Anti-LC3B antibody was purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). The Go6976 and bafilomycin A1 (BAF1) were obtained from Selleck, Inc. (Houston, TX, USA). Anti-phospho-PKC-α antibody was purchased from Millipore (Billerica, MA, USA). Diacylglycerol (DAG) enzyme-linked immunosorbent assay (ELISA) kit was purchased from USCN, Inc. (Wuhan, China). MitoTracker® Red CMXRos was purchased from Invitrogen, Inc. (Paisley, UK). Bromodeoxyuridine (BRDU) was purchased from Sigma-Aldrich Co. (St. Louis, MO, USA). Anti-PKC-α antibody was purchased from Bioworld Technology, Inc. (St. Louis Park, MN, USA). Anti-Mfn1, Mfn2, Opa1, Nix, Parkin and ULK1 antibodies were purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, CA, USA). Alexa Fluor 488 was purchased from Jackson ImmunoResearch Inc. (West Grove, PA, USA). Anti-FUNDC1 polyclonal antibody was purchased from Aviva PLC, Inc. (San Diego, CA, USA). Cell counting kit-8 (CCK-8) was purchased from Dojindo Molecular Technologies, Inc. (Kumamoto, Japan). LDH cytotoxicity assay kit was purchased from Keygen Biotech, Co. (Nanjing, China).
Go6976
Manufactured by Selleck Chemicals
Sourced in China
Go6976 is a selective inhibitor of protein kinase C (PKC) enzyme. It has a high degree of selectivity for the PKC isoform.
Automatically generated - may contain errors
Lab products found in correlation
Ly294002, by Selleck Chemicals (3 mentions)
Bromodeoxyuridine (brdu), by Merck Group (1 mentions)
Bafilomycin a1, by Selleck Chemicals (1 mentions)
Anti lc3b antibody, by Cell Signaling Technology (1 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
Mitotracker red cmxros, by Thermo Fisher Scientific (1 mentions)
Ldh cytotoxicity assay kit, by Keygen Biotech (1 mentions)
Alexa fluor 488, by Jackson ImmunoResearch (1 mentions)
Anti gsk3β, by Santa Cruz Biotechnology (1 mentions)
Torin1, by Selleck Chemicals (1 mentions)
Gapdh antibody, by Santa Cruz Biotechnology (1 mentions)
Anti pd l1 pe, by Thermo Fisher Scientific (1 mentions)
Igg pe, by Thermo Fisher Scientific (1 mentions)
Bafilomycin baf, by Merck Group (1 mentions)
Foxp3 antibody, by Thermo Fisher Scientific (1 mentions)
Recombinant human pd 1 fc protein, by R&D Systems (1 mentions)
Lysotracker, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Interleukin 2 (il 2), by R&D Systems (1 mentions)
Monensin, by Merck Group (1 mentions)
6 protocols using go6976
1
Mitochondrial Quality Control Assays
2
Investigating PD-L1 Regulation and Signaling
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SA-49 was synthesized as described previously and dissolved in DMSO [21 (link)]. LY294002, Go6976, 5Z-7-Oxozeaenol and Torin1 were purchased from Selleck (Beijing, China). Cycloheximide (CHX), MG132, and Bafilomycin (Baf) were purchased from Sigma (St. Louis, MO, USA). Antibodies against PD-L1, TFEB, MITF, H3, PKCα, p-GSK3β (Ser9), cleaved caspase 9 and 3 were purchased from Cell Signaling (Danvers, MA, USA). Anti-GSK3β and GAPDH antibodies were purchased from Santa Cruz (Santa Cruz, CA, USA). Anti-PD-L1-PE, IgG-PE and FoxP3 antibodies were purchased from eBioscience (San Diego, CA, USA). Antibodies against p-PKCα (T638), CD3 and Ki67 were obtained from Abcam (Cambridge, MA, USA). The probes LysoTracker and DAPI were purchased from Invitrogen (Carlsbad, CA, USA). Human PD-1 Fc recombinant protein and IL-2 were purchased from R&D Systems (Minneapolis, MN, USA).
3
LRRK2 Modulation and Astrocytic Glutamate Transporter Regulation
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The following compounds were used: dibutyryl cAMP (dbcaMP; 500 μM; 10 days, ChemCruz; #B0218) was exclusively applied to primary striatal LRRK2 WT and G2019S astrocytes to stimulate Glt-1 expression [103 (link)]; LRRK2 kinase inhibitor rel-3-[6-[(2R,6S)-2,6-dimethyl-4-morpholinyl]-4-pyrimidyl]-5-[(1-methylcyclopropyl)oxy]-1H-indazole (MLi-2; 200 nM, 90 min application for the experiments on transfected cells; 120 min application for electrophysiological recordings in oocytes, Tocris; Cat. #5756) [16 (link), 40 (link)]; PKC activator phorbol 12-myristate 13-acetate (TPA; 400 nM, 20 min, Sigma-Aldrich; Cat. #SLBX8889) [103 (link)]; PKC inhibitor Go 6976 (10 µM, 1, 10 and 90 min, Selleckchem) [26 (link), 96 (link)]; recycling inhibitor monensin (35 µM, 40 min; Sigma-Aldrich; Cat. #M5273) [47 (link)]; proteasome inhibitor MG132 (20 µM, 8 h; Sigma-Aldrich; Cat. #M5273 [74 (link)]); vacuolar H+-ATPase inhibitor bafilomycin A1 (Baf A1; 100 nM, 60 min, Selleck.EU; Cat. #S1413 [63 (link)]). For the investigation of Glt-1 recycling kinetics, MLi-2 (200 nM) was acutely applied to LRRK2 WT and LRRK2 G2019S astrocytes 90 min prior to further pharmacological manipulations. For TIRFM experiment and for the treatment of organotypic slices, MLi-2 (200 nM) was acutely dissolved into the medium 90 min prior to immunofluorescence experiments.
4
Cytokine-Induced Signaling Pathway Inhibition
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Recombinant human cytokines IL-17A/F (carrier-free), TNF-α and IFNγ were all obtained from R&D Systems (Oakville, ON, CA). Pharmacological inhibitors, phosphoinositide 3-kinase (PI3K) inhibitor LY294002, protein kinase-C (PKC) inhibitor GO6976 and MAPK/ERK kinase (MEK) inhibitor PD98059 were obtained from SelleckChem (Burlington, ON, CA). The inhibitors were used at a concentration range according to the manufacturer’s instructions. HBEC-3KT cells were pre-treated with the selected inhibitors reconstituted in DMSO (then diluted in airway epithelial cells basal medium containing 6 mM L-glutamine without growth factors to a final dilution of < 1:2000 v/v) one hour prior to cytokine stimulation. Antibodies specific to anti-mouse LCN-2, IL-17A/F and TNF-α, and anti-human Elafin were all obtained from Abcam (Toronto, ON, Canada). Anti-human actin antibody was obtained from Millipore (Burlington, MA, USA). HRP-linked purified anti-rabbit IgG, anti-goat IgG, and anti-mouse IgG-secondary antibodies were all obtained from Cell Signaling Technology (distributed by New England Biolabs, ON, Canada).
5
Evaluating (+)-Borneol's Impact on PMA-Induced NETosis
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(+)-Borneol was obtained from the Simcere Pharmaceutical Group. Polymorphprep was obtained from Axis–Shield. RPMI-1640 was obtained from Gibco. PMA and the NETosis assay kit were obtained from Cayman Chemical. HEPES buffer, enhanced cell counting kit-8 (CCK-8), and ROS assay kit were obtained from Beyotime Biotechnology. The Quant-iT PicoGreen dsDNA Assay Kit was obtained from Invitrogen. SYTOX Green nucleic acid stain and Calcein Blue AM were obtained from Maokang Biotechnology. Diphenyleneiodonium chloride (DPI) and Go6976 were obtained from Selleck. C29 and TAK-242 were obtained from MedChemExpress.
(+)-Borneol was dissolved in DMSO (80 mg/ml) and then diluted with RPMI-1640 to different concentrations (1.56, 6.25, 25, 100, and 400 μM). PMA (1 mg/ml in DMSO) was diluted with RPMI-1640 to 100 nM. The vehicle group had no (+)-borneol or inhibitor but the same PMA and DMSO level as the 400-μM (+)-borneol group. In the control group, neutrophils were treated without PMA.
(+)-Borneol was dissolved in DMSO (80 mg/ml) and then diluted with RPMI-1640 to different concentrations (1.56, 6.25, 25, 100, and 400 μM). PMA (1 mg/ml in DMSO) was diluted with RPMI-1640 to 100 nM. The vehicle group had no (+)-borneol or inhibitor but the same PMA and DMSO level as the 400-μM (+)-borneol group. In the control group, neutrophils were treated without PMA.
6
Resibufogenin-mediated cell signaling
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Resibufogenin was purchased from Shanghai Standard Technology (Shanghai, China) and dissolved in DMSO. Go6983, Go6976, Go6850, Ro31‐8220, and LY294002 were purchased from Selleck (Beijing, China). Rottlerin and Z‐VAD‐FMK were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The GAPDH antibody was obtained from Sigma‐Aldrich (St. Louis, MO, USA). Antibodies against p‐p65 (Ser536), p65, p‐GSK‐3α (Ser21), GSK‐3α, p‐GSK‐3β (Ser9), GSK‐3β, p‐IKKα/β (Ser176/180), IKKα, TAK1, TAB 1, TAB 2, cleaved PARP‐1, cleaved caspase9, cleaved caspase3, p‐Akt (Ser473), p‐PKCα/β (Thr638/641), PKCα, p100/p52, p‐GS (Ser641), GS, and Notch1 were purchased from Cell Signaling (Danvers, MA, USA). Anti‐c‐FLIP antibody was purchased from ENZO (Farmingdale, NY, USA).
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