@article{mottin_caesar_brodsky_mesquita_oliveira_noske_sousa_ramos_jarmer_loh_et al._2022, title={Chalcones from Angelica keiskei (ashitaba) inhibit key Zika virus replication proteins}, volume={120}, ISSN={["1090-2120"]}, DOI={10.1016/j.bioorg.2022.105649}, abstractNote={Zika virus (ZIKV) is a dangerous human pathogen and no antiviral drugs have been approved to date. The chalcones are a group of small molecules that are found in a number of different plants, including Angelica keiskei Koidzumi, also known as ashitaba. To examine chalcone anti-ZIKV activity, three chalcones, 4-hydroxyderricin (4HD), xanthoangelol (XA), and xanthoangelol-E (XA-E), were purified from a methanol-ethyl acetate extract from A. keiskei. Molecular and ensemble docking predicted that these chalcones would establish multiple interactions with residues in the catalytic and allosteric sites of ZIKV NS2B-NS3 protease, and in the allosteric site of the NS5 RNA-dependent RNA-polymerase (RdRp). Machine learning models also predicted 4HD, XA and XA-E as potential anti-ZIKV inhibitors. Enzymatic and kinetic assays confirmed chalcone inhibition of the ZIKV NS2B-NS3 protease allosteric site with IC50s from 18 to 50 µM. Activity assays also revealed that XA, but not 4HD or XA-E, inhibited the allosteric site of the RdRp, with an IC50 of 6.9 µM. Finally, we tested these chalcones for their anti-viral activity in vitro with Vero cells. 4HD and XA-E displayed anti-ZIKV activity with EC50 values of 6.6 and 22.0 µM, respectively, while XA displayed relatively weak anti-ZIKV activity with whole cells. With their simple structures and relative ease of modification, the chalcones represent attractive candidates for hit-to-lead optimization in the search of new anti-ZIKV therapeutics.}, journal={BIOORGANIC CHEMISTRY}, author={Mottin, Melina and Caesar, Lindsay K. and Brodsky, David and Mesquita, Nathalya C. M. R. and Oliveira, Ketllyn Zagato and Noske, Gabriela Dias and Sousa, Bruna K. P. and Ramos, Paulo R. P. S. and Jarmer, Hannah and Loh, Bonnie and et al.}, year={2022}, month={Mar} }
 @article{cortes_brodsky_chen_pridgen_odle_snider_cruse_putikova_masuda_doyle_et al._2022, title={Immunologic and pathologic characterization of a novel swine biomedical research model for eosinophilic esophagitis}, volume={3}, ISSN={["2673-6101"]}, DOI={10.3389/falgy.2022.1029184}, abstractNote={Eosinophilic esophagitis (EoE) is a chronic allergy-mediated condition with an increasing incidence in both children and adults. Despite EoE's strong impact on human health and welfare, there is a large unmet need for treatments with only one recently FDA-approved medication for EoE. The goal of this study was to establish swine as a relevant large animal model for translational biomedical research in EoE with the potential to facilitate development of therapeutics. We recently showed that after intraperitoneal sensitization and oral challenge with the food allergen hen egg white protein (HEWP), swine develop esophageal eosinophilia—a hallmark of human EoE. Herein, we used a similar sensitization and challenge treatment and evaluated immunological and pathological markers associated with human EoE. Our data demonstrate that the incorporated sensitization and challenge treatment induces (i) a systemic T-helper 2 and IgE response, (ii) a local expression of eotaxin-1 and other allergy-related immune markers, (iii) esophageal eosinophilia (>15 eosinophils/0.24 mm2), and (iv) esophageal endoscopic findings including linear furrows and white exudates. Thereby, we demonstrate that our sensitization and oral challenge protocol not only induces the underlying immune markers but also the micro- and macro-pathological hallmarks of human EoE. This swine model for EoE represents a novel relevant large animal model that can drive translational biomedical research to develop urgently needed treatment strategies for EoE.}, journal={FRONTIERS IN ALLERGY}, author={Cortes, Lizette M. and Brodsky, David and Chen, Celine and Pridgen, Tiffany and Odle, Jack and Snider, Douglas B. and Cruse, Glenn and Putikova, Arina and Masuda, Mia Y. and Doyle, Alfred D. and et al.}, year={2022}, month={Nov} }
 @article{gulledge_collette_mackey_johnstone_moazami_todd_moeser_pierce_cech_laster_2018, title={Mast cell degranulation and calcium influx are inhibited by an Echinacea purpurea extract and the alkylamide dodeca-2E,4E-dienoic acid isobutylamide}, volume={212}, ISSN={0378-8741}, url={http://dx.doi.org/10.1016/J.JEP.2017.10.012}, DOI={10.1016/J.JEP.2017.10.012}, abstractNote={Native Americans used plants from the genus Echinacea to treat a variety of different inflammatory conditions including swollen gums, sore throats, skin inflammation, and gastrointestinal disorders. Today, various Echinacea spp. preparations are used primarily to treat upper respiratory infections. The goal of this study was to evaluate the effects of an ethanolic E. purpurea (L) Moench root extract and the alkylamide dodeca-2E,4E-dienoic acid isobutylamide (A15) on mast cells, which are important mediators of allergic and inflammatory responses. Inhibition of mast cell activation may help explain the traditional use of Echinacea. A15 was evaluated for its effects on degranulation, calcium influx, cytokine and lipid mediator production using bone marrow derived mast cells (BMMCs) and the transformed rat basophilic leukemia mast cell line RBL-2H3. Methods included enzymatic assays, fluorimetry, ELISAs, and microscopy. A root extract of E. purpurea, and low and high alkylamide-containing fractions prepared from this extract, were also tested for effects on mast cell function. Finally, we tested A15 for effects on calcium responses in RAW 264.7 macrophage and Jurkat T cell lines. A15 inhibited ß-hexosaminidase release from BMMCs and RBL-2H3 cells after treatment with the calcium ionophore A23187 by 83.5% and 48.4% at 100 µM, respectively. Inhibition also occurred following stimulation with IgE anti-DNP/DNP-HSA. In addition, A15 inhibited 47% of histamine release from A23187-treated RBL-2H3 cells. A15 prevented the rapid rise in intracellular calcium following FcεRI crosslinking and A23187 treatment suggesting it acts on the signals controlling granule release. An E. purpurea root extract and a fraction with high alkylamide content derived from this extract also displayed these activities while fractions with little to no detectable amounts of alkylamide did not. A15 mediated inhibition of calcium influx was not limited to mast cells as A23187-stimulated calcium influx was blocked in both RAW 264.7 and Jurkat cell lines with 60.2% and 43.6% inhibition at 1 min post-stimulation, respectively. A15 also inhibited the release of TNF-α, and PGE2 to a lesser degree, following A23187 stimulation indicating its broad activity on mast cell mediator production. These findings suggest that Echinacea extracts and alkylamides may be useful for treating allergic and inflammatory responses mediated by mast cells. More broadly, since calcium is a critical second messenger, the inhibitory effects of alkylamides on calcium uptake would be predicted to dampen a variety of pathological responses, suggesting new uses for this plant and its constituents.}, journal={Journal of Ethnopharmacology}, publisher={Elsevier BV}, author={Gulledge, Travis V. and Collette, Nicholas M. and Mackey, Emily and Johnstone, Stephanie E. and Moazami, Yasamin and Todd, Daniel A. and Moeser, Adam J. and Pierce, Joshua G. and Cech, Nadja B. and Laster, Scott M.}, year={2018}, month={Feb}, pages={166–174} }
 @article{devlin_halvorsen_miller_laster_2017, title={Il-10 deficient mice express IFN-gamma mRNA and clear Leptospira interrogans from their kidneys more rapidly than normal C57BL/6 mice}, volume={222}, ISSN={["1878-3279"]}, DOI={10.1016/j.imbio.2017.02.004}, abstractNote={Leptospira interrogans (L. interrogans), the causative agent of leptospirosis, is a widespread zoonotic spirochete that lives a dual lifestyle. L. interrogans infects mice, rats, and wildlife in a persistent and asymptomatic fashion, while also causing productive and acute infections in other mammals such as humans and hamsters. Infections in humans can be fatal, accompanied by a cytokine storm and shock-like symptoms. Production of IL-10 has been noted in both rodent and human infections which has led a number of investigators to hypothesize that IL-10 plays a role in the pathogenesis of this disease. To test this hypothesis we have compared bacteremia and the cytokine response of normal and IL-10 deficient C57Bl/6 mice following ip infection with L. interrogans. In normal mice bacterial 16s mRNA was detected in both lung and kidney tissues within a day after infection. Levels of 16s mRNA then dropped in both organs with complete elimination from the lung by day 3 but persistence in the kidney for 7days after infection. In contrast, in IL-10 deficient mice, the organism was eliminated more rapidly from the kidney. We found that infection of both control and IL-10 deficient mice produced similar levels of a number of pro-inflammatory cytokine mRNAs. On the other hand, IFN-γ mRNA was only induced in IL-10 deficient mice. These results support the hypothesis that L. interrogans ability to induce IL-10, which in turn prevents production of IFN-γ and inhibits T cell immunity, may contribute to the persistent growth of this microorganism in the murine kidney.}, number={5}, journal={IMMUNOBIOLOGY}, author={Devlin, Amy A. and Halvorsen, Priya J. and Miller, Jennifer C. and Laster, Scott M.}, year={2017}, month={May}, pages={768–777} }
 @article{lila_schneider_devlin_plundrich_laster_foegeding_2017, title={Polyphenol-enriched berry extracts naturally modulate reactive proteins in model foods}, volume={8}, ISSN={2042-6496 2042-650X}, url={http://dx.doi.org/10.1039/C7FO00883J}, DOI={10.1039/C7FO00883J}, abstractNote={Healthy foods like polyphenol-rich berries and high quality edible proteins are in demand in today's functional food marketplace, but it can be difficult to formulate convenient food products with physiologically-relevant amounts of these ingredients and still maintain product quality. In part, this is because proteins can interact with other food ingredients and precipitate destabilizing events, which can disrupt food structure and diminish shelf life. Proteins in foods can also interact with human receptors to provoke adverse consequences such as allergies. When proteins and polyphenols were pre-aggregated into stable colloidal particles prior to use as ingredients, highly palatable food formulations (with reduced astringency of polyphenols) could be prepared, and the overall structural properties of food formulations were significantly improved. All of the nutritive and phytoactive benefits of the proteins and concentrated polyphenols remained highly bioavailable, but the protein molecules in the particle matrix did not self-aggregate into networks or react with other food ingredients. Both the drainage half-life (a marker of structural stability) and the yield stress (resistance to flow) of model foams made with the protein-polyphenol particles were increased in a dose-dependent manner. Of high significance in this complexation process, the reactive allergenic epitopes of certain proteins were effectively blunted by binding with polyphenols, attenuating the allergenicity of the food proteins. Porcine macrophages produced TNF-α proinflammatory cytokine when provoked with whey protein, but, this response was blocked completely when the cells were stimulated with particles that complexed whey protein with cinnamon-derived polyphenols. Cytokine and chemokine production characteristic of allergic reactions were blocked by the polyphenols, allowing for the potential creation of hypoallergenic protein-berry polyphenol enriched foods.}, number={12}, journal={Food & Function}, publisher={Royal Society of Chemistry (RSC)}, author={Lila, Mary Ann and Schneider, Maggie and Devlin, Amy and Plundrich, Nathalie and Laster, Scott and Foegeding, E. Allen}, year={2017}, pages={4760–4767} }
 @article{kaur_oberhofer_juzumaite_raja_gulledge_kao_faeth_laster_oberlies_cech_2016, title={Secondary metabolites from fungal endophytes of Echinacea purpurea suppress cytokine secretion by macrophage-type cells}, volume={11}, DOI={10.1177/1934578x1601100827}, abstractNote={Botanical extracts of Echinacea purpurea have been widely used for the treatment of upper respiratory infections. We sought to chemically examine fungal endophytes inhabiting E. purpurea, and to identify compounds produced by these endophytes with in vitro cytokine-suppressive activity. Twelve isolates from surface sterilized seeds of E. purpurea were subjected to fractionation and major components were isolated. Sixteen secondary metabolites belonging to different structural classes were identified from these isolates based on NMR and mass spectrometry data. The compounds were tested for their influence on cytokine secretion by murine macrophage-type cells. Alternariol (1), O-prenylporriolide (4), porritoxin (10) β-zearalenol (13), and (S)-zearalenone (14) inhibited production of TNF-α from RAW 264.7 macrophages stimulated with LPS in the absence of any significant cytotoxicity. This is the first report of a cytokine-suppressive effect for 4. The results of this study are particularly interesting given that they show the presence of compounds with cytokine-suppressive activity in endophytes from a botanical used to treat inflammation. Future investigations into the role of fungal endophytes in the biological activity of E. purpurea dietary supplements may be warranted.}, number={8}, journal={Natural Product Communications}, author={Kaur, A. and Oberhofer, M. and Juzumaite, M. and Raja, H. A. and Gulledge, T. V. and Kao, D. N. and Faeth, S. H. and Laster, S. M. and Oberlies, N. H. and Cech, N. B.}, year={2016}, pages={1143–1146} }
 @article{leyte-lugo_todd_gulledge_juzumaite_carter_laster_cech_2015, title={Cytokine-Suppressive Activity of a Hydroxylated Alkylamide from Echinacea purpurea}, volume={2}, ISSN={2199-157X}, url={http://dx.doi.org/10.1055/S-0035-1557764}, DOI={10.1055/S-0035-1557764}, abstractNote={Abstract Echinacea purpurea has been widely used to treat upper respiratory tract infections. Bioassay-guided fractionation of hexane and ethanol extracts of this plant yielded 4[(2-methylbutyl)amino]-4-oxo-2-butenoic acid (1) and 8,11-dihidroxy-dodeca-2E,4E,9E-trienoic acid isobutylamide (2), both of which are new to Echinacea. Compound 2 suppressed the production of TNF-α from RAW 264.7 cells with an IC50 value of 6.8 µM, while 1 was inactive. Neither compound showed cytotoxicity at concentrations up to 100 µM. These findings support a growing body of literature demonstrating that E. purpurea contains an array of compounds that suppress cytokine secretion by macrophage-type cells in vitro.}, number={01}, journal={Planta Medica Letters}, publisher={Georg Thieme Verlag KG}, author={Leyte-Lugo, Martha and Todd, Daniel and Gulledge, Travis and Juzumaite, Monika and Carter, Frederick and Laster, Scott and Cech, Nadja}, year={2015}, month={Aug}, pages={e25–e27} }
 @article{todd_gulledge_britton_oberhofer_leyte-lugo_moody_shymanovich_grubbs_juzumaite_graf_et al._2015, title={Ethanolic Echinacea purpurea Extracts Contain a Mixture of Cytokine-Suppressive and Cytokine-Inducing Compounds, Including Some That Originate from Endophytic Bacteria}, volume={10}, ISSN={["1932-6203"]}, DOI={10.1371/journal.pone.0124276}, abstractNote={Echinacea preparations, which are used for the prevention and treatment of upper respiratory infections, account for 10% of the dietary supplement market in the U.S., with sales totaling more than $100 million annually. In an attempt to shed light on Echinacea's mechanism of action, we evaluated the effects of a 75% ethanolic root extract of Echinacea purpurea, prepared in accord with industry methods, on cytokine and chemokine production from RAW 264.7 macrophage-like cells. We found that the extract displayed dual activities; the extract could itself stimulate production of the cytokine TNF-α, and also suppress production of TNF-α in response to stimulation with exogenous LPS. Liquid:liquid partitioning followed by normal-phase flash chromatography resulted in separation of the stimulatory and inhibitory activities into different fractions, confirming the complex nature of this extract. We also studied the role of alkylamides in the suppressive activity of this E. purpurea extract. Our fractionation method concentrated the alkylamides into a single fraction, which suppressed production of TNF-α, CCL3, and CCL5; however fractions that did not contain detectable alkylamides also displayed similar suppressive effects. Alkylamides, therefore, likely contribute to the suppressive activity of the extract but are not solely responsible for that activity. From the fractions without detectable alkylamides, we purified xanthienopyran, a compound not previously known to be a constituent of the Echinacea genus. Xanthienopyran suppressed production of TNF-α suggesting that it may contribute to the suppressive activity of the crude ethanolic extract. Finally, we show that ethanolic extracts prepared from E. purpurea plants grown under sterile conditions and from sterilized seeds, do not contain LPS and do not stimulate macrophage production of TNF-α, supporting the hypothesis that the macrophage-stimulating activity in E. purpurea extracts can originate from endophytic bacteria. Together, our findings indicate that ethanolic E. purpurea extracts contain multiple constituents that differentially regulate cytokine production by macrophages.}, number={5}, journal={PLOS ONE}, author={Todd, Daniel A. and Gulledge, Travis V. and Britton, Emily R. and Oberhofer, Martina and Leyte-Lugo, Martha and Moody, Ashley N. and Shymanovich, Tatsiana and Grubbs, Laura F. and Juzumaite, Monika and Graf, Tyler N. and et al.}, year={2015}, month={May} }
 @article{moazami_gulledge_laster_pierce_2015, title={Synthesis and biological evaluation of a series of fatty acid amides from Echinacea}, volume={25}, ISSN={["1464-3405"]}, DOI={10.1016/j.bmcl.2015.06.024}, abstractNote={Alkylamides are lipophilic constituents of Echinacea and possess numerous biological activities. Although significant effort has been focused on the study of crude Echinacea extracts, very little is known regarding the activities of the individual constituents that make up these herbal treatments. Herein we explore the SAR of simple alkylamides found in Echinacea extracts with respect to their ability to decrease the production of the pro-inflammatory mediator TNF-α. Our results have revealed the key structural requirements for activity and provide lead compounds for further investigation of these poorly understood molecules.}, number={16}, journal={BIOORGANIC & MEDICINAL CHEMISTRY LETTERS}, author={Moazami, Yasamin and Gulledge, Travis V. and Laster, Scott M. and Pierce, Joshua G.}, year={2015}, month={Aug}, pages={3091–3094} }
 @article{kao_kaur_raja_el-elimat_oberhofer_juzumaite_gulledge_laster_cech_faeth_et al._2014, title={Chemical investigation of fungal endophytes from Echinacea purpurea}, volume={80}, ISSN={0032-0943 1439-0221}, url={http://dx.doi.org/10.1055/S-0034-1382406}, DOI={10.1055/S-0034-1382406}, abstractNote={Botanical extracts of Echinacea purpurea have been traditionally used for treatment of respiratory infections. Recently, efforts to chemically examine the fungal endophytes inhabiting these plants were undertaken to investigate the role of endophyte colonization on the bioactivity of such medicinal herbs. In this study, twenty-four fungal isolates belonging to fourteen different OTUs were obtained in axenic culture from surface sterilized seeds and root segments of Echinacea. Most of the fungal endophytes belonged to the Ascomycota, whereas one isolate showed phylogenetic affinities to mushroom forming Basidiomycota. Solid-substrate fermentation extracts of the fungal cultures were subjected to various separation techniques after dereplication. Approximately twenty-five secondary metabolites belonging to different structural classes were encountered. The known compounds were identified by comparison of their NMR and mass spectrometry data. Sixteen compounds were tested for their ability to regulate production of the inflammatory cytokine TNF-α. Four compounds were found to inhibit production of TNF-α from RAW 264.7 macrophages stimulated with LPS in the absence of any detectable cytotoxicity, while none were found to induce TNF production.}, number={10}, journal={Planta Medica}, publisher={Georg Thieme Verlag KG}, author={Kao, D and Kaur, A and Raja, HA and El-Elimat, T and Oberhofer, M and Juzumaite, M and Gulledge, TV and Laster, SM and Cech, NB and Faeth, SH and et al.}, year={2014}, month={Jul} }
 @article{britton_todd_leyte-lugo_gulledge_grubbs_oberhofer_faeth_laster_cech_2014, title={Unraveling immunomodulatory activities of complex Echinacea purpurea extracts}, volume={80}, ISSN={0032-0943 1439-0221}, url={http://dx.doi.org/10.1055/S-0034-1382726}, DOI={10.1055/S-0034-1382726}, abstractNote={Echinacea purpurea has a long history of traditional use for the treatment of infection, but there remains considerable controversy surrounding this plant's mechanism of action. Recent studies have suggested that crude Echinacea extracts may contain a mixture of anti-inflammatory and pro-inflammatory compounds, and our goal with this research was to separate such extracts and identify specific bioactive constituents. In addition, we tested the hypothesis that in vitro pro-inflammatory effects of E. purpurea crude extracts could be caused by bacterial endophytes (bacteria living asymptomatically within the plant tissues). To test our hypothesis, E. purpurea plants with and without bacterial endophytes were grown in sterile chambers and tested for immunomodulatory effects on macrophage-type cells in vitro. Bioactivity guided fractionation was also employed to identify anti-inflammatory compounds from crude E. purpurea extracts. Our results clearly demonstrate that bacterial endophytes play a role in the in vitro pro-inflammatory activity of ethanolic E. purpurea extracts. Additionally, we have identified a previously unreported anti-inflammatory constituent of E. purpurea, 4-[(2-methylbutyl)amino-4-oxo-2-butenoic acid.}, number={10}, journal={Planta Medica}, publisher={Georg Thieme Verlag KG}, author={Britton, ER and Todd, DA and Leyte-Lugo, M and Gulledge, T and Grubbs, LF and Oberhofer, M and Faeth, SH and Laster, SM and Cech, NB}, year={2014}, month={Jul} }
 @article{pollara_spesock_pickup_laster_petty_2012, title={Production of prostaglandin E-2 in response to infection with modified vaccinia Ankara virus}, volume={428}, ISSN={["0042-6822"]}, DOI={10.1016/j.virol.2012.03.019}, abstractNote={Prostaglandin E₂ (PGE₂) is an arachidonic acid (AA)-derived signaling molecule that can influence host immune responses to infection or vaccination. In this study, we investigated PGE₂ production in vitro by cells infected with the poxvirus vaccine strain, modified vaccinia Ankara virus (MVA). Human THP-1 cells, murine bone marrow-derived dendritic cells, and murine C3HA fibroblasts all accumulated PGE₂ to high levels in culture supernatants upon infection with MVA. We also demonstrated that MVA induced the release of AA from infected cells, and this was, most unusually, independent of host cytosolic phospholipase A₂ activity. The accumulation of AA and PGE₂ was dependent on viral gene expression, but independent of canonical NF-κB signaling via p65/RelA. The production of PGE₂ required host cyclooxygenase-2 (COX-2) activity, and COX-2 protein accumulated during MVA infection. The results of this study provide insight into a novel aspect of MVA biology that may affect the efficacy of MVA-based vaccines.}, number={2}, journal={VIROLOGY}, author={Pollara, Justin J. and Spesock, April H. and Pickup, David J. and Laster, Scott M. and Petty, Ian T. D.}, year={2012}, month={Jul}, pages={146–155} }
 @article{cecil_davis_cech_laster_2011, title={Inhibition of H1N1 influenza A virus growth and induction of inflammatory mediators by the isoquinoline alkaloid berberine and extracts of goldenseal (Hydrastis canadensis)}, volume={11}, ISSN={1567-5769}, url={http://dx.doi.org/10.1016/j.intimp.2011.06.002}, DOI={10.1016/j.intimp.2011.06.002}, abstractNote={In this study we tested whether the isoquinoline alkaloid berberine can inhibit the growth of influenza A. Our experiments showed strong inhibition of the growth of H1N1 influenza A strains PR/8/34 or WS/33 in RAW 264.7 macrophage-like cells, A549 human lung epithelial-derived cells and murine bone marrow derived macrophages, but not MDCK canine kidney cells. Studies of the mechanism underlying this effect suggest that berberine acts post-translationally to inhibit virus protein trafficking/maturation which in turn inhibits virus growth. Berberine was also evaluated for its ability to inhibit production of TNF-α and PGE(2) from A/PR/8/34 infected-RAW 264.7 cells. Our studies revealed strong inhibition of production of both mediators and suggest that this effect is distinct from the anti-viral effect. Finally, we asked whether berberine-containing ethanol extracts of goldenseal also inhibit the growth of influenza A and production of inflammatory mediators. We found strong effectiveness at high concentrations, although upon dilution extracts were somewhat less effective than purified berberine. Taken together, our results suggest that berberine may indeed be useful for the treatment of infections with influenza A.}, number={11}, journal={International Immunopharmacology}, publisher={Elsevier BV}, author={Cecil, Chad E. and Davis, Jeanine M. and Cech, Nadja B. and Laster, Scott M.}, year={2011}, month={Nov}, pages={1706–1714} }
 @article{cech_kandhi_davis_hamilton_eads_laster_2010, title={Echinacea and its alkylamides: Effects on the influenza A-induced secretion of cytokines, chemokines, and PGE(2) from RAW 264.7 macrophage-like cells}, volume={10}, ISSN={["1878-1705"]}, DOI={10.1016/j.intimp.2010.07.009}, abstractNote={The goal of this study was to determine whether extracts and isolated alkylamides from Echinacea purpurea would be useful for prevention of the inflammatory response that accompanies infections with H1N1 influenza A. Seventeen extracts and 4 alkylamides were tested for the ability to inhibit production of cytokines, chemokines, and PGE2 from RAW 264.7 macrophage-like cells infected with the H1N1 influenza A strain PR/8/34. The alkylamides undeca-2Z,4E-diene-8,10-diynic acid isobutylamide, dodeca-2E,4E,8Z,10E/Z-tetraenoic acid isobutylamide, dodeca-2E,4E-dienoic acid isobutylamide, and undeca-2E-ene-8,10-diynoic acid isobutylamide suppressed production of TNF-α and PGE2 from infected cells. Dodeca-2E,4E-dienoic acid isobutylamide was especially effective at inhibiting production of these mediators and also strongly inhibited production of G-CSF, CCL2/MCP-1, CCL3/MIP-1α and CCL5/RANTES. In contrast, the ethanol extracts (75%), which were prepared from dormant roots of E. purpurea grown in different locations throughout North Carolina, displayed a range of effects from suppression to stimulation of mediator production. Precipitation of the extracts with ethanol removed the stimulatory activity, however, even after precipitation; many of the extracts did not display any suppressive activity. Analysis of the extracts revealed slight variations in concentration of alkylamides, caftaric acid, and cichoric acid, but the activity of the extracts did not strongly correlate with concentrations of these compounds. Our in vitro experiments suggest that E. purpurea extracts have the potential for use in alleviating the symptoms and pathology associated with infections with influenza A; however, further study will be necessary to define procedures necessary to unmask the alkylamide activity in crude extracts.}, number={10}, journal={INTERNATIONAL IMMUNOPHARMACOLOGY}, author={Cech, Nadja B. and Kandhi, Vamsikrishna and Davis, Jeanine M. and Hamilton, Amy and Eads, Dawn and Laster, Scott M.}, year={2010}, month={Oct}, pages={1268–1278} }
 @article{pollara_laster_petty_2010, title={Inhibition of poxvirus growth by Terameprocol, a methylated derivative of nordihydroguaiaretic acid}, volume={88}, ISSN={["0166-3542"]}, DOI={10.1016/j.antiviral.2010.09.017}, abstractNote={Terameprocol (TMP) is a methylated derivative of nordihydroguaiaretic acid, a phenolic antioxidant originally derived from creosote bush extracts. TMP has previously been shown to have antiviral and anti-inflammatory activities, and has been proven safe in phase I clinical trials conducted to evaluate TMP as both a topical and parenteral therapeutic. In the current study, we examined the ability of TMP to inhibit poxvirus growth in vitro, and found that TMP potently inhibited the growth of both cowpox virus and vaccinia virus in a variety of cell lines. TMP treatment was highly effective at reducing infectious virus yield in multi-step virus growth assays, but it did not substantially inhibit the synthesis of infectious progeny viruses in individual infected cells. These contrasting results showed that TMP inhibits poxvirus growth in vitro by preventing the efficient spread of virus particles from cell to cell. The canonical mechanism of poxvirus cell-to-cell spread requires morphogenesis of cell-associated, enveloped virions. The virions then trigger the formation of actin tails to project them from the cell surface. The number of actin tails present at the surface of poxvirus-infected cells was reduced dramatically by treatment with TMP. Whether TMP inhibits poxvirus morphogenesis, or subsequent events required for actin tail formation, remains to be determined. The results of this study, together with the clinical safety record of TMP, support further evaluation of TMP as a poxvirus therapeutic.}, number={3}, journal={ANTIVIRAL RESEARCH}, author={Pollara, Justin J. and Laster, Scott M. and Petty, Ian T. D.}, year={2010}, month={Dec}, pages={287–295} }
 @article{oyegunwa_sikes_wilson_scholle_laster_2010, title={Tetra-O-methyl nordihydroguaiaretic acid (Terameprocol) inhibits the NF-kappa B-dependent transcription of TNF-alpha and MCP-1/CCL2 genes by preventing RelA from binding its cognate sites on DNA}, volume={7}, ISSN={["1476-9255"]}, DOI={10.1186/1476-9255-7-59}, abstractNote={Tetra-O-methyl nordihydroguaiaretic acid, also known as terameprocol (TMP), is a naturally occurring phenolic compound found in the resin of the creosote bush. We have shown previously that TMP will suppress production of certain inflammatory cytokines, chemokines and lipids from macrophages following stimulation with LPS or infection with H1N1 influenza virus. In this study our goal was to elucidate the mechanism underlying TMP-mediated suppression of cytokine and chemokine production. We focused our investigations on the response to LPS and the NF-κB protein RelA, a transcription factor whose activity is critical to LPS-responsiveness.Reporter assays were performed with HEK293 cells overexpressing either TLR-3, -4, or -8 and a plasmid containing the luciferase gene under control of an NF-κB response element. Cells were then treated with LPS, poly(I:C), or resiquimod, and/or TMP, and lysates measured for luciferase activity.RAW 264.7 cells treated with LPS and/or TMP were used in ChIP and EMSA assays. For ChIP assays, chromatin was prepared and complexes precipitated with anti-NF-κB RelA Ab. Cross-links were reversed, DNA purified, and sequence abundance determined by Q-PCR. For EMSA assays, nuclear extracts were incubated with radiolabeled probes, analyzed by non-denaturing PAGE and visualized by autoradiography.RAW 264.7 cells treated with LPS and/or TMP were also used in fluorescence microscopy and western blot experiments. Translocation experiments were performed using a primary Ab to NF-κB RelA and a fluorescein-conjugated secondary Ab. Western blots were performed using Abs to IκB-α and phospho-IκB-α. Bands were visualized by chemiluminescence.In reporter assays with TLR-3, -4, and -8 over-expressing cells, TMP caused strong inhibition of NF-κB-dependent transcription.ChIP assays showed TMP caused virtually complete inhibition of RelA binding in vivo to promoters for the genes for TNF-α, MCP-1/CCL2, and RANTES/CCL5 although the LPS-dependent synthesis of IκB-α was not inhibited. EMSA assays did not reveal an effect of TMP on the binding of RelA to naked DNA templates in vitro.TMP did not inhibit the nuclear translocation of NF-κB RelA nor the phosphorylation of IκB-α.TMP acts indirectly as an inhibitor of NF-κB-dependent transcription by preventing RelA from binding the promoters of certain key cytokine and chemokine genes.}, journal={JOURNAL OF INFLAMMATION-LONDON}, author={Oyegunwa, Akinbolade O. and Sikes, Michael L. and Wilson, Jason R. and Scholle, Frank and Laster, Scott M.}, year={2010}, month={Dec} }
 @article{eads_hansen_oyegunwa_cecil_culver_scholle_petty_laster_2009, title={Terameprocol, a methylated derivative of nordihydroguaiaretic acid, inhibits production of prostaglandins and several key inflammatory cytokines and chemokines}, volume={6}, ISSN={["1476-9255"]}, DOI={10.1186/1476-9255-6-2}, abstractNote={Extracts of the creosote bush, Larrea tridentata, have been used for centuries by natives of western American and Mexican deserts to treat a variety of infectious diseases and inflammatory disorders. The beneficial activity of this plant has been linked to the compound nordihydroguaiaretic acid (NDGA) and its various substituted derivatives. Recently, tetra-O-methyl NDGA or terameprocol (TMP) has been shown to inhibit the growth of certain tumor-derived cell lines and is now in clinical trials for the treatment of human cancer. In this report, we ask whether TMP also displays anti-inflammatory activity. TMP was tested for its ability to inhibit the LPS-induced production of inflammatory lipids and cytokines in vitro. We also examined the effects of TMP on production of TNF-alpha in C57BL6/J mice following a sublethal challenge with LPS. Finally, we examined the molecular mechanisms underlying the effects we observed.RAW 264.7 cells and resident peritoneal macrophages from C57BL6/J mice, stimulated with 1 mug/ml LPS, were used in experiments designed to measure the effects of TMP on the production of prostaglandins, cytokines and chemokines. Prostaglandin production was determined by ELISA. Cytokine and chemokine production were determined by antibody array and ELISA.Western blots, q-RT-PCR, and enzyme assays were used to assess the effects of TMP on expression and activity of COX-2.q-RT-PCR was used to assess the effects of TMP on levels of cytokine and chemokine mRNA.C57BL6/J mice injected i.p. with LPS were used in experiments designed to measure the effects of TMP in vivo. Serum levels of TNF-alpha were determined by ELISA.TMP strongly inhibited the production of prostaglandins from RAW 264.7 cells and normal peritoneal macrophages. This effect correlated with a TMP-dependent reduction in levels of COX-2 mRNA and protein, and inhibition of the enzymatic activity of COX-2.TMP inhibited, to varying degrees, the production of several cytokines, and chemokines from RAW 264.7 macrophages and normal peritoneal macrophages. Affected molecules included TNF-alpha and MCP-1. Levels of cytokine mRNA were affected similarly, suggesting that TMP is acting to prevent gene expression.TMP partially blocked the production of TNF-alpha and MCP-1 in vivo in the serum of C57BL6/J mice that were challenged i.p. with LPS.TMP inhibited the LPS-induced production of lipid mediators and several key inflammatory cytokines and chemokines, both in vitro and in vivo, raising the possibility that TMP might be useful as a treatment for a variety of inflammatory disorders.}, journal={JOURNAL OF INFLAMMATION-LONDON}, author={Eads, D. and Hansen, R. L. and Oyegunwa, A. O. and Cecil, C. E. and Culver, C. A. and Scholle, F. and Petty, I. T. D. and Laster, S. M.}, year={2009}, month={Jan} }
 @article{culver_laster_2007, title={Adenovirus type 5 exerts multiple effects on the expression and activity of cytosolic phospholipase A(2), cyclooxygenase-2, and prostaglandin synthesis}, volume={179}, ISSN={["1550-6606"]}, DOI={10.4049/jimmunol.179.6.4170}, abstractNote={In this study, we examine how infection of murine and human fibroblasts by adenovirus (Ad) serotype 5 (Ad5) affects the expression and activity of cytosolic phospholipase A2 (cPLA2), cyclooxygenase-2 (COX-2), and production of PGs. Our experiments showed that infection with Ad5 is accompanied by the rapid activation of cPLA2 and the cPLA2-dependent release of [3H]arachidonic acid ([3H]AA). Increased expression of COX-2 was also observed after Ad infection, as was production of PGE2 and PGI2. Later, however, as the infection progressed, release of [3H]AA and production of PGs stopped. Late-stage Ad5-infected cells also did not release [3H]AA or PGs following treatment with a panel of biologically diverse agents. Experiments with UV-inactivated virus confirmed that Ad infection is accompanied by the activation of a host-dependent response that is later inhibited by the virus. Investigations of the mechanism of suppression of the PG pathway by Ad5 did not reveal major effects on the expression or activity of cPLA2 or COX-2. We did note a change in the intracellular position of cPLA2 and found that cPLA2 did not translocate normally in infected cells, raising the possibility that Ad5 interferes with the PG pathway by interfering with the intracellular movement of cPLA2. Taken together, these data reveal dynamic interactions between Ad5 and the lipid mediator pathways of the host and highlight a novel mechanism by which Ad5 evades the host immune response. In addition, our results offer insight into the inflammatory response induced by many Ad vectors lacking early region gene products.}, number={6}, journal={JOURNAL OF IMMUNOLOGY}, author={Culver, Carolyn A. and Laster, Scott M.}, year={2007}, month={Sep}, pages={4170–4179} }
 @article{maia_culver_laster_2006, title={Evidence against calcium as a mediator of mitochondrial dysfunction during apoptosis induced by arachidonic acid and other free fatty acids}, volume={177}, ISSN={["0022-1767"]}, DOI={10.4049/jimmunol.177.9.6398}, abstractNote={Apoptosis is often accompanied by activation of phospholipase A2, causing release of free fatty acids (FFAs), which in turn are thought to contribute to the loss of mitochondrial transmembrane potential (Δψm). In these experiments, we asked whether calcium plays a role as an intermediate in this process. A total of 14 FFAs were compared for their ability to cause loss of Δψm and for their ability to affect levels of intracellular calcium. Among the FFAs, unsaturated FFAs tended to induce apoptosis while saturated FFAs did not. Arachidonic acid (AA) was most damaging, causing loss of Δψm and cell death in 8–10 h while linoleic acid, γ-linolenic acid, and docosapentaenoic also strongly induced apoptosis. Effects of the FFAs on levels of intracellular calcium were very different. Many caused strong calcium responses; however, the ability to induce a strong calcium response was not predictive of ability to induce apoptosis, and overall, we did not find a correlation between apoptosis and calcium induction. Also, verapamil and TMB-8 were able to block the calcium response, but these inhibitors did not prevent loss of Δψm, indicating that the calcium response is not necessary for FFA-induced loss of Δψm. In contrast, we found that cyclosporine A could inhibit the AA-induced loss of Δψm with both whole cells and isolated mitochondria, confirming that the antimitochondrial effects of FFA can stem from direct effects on the mitochondrial permeability transition pore. Finally, we show that the strong apoptosis-inducing activity of AA may stem from its ability to selectively induce its own release.}, number={9}, journal={JOURNAL OF IMMUNOLOGY}, author={Maia, Rita C. and Culver, Carolyn A. and Laster, Scott M.}, year={2006}, month={Nov}, pages={6398–6404} }
 @article{culver_michalowski_maia_laster_2005, title={The anti-apoptotic effects of nordihydroguaiaretic acid: Inhibition of cPLA(2) activation during TNF-induced apoptosis arises from inhibition of calcium signaling}, volume={77}, ISSN={["1879-0631"]}, DOI={10.1016/j.lfs.2005.03.023}, abstractNote={Nordihydroguaiaretic acid (NDGA) is a plant lignan produced by Larrea tridentata, the creosote bush of the American southwest. In this report we examine the mechanism underlying the ability of NDGA to inhibit TNF-induced apoptosis. Our results show that NDGA blocks many key indicators of apoptosis. Caspase cleavage, mitochondrial inactivation, externalization of phosphatidyl serine, and (51)Cr-release were all blocked by low micromolar concentrations of NDGA. NDGA also inhibited the cPLA(2)-dependent release of (3)H-arachidonic acid. We investigated this activity and found that NDGA prevented the rise in intracellular calcium necessary for the apoptotic activation of cPLA(2). On the other hand, NDGA did not interfere with the TNF-induced phosphorylation of cPLA(2), indicating that NDGA does not block all TNF-dependent signaling. Finally, we asked whether the anti-apoptotic effect of NDGA could be attributed to its anti-oxidant activity. Comparison with the effects of butylated hydroxyanisole (BHA) did not completely support this hypothesis. While BHA strongly inhibited caspase activation and partially blocked the release of (51)Cr, it was unable to significantly block the calcium response or the release of (3)H-arachidonic acid associated with TNF-induced apoptosis. The anti-oxidant activity of NDGA may, therefore, explain some but not all of its anti-apoptotic activity.}, number={19}, journal={LIFE SCIENCES}, author={Culver, CA and Michalowski, SA and Maia, RC and Laster, SA}, year={2005}, month={Sep}, pages={2457–2470} }
 @article{draper_harris_culver_laster_2004, title={Calcium and its role in the nuclear translocation and activation of cytosolic phospholipase A(2) in cells rendered sensitive to TNF-induced apoptosis by cycloheximide}, volume={172}, ISSN={["1550-6606"]}, DOI={10.4049/jimmunol.172.4.2416}, abstractNote={In these experiments, we investigated the role of calcium as a second messenger in the apoptotic activation of cytosolic phospholipase A2 (cPLA2). As our model, we used a murine fibroblast cell line (C3HA) that was induced to undergo apoptosis by a combination of TNF and cycloheximide. Using fura 2 Ca2+ imaging, we found strong evidence for an intracellular calcium response after 1 h of treatment, which correlated with the onset of phosphatidylserine externalization, but preceded effector procaspase processing by several hours. The response was strongest in the perinuclear region, where mean levels rose 83% (144 ± 14 nM in untreated cells vs 264 ± 39 nM in treated), while cells displaying morphological evidence of apoptosis had the highest levels of calcium (250–1000 nM). Verapamil blocked this response, indicating an extracellular source for the calcium. Fluorescence microscopy revealed a pattern of nuclear translocation of cPLA2 during apoptosis, which was also blocked by verapamil, indicating an important role for calcium in this process. In addition, we found that verapamil prevented the release of [3H]arachidonic acid from C3HA cells induced to undergo apoptosis by the chemotherapeutic agents vinblastine, melphalan, and cis-platinum. Together, these data suggest that calcium is important for cPLA2 activation by diverse apoptotic stimuli.}, number={4}, journal={JOURNAL OF IMMUNOLOGY}, author={Draper, DW and Harris, VG and Culver, CA and Laster, SM}, year={2004}, month={Feb}, pages={2416–2423} }
 @article{feng_tompkins_xu_brown_laster_zhang_mccaw_2002, title={Thymocyte and peripheral blood T lymphocyte subpopulation changes in piglets following in utero infection with porcine reproductive and respiratory syndrome virus}, volume={302}, ISSN={["0042-6822"]}, DOI={10.1006/viro.2002.1650}, abstractNote={Piglets infected in utero with porcine reproductive and respiratory syndrome virus (PRRSV) are born severely immunocompromised. In this article we more closely examine the effects of in utero PRRSV infection on circulating and thymic T cell populations. Numbers of CD4+, CD8+, and dual-positive lymphocytes were quantitated in circulation and in the thymus during the 2 weeks following birth. At birth we found that the number of circulating lymphocytes was suppressed by 60%. Lymphocyte numbers were also suppressed by 42% at day 7, but by day 14 the number of lymphocytes had rebounded and was actually 47% greater than controls. At birth and day 7, a drop in the number of CD4+ cells could partially explain the suppression we observed, while the rebound in total lymphocyte numbers seen at day 14 was due to a nearly fourfold increase in the number of circulating CD8+ cells. As a result, the normal CD4+:CD8+ ratio of between 1.4 and 2.2 for neonatal pigs was reduced to 0.1-0.5. The thymuses of infected piglets were found to be 50% smaller than those of control pigs and were characterized by cortical involution and severe cortical depletion of thymocytes. Analysis of the population of thymocytes revealed that double-positive thymocytes were suppressed to a greater degree than either single positive subpopulation. In addition, we show that the number of thymocytes undergoing apoptosis was increased twofold in piglets infected with PRRSV. Taken together, these results help explain the dramatic immunosuppression observed in neonatal animals infected in utero with PRRSV.}, number={2}, journal={VIROLOGY}, author={Feng, WH and Tompkins, MB and Xu, JS and Brown, TT and Laster, SM and Zhang, HX and McCaw, MB}, year={2002}, month={Oct}, pages={363–372} }
 @article{feng_laster_tompkins_brown_xu_altier_gomez_benfield_mccaw_2001, title={In utero infection by porcine reproductive and respiratory syndrome virus is sufficient to increase susceptibility of piglets to challenge by Streptococcus suis type II}, volume={75}, ISSN={["1098-5514"]}, DOI={10.1128/JVI.75.10.4889-4895.2001}, abstractNote={ABSTRACT Porcine reproductive and respiratory syndrome (PRRS) consistently elevates the frequency of disease and mortality in young pigs. Many different secondary bacterial diseases occur in PRRS virus (PRRSV)-infected pigs. However, to date, establishing a reproducible experimental model of PRRSV infection in weaned pigs, with subsequent clinical disease following secondary bacterial challenge, has been difficult. PRRSV is frequently isolated during outbreaks from weak-born piglets affected by secondary bacterial diseases. This study was performed to investigate the potential role of intrauterine PRRSV infection on piglet susceptibility to secondary bacterial infection. PRRSV-free pregnant sows were intranasally infected at 98 days of gestation with PRRSV strain SD 23983. All piglets born to the PRRSV-infected sows were viremic. Piglets were removed from the sows at birth and deprived of colostrum. Piglets from PRRSV-infected and noninfected sows were randomly assigned to Streptococcus suis challenge or control subgroups. At 5 days of age, piglets were challenged intranasally with strain MN 87555 of S. suis type II. Total and differential leukocyte counts were performed on blood samples collected at 3 days of age. The numbers of leukocytes, lymphocytes, and monocytes were significantly reduced in the PRRSV-infected piglets. Lesions were observed in bone marrow, brain, lung, heart, spleen, lymph node, tonsil, and thymus of PRRSV-infected piglets. Thymus/body weight ratios of in utero PRRSV-infected piglets were significantly reduced compared to those of non-PRRSV-infected piglets, and thymic lesions were characterized by severe cortical depletion of thymocytes. Lesions were not observed in piglets born to PRRSV-free sows. Overall, 20 out of 22 piglets in the PRRSV-S. suis dual-infection group died within 1 week after challenge with S. suis (10 of 11 in each of two trials). This contrasts with 1 of 18 piglets in the PRRSV-infection-only group and 5 of 23 piglets in the S. suis-challenge-only group (1 of 12 in trial 1 and 4 of 11 in trial 2). No piglets died in the uninfected control groups. Most of the piglets in the PRRSV-S. suis dual-infection group developed suppurative meningitis. S. suis type II was recovered from their brains and joints. These results indicate that in utero infection by PRRSV makes piglets more susceptible to infection and disease following challenge by S. suis type II. In utero infection by PRRSV may provide a useful model to study the interaction between PRRSV and bacterial coinfections in piglets.}, number={10}, journal={JOURNAL OF VIROLOGY}, author={Feng, WH and Laster, SM and Tompkins, M and Brown, T and Xu, JS and Altier, C and Gomez, W and Benfield, D and McCaw, MB}, year={2001}, month={May}, pages={4889–4895} }
 @article{wolf_laster_1999, title={Characterization of arachidonic acid-induced apoptosis}, volume={30}, DOI={10.1007/bf02738119}, abstractNote={Tumor necrosis factor (TNF) can induce apoptosis in a number of different cell types. This response often depends on the activity of cytosolic phospholipase A2 (cPLA2), which catalyzes the release of arachidonic acid from the sn-2 position of membrane phospholipids. In this study, we investigate the ability of arachidonic acid itself to cause cell death. We show that in assays with 10% fetal bovine serum (FBS) arachidonic acid will not kill, nor does act synergistically with TNF. In contrast, by lowering the concentration of FBS to 2%, it is possible to use arachidonic acid to induce cell death. Arachidonic acid-induced cell death was judged to be apoptotic based on morphology and the cleavage of poly(ADP)ribose polymerase. Arachidonic acid was able to kill all cell lines tested including two human melanoma-derived cell lines, and susceptibility to arachidonic acid was not influenced by adenovirus gene products that control susceptibility to TNF. Finally, we show that arachidonic acid is unique among 20 carbon fatty acids for its ability to induce apoptosis and that several other unsaturated, but not saturated fatty acids can also induce apoptosis.}, number={3}, journal={Cell Biochemistry and Biophysics}, author={Wolf, L. A. and Laster, S. M.}, year={1999}, pages={353–368} }
 @article{o'brien_piddington_voelkel-johnson_richards_hadley_laster_1998, title={Sustained phosphorylation of cytosolic phospholipase A(2) accompanies cycloheximide- and adenovirus-induced susceptibility to TNF}, volume={161}, number={3}, journal={Journal of Immunology}, author={O'Brien, J. B. and Piddington, D. L. and Voelkel-Johnson, C. and Richards, D. J. and Hadley, L. A. and Laster, S. M.}, year={1998}, pages={1525–1532} }