bluuebunny/crossref_metadata_2025_split huggingface dataset:
Regulation of Inducible Nitric Oxide Synthase (iNOS) by Cyclin‐Dependent Kinase 7 (CDK7) in Endothelial Cells
Authors: Shetty Shruthi , Spaulding Amelia M. , Baker Cassadie , Baine Stephen H. , Hoyt Dale G. 
Published: April 2020

E. coli endotoxin (LPS) combined with interferon‐γ (IFN) induces inducible nitric oxide synthase (iNOS), with production of nitric oxide (NO) and reactive nitrogen species in endothelial cells. Appropriate manipulation of iNOS during onset or resolution of inflammation may be useful. Since transcription is required for induction of iNOS by LPS/IFN, we hypothesized that depletion of cyclin dependent kinase (CDK) 7, a promoter of transcription, would antagonize induction. Mouse aortic endothelial cells (MAEC) were transfected with Non‐targeted (NT) control RNA or CDK7 siRNA to deplete the kinase, followed by stimulation with LPS/IFN for 12 h. Rather than suppressing, CDK7 depletion increased induction of iNOS, and its dimerization and NO production. The effect was post‐transcriptional in that mRNA induction was unaffected, while iNOS protein turnover was reduced in the absence of CDK7. Cellular calpain activity, which is required for iNOS protein degradation in MAEC, was unaffected by CDK7 depletion, suggesting an effect on iNOS itself. Indeed, iNOS in lysates of kinase‐deficient cells was resistant to in vitro digestion with exogenous calpain. In conjunction with protease resistance, the association of iNOS with calmodulin, known to inhibit its digestion by calpain, was increased in cells lacking CDK7. These results suggest that CDK7 normally restrains iNOS induction by limiting calmodulin binding and maintaining calpain‐mediated turnover. By regulating iNOS enzyme complexes, CDK7 may serve as a novel target for manipulation of vascular NO.Support or Funding InformationSupport was received from the Popat Patil Fellowship (SS).
Impact of Short, Long, and Intermittent Access Conditions on MDPV and Cocaine Self‐administration in Male and Female Rats
Authors: Doyle Michelle R. , Sulima Agnieszka , Rice Kenner C. , Collins Gregory T. 
Published: April 2020

Despite decades of research, the behavioral, pharmacological, and neurobiological determinants of one’s vulnerability to develop a substance use disorder are not well understood. When rats are allowed to self‐administer cocaine under short‐access conditions, patterns of drug intake tend to be well‐regulated; however, manipulating drug access conditions (e.g., long‐access and intermittent‐access) can produce neurobiological and behavioral changes thought to be related to addiction. In contrast to cocaine, when rats self‐administer the synthetic cathinone 3,4‐methylenedioxypyrovalerone (MDPV) under short access conditions, a subset display high levels of dysregulated drug‐taking behavior. However, MDPV has not been studied under extended access conditions, and it is unknown whether this MDPV phenotype is simply an earlier manifestation of the addiction‐like phenotype observed with cocaine, or whether the two phenotypes are qualitatively different. The current studies used male and female Sprague Dawley rats self‐administering MDPV (0.032 mg/kg/infusion) or cocaine (0.32 mg/kg/infusion) to test the hypotheses that (1) under short‐access conditions, rats self‐administering MDPV exhibit higher levels of drug taking, greater levels of responding when drug is not available, and reduced sensitivity to punishment by foot shock (i.e., addiction‐like behaviors); and (2) that 21 days of long‐ and intermittent‐access to MDPV or cocaine self‐administration will result in greater addiction‐like behaviors compared to short‐access self‐administration. After short‐access self‐administration, rats that self‐administer MDPV exhibit greater addiction‐related behaviors than rats self‐administering cocaine, as measured by the three criteria detailed above. Though long‐and intermittent‐access produce some changes in patterns of drug‐taking behavior, the access condition manipulation does not appear to systematically alter the addiction‐like behaviors in either rats self‐administering cocaine or MDPV. These results suggest that the phenotype displayed by rats self‐administering MDPV is not the same phenotype as that exhibited by rats with a history of self‐administering cocaine. Therefore, MDPV self‐administration may be a useful method to study individual differences in drug‐taking behavior under short‐access conditions.Support or Funding InformationThis work was supported by NIH/NIDA (R01 DA039146; GTC) and NIDA‐ and NIAAA‐IRPs (KCR).
The Therapeutic Potential of Apoptotic Mesenchymal Stem Cells in Muscle Regeneration
Authors: Huang Wenbai , Suggs Laura J. , Farrar Roger P. 
Published: April 2020

Macrophages are critical for muscle regeneration and have an unusually high level of responsiveness to their environment. Because of this sensitivity, their beneficial effects may be realized only upon certain activation conditions. Previous studies in our lab have shown that transplantation of classically activated macrophages and anti‐inflammatory macrophages sorted from injured muscle enhanced functional regeneration in ischemia‐reperfusion muscles. The phagocytosis of apoptotic cells that are present in injured muscle or from transplantation serves as a well‐known activation signal for macrophages to initiate a regenerative and anti‐inflammatory profile leading to enhanced muscle regeneration. The aim of this study is to investigate if the phagocytosis of apoptotic mesenchymal stem cells (MSCs) can polarize bone marrow‐derived macrophages (BMDMs) towards a regenerative and anti‐inflammatory phenotype and subsequently if apoptotic MSCs in different stages of apoptosis have differential effects. We induced apoptosis in MSCs and incubated them at different time intervals to obtain differential stages of apoptosis, which were characterized by the amount of externalized phosphatidylserine (PS) on their cell membrane. Apoptotic MSCs with different concentrations of PS were then co‐cultured with BMDMs for 48 hours. Cell surface markers and gene expression of characteristic of pro‐ and anti‐inflammatory polarized BMDMs were measured. Our results showed that apoptotic MSCs without incubation significantly increased the expression of F4/80 and CD206 on the cell surface of BMDMs. They also increased the gene expression of MRC‐1 and IL‐10 as well as decreased the expression of iNOS and IL‐1β in co‐cultured BMDMs. The extent of these effects exhibited a decreasing trend when incubation time of apoptotic MSCs increased. In conclusion, the extent of MSC apoptosis as characterized by PS expression resulted in differential anti‐inflammatory and regenerative effects on BMDMs, wherein the apoptotic MSCs in the earlier stage of apoptosis result in the greatest increase in anti‐inflammatory phenotype, which reveals their therapeutic potential in muscle regeneration.Support or Funding InformationThis project is funded by National Institutes of Health Grant 2R01EB015007‐05A1 (to Laura J. Suggs and Roger P. Farrar)
Anticancer Activity of Curcumin and Curcumin Degradation Products in Combination with Silibinin
Authors: Sayyed Alhan , Ezekiel Uthayashanker 
Published: April 2020

Colorectal cancer (CRC) is the third most commonly diagnosed cancer in men and women in the United States. Several studies have shown that a diet high in fruits, vegetables, and spices is strongly associated with reduced risk of developing colorectal cancer. Fruits and vegetables contain a wide variety of phytochemicals that offer protection against colorectal cancer. The anticancer activity of healthy diets may be attributable to a synergy among various phytochemicals present in the diet. We have previously shown that phytochemicals derived from turmeric (e.g. curcumin) and milk thistle (e.g. silibinin) have exhibited synergistic anticancer activity against colorectal cancer cells. In the present study we hypothesize that the curcumin degradation products, tran‐ferulic acid, vanillylidenacetone, ferulic aldehyde, and vanillin in combination with silibinin will exhibit synergistic anticancer activity. Our results indicate that the degradation products of curcumin plus silibinin did not exhibit synergistic anticancer activity. We found that only curcumin plus silibinin exhibited synergistic anticancer activity.Support or Funding InformationAS is supported by the DeNardo Education and Research Foundation grant.
Science Communication Boot Camp: An American Association for Anatomy program to empower members with skills in effective public communication strategies
Authors: Organ Jason , Hoffmann-Longtin Krista 
Published: April 2020

As scientific fields have become more complex in the 21st century, some Americans have given up trying to understand what we do, leading to science illiteracy. Although scientists have reached consensus on many issues, such as climate change accelerated by humans, and evolution, the public and (American) elected officials have not always accepted this consensus. According to the Pew Center, there is a wide gap between scientists’ and the public’s perception of how science is used to inform government policy. Without expert voices communicating science to the public and policy‐makers, fields like healthcare, climate science, and evolutionary biology—all fields that are inextricably linked to anatomy—have been overrun with conspiracy theories and misinformation.The timing is right to make science communication a fundamental part of doing science. And to do that, we must train our students/faculty in the skills necessary to be effective communicators. Over the last several years, the American Association for Anatomy (AAA) has prioritized professional development in science communication for researchers, educators, and practitioners at its annual meeting. Furthermore, AAA recently adopted a new strategic plan which brings science communication front and center, where the public is engaged through the promotion of anatomical sciences. To support this new science communication initiative, AAA hosted its inaugural Science Communication Boot Camp (SCBC) on the campus of Indiana University School of Medicine in Indianapolis in July 2019. This three‐day program brought participants together to learn effective communication skills from the faculty of the Alan Alda Center for Communicating Science and from representatives of the Federation of American Societies for Experimental Biology science policy staff. This talk reports on the SCBC program and demonstrates some of the hands‐on activities employed.Support or Funding InformationThis study was granted exempt status by the Institutional Review Board of Indiana University School of Medicine. Funding was provided by the AAA Innovations Grant Program and the Indiana University School of Medicine Office of Faculty Affairs, Professional Development, and Diversity.
Comparison of Wire and Disc Leads to Activate the Inspiratory Muscles in Dogs
Authors: Kowalski Krzysztof E. , DiMarco Anthony F. 
Published: April 2020

ObjectiveHF‐SCS is a unique method of inspiratory muscle activation which results in coincident activation of the diaphragm and inspiratory intercostal muscles via spinal cord pathways and has the potential to provide respiratory support in ventilator dependent persons with spinal cord injury (SCI). The purpose of the present study was to examine the effectiveness of wire leads, which can be placed using minimally invasive techniques, to activate the inspiratory muscles.MethodsIn 5 anesthetized dogs, in separate trials, disc and wire leads were inserted onto the ventral epidural space at the T1–T4 spinal cord levels. The effects of electrical stimulation with disc, single wire, and two wire leads placed in parallel were compared.Outcome measuresAirway pressure generation following stimulation with disc and various configurations of wire leads were compared.ResultsSeveral different configurations of wire leads resulted in airway pressures that were similar to those generated with monopolar stimulation with disc leads (D). For example, monopolar stimulation (3mA, 300Hz, 0.2ms) with parallel wire leads in which each of the electrode contacts at the T2 were connected together and functioned as a common cathode (Y‐connection) and monopolar stimulation (separate cathode) at T2 both with a remote ground resulted in airway pressures that were 91±6 and 92±4%, respectively, of those achieved with D (NS for both). Bipolar stimulation (3mA, 300Hz, 0.2ms) with parallel wire leads using Y‐connection and with a separate cathode at the T2 and T3 level resulted in airway pressures that were 96 ± 4 and 94 ± 4%, respectively, of the pressures achieved with D (NS for both).ConclusionThese results suggest that specific configurations of wire leads, which can be placed via minimally invasive techniques, result in comparable activation of the inspiratory muscles compared to disc leads and may be a useful technique to restore ventilatory support in persons with SCI.Support or Funding InformationSupportNIH‐NINDS (R01NS064157)DisclosureDr. DiMarco is a founder of and has a significant financial interest in Synapse BioMedical, a manufacturer of diaphragm pacing systems and holds patents for spinal cord stimulation to restore cough and respiration (5,678,535; 5,911,218; 5,999,855; 8,751,004). Drs. DiMarco and Kowalski hold the U.S. patents for technology related to the content of this manuscript: Respiratory Muscle Activation by Spinal Cord Stimulation (8,352,036).
Impaired Mitochondrial Respiration in Hermansky‐Pudlak Syndrome 1‐Defective Alveolar Type 2 Cells is Associated with Enhanced Mitochondrial Fission
Authors: Kook Seunghyi , Meng Shufang , Rasmussen Megan L. , Trenary Irina , Young Jamey D. , Gama Vivian , Guttentag Susan H. 
Published: April 2020

Alveolar epithelial injury is central to the pathogenesis of pulmonary fibrosis in lung diseases across the lifespan, from bronchopulmonary dysplasia and childhood interstitial lung disease to idiopathic pulmonary fibrosis. Disturbances in mitochondrial structure that alter function have been associated with lung epithelial cell injury, yet molecular mechanisms regulating mitochondrial dynamics in the setting of pulmonary fibrosis remain unclear. Hermansky‐Pudlak Syndrome (HPS) patients, especially those with mutations in HPS1 and HPS4, develop fibrosis in earlier age than patients with idiopathic pulmonary fibrosis (IPF), suggesting that loss of HPS gene function drives alveolar epithelial injury. Here we show enhanced mitochondrial fragmentation in HPS1‐defective alveolar type 2 (AT2) cells associated with impaired phosphorylation of Dynamin‐related protein 1 (DRP1) at Serine (Ser)637 (inactivating phosphorylation). Ectopic expression of constitutively active GTPase Rab32, Rab32(Q85L), designed to bypass the guanine nucleotide exchange factor (GEF) function of HPS1‐HPS4 complex (BLOC‐3), in MLE15/DHPS1 cells restores mitochondrial architecture and increases p‐Ser637‐Drp1. In addition, MLE15/DHPS1 cells exhibit markedly reduced mitochondrial respiration, signified by reduced oxygen consumption (OCR), and robust glycolysis, indicated by a brisk increase in extracellular acidification rate (ECAR) after addition of glucose and accumulation of lactate in culture media. Despite robust glycolysis, MLE15/DHPS1 cells have lower total cellular ATP than MLE15/WT cells. Moreover, MLE15/DHPS1 cells demonstrate dependency on glucose as a nutrient source, reflected in activation of the energy sensor 5’ AMP‐activated protein kinase (AMPK) and cell death upon decreasing glucose in culture media. Taken together, our results suggest that HPS1, functioning in a heterodimer with HPS4 as an activator of Rab32, plays a central role in balancing mitochondrial fission/fusion in AT2 cells, while loss of HPS1 disrupts oxidative phosphorylation and drives metabolic reprogramming in AT2 cells.Support or Funding InformationNIH (GM108807/SG). SG is the Julia Carell Stadler Professor of Pediatrics. NIH/NIGMS 1R35GM128915‐01 (to VG); NIH/NCI 1R21 CA227483‐01A1 (to VG). NIH/R01 DK106348 and U01 CA235508 (to JY)
Examination of the demyelination process in mGFAP‐driven conditional transient receptor potential ankyrin 1 (TRPA1) receptor knockout mice
Authors: Pintér Erika , Böcskei Kata , Kriszta Gábor , Sándor Zoltán , Nemes Balázs , Ács Péter , Komoly Sámuel , Berente Zoltán 
Published: April 2020

Transient receptor potential ankyrin 1 (TRPA1) receptors are non‐selective cation channels responsive to a variety of exogenous irritants and endogenous stimuli including products of oxidative stress. It is mainly expressed by primary sensory neurons however, expression of TRPA1 by astrocytes and oligodendrocytes has recently been detected in the mouse brain. Genetic deletion of TRPA1 was shown to attenuate cuprizone‐induced oligodendrocyte apoptosis and myelin loss in mice. In the present study we investigated the astrocyte‐specific mGFAP‐driven conditional TRPA1 gene deficient mice in the cuprizone model. Astrocyte‐specific knockout animals were generated by crossbreeding GFAP‐Cre+/− and floxed TRPA1 (TRPA1Fl/Fl) mice. Cuprizone was administered during 6 weeks and demyelination was followed by magnetic resonance imaging (MRI). At the end of the treatment, myelin loss and glial activation were also investigated by histological methods. The results of the MRI showed significantly increased signal intensity in cuprizone‐treated GFAP‐Cre−/− TRPA1Fl/Fl mice compared to GFAP‐Cre+/− TRPA1Fl/− and GFAP‐Cre+/− TRPA1Fl/Fl animals on week 3 and week 4. However, by week 6 of the treatment the difference was not detectable by either the MRI or the histological methods. According to the currently available data, we presume that TRPA1 receptors localized on astrocytes can be activated by reactive oxygen species in response to cuprizone challenge and the consequent release of pro‐inflammatory mediators contributes to the progression of oligodendrocyte apoptosis. However, the global TRPA1 KO mice presented a more attenuated demyelination compared to the conditional KO mice, it could be hypothesized that TRPA1 receptors expressed by other cell types (neurons, oligodendrocytes) in the brain may also participate in the demyelination process.Support or Funding InformationThis research was funded by grants “The role of neuro‐inflammation in neurodegeneration: from molecules to clinics” (EFOP‐3.6.2‐16‐2017‐00008); Hungarian Brain Research Program 2. 2017‐1.2.1‐NKP‐2017‐00002. and the University of Pécs is acknowledged for a support by the 17886‐4/23018/FEKUTSTRAT excellence grant. G.K. was supported by Richter Gedeon Centenárium Foundation.
Assessing the Global Impact of an Adenylate Kinase Isoenzyme on AMPK and mTOR Signaling Pathways Through Metabolic Profiling in both Brain and Lung Cancer Cell Lines
Authors: Salladay-Perez Ivan Alexander , Lanning Nathan J. 
Published: April 2020

In a variety of human cancers, it is well observed that specific altered bioenergetic programs must be sustained in order to maintain the cancer cell phenotype. Well‐described genes play a role in sustaining energy flux for the cancer cell and recent studies add the regulation of AK4 expression, an adenylate kinase localized to the mitochondrial matrix, to the list. Our results demonstrate that in response to altered AK4 expression in cancer cells, intracellular ATP levels drastically increase. Secondly, we demonstrate that in response to perturbed AK4 expression in cancer cells, two critical homeostatic signaling pathways, the 5′ AMP‐activated protein kinase (AMPK) and a mammalian target of rapamycin (mTOR) pathways, are significantly altered. Both mTOR and AMPK proteins govern a metabolic pathway in which they sense cellular energy levels and nutrient status in response to regulate cancer cell growth and/or proliferation. Lastly, AK4 also has an impact on other bioenergetic pathways. For instance, AK4 can catalyze the reaction GTP + AMP ↔ GDP + ADP24 and sustain critical energy flux of nucleotides in the cell. Because AK4 expression appears to regulate cellular metabolic responses at multiple levels, further analyses by obtaining a metabolomic profile is necessary to highlight the global impact AK4 exerts on all the metabolites in the cancer cell. To do this, cell lines will be engineered without the AK4 gene in glioblastomas, lung adenocarcinomas, and HeLa cells then analyzed to identify all known metabolites in the system. It is hypothesized that without the expression of an AK4 protein there will be indirect or direct disruptions in the global nucleotide pools by either increasing nucleotide biosynthesis or decreasing nucleotide consumption. The outcome of this study will add to the small library of research on AK4 as well as provide a comprehensive understanding on the enzymatic activity and interactions where AK4 exerts its biological effects on cancer cell metabolism.Support or Funding InformationFunded by National Institutes of Health (NIH) National Institute of General Medical Sciences (NIGMS) Grant 1R15GM123382‐01.
Rapid Real‐Time Ubiquitination Assays as Alternatives to Western Blotting
Authors: de Silva Ruvindi , Page Richard C. 
Published: April 2020

The refolding and degradative pathways of the protein quality control system converge through the chaperoned ubiquitination complex formed by the interaction of the E3 ubiquitin ligase CHIP (
C‐terminus of
Hsp70
Interacting
Protein) and the ATP‐dependent chaperone Hsp70 (70 kilodalton heat shock protein). We have applied the CHIP/Hsp70 chaperoned ubiquitination system as a test for our newly developed ubiquitination assay, presenting a viable alternative to traditional western blots for detection of ubiquitination. Here we describe the cutting‐edge biolayer interferometry ubiquitination assay to rapidly assay ubiquitination in real time. Our BLI ubiquitination assay is recorded with 1‐second resolution and is the first assay to allow for determination of rate and extent of ubiquitination in real‐time. This assay detects formation of polyubiquitin chains directly on a biosensor‐bound target. Polyubiquitination of substrates using wild‐type components produces a robust signal. Reagent concentrations and quantities are similar to those used by traditional western blot‐based ubiquitination assays. Results are returned instantaneously and result in substantial time savings.Support or Funding InformationNIGMS R35 GM128595