Journal: Cell Death & Disease

Article Title: ELK3 destabilization by speckle-type POZ protein suppresses prostate cancer progression and docetaxel resistance

doi: 10.1038/s41419-024-06647-0

Figure Lengend Snippet: a Dephosphorylation of ELK3 prevent the interaction between ELK3 and SPOP. The cell lysates of HEK293T cells transiently transfected with indicated plasmids were treated with/without λ-phosphatase. The interaction between ELK3 and SPOP was evaluated by IP and WB. b CHK1/2 inhibition suppresses ELK3 destabilization. The cell lysates of HeLa cells treated with 5 μM of indicated inhibitor and CHX (10 μg/ml) for 12 h were used to evaluate the ELK3 protein levels by WB. c CHK1/2 inhibition abolishes ELK3 and SPOP interaction. The cell lysates of HEK293T cells transfected with indicated plasmids treated with 5 μM of indicated inhibitor for 12 h and MG132 (10 μM) for 4 h were used to evaluate the interaction between ELK3 and SPOP by IP and WB. d ELK3 interacts with CHK1 and CHK2. The cell lysates of HEK293T cells transiently transfected with indicated plasmids and treated with MG132 (10 μM) for 4 h were used to evaluate the interaction between ELK3 and CHK1 or CHK2 by IP and WB. e CHK2 facilitates ELK3 ubiquitination by SPOP. cell lysates of HEK293T cells transiently transfected with indicated plasmids and treated with MG132 (10 μM) for 4 h were used to evaluate the ELK3 ubiquitination by IP and WB. f CHK1/2 inhibition abrogates SPOP-mediated ELK3 ubiquitination. The cell lysates of HEK293T cells transiently transfected with indicated plasmids and treated with 5 μM of AZD7762 for 12 h and MG132 (10 μM) for 4 h were used to evaluate the SPOP-mediated ELK3 ubiquitination by IP and WB. g CHK1 knockdown increases ELK3 protein levels. The cell lysates of HeLa cells stably expressing sh-mock or sh-CHK1 or CHK2 were used to evaluate ELK3 protein levels by WB. h CHK1 and CHK2 phosphorylates ELK3. In vitro kinase assay using partially purified His-ELK3 and active CHK1 or CHK2 was conducted. ELK3 phosphorylation by CHK1 or CHK2 was evaluated RxxS/T antibody by WB. i ELK3 deg1 deletion abolishes CHK1- or CHK2-mediated phosphorylation. In vitro kinase assay using partial purified His-ELK3-wt or -∆Deg1 and active CHK1 or CHK2 was conducted. ELK3 phosphorylation by CHK1 or CHK2 was evaluated RxxS/T antibody by WB. j , k CHK2 phosphorylates ELK3 at Ser133 in cell system. The phosphorylation of ELK3 mediated by CHK2 in cell culture ( j ) and in vitro kinase assay system ( k ) was evaluated using phos-tag immunoblot analysis. Treatment with AZD7762 ( j ) and mutation of ELK3 Ser133 to Ala ( l ) abolished the phosphorylation of ELK3 induced by CHK2. l ELK3 phosphorylation at Ser133 is indispensable to interact with SPOP. ELK3 phosphorylation requirement for the interaction with SPOP was evaluated by IP and western blotting using cell lysates transiently expressing mock, His-ELK3-wt, His-ELK3-S133D, or His-ELK3-S133A.

Article Snippet: Subsequently, the partially purified ELK3-wt (1 μg) and ELK3-ΔDeg1 (1 μg) were combined with active CHK1 (cat. no.: C47-10H, SignalChem, Richmond, BC, Canada) and CHK2 (cat. no.: C48-10G, SignalChem) and cold ATP.

Techniques: De-Phosphorylation Assay, Transfection, Inhibition, Ubiquitin Proteomics, Knockdown, Stable Transfection, Expressing, In Vitro, Kinase Assay, Purification, Phospho-proteomics, Cell Culture, Western Blot, Mutagenesis

Journal: Frontiers in Immunology

Article Title: Neuronal substance P-driven MRGPRX2-dependent mast cell degranulation products differentially promote vascular permeability

doi: 10.3389/fimmu.2024.1477072

Figure Lengend Snippet: Binding of compound 48/80 or ciprofloxacin to MRGPRX2 strongly promotes PMC degranulation and murine vascular permeability more than the binding to Mrgprb2. (A) Surface expression levels of FcεRIα and c-Kit (upper panel) and MRGPRX2 (middle panel) and expression levels of tdTomato (lower panel) in the PMCs from WT, Mrgprb2-KO (b2-KO), and MRGPRX2-KI (X2-KI) mice. Control staining is shown in the shaded histograms. (B, D, F) Percentages of surface CD63-positive cells in WT, Mrgprb2-KO, and MRGPRX2-KI PMCs and BMMCs (B) and PMCs (D, F) after treatment with the indicated concentrations of compound 48/80 (B) , 10 μg/mL ciprofloxacin (D) , or 10 μg/mL icatibant (F) . Data are representative of three independent experiments and indicate the mean ± SD. * P < 0.05 and ** P < 0.01. (C, E, G) Quantification of the Evans blue dye that extravasated into the ear skin in WT, Mrgprb2-KO, and MRGPRX2-KI mice after treatment with indicated amounts of compound 48/80 (C) , ciprofloxacin (E) , and 1.75 μg icatibant (G) or phosphate-buffered saline (PBS). n = 5-9; ± SD. * P < 0.05 and ** P < 0.01.

Article Snippet: Compound 48/80 and capsaicin (Sigma-Aldrich, St Louis, MO), SP (Peptide Institute, Inc., Osaka, Japan), Cetirizine (TCI AMERICA, Portland, OR), TY-51469 (MedChemExpress, Monmouth Junction, NJ), RWJ-56110 (Tocris Bioscience, Ellisville, MO), AZ3451, I-191, AMG-517 (Selleck Chemicals LLC), AMG-9810 (Cayman Chemical, Ann Arbor, MI), DAPI (4’,6-diamidino-2-phenylindole) (FujifilmWako, Osaka, Japan), and Piperine (TCI, Tokyo, Japan) were used.

Techniques: Binding Assay, Permeability, Expressing, Control, Staining, Saline

Journal: Chemistry Central Journal

Article Title: Novel enaminone derived from thieno [2,3- b ] thiene: Synthesis, x-ray crystal structure, HOMO, LUMO, NBO analyses and biological activity

doi: 10.1186/s13065-015-0100-9

Figure Lengend Snippet: The natural atomic charges calculated at the B3LYP/6-311G (d, p)

Article Snippet: BD (2) C49-C51 , BD*(2) O4-C48 , 26.57.

Techniques:

Journal: Chemistry Central Journal

Article Title: Novel enaminone derived from thieno [2,3- b ] thiene: Synthesis, x-ray crystal structure, HOMO, LUMO, NBO analyses and biological activity

doi: 10.1186/s13065-015-0100-9

Figure Lengend Snippet: The second order perturbation energies E (2) (kcal/mol) of the most important charge transfer interactions in the studied compound using B3LYP method

Article Snippet: BD (2) C49-C51 , BD*(2) O4-C48 , 26.57.

Techniques:

Journal: Molecules

Article Title: Formononetin Inhibits Mast Cell Degranulation to Ameliorate Compound 48/80-Induced Pseudoallergic Reactions

doi: 10.3390/molecules28135271

Figure Lengend Snippet: FNT inhibits C48/80-induced cell degranulation in RBL-2H3. ( A ) Chemical structure of FNT. ( B ) Cell viability of RBL-2H3 cells treated with 0–100 µM FNT was determined using CCK-8 assays. ( C – F ) The inhibitory effect of FNT on the secretion of β-Hex ( C ), histamine ( D ), and the transcriptional level of proinflammatory factors TNFα ( E ) and IL-13 ( F ) in C48/80-mediated RBL-2H3 cells. The data are expressed as the mean ± SDs (n = 3). ### p < 0.001 vs. the control group; * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. the model group. FNT, formononetin; β-Hex, β-hexosaminidase; HIS, histamine; Dexa, dexamethasone.

Article Snippet: C48/80 was purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: CCK-8 Assay

Journal: Molecules

Article Title: Formononetin Inhibits Mast Cell Degranulation to Ameliorate Compound 48/80-Induced Pseudoallergic Reactions

doi: 10.3390/molecules28135271

Figure Lengend Snippet: FNT inhibits C48/80-mediated morphological changes in RBL-2H3 cells. RBL-2H3 cells were pre-treated with or without the indicated doses of FNT for 2 h and then stimulated with C48/80 for 10 min. ( A , B ) Representative morphological changes in RBL-2H3 cells were shown via toluidine blue staining. Red arrows indicate the cells that become irregular in shape and release purple particles. ( C , D ) FITC-phalloidin staining showed morphological changes in cells. F-actin cytoskeleton was stained with green fluorescence. Red arrow indicates oval cells in response to F-actin cytoskeleton changes. Means ± SDs (n = 3) are shown; ### p < 0.001 vs. the control group; ** p < 0.01, and *** p < 0.001 vs. the model group. Abbreviations as in .

Article Snippet: C48/80 was purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Staining, Fluorescence

Journal: Molecules

Article Title: Formononetin Inhibits Mast Cell Degranulation to Ameliorate Compound 48/80-Induced Pseudoallergic Reactions

doi: 10.3390/molecules28135271

Figure Lengend Snippet: FNT inhibits C48/80-increased NF-κB activity in RBL-2H3 cells. RBL-2H3 cells were pre-treated (or not) with different doses of FNT for 2 h separately and then stimulated with C48/80 for 10 min, the whole cell lysate was taken. ( A ) Western blot analysis of NF-κB pathway signaling molecules (p-p65, p65, p-IκBα, and IκBα) in C48/80-stimulated RBL-2H3 cells treated with 10, 20, or 40 µM FNT. ( B , C ) Quantification of the proteins. ( D ) RBL-2H3 cells transfected with pNF-κB-Luc plasmid were treated with FNT for 4h, and the selective inhibition of NF-κB pathway via FNT was detected using dual luciferase reporter gene assay. Means ± SDs (n = 3) are shown; ### p < 0.001 vs. the control group; * p < 0.05, and *** p < 0.001 vs. the model group. Abbreviations as in .

Article Snippet: C48/80 was purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Activity Assay, Western Blot, Transfection, Plasmid Preparation, Inhibition, Luciferase, Reporter Gene Assay

Journal: Molecules

Article Title: Formononetin Inhibits Mast Cell Degranulation to Ameliorate Compound 48/80-Induced Pseudoallergic Reactions

doi: 10.3390/molecules28135271

Figure Lengend Snippet: FNT suppresses C48/80-induced cell degranulation in primary BMMCs. ( A ) Cell viability of BMMCs treated with 0–100 µM FNT was determined using CCK-8 assays. ( B – E ) The inhibitory effect of FNT on the secretion of β-Hex ( C ), histamine ( D ), and RBL-2H3 cells were pre-treated (or not) with different doses of FNT for 2 h separately and then stimulated with C48/80 for 2h. and the transcriptional level of inflammatory factors TNFα ( E ) and IL-13 ( F ) in C48/80-stimulated BMMCs. The data are expressed as the mean ± SDs (n = 3). ### p < 0.001 vs. the control group; * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. the model group. Abbreviations as in .

Article Snippet: C48/80 was purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: CCK-8 Assay

Journal: Molecules

Article Title: Formononetin Inhibits Mast Cell Degranulation to Ameliorate Compound 48/80-Induced Pseudoallergic Reactions

doi: 10.3390/molecules28135271

Figure Lengend Snippet: FNT attenuates C48/80-mediated pseudoallergic reactions in PCA Mice. ( A ) Representative images showed the diffusion of Evans blue from paws of PCA mice. ( B ) Representative pictures of pathological changes in PCA mice. H&E staining showed the degree of telangiectasia, and toluidine blue staining showed MC infiltration. Small blue particles indicated by arrows in toluidine blue staining are infiltrating mast cells. ( C ) Evans blue diffusion quantified at 620 nm. ( D ) Quantification of the change in paw thickness after stimulation. The data are shown as the mean ± SDs (n = 6); ## p < 0.01, ### p < 0.001 vs. the control group, * p < 0.05, ** p < 0.01, *** p < 0.001 vs. the model group. Abbreviations as in .

Article Snippet: C48/80 was purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Diffusion-based Assay, Staining

Journal: Molecules

Article Title: Formononetin Inhibits Mast Cell Degranulation to Ameliorate Compound 48/80-Induced Pseudoallergic Reactions

doi: 10.3390/molecules28135271

Figure Lengend Snippet: FNT attenuated C48/80-mediated pseudoallergic reactions in ASA mice. ( A ) The protocol for ASA experiment. ( B ) Changes in rectal temperature after injecting with C48/80 were measured every 5 min over the next 30 min in each group. ( C ) Effect of FNT on the release of histamine in ASA mice (determined using ELISA). The data are shown as the mean ± SDs (n = 6); ## p < 0.01, ### p < 0.001 vs. the control group, * p < 0.05, ** p < 0.01, *** p < 0.001 vs. the model group. Abbreviations as in .

Article Snippet: C48/80 was purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Oncotarget

Article Title: An extract from the Atlantic brown algae Saccorhiza polyschides counteracts diet-induced obesity in mice via a gut related multi-factorial mechanisms

doi: 10.18632/oncotarget.18113

Figure Lengend Snippet: Food intake and fecal excretion is increased in high fat diet fed mice supplemented with a brown algae extract (BAE)

Article Snippet: Fecal caloric value was determined with a bomb calorimeter (C 7000, cooler C7002, oxygen filling station C48, IKA ® -Werke GmbH & Co. KG, Staufen, Germany) and apparent food digestibility was calculated as (total energy intake - fecal energy excretion)/total energy intake*100.

Techniques: Algae

Journal: Nucleic Acids Research

Article Title: Kissing-loop interaction between 5′ and 3′ ends of tick-borne Langat virus genome ‘bridges the gap’ between mosquito- and tick-borne flaviviruses in mechanisms of viral RNA cyclization: applications for virus attenuation and vaccine development

doi: 10.1093/nar/gkw061

Figure Lengend Snippet: Immunogenicity and protective efficacy of C48–124(2)/9/1-E5 virus in 3-week old C3H mice

Article Snippet: To evaluate the immunogenicity of miRNA-targeted viruses, groups of five 3-week-old female C3H mice (Taconic) were infected intraperitoneally with 10 5 pfu of C48–124(2)/9/1-E5 virus or mock inoculated with L-15 medium supplemented with 1× SPG and monitored for signs of neurotropic disease daily until 28 dpi.

Techniques: Immunopeptidomics, Virus