this class of enzymes.Supported by NIH grants R00NS061803, P20RR020171, and R01NS070899, and University of Kentucky College of Medicine startup funds to M.S.G.
Tuberculosis and Epigenetics: Functional Noncoding RNA in the Host‐Pathogen Interaction
Authors: Wissler Josef H 
Published: June 2021

OBJECTIVENoncoding ncRNAs, the main RNA transcript fraction, were investigated from tuberculin‐activated mononuclear cells [MMC] for functions in host‐pathogen interactions and phenotype changes upon transfer of the tuberculin reaction as a feature of transmissible adaptive cell‐mediated immunity.METHODSAnn. N. Y. Acad. Sci. 1022 : 163–184, 2004; 1137 : 316–342, 2008; OWWZ 2010: http://www.owwz.de/868.html?&L=1RESULTSThe MMC activation by purified protein derivative tuberculin [PPD] and assays in use represent in vitro correlatives of delayed‐type [−IV] hypersensitivity reactions including Koch‐Metchnikoff‐ and Jones‐Mote types. These concern induration, swelling and endothelial infiltration of memory MMC into skin lesion sites within 24 to 72 hours. PPD‐activated cells form ncRNA. Some modified, edited, redox‐ and metalloregulated functional small hairpin fncRNA [<200n] were isolated and sequenced. Modified nucleosides are e.g. isoguanosine / crotonoside, adenosine‐N1‐oxide of 151Da base families. fncRNAs regulate phenotypic adaptation by complementary interaction with defined conserved homologous helix‐nucleating Mendelian consensus domains shared in epigenetic regulator proteins entangled in tolerated growth, vascularization, metabolic syndromes, cancer, epi‐ and genetic information indexing of the epigenome.CONCLUSIONSfncRNA from PPD‐activated cells are considered useful for better evaluations of test systems of infectious diseases, nuclear antibodies, aberrant histone codes in pathogenesis of autoimmune disease [e.g. SLE] and tolerance of biomaterials. Their features suggest that immune cells beside known protein recognition systems, may be armed by additional arsenals of recognition entities in terms of found fncRNAs
Measured vs. Estimated Resting Energy Expenditure in a Clinic Population of Obese Youth
Authors: Henes Sarah , Hickner Robert , Cummings Doyle , Houmard Joseph , Kolasa Kathryn , Lazorick Suzanne , Collier David 
Published: June 2021

The objective of this study was to compare measured resting energy expenditure (MREE) using portable indirect calorimetry in obese youth with that estimated from six published predictive equations used by clinicians to determine caloric targets for weight loss.MREE was measured in fasting obese youth who were referred to a multidisciplinary Healthy Weight Clinic (n=80, mean age 12.18 yrs ± 0.31 SE, mean BMI 37.39 ± 1.07 SE, mean BMI z score 2.47± 0.04 SE). The mean caloric difference between MREE and each predictive equation was calculated. The Harris Benedict equation demonstrated the lowest mean caloric difference (197.94 kcal/d ± 23.26 SE) compared to MREE. While there was a strong correlation (p < 0.05) between MREE and estimations from each of the predictive equations, there was substantial variability between estimations and discrepancies by gender. Mean measured and estimated caloric differences were greater for males (634.12 kcal/d± 69.45 SE) than for females (254.51 kcal/d± 25.92 SE)(p<0.05). No significant racial differences were evident. These data demonstrate the inconsistent findings among predictive equations and suggest a stronger role for portable indirect calorimetry with obese youth in the clinical setting. When indirect calorimetry is not available, the Harris Benedict equation may be the best predictive equation to use in obese youth aged 7–18 who are >99th percentile for age and gender.