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Staurosporine

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Staurosporine is a chemical compound used in scientific research. It is a protein kinase inhibitor that can modulate various cellular processes. The core function of Staurosporine is to inhibit protein kinases, which are enzymes that play a role in signal transduction pathways within cells.

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Lab products found in correlation

Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions) Streptomycin, by Thermo Fisher Scientific (2 mentions) L glutamine, by Thermo Fisher Scientific (2 mentions) Penicillin, by Thermo Fisher Scientific (2 mentions) High purity methanol, by Honeywell (2 mentions) Formic acid, by Merck Group (2 mentions) Rwpe 1, by Thermo Fisher Scientific (1 mentions) Rwpe 1, by American Type Culture Collection (1 mentions) Spectramax m5 microplate reader, by Molecular Devices (1 mentions) Actinomycin d, by Santa Cruz Biotechnology (1 mentions) Rhtgf β1, by Thermo Fisher Scientific (1 mentions) Triton x 100, by Merck Group (1 mentions) Paraformaldehyde (pfa), by Merck Group (1 mentions) Dimethyl sulfoxide (dmso), by Merck Group (1 mentions) S camptothecin, by Merck Group (1 mentions) Sodium phosphate dibasic, by Merck Group (1 mentions) Potassium phosphate monobasic, by Merck Group (1 mentions) Hoechst 33342, by AAT Bioquest (1 mentions) Mln4924, by Merck Group (1 mentions) Gw4869, by Merck Group (1 mentions)

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Staurosporine (STS) (3 citations)

17 protocols using staurosporine

1

RWPE-1 Cell Culture and CTRP3 Treatment

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The human prostate epithelial cell line RWPE-1 was purchased from the American Type Culture Collection (ATCC Number CRL-11609). RWPE-1 cells were maintained in keratinocyte serum-free medium (KSFM; GIBCO Laboratories, Grand Island, NY) supplemented with 50 mg/L bovine pituitary extract, 5% l-glutamine and 5 μg/L EGF. RWPE-1 cells were maintained in a humidified incubator (5% CO2) at 37°C.
Cells were treated with various concentrations of CTRP3 as indicated and analyzed as described below. In the control experiments, 0.1 M phosphate buffer (pH 7.2) containing 0.1% gelatin and 40% glycerol was added to the culture.
To detect the effects of a PKC inhibitor, staurosporine (Santa Cruz, CA, USA), on CTRP3-induced proliferation, anti-apoptosis and change of cell cycle, RWPE-1 cells were pretreated with 0.25μmol/L of staurosporine before and during the stimulation with 10 μg/mL CTRP3.
Hou Q., Lin J., Huang W., Li M., Feng J, & Mao X. (2015). CTRP3 Stimulates Proliferation and Anti-Apoptosis of Prostate Cells through PKC Signaling Pathways. PLoS ONE, 10(7), e0134006.
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2

Cytotoxicity Assay with Inhibitors

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HeLa cells (1 × 104 cells) were seeded on 96-well plates and incubated for 1 day before exposure (24 h) to the indicated concentrations of staurosporine and actinomycin D (Santa Cruz Biotechnology). PARP inhibitor and caspase inhibitor were added for 30 min before staurosporine and actinomycin D exposure. Cell numbers were determined using cell counting reagent SF (Nacalai Tesque) according to the manufacturer's instructions, by measuring absorbance at 450 nm (SpectraMax M5 Microplate Reader; Molecular devices).
Mashimo M., Onishi M., Uno A., Tanimichi A., Nobeyama A., Mori M., Yamada S., Negi S., Bu X., Kato J., Moss J., Sanada N., Kizu R, & Fujii T. (2020). The 89-kDa PARP1 cleavage fragment serves as a cytoplasmic PAR carrier to induce AIF-mediated apoptosis. The Journal of Biological Chemistry, 296, 100046.
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3

Photoencapsulation of Spheroids and Fibers

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Spheroids were harvested and centrifuged to remove residual single cells. Spheroids (6,000 spheroids mL−1) and fiber segments (variable density) were simultaneously photoencapsulated in GelMA. Studies were cultured in complete MCF10A media for 2 or 4 days, replenishing media every other day. For matrix metalloproteinase (MMP) inhibition studies, gels were cultured in complete MCF10A media containing marimastat (50 mM). For EMT studies, gels were cultured in complete MCF10A media containing rhTGF-β1 (Invitrogen). For apoptosis studies, gels were cultured in complete MCF10A media containing staurosporine (Santa Cruz Biotechnology) and left on a rocker plate at 0.33 Hz for 6 hrs to enhance diffusive transport.
Hiraki H.L., Matera D.L., Wang W.Y., Prabhu E.S., Zhang Z., Midekssa F., Argento A.E., Buschhaus J.M., Humphries B.A., Luker G.D., Pena-Francesch A, & Baker B.M. (2022). Fiber density and matrix stiffness modulate distinct cell migration modes in a 3D stroma mimetic composite hydrogel. Acta biomaterialia, 163, 378-391.
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4

Cytotoxicity Assay Reagents Protocol

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(S)-(+)-camptothecin (#C9911), Triton X-100 (#X100), paraformaldehyde (#P6148), potassium phosphate monobasic (#P5655), sodium phosphate dibasic (#S5136), and dimethyl sulfoxide (DMSO, #D8418) were purchased from Sigma-Aldrich (Prague, Czech Republic); staurosporine (#3510A) was purchased from Santa Cruz Biotechnology (Heidelberg, Germany); Hoechst 33342 (#17530) was purchased from AAT Bioquest (Sunnyvale, CA, USA); and fetal bovine serum (#16000–036) was purchased from Thermo Fisher Scientific (Waltham, MA, USA). Hydrogen peroxide (30% solution, #23980–11000), potassium chloride (#60130G1000), sodium chloride (#71380G1000), isopropyl alcohol (#17510), nitric acid 65% (#18980) and sulfuric acid 96% (#20450) were obtained from Penta (Prague, Czech Republic).
Filipova M., Elhelu O.K., De Paoli S.H., Fremuntova Z., Mosko T., Cmarko D., Simak J, & Holada K. (2018). An effective “three-in-one” screening assay for testing drug and nanoparticle toxicity in human endothelial cells. PLoS ONE, 13(10), e0206557.
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5

Glycosidase-mediated Protein Deglycosylation

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MG132, Z-VAD-FMK (Selleckchem); MLN4924, GW4869 (Sigma); chloroquine (Invivogen); concanamycin A, rapamycin (Enzo); spiroepoxide (Santa Cruz); staurosporine (CST). Recombinant glycosidases PNGaseF and EndoH, and the substrate recombinant RNaseB protein were purchased from New England Biolabs.
Ding S., Zhu S., Ren L., Feng N., Song Y., Ge X., Li B., Flavell R.A, & Greenberg H.B. (2018). Rotavirus VP3 targets MAVS for degradation to inhibit type III interferon expression in intestinal epithelial cells. eLife, 7, e39494.
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6

Apoptosis Signaling in Human Cell Lines

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Unless stated otherwise, standard laboratory procedures were used for manipulating DNA, RNA, oligonucleotides and proteins (32 ). Oligonucleotides listed in Supplementary Data were synthesized by Eurofins MWG. Human HeLa, HEK293, MRC5 and fibroblast cells and mouse NIH 3T3 cells were grown in Dulbecco's modified Eagle medium (DMEM) supplemented with 10% fetal calf serum (Invitrogen). Apoptosis was induced by treatment of HeLa cells with 1 μM staurosporine (Sigma) for 4 h and caspase activity was inhibited with 60 μM of z-DEVD-FMK (Santa Cruz) added 1 h before staurosporine administration. Expression plasmids (jetPRIME, Ozyme), siRNAs and in vitro transcribed mt tRNAs (Lipofectamine 2000, Invitrogen) were transfected into HeLa cells by using the indicated transfection reagents as recommended by the suppliers.
Marnef A., Jády B.E, & Kiss T. (2015). Human polypyrimidine tract-binding protein interacts with mitochondrial tRNAThr in the cytosol. Nucleic Acids Research, 44(3), 1342-1353.
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7

Apoptosis Induction and Detection

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The cells were treated with DMSO or idasanutlin for 48 h, or with Staurosporine (Santa Cruz Biotechnology, Dallas, TX, USA), a well-known inducer of apoptosis, for 16 h. Following this treatment, the cells were trypsinized and collected together with growth media and PBS, which was used for washes, to avoid any loss of detached apoptotic cells. The cells were centrifuged, washed with PBS, centrifuged again, suspended with annexin-binding buffer (BioLegend) and stained with 5 µg/mL of FITC-conjugated annexin V (BioLegend) and 2.5 µg/mL 7-Aminoactinomycin D (7-AAD, BioLegend) for 15 min at room temperature. The samples were analyzed using BD FACSVerse flow cytometer (Becton Dickinson) with appropriate color compensation.
Skalniak L., Kocik J., Polak J., Skalniak A., Rak M., Wolnicka-Glubisz A, & Holak T.A. (2018). Prolonged Idasanutlin (RG7388) Treatment Leads to the Generation of p53-Mutated Cells. Cancers, 10(11), 396.
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8

Quantification of Cell Damage and Survival

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We evaluated cell damage and cell survival 48 h after the infection of SeV vectors. For quantification of cell damage, we collected the supernatant and measured the signal counts of adenylate kinase (ToxiLight bioassay kit; Lonza, Basel, Switzerland) that originated from damaged cells. For quantification of cell survival, we used the commercially available WST‐8 kit (Cell Counting Reagents; Nacalai tesque) and measured the absorbance at 450 nm. We used the condition of 48‐hour incubation with 1 μM staurosporine (SantaCruz Biotechnology) and 1‐h incubation with 1% Triton‐X (Nacalai tesque).
Tan G.W., Kondo T., Imamura K., Suga M., Enami T., Nagahashi A., Tsukita K., Inoue I., Kawaguchi J., Shu T, & Inoue H. (2021). Simple derivation of skeletal muscle from human pluripotent stem cells using temperature‐sensitive Sendai virus vector. Journal of Cellular and Molecular Medicine, 25(20), 9586-9596.
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9

Cell Cycle and Death Analysis of Transfected K1 Cells

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K1 cells were transfected as described above. For cell cycle experiments, 6 hours after transfection, cells were synchronized by serum starvation (1%FBS, DMEM: F12 for 12h) and then stimulated (10% FBS, DMEM: F12 for 6 hr), fixed and permeabilized. The fraction of cells in each phase of cell cycle (G1, S, and G2/M) was determined by DNA content analysis using propidium iodide staining (PI—50μg/mL) followed by flow cytometry. For cell death analysis, 24 hours after transfection cells were incubated in fresh medium for two additional hours in the absence or presence of staurosporine (Santa Cruz Biotechnology) at selected final concentrations (5nM and 50nM). Stages of cell death were determined by flow cytometry, based on the combined cell staining with PI (2μg/mL) (Sigma, USA) and Annexin V (0.75μg/mL) (BioLegend, San Diego, CA; USA). Acquisition parameters were defined with the software BD Cell Quest; cells were acquired in the flow cytometer (FACScalibur–Becton Dickinson); acquired data was analysed using FlowJo (Tree Star Inc, USA) software.
Faria M., Matos P., Pereira T., Cabrera R., Cardoso B.A., Bugalho M.J, & Silva A.L. (2017). RAC1b overexpression stimulates proliferation and NF-kB-mediated anti-apoptotic signaling in thyroid cancer cells. PLoS ONE, 12(2), e0172689.
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10

Isolation and culture of primary murine keratinocytes

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Primary keratinocytes were isolated from newborn pups expressing Keratin 5 promoter-driven Cre recombinase and Ptch1flox/flox alleles, using standard techniques and as previously described (Peterson et al., 2015 (link)). Cells were grown in complete Cnt-57 keratinocyte media for 3–4 days at 37° C/5% CO2, then either maintained in complete media or switched to basal starve media (CnT-BM1.500) for an additional 2 days to induce Hh activation. Subsequently, cells were treated with either 2 µg/mL vismo for 2 days or 1 µM staurosporine (Santa Cruz) for 4.5 hours, both diluted into starve media. ASZ cells (Aszterbaum et al., 1999 (link)) were cultured in 154-CF media with 4% fetal bovine serum (6.7 parts chelexed:1 part unchelexed). These cells were starved in 154-CF media lacking serum, and treated with vismo or staurosporine, similar to primary keratinocytes. Sex of cells was not determined, as this is not a known determining factor for Hh signaling in vitro. ASZ cells were authenticated based on epithelial morphology and robust Hh pathway activity upon serum starvation, as previously reported (Aszterbaum et al., 1999 (link)).
Eberl M., Mangelberger D., Swanson J.B., Verhaegen M.E., Harms P.W., Frohm M.L., Dlugosz A.A, & Wong S.Y. (2018). Tumor architecture and Notch signaling modulate drug response in basal cell carcinoma. Cancer cell, 33(2), 229-243.e4.
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