HSC3 and HSC4 human OSCC cell lines were purchased from Dainihon Pharmaceutical Co. (Tokyo, Japan). Cells were maintained in Dulbecco’s modified essential medium (DMEM; Wako) containing 10% fetal bovine serum (Sigma Chemical Co., St. Louis, MO, USA) under 5% CO2 and 37°C conditions. The glucose concentration in the regular medium was 100 mg/dl. To mimic hyperglycemic conditions, we used DMEM with high glucose concentration (450 mg/ml, WAKO). A-II (1 ng/ml, Abgent, San Diego, CA, USA), LOS (2 μg/ml, Wako), importazole (IPZ, 20 μM, Sigma) and leptomycin B (LMB, 2 μM, Sigma) were used for cell treatment.
Importazole ipz
Manufactured by Merck Group
Sourced in United States
Importazole (IPZ) is a laboratory equipment product designed for use in research and analytical applications. It serves as a critical component in various scientific workflows. The core function of Importazole is to facilitate the import and processing of data from external sources, enabling seamless integration with other laboratory systems and equipment.
Automatically generated - may contain errors
Lab products found in correlation
Dulbecco modified eagle medium (dmem), by Fujifilm (1 mentions)
Leptomycin b lmb, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Lactacystin, by Enzo Life Sciences (1 mentions)
Doxycycline, by Merck Group (1 mentions)
Bafilomycin a1, by InvivoGen (1 mentions)
Q vd oph, by Merck Group (1 mentions)
Ionomycin, by Merck Group (1 mentions)
Blasticidin, by Thermo Fisher Scientific (1 mentions)
Htert rpe1, by American Type Culture Collection (1 mentions)
Wi 38, by American Type Culture Collection (1 mentions)
Crl 1474, by American Type Culture Collection (1 mentions)
Hek293ft, by Thermo Fisher Scientific (1 mentions)
Hff 1, by American Type Culture Collection (1 mentions)
β actin antibody, by Cell Signaling Technology (1 mentions)
Phycoerythrin pe labeled anti human pd l1 monoclonal antibody, by BioLegend (1 mentions)
Pe conjugated anti mouse igg2b antibody, by BioLegend (1 mentions)
Gapdh d16h11 xp rabbit monoclonal antibody, by Cell Signaling Technology (1 mentions)
Horseradish peroxidase hrp conjugated anti rabbit igg antibody, by Cell Signaling Technology (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
4 protocols using importazole ipz
1
Comparison of Oral Squamous Cell Carcinoma Cell Lines
2
Modulating KPNB1 and Proteolytic Pathways
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For KPNB1 inhibition, transfected cells were incubated with 16 µM importazole (IPZ; Sigma-Aldrich) for 48 h. To turn off the expression of Tet-off system, 4.5 µM doxycycline (Sigma-Aldrich) was applied. Activation of endogenous calpains was performed as previously described [18 (link)]. Briefly, calpains were activated by incubating transfected cells with 1 µM ionomycin (Sigma-Aldrich) and 5 mM CaCl2 for 1 h. To block major proteolytic pathways, transfected cells were treated with 10 µM calpain inhibitor III (CI-III; Sigma-Aldrich), 10 µM broad-spectrum caspase inhibitor Q-VD-OPh (Sigma-Aldrich), 50 nM autophagy inhibitor bafilomycin A1 (InvivoGen, Toulouse, France), or 10 µM proteasomal inhibitor lactacystin (Enzo Life Sciences, Lausen, Switzerland) for 18 h prior to harvesting.
3
Culturing Human Cell Lines and Drug Treatments
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Human foreskin fibroblasts (HFF-1) from the American Type Culture Collection (ATCC, Manassas, VA), hTERT-immortalized retinal pigment epithelial cells (hTERT-RPE1; ATCC), and human embryonic kidney cells, 293FT (HEK-293FT; Invitrogen) were cultured as previously described (Kalab et al., 2006 (link); Hasegawa et al., 2013 (link)). Human colorectal carcinoma HCT116 cells were cultured in DMEM with 10% fetal bovine serum (FBS). Normal human fibroblasts CRL1474 (ATCC) were cultured in MEM with 15% FBS and normal lung fibroblasts Wi38 (ATCC) in MEM with 10% FBS. Media of cells expressing RCC1-V5 was supplemented with 10 μg/ml blasticidin (Invitrogen). All cells were grown in a 37°C humidified incubator with 5% CO2 and in media supplemented with 100 U/ml penicillin and 10 μg/ml streptomycin. Cells were treated with Importazole (IPZ; Sigma-Aldrich, St. Louis, MO; final concentration 40 μM) and KPT-330 (Karyopharm, Newton, MA; final concentration 500 nM) by supplementing their complete cell culture with aliquots of 10 mM solutions of the drugs in dimethyl sulfoxide.
4
Regulation of PD-L1 by KPNB1 and IRF1
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The KPNB1 inhibitor importazole (IPZ) and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich (St. Louis, MO, USA). β-actin antibody (#4967), GAPDH (D16H11) XP® rabbit monoclonal antibody (#5174), IRF1 (D5E4) XP® rabbit monoclonal antibody (#8478), KPNA2 rabbit antibody (#14372), and anti-rabbit IgG horseradish peroxidase (HRP)-conjugated antibody (#7074) were purchased from Cell Signaling Technology Japan, K.K. (Tokyo, Japan). Phycoerythrin (PE)-labeled anti-human PD-L1 monoclonal antibody (#329706) and PE-conjugated anti-mouse IgG2b antibody (#400314) were purchased from BioLegend (San Diego, CA, USA). Ambion Silencer® Select Pre-designed small interfering RNA (siRNA) against the genes encoding IRF1 (cat. no. s7501) and KPNA2 (cat. no. s7922), and Silencer® Select Negative #1 Control siRNA (cat. no. AM4611) were purchased from Thermo Fisher Scientific (Waltham, MA, USA).
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