bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tempol Treatment Inhibits Inflammatory Process and Improves Morphological Aspects in Early and Late‐stage of Prostate Cancer Progression in the Ventral Lobe of TRAMP Model.
Authors: Rossetto Isabela Maria Urra , Kido Larissa Akemi , Santos Felipe Rabelo , Montico Fabio , de Almeida Lamas Celina , Cagnon Valéria Helena Alves 
Published: April 2020

Tempol (4‐hydroxy‐TEMPO) is a SOD mimetic and member of the nitroxide compounds family. It is used as a co‐adjuvant or protective factor in cancer treatment due to its anti‐oxidant properties. However, in prostate cancer (PCa) Tempol effects are still poorly understood. Its known that PCa progression requires a sustained inflammation, which generates a broad spectrum of reactive nitrogen and oxygen species, leading to uncontrolled cellular proliferation. We aimed to evaluate the Tempol effects in different stages of PCa progression (a) in vivo using a transgenic adenocarcinoma of the mouse prostate model (TRAMP); (b) and in vitro using human PCa cell lines with different genetic background. For the early‐stage analysis, 8‐weeks‐old mice (TPL1 group) were treated orally with 50 mg/kg of Tempol during 4 weeks. For the late‐stage analysis, 16‐weeks‐old mice were treated with 50mg/Kg (TPL2 group) or 100mg/Kg (TPL3 group) of Tempol during the same period. All control groups received water as vehicle. The ventral lobe of the prostate was collected and submitted to the histopathological, immunohistochemical and Western blotting analysis. PC‐3 and LnCaP cells were exposed to different doses of Tempol (0.5, 1.0 and 2.0 mM) and cell viability assay was performed. In vivo, Tempol treatment promoted a great improvement in the prostatic epithelium in all the analyzed cancer stages, reducing significantly premalignant (high‐grade prostatic intraepithelial neoplasia) and malignant lesions (well differentiated prostatic adenocarcinoma). The PCNA immunostaining confirmed the cellular proliferation decrease between the control and treated groups. Tempol also reduced COX‐2 and NFkB, especially in the early‐stage, whereas iNOS increased in this period. In the late‐stage, both doses of Tempol reduced NFkB levels concomitant with IkB‐α increase, suggesting an inhibitory effect of Tempol on NFkB pathway in association with its anti‐oxidant role. In vitro, Tempol decreased cell viability of both cell lines, being the 1.0 mM dose the most efficient and nontoxic concentration. In conclusion, Tempol had a remarkable action in the prostate, delaying cancer progression in the ventral lobe of TRAMP model and inhibiting PC‐3 and LnCaP growth. This SOD mimetic presented great capability of inhibiting the inflammation process, and consequently the cell proliferation. Its action was effective in different stages of PCa progression, particularly in the early stage, indicating that Tempol treatment can be explored as a preventive approach. Finally, for the late PCa stage, Tempol may be considered as an interesting adjuvant therapy.Support or Funding InformationSão Paulo Research Foundation, FAPESP, process 2018/21647‐6 and FAEPEX/UNICAMP, process 2202/19
Targeted proteomics analysis revealed the imbalanced allele‐specific protein expression of carboxylesterase1 in human livers
Authors: Her Lucy , Shi Jian , Wang Xinwen , Zhu Hao-Jie 
Published: April 2020

Carboxylesterase 1 (CES1) is the primary hepatic hydrolase and is responsible for metabolizing many medications. Its clinical significance has been well demonstrated in various clinical trials with methylphenidate, clopidogrel, oseltamivir, and ACE inhibitors. CES1 expression varies markedly among individuals, which is a major contributing factor to the interindividual variability in the pharmacokinetics and pharmacodynamics of CES1 substrate drugs. Identifying genetic variants responsible for CES1 expression regulation could lead to the optimization of pharmacotherapy of medications metabolized by CES1. The analysis of allele‐specific expression is a powerful tool to determine regulatory genetic variants because this assay is less susceptible to the confounding effect of nongenetic factors (e.g. inducers) compared to those conventional gene expression‐based approaches (e.g. GWAS). In the present study, we developed a targeted proteomics method to measure the allele‐specific protein expression (ASPE) of CES1 in human livers using a heavy stable isotope‐labeled QconCAT internal standard. The CES1 nonsynonymous variant S75N (rs2307240) was utilized to differentiate the protein expressions of the two CES1 alleles. The QconCAT construct contained the sequences of both the wild type peptide (FTPPQPAEPW
SFVK) and its S75N variant (FTPPQPAEPW
NFVK), which ensures that the amounts of the two peptides released from the QconCAT internal standard are equal. Another essential feature of the QconCAT protein is that the 75S and 75N peptides are flanked with several native CES1 amino acids, such that the digestion efficiency of the QconCAT peptides will be the same of the native CES1. The method was successfully applied to the determination of the ASPE of CES1 in 32 S75N heterozygous liver samples. The ASPE ratios of 75S to 75N ranged from 0.80 to 1.47, indicating the presence of cis‐regulatory genetic variants of the CES1 gene. Further studies have been carried out to determine the variants responsible for the imbalanced ASPE of CES1 in human livers.Support or Funding InformationThis work was supported in part by the National Heart, Lung, and Blood Institute [Grant R01HL126969]
Chronic Alcohol Drinking Increases L‐Type Voltage‐Gated Calcium Channel Cav1.2 in the Substantia Nigra
Authors: Rodriguez Eric A. , Yamamoto Bryan K 
Published: April 2020

Studies report that 77 % of methamphetamine (Meth)‐dependent adults in the United States also have alcohol use disorder. This comorbidity is associated with negative cognitive and behavioral effects. In addition, the serial exposure of alcohol and then methamphetamine (Meth) to adult male Sprague Dawley rats is neurotoxic and marked by a loss of tyrosine hydroxylase‐positive dopamine (DA) cells in the substantia nigra pars compacta (SNpc) that is absent after either drug alone. Dopamine cell loss was associated with caspase‐3 mediated apoptosis after Meth exposure to alcohol drinking rats; however, the effect of alcohol that sensitizes DA cells to Meth‐induced apoptosis remains unknown. Adult male Sprague Dawley rats had access to either two bottles of water or a choice of 10% alcohol and water every other day over 28 days. Rats with access to alcohol every other day had significantly increased L‐type voltage gated calcium channel Cav1.2 protein in the substantia nigra (SN) (33 ± 1.4 %) at 1 day after the last day of alcohol drinking compared to water drinking only controls (p<0.05). The increases in Cav1.2 were region‐specific since no such changes were detected in tissue from other dopaminergic regions including the dorsal striatum, ventral tegmental area, and nucleus accumbens. Furthermore, the cyclooxygenase‐2 selective and prostaglandin synthesis inhibitor, nimesulide (7.5mg/kg i.p., b.i.d.) administered on days when alcohol was not available, blocked the increases in Cav1.2 at 1 day after the alcohol drinking paradigm. Nimesulide administered only during the alcohol drinking paradigm also blocked the loss of DA cells in the SN pars compacta measured 1 week after exposure of Meth (10 mg/kg q 2 hrs X 4 injections). Although previous studies by others showed chronic alcohol exposure increases Cav1.2 in other brain regions, this is the first evidence of increases in the SN and that the increases occur in a prostaglandin synthesis‐dependent manner. The results are consistent with the excitotoxic effects produced calcium dysregulation and suggest that prostaglandins and disruption of calcium homeostasis produced by voluntary alcohol drinking sensitize DA cells in the SN to the neurodegenerative effects produced by the serial exposure to alcohol and Meth.Support or Funding InformationThis work is supported by the National Institutes of Health [DA042737].
Function of Mte1/ZGRF1 in Homologous Recombination
Authors: Durham Jesse 
Published: April 2020

Fanconi anemia (FA) is a human genetic disorder that causes organ defects, physical abnormalities, bone marrow failure, and increased risk of malignancy. These phenotypes result from an impaired response to DNA damage due to the dysfunction of a cluster of FA proteins. Among these, human FANCM (budding yeast ortholog, Mph1) functions in DNA replication fork (RF) repair and DNA double‐strand break (DSB) repair by homologous recombination (HR). In a recent study, Mte1 in Saccharomyces cerevisiae was found to protect the Rad51‐made D‐loop, essential for HR, from disassociation by Mph1 (Xue et al. 2016). Our preliminary data showed that Mte1 interacts with Rad51 in vitro. However, the biological role of this interaction remains unknown. The goal of this project is to examine the effects of 2 different binding site mutants of Mte1, where mutations are hypothesized to affect binding of Rad51. Quick change site directed mutagenesis will be used to create the 3 mutants by changing 2 different phenylalanine side groups to alanine at positions 156, 169, and 156/169 combined. The Rad51 interaction of each mutant will be evaluated via in vitro pull down assay. The D‐loop binding activity of each mutant will be evaluated via electrophoresis mobility shift assays (EMSA). Mutation effects on Rad51‐made D‐loop formations can direct further studies investigating proteins required for HR. Evidence suggest that human ZGRF1 is functionally related to Mte1 due to N‐terminus conservation between the two. However, little is known about the role and function that ZGRF1 plays in HR in humans.Support or Funding InformationResearch supported by NIH R21 #ES028792 awarded to XX. JD supported by NIH R25 Bridge to Doctorate Program (GM102783)
Developing a Blood Test to Support Treatment of Surgically Induced Type I Diabetes
Authors: Dailey Allyson , Kanak Mazhar , Couch Robin 
Published: April 2020

In this pilot‐scale investigation, serum was collected from nine study participants, each of which having underwent a procedure of total pancreatectomy with islet autotransplantation. The procedure was a success with 6 of the patients (i.e. they are not dependent on insulin injections), whereas the insulin‐producing islet cells did not successfully autotransplant in 3 of the patients (and thus they are dependent on insulin injections). Each of the nine patients have provided four serum samples; one obtained prior to surgery, one immediately following surgery, one 1 hr following islet autotransplantation, and one 7 days following islet autotransplantation. Following organic extraction, the serum extracts were analyzed for metabolite content using liquid chromatography quadrupole time of flight mass spectrometry (LC‐QToF). A differential comparison of the serum metabolites present in each of the samples was used to define molecular features that may serve as biomarkers of patient outcome. Subsequent tandem mass spectrometry (LC‐MS/MS), along with comparison to molecular libraries and purchased authentic standards, will identify the molecular features, and permit an assessment of their suitability for use in a predictive diagnostic test to determine patient suitability/compatibility with islet autotransplantation.Support or Funding Information4‐VA Funds
Effects of diet and torpor on skeletal muscle relaxation in hibernating animals
Authors: Krishnan Jishnu K.S. , Rice Sarah , Mikes Monica , Hunstiger Moriah , Drew Kelly , Oliver S. Ryan 
Published: April 2020

Arctic ground squirrels (AGS) undergo extreme temperature fluctuations during the hibernation season (body temperature, Tb, as low as −3 °C), where they mostly remain physically inactive. During interbout arousals, once Tb reaches ~16°C, hibernators recruit skeletal muscle (SkM) for shivering thermogenesis to attain Tb of ~37°C. Dietary polyunsaturated fatty acids (PUFA) are known to affect both SkM function and metabolism. Recent studies in cardiac muscle of hibernators have shown an increased level of ω‐6 in ω‐6:ω‐3 PUFA ratio are correlated to sarco/endoplasmic reticulum Ca‐ATPase (SERCA) activity with respect to torpor and hibernation status. SERCA catalyzes the re‐uptake of Ca2+ back into the sarcoplasmic reticulum (SR), which was previously bound to actin/myosin filaments thus completing the calcium cycle and eliciting muscle relaxation. We hypothesized that diet (increased ω‐6:ω‐3 PUFA ratios) and torpor status play essential roles in the regulation of the SERCA pump, which may improve SkM performance during hibernation. An Ex‐vivo functional assay was used to characterize performance alterations in SkM (diaphragm) collected from AGS fed the following diets (ω‐6:ω‐3, 5:1 control diet versus ω‐6:ω‐3, 1:1 test diet) and collected at 3 timepoints of torpor (early torpor, late torpor, and arousal). Additionally, the muscles were assessed for their response to hypothermic temperature stress (4 °C; 15 °C; 25 °C and 37 °C) to determine functional resilience. To our knowledge this is the first study correlating torpor status, diet, and effects of hypothermic temperature stress on SkM functional performance in hibernators (AGS). Data suggest: 1) diet and torpor significantly alter relaxation kinetics (calcium re‐absorption), but not force development (rate of calcium release), and 2) diet, torpor status, and inherent temperature resilience (4°C; 15°C) affect the muscles ability to produce force. This evidence supports the interpretation that functional properties of SkM can be altered by membrane lipid composition, ambient temperature, and hibernation status to control performance.Support or Funding InformationThis work was supported by the INBRE IDeA Faculty Pilot Grant (University of Alaska Fairbanks ) from NIH NIGMS grant number P20GM103395.
In Vitro Reconstitution of Centralspindlin‐Mediated Pebble/RhoGEF Transport
Authors: Warecki Brandt , Tao Li 
Published: April 2020

Cytokinesis is the last step of cell division. Failure in regulation of cytokinesis is the cause of several human diseases, including cancer. Appropriate formation of the cleavage furrow during cytokinesis requires accurate regulation of the small GTPase Rho. Multiple experiments have demonstrated that Rho activity is modulated at the central spindle by RhoGEF (Pebble in Drosophila) and the conserved motor complex centralspindlin. However, perhaps due to the necessity of these proteins in in vivo systems, the precise mechanism for how centralspindlin and RhoGEF cooperate to activate Rho at the central spindle remains poorly understood. Here, we use the baculovirus expression system to generate recombinant Drosophila versions of the centralspindlin components Pavarotti/Kinesin‐6 and Tumbleweed/RacGAP as well as Pebble/RhoGEF. Using these purified proteins, we reconstitute key events in cytokinesis and test models for Rho activation that have been based on in vivo studies. We observe Pavarotti/Kinesin‐6‐mediated microtubule bundling and show that the recombinant centralspindlin holoenzyme recruits Pebble/RhoGEF to microtubules in vitro. Furthermore, we demonstrate that centralspindlin transports Pebble/RhoGEF along microtubules in a plus‐end directed fashion. Taken together, our studies support a model in which centralspindlin motor activity delivers Pebble/RhoGEF to the central spindle where Pebble/RhoGEF activates Rho, leading to cleavage furrow formation and proper cytokinesis.Support or Funding InformationNIH INBRE P20GM103466‐17
Exercise training upregulates Dicer to modulate hippocampal miRNA expression in 5xFAD mice
Authors: Dungan Cory M. , Valentino Taylor R. , Vechetti Ivan J. , Zdunek Christopher J. , Bachstetter Adam D. , Lin Ai-Ling , McCarthy John J. , Peterson Charlotte A. 
Published: April 2020

Changes to cerebral miRNA expression have been implicated in the progression of Alzheimer’s disease (AD), as miRNAs that regulate the expression of genes involved in amyloid beta (Aβ) processing are dysregulated in those that suffer from AD. Exercise training improves cognitive function and reduces Aβ plaque burden, however, the mechanisms are not fully understood. Using our progressive weighted wheel running (PoWeR) exercise program, which utilizes an unbalanced running wheel to promote physiological adaptations similar to those seen in humans, we assessed the effect of 20 weeks of voluntary exercise training on changes in cognitive function and hippocampal miRNA expression in female 5xFAD mice; a model that expresses human APP and PSEN1 transgenes with a total of five AD‐linked mutations. Two month old female 5xFAD mice were PoWeR trained for 20 weeks, running ~7km/day, while groups of female wild‐type (B6SJLF1/J) and 5xFAD served as sedentary controls. At the end of the training period, mice performed cognitive (Morris water maze, novel object recognition) and physical (rotor rod and balance beam) function tests. The hippocampus was removed to assess miRNA abundance via Nanostring and gene expression with RT‐qPCR. Exercise training improved cognitive and physical function, in addition to having a significant effect on the expression of the miRNA processing enzyme, Dicer. Specifically, Dicer was lower in sedentary 5xFAD hippocampus, when compared to sedentary WT mice, but was robustly elevated following exercise. Correspondingly, ~45% of the miRNAs that were dysregulated in sedentary 5xFAD mice were restored towards wild‐type levels following exercise. Specifically, miR‐29, which is a validated target of the APP processing protein, BACE1, was significantly higher in exercised 5xFAD mice. Accordingly, BACE1 expression was lower in these mice. Based on these results, we conclude that exercise training alters cerebral miRNA expression via Dicer to affect APP processing.Support or Funding InformationNIH AG049806 to J.J.M and C.A.P.NIH P30 AG028383 to C.M.D.
Evidence of Degradation of Endothelial Glycocalyx in African Children with Falciparum Malaria
Authors: Bush Margaret A. , Florence Salvatore M. , Kalingonji Ayam R. , Chen Youwei , Yeo Tsin W. , Anstey Nicholas M. , Rubach Matthew P. , Granger Donald L. , Mwaikambo Esther D. , Weinberg J. Brice 
Published: April 2020

The microvasculature and endothelium play a pivotal role in pathophysiology of malaria. The endothelial glycocalyx (eGC) is a layer on the luminal side of vascular endothelial cells with involvement in homeostatic functions including regulation of vascular permeability, cellular adhesion and blood flow‐mediated nitric oxide formation. Breakdown products of eGC degradation include glycosaminoglycans (GAG) and syndecan‐1, a core protein. The objective of this study was to evaluate a possible role of eGC breakdown in the pathogenesis of malaria. We prospectively evaluated measures of endothelial glycocalyx integrity in children with severe and moderately severe malaria and healthy controls, and determined the relationship between glycocalyx integrity and markers of malaria severity, microvascular reactivity, and endothelial activation. At the Hubert Kairuki Medical Center in Dar es Salaam, Tanzania we studied 146 subjects, aged 2–11 years, including 60 healthy controls (HC), 49 patients with moderately severe malaria (MSM) and 37 patients meeting WHO criteria for severe malaria (SM). Ethics committee approvals were obtained prior to study initiation. We assessed eGC integrity biochemically by measuring plasma syndecan‐1 (ELISA) and total urinary glycosaminoglycan (GAG) breakdown products (dimethylmethylene blue assay). Angiopoietin‐2 has been reported to cause breakdown of eGC and was measured by ELISA. Quantitative measures of eGC were assessed using non‐invasive, side‐stream dark field (SDF) microscopy of the pinna and axilla. Levels of urinary GAG were increased in malaria patients compared with HC (mean±SEM 4.3±0.4 g/mol creatinine); MSM (12.4±1.0); SM (13.4±1.0); p<0.0001. Plasma levels of angiopoietin‐2 and syndecan‐1 were also elevated in MSM and SM compared with HC (p<0.0001), and significantly correlated with each other. For HC compared with MSM, imaging results showed eGC degradation [an increase in the perfused boundary region (mean±SEM 1.55±0.04 microns vs.1.68±0.05; p<0.05)], increased microvascular density (549± 34 μm/mm2 vs. 649 ±38; p<0.05)], and a decrease in the extent of RBC filling (79.1± 0.98% vs. 74.9±1.02%; p<0.01). In conclusion, these results provide evidence for degradation of the eGC that could contribute to pathogenesis of vascular dysfunction in malaria. Decreased nitric oxide formation, endothelial activation and increased adhesion of infected RBC are potential consequences of the eGC degradation and vascular dysfunction. Administration of an agent that prevents or repairs eGC degradation may be helpful as adjunctive therapy for malaria.Support or Funding InformationNational Institutes of Health (R01 HL130783)
Evaluation of Sucrose‐enriched Diet Consumption in the Development of Risk Factors Associated to Type 2 Diabetes, Atherosclerosis and Non‐alcoholic Fatty Liver Disease in a Murine Model
Authors: Plazas-Guerrero Carolina Gabriela , de Jesùs Acosta-Cota Selene , Castro-Sánchez Francisco Humberto , Ríos-Burgueño Efrén Rafael , Sarmiento-Sánchez Juan Ignacio , Picos-Corrales Lorenzo Antonio , de Jesús Vergara-Jiménez Marcela , Osuna-Martìnez Ulises 
Published: April 2020

IntroductionOverconsumption of sucrose has been associated with negative metabolic effects related to non‐communicable diseases. However, this relationship continues to be a controversial topic and further studies are needed to generate scientific evidence to help clarify existing questions and which can be used in the future for the implementation of prevention and control strategies of this important health problem.ObjectiveThe aim of this study was to evaluate the sucrose‐enriched diet consumption in the development of risk factors associated to type 2 diabetes, atherosclerosis and non‐alcoholic fatty liver disease in a murine model.MethodologyTwelve healthy male Wistar rats were randomly divided into two experimental groups (n=6 each) which received one of two different diets for twenty five weeks. Control group (CG) was fed with a standard laboratory diet and tap water. Sucrose group (SG) received the same laboratory diet and water enriched with 50% w/v of sucrose. Body weight, food, macronutrient and total caloric intake were determined. After intervention period, blood samples, hepatic and aortic tissues were collected. Glucose, insulin, total cholesterol, triglycerides, very low‐density lipoprotein cholesterol, low‐density lipoprotein cholesterol and high‐density lipoprotein cholesterol levels were determined in serum. Insulin resistance was calculated and total hepatic cholesterol was measured. Histopathological analysis was made for liver and aortic tissues. Results of parametric tests were expressed as mean values ± standard errors of the mean and for non‐parametric tests, results were expressed as medians and ranks. A p value of < 0.05 was considered significant.ResultsSucrose‐enriched diet‐fed rats showed a decrease in food, lipids and protein intake as well as in serum total cholesterol levels, an increase in carbohydrates intake, glucose, insulin, triglycerides, very low‐density lipoprotein cholesterol and high‐density lipoprotein cholesterol levels, the development of insulin resistance, a higher percentage of steatosis and a greater degree of non‐alcoholic steatohepatitis. No significant differences were found in body weight, total caloric intake, low‐density lipoprotein cholesterol and hepatic cholesterol between the experimental groups. In aortic tissue samples, both groups showed a slight wall thickening.ConclusionOur results show that sucrose‐enriched diet consumption during twenty five weeks contribute to the development of risk factors associated to type 2 diabetes, atherosclerosis and non‐alcoholic fatty liver disease in male Wistar rats.