Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: Colchicine suppresses cholesterol crystal-induced endothelial cell pyroptosis. (a–c) Impacts of increasing cholesterol crystals and colchicine concentrations on the viability of HUVECs. Endothelial cells treated with increasing doses of (a) cholesterol crystals (0-2.0 mg/ml) and (b) colchicine (0-100 nM) and (c) cholesterol crystal (0.5 mg/ml) and different concentrations of colchicine (0-10 nM) for 24 h. The cell viability was detected using a CCK8 assay. (d) Pyroptotic cell death was measured with Hoechst 33342 (blue)/PI (red) double-fluorescent staining (scale bars = 100 μ m). (e) The LDH release was evaluated with a cytotoxicity detection LDH kit. (f) Transmission electron microscopy (TEM) was used to observe the pyroptotic cell morphology (scale bar TEM = 2 μ m). Data was expressed as the mean ± SD of three separate experiments. ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001 vs. the control group. # P < 0.05, ## P < 0.01, and ### P < 0.001 vs. the cholesterol crystal group.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques: CCK-8 Assay, Staining, Transmission Assay, Electron Microscopy, Control

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: Colchicine alleviates NLRP3 inflammasome activation relevant in pyroptosis. HUVECs were treated for 24 h with different doses of colchicine (0–10 nM) in the presence of cholesterol crystal (0.5 mg/ml) or were left untreated (control). (a) Immunofluorescence was used for qualitative detection of NLRP3 (400x). (b) Western blotting was dedicated to examine the protein expression levels of NLRP3, ASC-1, procaspase-1, caspase-1, IL-18, and IL-1 β . (c) Quantitative analysis of pyroptosis-associated protein expression. (d–i) Real-time PCR analysis of the mRNA levels of GSDMD and proinflammatory cytokines (IL-18, IL-1 β , IL-6, IL-8, and MCP-1). Data was expressed as the mean ± SD of three separate experiments. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001 vs. the control group. # P < 0.05, ## P < 0.01, ### P < 0.001, and #### P < 0.0001 vs. the cholesterol crystal group.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques: Activation Assay, Control, Immunofluorescence, Western Blot, Expressing, Real-time Polymerase Chain Reaction

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: Colchicine plays an antipyroptosis role by inhibiting intracellular oxidative stress. HUVECs were treated for 10 or 24 h with different doses of colchicine (0–10 nM) in the presence of cholesterol crystal (0.5 mg/ml) or were left untreated (control). (a) Intracellular ROS level was detected using a DCFH-DA probe. (b) JC-1 Staining Kit was used for the detection of mitochondrial membrane potential. (c, d) The indicators of oxidative stress GSSG and MDA were tested by assay kits in endothelial cells. Endothelial cells treated with increasing doses of H 2 O 2 (0-400 μ M); (e) the cell viability was detected using a CCK8 assay. HUVECs were subjected to H 2 O 2 (300 μ M), cholesterol crystal (0.5 mg/ml) and colchicine (10 nM), or N-acetyl-cysteine (NAC, 5 mM) for 24 hrs. (f) The cell viability was detected using a CCK8 assay. Pyroptotic cell death was evaluated with (g) LDH release and (h) Hoechst 33342/PI staining. Scale bars = 100 μ m. Data was expressed as the mean ± SD of three separate experiments. ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001 vs. the control group. # P < 0.05, ## P < 0.01, ### P < 0.001, and #### P < 0.0001 vs. the cholesterol crystal group. & P < 0.05, &&&& P < 0.0001 vs. the H 2 O 2 group.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques: Control, Staining, Membrane, CCK-8 Assay

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: Colchicine plays an antipyroptosis role by inhibiting intracellular oxidative stress. HUVECs were subjected to H 2 O 2 (300 μ M), cholesterol crystal (0.5 mg/ml) and colchicine (10 nM), or N-acetyl-cysteine (NAC, 5 mM) for 10 h or 24 h. (a) ROS level was detected using a DCFH-DA probe, and (b) mitochondrial membrane potential was tested by JC-1 (scale bars = 100 μ m). (c, d) The indicators of oxidative stress GSSG and MDA were tested by assay kits. (e) Western blotting was dedicated to examine the protein expression levels of NLRP3, ASC-1, procaspase-1, and caspase-1. (f–h) Quantitative analysis of pyroptosis-associated protein expression. (i, j) Real-time PCR analysis of the mRNA levels of GSDMD and proinflammatory cytokines (IL-18 and IL-1 β ). Data was expressed as the mean ± SD of three separate experiments. ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001 vs. the control group. ## P < 0.01, ### P < 0.001, and #### P < 0.0001 vs. the cholesterol crystal group. & P < 0.05, && P < 0.01, &&& P < 0.001, and &&&& P < 0.0001 vs. the H 2 O 2 group.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques: Membrane, Western Blot, Expressing, Real-time Polymerase Chain Reaction, Control

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: Colchicine reverses the downregulation of various antioxidant enzymes and promotes the activation of AMPK-SIRT1 signaling in endothelial cells. HUVECs were treated for 24 h with different doses of colchicine (0–10 nM) in the presence of cholesterol crystal (0.5 mg/ml) or were left untreated (control). (a) Western blotting was dedicated to examine the protein expression levels of SIRT1, p-AMPK α , AMPK α , HO-1, SOD-2, and SOD-1. (b–f) Quantitative analysis of AMPK/SIRT1 pathway-associated protein expression. Data was expressed as the mean ± SD of three separate experiments. ∗ P < 0.05, ∗∗ P < 0.01 vs. the control group. # P < 0.05, ## P < 0.01, and ### P < 0.001 vs. the cholesterol crystal group.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques: Activation Assay, Control, Western Blot, Expressing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: Silencing of AMPK-SIRT1 signaling eliminates the protective effect of colchicine on endothelial cells. HUVECs were transfected with siRNA targeting AMP α 1 (si-AMPK α 1) and SIRT1 (si-SIRT1) or a control siRNA (Si NC) or were not transfected (-). Cells were then added to cholesterol crystal (0.5 mg/ml) in the presence of colchicine (10 nM) for 10 or 24 hrs. Real-time PCR analysis of the mRNA expression of AMPK α and SIRT1 after transfection with si-AMPK α 1, si-SIRT1, and si-NC for 24 hrs without (a) or with (b) the presence of cholesterol crystals and colchicine. (c) Intracellular ROS level was detected using a DCFH-DA probe, and (d) mitochondrial membrane potential was tested by JC-1. (e) Pyroptotic cell death was measured with Hoechst 33342 (blue)/PI (red) double-fluorescent staining. (f, g) The indicators of oxidative stress GSSG and MDA were tested by assay kits. (h) Real-time PCR analysis of the mRNA levels of GSDMD and proinflammatory cytokines (IL-18, IL-1 β , IL-6, IL-8, and MCP-1). Scale bars = 100 μ m. Data was expressed as the mean ± SD of three independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001 vs. the si-NC group or CC+COL+si-NC group.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques: Transfection, Control, Real-time Polymerase Chain Reaction, Expressing, Membrane, Staining

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: Silencing of AMPK-SIRT1 signaling eliminates the protective effect of colchicine on endothelial cells. HUVECs were transfected with siRNA targeting AMP α 1 (si-AMPK α 1) and SIRT1 (si-SIRT1) or a control siRNA (si-NC) or were not transfected (-). Cells were then added to cholesterol crystal (0.5 mg/ml) in the presence of colchicine (10 nM) for 24 hrs. (a) Western blotting was dedicated to examine the protein expression levels of SIRT1, p-AMPK α , AMPK α , HO-1, SOD-2, SOD-1, NLRP3, ASC-1, procaspase-1, and caspase-1 in endothelial cells. (b–i) Quantitative analysis of pyroptosis-associated protein and AMPK/SIRT1 pathway-associated protein expression. Values were expressed as the mean ± SD of three independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001 vs. the CC+COL+si-NC group.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques: Transfection, Control, Western Blot, Expressing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Colchicine Alleviates Cholesterol Crystal-Induced Endothelial Cell Pyroptosis through Activating AMPK/SIRT1 Pathway

doi: 10.1155/2020/9173530

Figure Lengend Snippet: A model diagram showing the possible mechanism of colchicine alleviating cholesterol crystal-induced endothelial cell pyroptosis through activating the AMPK/SIRT1 pathway.

Article Snippet: Cholesterol (C3045) and total ROS scavenger N-acetyl-L-cysteine (NAC, A7250) were purchased from Sigma-Aldrich; colchicine (HY-16569) was obtained from MCE.

Techniques:

Journal: The Pharmacogenomics Journal

Article Title: Genetic screening reveals a link between Wnt signaling and antitubulin drugs

doi: 10.1038/tpj.2015.50

Figure Lengend Snippet: Survival curves and drug correlations. ( a ) Survival curves for RH23; means±s.e.m. ( b ) P -values for drug correlations across RH panel on a negative logarithmic scale. The thin ends of the wedges correspond to lowest drug concentrations. 6 MP, 6-mercaptopurine; COL, colchicine; DMSO, dimethyl sulfoxide; MTX, methotrexate; PX, paclitaxel; RH, radiation hybrid.

Article Snippet: Methotrexate (MTX), 6-mercaptopurine, paclitaxel (PX; Taxol) and colchicine (COL) were obtained from Tocris Bioscience (Bristol, UK).

Techniques:

Journal: Viruses

Article Title: Hepatitis C Virus-Induced FUT8 Causes 5-FU Drug Resistance in Human Hepatoma Huh7.5.1 Cells

doi: 10.3390/v11040378

Figure Lengend Snippet: Inhibition of P-gp and MRP1 increased the 5-FU drug sensitivity after HCV infection. ( A , C ) Western blot analysis of P-gp and MRP1 expression in Huh7.5.1 cells in the presence or absence of NSC23925 (P-gp inhibitor) and MK571 (MRP1 inhibitor). Western blot quantification analysis is also listed as ( B , D ). ( E – H ) Analysis of the cytotoxicity of drugs (5-FU/MTX/ADR/CDDP) in HCVcc-infected Huh7.5.1 cells in the presence of P-gp and MRP1 inhibitors.

Article Snippet: P-glycoprotein (P-gp) inhibitor NSC23925 and multidrug resistance-associated protein-1 (MRP1) inhibitor MK571 were purchased from MedChemExpress (MCE) (Monmouth Junction, NJ, USA) and dissolved in dimethyl sulfoxide (DMSO).

Techniques: Inhibition, Infection, Western Blot, Expressing

Journal: Oncotarget

Article Title: Picropodophyllin causes mitotic arrest and catastrophe by depolymerizing microtubules via Insulin-like growth factor-1 receptor-independent mechanism

doi:

Figure Lengend Snippet: Hep3B and Huh7 cells were treated with either 0.5 μM PPP or 0.05 μM colchicine, followed by visualization of b-tubulin by immunofluorescence using an Alexa-conjugated b-tubulin binding antibody. After 8 h treatment microtubule organization appeared intact in vehicle-treated control cells (DMSO) (left panels) and in PPP-treated cells (middle panels). Microtubule disorganization was observed following treatment with 0.05 μM colchicine (right panels) (A). The fraction of cells with disorganized microtubules at different time points were calculated and depicted in a histogram (B). Data represent mean ± SD, n=3, and considered significant at p ≤ 0.05. (*) both between colchicine- and PPP-treated cells and between PPP- and vehicle-treated cells.

Article Snippet: PPP and picropodophyllotoxin (PPT) were synthesized as described [ ], and similar to colchicine, nocodazole, MG-132 (Sigma, St Louis, MO), S-Trityl-L-Cysteine (STLC) (Tocris Bioscience, Ellisville, MO) and BI 2536 (Selleckchem, Houston, TX) were dissolved in dimethylsulphoxide (DMSO).

Techniques: Immunofluorescence, Binding Assay, Control

Journal: Nature Biomedical Engineering

Article Title: PET-based tracking of CAR T cells and viral gene transfer using a cell surface reporter that binds to lanthanide complexes

doi: 10.1038/s41551-025-01415-7

Figure Lengend Snippet: A schematic representation of the DTPA-R reporter gene system composed of the reporter protein DTPA-R and the cognate PET reporter probe [ 18 F]F-DTPA•metal. a , A molecular model (PyMol) based on the crystal structure of the anticalin/CHX-A″-DTPA•Y complex (PDB ID: 4IAX) and the NMR structure of the CD4 transmembrane domain (PDB ID: 2KLU). b , The design of the coding region for the anticalin-based reporter protein with a promoter, the Lcn2 signal peptide (sp), the mature anticalin, the V5-tag, the CD4 transmembrane domain and, optionally, a fluorescent protein. c , d , The chemical structure of the PET radioligand [ 18 F]F-Nic-Glu 2 -PEG 4 -CHX-A″-DTPA•metal, dubbed [ 18 F]F-DTPA ( c ), and the ligand moiety of [ 18 F]F-colchicine ( d ). e , Fluorescence microscopy of PC3 DTPA-R and PC3 Colchi-R cells stained with Hoechst 33342 (cell nucleus) and an AlexaFluor488-conjugated anti-V5-tag antibody (reporter protein). f – h , Experiments conducted with Jurkat lines, created by retroviral transduction, and FACS isolation of the 10% highest-expressing clones: quantification of reporter protein surface densities by flow cytometry with MESF beads (four biological replicates; mean with s.d.; statistical analysis: multiple unpaired Student’s t -test) ( f ); analysis of transduced Jurkat cell lines for their proliferation kinetics using the CFSE assay (median doubling time is shown in black bars and the quartiles as white lines) ( g ); isolation of transgenic Jurkat DTPA-R cells from a 5:95 mixture with wild-type Jurkat cells using the anti-V5-tag antibody for MACS ( h ).

Article Snippet: The reaction product was cleaved from the resin and conjugated to NH 2 -CHX-A′′-DTPA (Macrocyclics) or deacetylcolchicine, which was prepared from colchicine (BOC Sciences) according to Bagnato et al. .

Techniques: Fluorescence, Microscopy, Staining, Retroviral, Transduction, Isolation, Expressing, Clone Assay, Flow Cytometry, CFSE Assay, Transgenic Assay

Journal: Nature Biomedical Engineering

Article Title: PET-based tracking of CAR T cells and viral gene transfer using a cell surface reporter that binds to lanthanide complexes

doi: 10.1038/s41551-025-01415-7

Figure Lengend Snippet: a , Jurkat cell lines transduced with reporter proteins containing the anticalins binding CHX-A”-DTPA•metal or colchicine, or the single chain variable fragment (scFv) binding DOTA•metal complexes were stained with AlexaFluor488-conjugated SV5-Pk1 anti-V5-tag antibody. Subsequently, the fluorescence was quantified by flow cytometry. The median is shown as black bars, quartiles as white bars, and 100,000 total events were analysed each. b , Average number of genomic transgene insertions by retroviral gene transfer quantified by ddPCR. c , The C825 scFv-based reporter genes show similarly high mRNA levels in qPCR comparable to DTPA-R. d , While a linear correlation between mRNA level and surface-accessible protein levels was found for anticalin-based reporters (dots), the C825 scFv-based reporters showed low reporter protein densities on the cell surface (triangles), hinting towards a superior folding and/or stability of DTPA-R compared to scFv-Rs.

Article Snippet: The reaction product was cleaved from the resin and conjugated to NH 2 -CHX-A′′-DTPA (Macrocyclics) or deacetylcolchicine, which was prepared from colchicine (BOC Sciences) according to Bagnato et al. .

Techniques: Transduction, Binding Assay, Staining, Fluorescence, Flow Cytometry, Retroviral

Journal: Nature Biomedical Engineering

Article Title: PET-based tracking of CAR T cells and viral gene transfer using a cell surface reporter that binds to lanthanide complexes

doi: 10.1038/s41551-025-01415-7

Figure Lengend Snippet: a , The radioligand [ 18 F]F-Nic-Glu 2 -PEG 4 -colchicine (=[ 18 F]F-colchicine) was produced by direct radio-fluorination of the precursor molecule TMA-Nic-Glu 2 -PEG4-colchicine, followed by PSH-H + cartridge purification and subsequent preparative HPLC (250 × 10 mm C18 reversed-phase, isocratic), followed by a C18 cartridge purification. b , Quality control was performed by analytical HPLC using a Chromolith C18 reversed-phase column with a gradient elution (5–55% MeCN, 0.1% TFA). In the reaction product, free fluorine and a single fluorinated product were visible. Cartridge purification on PSH-matrix removed free fluorine, and the HPLC further increased purity by isolating the [ 18 F]F-colchicine radioligand from minor impurities. c , Binding studies of [ 18 F]F-colchicine were conducted on untransduced PC3 cells and sub-cell lines expressing Colchi-R and DTPA-R. The binding of [ 18 F]F-colchicine was highly specific for PC3 Colchi-R cells when compared to the PC3 DTPA-R control cells (510-fold), and the binding to the Colchi-R reporter protein on PC3 cells could be blocked by the previous addition of a high molar excess of 200 µM colchicine, which saturated binding sites (390-fold). d,e , Finally, the pharmacokinetic profile of [ 18 F]F-colchicine was tested in a dynamic PET study. To this end, CD1-nude mice bearing a PC3 DTPA-R and a PC3 Colchi-R xenograft tumour above the shoulders, were injected i.v. with the radioligand. d , In the 2–3 min time frame [ 18 F]F-colchicine is predominantly accumulated in the liver, indicating a mainly hepato-biliary excretion profile. e , VOI quantification of the last time frame of the dynamic PET scan (t = 75–90 min) revealed that 69.7 %ID was present in the gastrointestinal tract (gut), while only 0.7 %ID was taken up in the PC3 Colchi-R tumour and 19.8 %ID was excretion via the renal route and accumulated in the bladder.

Article Snippet: The reaction product was cleaved from the resin and conjugated to NH 2 -CHX-A′′-DTPA (Macrocyclics) or deacetylcolchicine, which was prepared from colchicine (BOC Sciences) according to Bagnato et al. .

Techniques: Produced, Purification, Control, Binding Assay, Expressing, Injection