Journal: International Journal of Molecular Sciences

Article Title: Evaluation of Stable LifeAct-mRuby2- and LAMP1-NeonGreen Expressing A549 Cell Lines for Investigation of Aspergillus fumigatus Interaction with Pulmonary Cells

doi: 10.3390/ijms22115965

Figure Lengend Snippet: Infection of A549 and LifeAct-mRuby2 cells with A. fumigatus results in formation of actin rings around internalized fungal conidia. ( a ) Actin rings in A549 cells (white arrows). Immunofluorescence: Phalloidin-Alexa 555 (actin) in red, GFP-conidia in green, ( b ) actin rings in LifeAct-mRuby2-expressing cells (white arrows). GFP-conidia in green, ( c ) actin rings in LifeAct-mRuby2-expressing cells (white arrows). Immunofluorescence: Phalloidin-Alexa 350 (actin) in blue, GFP-conidia in green, ( d ) actin rings (white arrows) in A549 and LifeAct-mRuby2 cells visualized by phalloidin-Alexa 488 (green), Dapi in blue. ( e ) Expression and phosphorylation of cofilin in cells treated with gliotoxin. Red asterisk points on phosphorylation of cofilin after treatment with gliotoxin in control A549 cells. GT: Gliotoxin. Scale bar 10 µm. See also .

Article Snippet: Gliotoxin (MedChemExpress, Monmouth Junction, NJ, USA) was applied for 30 min, with a concentration of 50 ng/mL as previously described [ ].

Techniques: Infection, Immunofluorescence, Expressing, Phospho-proteomics, Control

Journal: The Journal of Biological Chemistry

Article Title: Decreased Superoxide Production in Macrophages of Long-lived p66Shc Knock-out Mice *

doi: 10.1074/jbc.M109.017491

Figure Lengend Snippet: 600,000 peritoneal macrophages preincubated or not, as indicated, with gliotoxin were resuspended in KRP medium, containing H2HFF-BSA dye according to the dye manufacturer's instructions. Initial readings at excitation 480 nm and emission 520 nm were taken during the first 12 min. PMA was added at the point shown by the arrow, and readings were taken during the next 30 min. Error bars, S.E. (n = 2).

Article Snippet: Antibodies and Reagents Diphenyleneiodonium (DPI) 2 and gliotoxin were purchased from Axxora LLC (San Diego, CA), phorbol 12-myristate 13-acetate (PMA) was from Enzo Life Sciences International Inc. (Plymouth Meeting, PA); N -formyl-Met-Leu-Phe (fMLP) was from Tocris Bioscience (Ellisville, MO); arachidonic acid (AA) was from Acros Organics (Morris Plains, NJ); OxyBURST Green H 2 HFF-BSA dye was purchased from Molecular Probes, Inc. (Eugene, OR); goat anti-p22phox antibody was from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA); rabbit anti-p40phox polyclonal and rabbit anti-p67phox polyclonal antibodies were from Upstate Cell Signaling Solutions (Lake Placid, NY); rabbit anti-phospho-p40phox polyclonal antibody was from Cell Signaling Technology Inc. (Danvers, MA); goat anti-p47phox polyclonal, goat anti-Rac2 polyclonal, mouse anti-PKCδ polyclonal, and rabbit anti-phospho-PKCδ monoclonal antibody were from Abcam Inc. (Cambridge, MA); rabbit anti-Shc polyclonal antibodies and mouse anti-gp91phox antibody were from BD Biosciences; Rac/Cdc42 assay reagent PAK-1 p21-binding domain-agarose conjugate and mouse monoclonal anti-Rac1 antibody were from Millipore Inc. (Temecula, CA); and goat anti-rabbit monoclonal antibody labeled with infrared (IR) dye 700CW, donkey anti-mouse monoclonal antibody labeled with IR dye 800CW, and donkey anti-goat polyclonal antibodies conjugated with IR dye 800CW were from Li-Cor Biosciences (Lincoln, NE).

Techniques:

Journal: The Journal of Biological Chemistry

Article Title: Decreased Superoxide Production in Macrophages of Long-lived p66Shc Knock-out Mice *

doi: 10.1074/jbc.M109.017491

Figure Lengend Snippet: Total protein was extracted from mouse peritoneal macrophages of control and p66Shc(−/−) mice, mock-treated or treated with PMA or pretreated with gliotoxin and treated with PMA, as indicated. Blots were probed with rabbit polyclonal antibody to p40phox, p47phox, p67phox, and gp91phox. At the same time, blots were probed with mouse anti-actin β. Bands of the NAD(P)H-oxidase components and β-actin are indicated.

Article Snippet: Antibodies and Reagents Diphenyleneiodonium (DPI) 2 and gliotoxin were purchased from Axxora LLC (San Diego, CA), phorbol 12-myristate 13-acetate (PMA) was from Enzo Life Sciences International Inc. (Plymouth Meeting, PA); N -formyl-Met-Leu-Phe (fMLP) was from Tocris Bioscience (Ellisville, MO); arachidonic acid (AA) was from Acros Organics (Morris Plains, NJ); OxyBURST Green H 2 HFF-BSA dye was purchased from Molecular Probes, Inc. (Eugene, OR); goat anti-p22phox antibody was from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA); rabbit anti-p40phox polyclonal and rabbit anti-p67phox polyclonal antibodies were from Upstate Cell Signaling Solutions (Lake Placid, NY); rabbit anti-phospho-p40phox polyclonal antibody was from Cell Signaling Technology Inc. (Danvers, MA); goat anti-p47phox polyclonal, goat anti-Rac2 polyclonal, mouse anti-PKCδ polyclonal, and rabbit anti-phospho-PKCδ monoclonal antibody were from Abcam Inc. (Cambridge, MA); rabbit anti-Shc polyclonal antibodies and mouse anti-gp91phox antibody were from BD Biosciences; Rac/Cdc42 assay reagent PAK-1 p21-binding domain-agarose conjugate and mouse monoclonal anti-Rac1 antibody were from Millipore Inc. (Temecula, CA); and goat anti-rabbit monoclonal antibody labeled with infrared (IR) dye 700CW, donkey anti-mouse monoclonal antibody labeled with IR dye 800CW, and donkey anti-goat polyclonal antibodies conjugated with IR dye 800CW were from Li-Cor Biosciences (Lincoln, NE).

Techniques:

Journal: PLOS ONE

Article Title: In silico exploration of deep-sea fungal metabolites as inhibitor of Ebola and Marburg VP35 and VP40

doi: 10.1371/journal.pone.0307579

Figure Lengend Snippet: Selection of best inhibitor for protein based on physiochemical properties of protein-ligand complexes.

Article Snippet: A model system was used in which Gliotoxin was used as a ligand with Marburg virus VP40 Dimer, Austinol with matrix protein VP40 from Ebola virus Sudan, Ozazino-cyclo-(2,3-dihydroxyl-trp-tyr) with Ebola VP35 Interferon Inhibitory Domain, and Dehydroaustinol with VP35 from Marburg virus.

Techniques: Selection, Virus

Journal: PLOS ONE

Article Title: In silico exploration of deep-sea fungal metabolites as inhibitor of Ebola and Marburg VP35 and VP40

doi: 10.1371/journal.pone.0307579

Figure Lengend Snippet: Glide score of protein-ligand complexes.

Article Snippet: A model system was used in which Gliotoxin was used as a ligand with Marburg virus VP40 Dimer, Austinol with matrix protein VP40 from Ebola virus Sudan, Ozazino-cyclo-(2,3-dihydroxyl-trp-tyr) with Ebola VP35 Interferon Inhibitory Domain, and Dehydroaustinol with VP35 from Marburg virus.

Techniques: Virus

Journal: PLOS ONE

Article Title: In silico exploration of deep-sea fungal metabolites as inhibitor of Ebola and Marburg VP35 and VP40

doi: 10.1371/journal.pone.0307579

Figure Lengend Snippet: (a) The ligand Gliotoxin is docked with Marburg virus VP40 dimer (b) The interactive 3D viewer illustrating selected binding modes (c) Residue wise interaction between ligand and protein obtained through Discovery Studio.

Article Snippet: A model system was used in which Gliotoxin was used as a ligand with Marburg virus VP40 Dimer, Austinol with matrix protein VP40 from Ebola virus Sudan, Ozazino-cyclo-(2,3-dihydroxyl-trp-tyr) with Ebola VP35 Interferon Inhibitory Domain, and Dehydroaustinol with VP35 from Marburg virus.

Techniques: Virus, Binding Assay, Residue

Journal: PLOS ONE

Article Title: In silico exploration of deep-sea fungal metabolites as inhibitor of Ebola and Marburg VP35 and VP40

doi: 10.1371/journal.pone.0307579

Figure Lengend Snippet: (a) The negative binding energy for the Marburg virus VP40 dimer and Gliotoxin complex represents the stability in the structure of the protein after the attachment of Gliotoxin ligand. (b) The decrease in the enthalpy makes the complex stable while the entropy is showing an unfavorable interaction protein between the protein and the ligand Gliotoxin.

Article Snippet: A model system was used in which Gliotoxin was used as a ligand with Marburg virus VP40 Dimer, Austinol with matrix protein VP40 from Ebola virus Sudan, Ozazino-cyclo-(2,3-dihydroxyl-trp-tyr) with Ebola VP35 Interferon Inhibitory Domain, and Dehydroaustinol with VP35 from Marburg virus.

Techniques: Binding Assay, Virus

Journal: PLOS ONE

Article Title: In silico exploration of deep-sea fungal metabolites as inhibitor of Ebola and Marburg VP35 and VP40

doi: 10.1371/journal.pone.0307579

Figure Lengend Snippet: The toxicity profiles of the identified compounds.

Article Snippet: A model system was used in which Gliotoxin was used as a ligand with Marburg virus VP40 Dimer, Austinol with matrix protein VP40 from Ebola virus Sudan, Ozazino-cyclo-(2,3-dihydroxyl-trp-tyr) with Ebola VP35 Interferon Inhibitory Domain, and Dehydroaustinol with VP35 from Marburg virus.

Techniques: Drug discovery

Journal: Scientific Reports

Article Title: KIF1Bβ increases ROS to mediate apoptosis and reinforces its protein expression through O 2 − in a positive feedback mechanism in neuroblastoma

doi: 10.1038/s41598-017-17192-6

Figure Lengend Snippet: Gliotoxin induces O 2 − to increase KIF1Bβ protein expression and apoptosis in neuroblastoma cells. (A) Flow cytometric analysis of intracellular ROS in CHP212 and (B) SK-N-SH cells after 1 hour of Gliotoxin treatment at 50 nM and 300 nM respectively. (C) Fold change in intracellular O 2 − determined using lucigenin-based chemiluminescence assay in CHP212 and (D) SK-N-SH cells after 24 hours (50 nM) and 12 hours (300 nM) of Gliotoxin treatment respectively (mean ± SD; n = 3; *P < 0.05; **P < 0.01). (E) Immunoblot analysis of CHP212 and (F ) SK-N-SH cells after 24 hours (50 nM) and 12 hours (300 nM) of Gliotoxin treatment respectively. Right – corresponding densitometry for KIF1Bβ and cleaved caspase-3 (CC3) expression (mean ± SD; n = 3; *P < 0.05; **P < 0.01). (G) Corresponding flow cytometric analysis of Propidium iodide/Annexin V (PI/AV) apoptotic cell staining in CHP212 and (H) SK-N-SH cells after 24 hours (50 nM) and 12 hours (300 nM) of Gliotoxin treatment respectively (mean ± SD; n = 3; **P < 0.01; ***P < 0.001). (I) Crystal violet staining to determine colony formation ability of CHP212 and (J) SK-N-SH cells that were treated with Gliotoxin for every 24 hours (50 nM) and 12 hours (300 nM) respectively, for several days.

Article Snippet: A stock solution of 1 mM Gliotoxin (ApexBio; Cat no. A4443) was prepared with DMSO and serial dilutions were prepared with PBS.

Techniques: Expressing, Chemiluminescence Immunoassay, Western Blot, Staining

Journal: Toxins

Article Title: Gliotoxin Aggravates Experimental Autoimmune Encephalomyelitis by Triggering Neuroinflammation

doi: 10.3390/toxins11080443

Figure Lengend Snippet: GTX enhances neuroinflammation and demyelination in EAE mice. Normal and EAE mice received three GTX doses (1.0 mg/kg). Inflammatory infiltration was evaluated in hematoxylin/eosin ( A ) and demyelination in luxol fast blue ( B ) stained sections of control (CTL) ( 1 ), gliotoxin (GTX) ( 2 ), EAE ( 3 ), and EAE/GTX ( 4 ) groups. The blood-spinal cord barrier permeability test (NaFlu uptake) was assessed in spinal cord homogenates obtained from non-EAE mice ( C ). NaFlu uptake results are representative of one experiment and the statistical analysis was performed between CTL vs. GTX groups ( n = 6), * p < 0.05.

Article Snippet: Gliotoxin (GTX) from Gliocladium fimbriatum (Merck Millipore Corporation, Darmstadt, Germany) was initially diluted in dimethyl sulfoxide solution (DMSO, 1.0 mg/mL, ≥99.5% GC, Sigma) and then adjusted with 0.9% sterile saline solution (SSS) to be intraperitoneally injected in mice (1.0 mg/kg).

Techniques: Staining, Control, Permeability