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Sp600125

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SP600125 is a laboratory compound that functions as a highly selective ATP-competitive inhibitor of the c-Jun N-terminal kinase (JNK) signaling pathway. It is commonly used in scientific research to study the role of JNK activation in various cellular processes and disease models.

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Lab products found in correlation

Sb203580, by Thermo Fisher Scientific (4 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions) Pd98059, by Thermo Fisher Scientific (3 mentions) Tgf β1, by Thermo Fisher Scientific (2 mentions) Nrk 49f, by Thermo Fisher Scientific (2 mentions) Cardamonin, by Merck Group (1 mentions) Anti phospho p38 mapk, by Cell Signaling Technology (1 mentions) Tnf α, by Thermo Fisher Scientific (1 mentions) Anti p38 mapk, by Cell Signaling Technology (1 mentions) Anti nf κb p65, by Cell Signaling Technology (1 mentions) Anti phospho jnk, by Cell Signaling Technology (1 mentions) Anti jnk, by Cell Signaling Technology (1 mentions) Anti p21, by Cell Signaling Technology (1 mentions) Anti parp, by Cell Signaling Technology (1 mentions) Cell counting kit 8 (cck8), by Dojindo Laboratories (1 mentions) Bay 11 7082, by Beyotime (1 mentions) Mg132, by Beyotime (1 mentions) N acetyl cysteine (nac), by Beyotime (1 mentions) Retinoic acid, by Thermo Fisher Scientific (1 mentions) Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (1 mentions)

15 protocols using sp600125

1

Cardamonin Inhibits NF-κB Signaling

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Cardamonin was purchased from Sigma-Aldrich (St Louis, MO, USA). BAY 11-7082 and MG132 and NAC were purchased from Beyotime Biotechnology (Haimen, China). TNF-α, SP600125, SB203580 were from PeproTech Inc. (Rocky Hill, NJ, USA). Cell Counting Kit-8 was from Dojindo Molecular Technologies (Kumamoto, Japan). Anti-NF-κB p65, anti-phospho-p65 anti-CDK4, anti-p21, anti-PARP, anti-JNK, anti-p38 MAPK, anti-phospho-JNK and anti-phospho-p38 MAPK were purchased from Cell Signaling Technology (Danvers, MA, USA).
Li Y., Qin Y., Yang C., Zhang H., Li Y., Wu B., Huang J., Zhou X., Huang B., Yang K, & Wu G. (2017). Cardamonin induces ROS-mediated G2/M phase arrest and apoptosis through inhibition of NF-κB pathway in nasopharyngeal carcinoma. Cell Death & Disease, 8(8), e3024-.
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2

Molecular Profiling of Glial Cells

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The JMJD3 antibodies were from CST (#3457) and Abcam(ab38113). The #3457 was for immunoblot assay and ab38113 was for IHC staining. The Olig2 antibody was from Abcam(ab109186). The GFAP antibody was from Abcam(ab7260). The PDGFR-a antibody was from Abcam(ab203491). FACS antibodies was from BD, CD133 (566596), CD140a (740148). The H3K27me3 antibody was from CST (#9733). The Histone3 antibody was from CST (#4499). The antibodies for SAPK/JNK signaling study were from CST, P-c-fos(#5348), P-JNK(#4668), JNK(#9252), P-c-jun(#91952), c-jun(9165). The actin antibody was from Abcam(ab6276). The specific inhibitors were from MedChemeExpress, GSK-J4(HY-15648b), SP600125(HY-10241), Vinblastine (HY-13780), Retinoic acid (HY-14649).
bFGF and EGF were from Peprotech (45033, 31509). DMEM/F12, N2, B27and TrypLE express was from Gibco. The collagenase A was from ROCHE (10103578001). Protease and Phosphatase inhibitor cocktail kit was from Sigma Aldrich (P8340, P2850). IHC DAB kit was from Abcam(ab94665). ECL kits were from Abcam(ab133406).
Bo-Yin Z., Qingsan Z., Yihang M., Fan Y., Yuhang Z, & Pengyu C. (2021). Unlocking the Recovery Potential: JMJD3 Inhibition-Mediated SAPK/JNK Signaling Inactivation Supports Endogenous Oligodendrocyte-Lineage Commitment Post Mammalian Spinal Cord Injury. Neurochemical Research, 46(4), 792-803.
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3

Investigating Fibrotic Effects of TGF-β1 and TSA

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Normal rat kidney fibroblast cells (NRK-49F, from American Type Culture Center, Manassas, VA) were cultured in Dulbecco’s Modified Eagle Medium supplemented with 2 mM L-glutamine, 1% non-essential amino acids and 5% fetal bovine serum (Gibco, Carlsbad, CA) in a 5% CO2, 37 °C humidified incubator. To examine the pro- or anti-fibrotic effects of TGF-β1 and TSA, subconfluent cultures of NRK-49F cells were first maintained quiescent in basal medium with 0.5% fetal bovine serum for 24 hrs, and then stimulated with 5 ng/ml TGF-β1 (PeproTech, Rocky Hill, NJ) in the presence or absence of TSA (Sigma-Aldrich, St Louis, MO). To investigate the roles of MAPKs and Notch signaling pathways in TGF-β1-mediated fibrogenesis, subconfluent NRK-49F cells were treated with the following inhibitors: 10 μM SB203580 (p38 inhibitor; Gibco, Frederick, MD), 20 μM PD98059 (ERK inhibitor; Gibco, Frederick, MD), 10 μM SP600125 (JNK inhibitor; Gibco, Camarillo, CA) and RO4929097 (γ-secretase inhibitor; Selleckchem, Houston, TX).
Tung C.W., Hsu Y.C., Cai C.J., Shih Y.H., Wang C.J., Chang P.J, & Lin C.L. (2017). Trichostatin A ameliorates renal tubulointerstitial fibrosis through modulation of the JNK-dependent Notch-2 signaling pathway. Scientific Reports, 7, 14495.
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4

Antioxidant and Apoptosis Signaling Assays

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Resveratrol, PD169316, FR180204, SP600125, anisomycin, curcumin and the fluorescent dye H2DCF-DA were provided by Gibco (Grand Island, NY, USA). Dimethyl sulfoxide (DMSO), H2O2, and the CCK8 kit were provided by Gibco (Grand Island, NY, USA). Antibodies against cleaved caspase-3, cleaved caspase-9, phospho-p38 (p-p38), phospho-ERK (p-ERK), or phospho-JNK (p-JNK) were provided by Abcam (Grand Island, USA). Annexin V/FITC kit was provided by eBioscience (Bender MedSystems, Vienna, Austria). Kits to assay catalytic activity of superoxide dismutase (SOD), catalase (CAT), and glutathione S-transferase (GSH) were obtained by Abcam (Grand Island, USA).
Ye M.J, & Meng N. (2021). Resveratrol acts via the mitogen-activated protein kinase (MAPK) pathway to protect retinal ganglion cells from apoptosis induced by hydrogen peroxide. Bioengineered, 12(1), 4878-4886.
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5

Phospho-regulatory Pathway Profiling

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All cell culture materials were obtained from Gibco Inc. SP600125 and AS601245, pharmacological inhibitors of JNK, were purchased from Calbiochem. Control (#6568) and SAPK/JNK (#6232) siRNA, SAPK/JNK Kinase Assay Kit (#8794, nonradioactive), Hoechst 33342 (#4082), Anti-c-Jun (#9165), Anti-Phospho-c-JunSer63 (#12598), Anti-Phospho-c-JunSer73 (#3270 S), Anti SAPK/JNK (#9252), Anti-Phospho-SAPK/JNKThr183/Tyr185 (#9251), Anti-Cyclin A (4656), Anti-Cyclin B1 (#12231) and Anti-Phospho-cdc-2Tyr15 (#4539) antibodies were obtained from Cell Signaling. All western blotting buffers and reagents were purchased from Bio-Rad. Anti-phospho-Histone H3Ser10 (06-570) antibody was obtained from Upstate. Anti-Vinculin Antibody (V9264) was purchased from Sigma-Aldrich. TransIT-X2® Dynamic Delivery System (MIR 6000) was purchased from Mirus Bio LLC. YO-PRO®-1 Iodide (491/509) was obtained from Life Technologies. Texas Red™-X Phalloidin and Click-iT™ EdU Alexa Fluor™ 488 Imaging Kit (C10337) were obtained from Thermo Fisher Scientific. Matrigel Basement Membrane Matrix (356230) was from BD Biosciences. Super Block reagent (#AAA125) was purchased from ScyTek Laboratories.
Bildik G., Akin N., Senbabaoglu F., Esmalian Y., Sahin G.N., Urman D., Karahuseyinoglu S., Ince U., Palaoglu E., Taskiran C., Arvas M., Guzel Y., Yakin K, & Oktem O. (2018). Endogenous c-Jun N-terminal kinase (JNK) activity marks the boundary between normal and malignant granulosa cells. Cell Death & Disease, 9(4), 421.
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6

JNK Inhibitor Administration in Mice

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3-week-old wild type and TgUsp14CA mice were given daily intraperitoneal injections of the JNK inhibitor SP600125 (Fisher Scientific, Rockford, IL) for two weeks. SP600125 was dissolved in DMSO at a concentration of 91 mM and this solution was directly administered at a dose of 16 mg/kg using a 25 μL Hamilton syringe (Hamilton Company, Reno, NV). This procedure was used to minimize the injection volume, as SP600125 is not soluble in aqueous solutions and DMSO was not tolerated at higher doses. Animals were weighed every other day to adjust doses and to monitor potential adverse effects of the injections. At the doses given, both DMSO and SP600125 were well tolerated.
Vaden J.H., Bhattacharyya B.J., Chen P.C., Watson J.A., Marshall A.G., Phillips S.E., Wilson J.A., King G.D., Miller R.J, & Wilson S.M. (2015). Ubiquitin-specific protease 14 regulates c-Jun N-terminal kinase signaling at the neuromuscular junction. Molecular Neurodegeneration, 10, 3.
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7

Monocyte Differentiation and Maintenance

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THP-1 monocytes (ATCC TIB-202) and primary monocytes were maintained in RPMI 1640 medium supplemented with 2 mM L-glutamine and 10% heat-inactivated fetal bovine serum (FBS) at 37 °C in a humidified atmosphere of 5% CO2. THP-PPM1A cells were generated previously21 (link). Human PBMCs were isolated from buffy coats by the Ficoll-Paque density centrifugation method. Monocytes were enriched by positive selection using anti-CD14 mAb-coated microbeads from Miltenyi Biotec (San Diego, CA) according to manufacturer’s protocol. Monocytes were then differentiated with 5 ng/ml GM-CSF (R&D Systems, Minneapolis, MN) for 5–7 days to obtain monocyte derived macrophages (MDM). fetal bovine serum was obtained from Life Technologies (Grand Island, NY). Phorbol ester 13-phorbol-12-myristate acetate (PMA), puromycin, etoposide, kanamycin and anisomycin were purchased from Sigma (St. Louis, MO). Sanguinarine and SP600125 were purchased from Fisher Scientific (Pittsburgh, PA). CellTiter-Glo luminescent cell viability assay kit was purchased from Promega (Madison, WI). Polyclonal PPM1A antibodies were purchased from Thermo Scientific (Rockford, IL). Monoclonal mouse antibodies to GAPDH and α-tubulin were purchased from Santa Cruz (Dallas, Texas) and Cell Signaling (Danvers, MA), respectively.
Schaaf K., Smith S.R., Duverger A., Wagner F., Wolschendorf F., Westfall A.O., Kutsch O, & Sun J. (2017). Mycobacterium tuberculosis exploits the PPM1A signaling pathway to block host macrophage apoptosis. Scientific Reports, 7, 42101.
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8

Characterization of Mutant GPCR Constructs

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All GPCR constructs were purchased from Missouri S&T cDNA Resource Center. The HA-dDRY-OX1R construct was generated using wildtype HA-OX1R and mutating Arg-144 (CGC) of the DRY motif to Ala (GCC) using the QuikChange II Site-Directed Mutagenesis Kit from Agilent Technologies. (Forward primer: 5′-GCTTCATCG CCCTGGACGCCTGGTATGCCATCTGC-3′, Reverse primer: 5′-GCAGATGGCATACCAGGCGTCCAGGGCGATGA AGC-3′). Note that mutation of the ‘DRY motif’ to abolish G protein-coupling has been described previously [15 (link)]. Dynorphin A (1-17) was purchased from American Peptide and orexin A from Tocris Bioscience. Rabbit phospho-p38, mouse p38 and rabbit phospho-JNK antibodies were purchased from Cell Signaling Technology, rat anti-HA from Roche and mouse anti-actin from Santa Cruz Biotechnology. Secondary antibodies were purchased from LI-COR Biosciences. Dynasore, H89 and norBNI were purchased from Tocris Bioscience, SP-600125 from Fisher Scientific, PKI from Santa Cruz Biotechnology and UBO-QIC was purchased from The University of Bonn.
Robinson J.D, & McDonald P.H. (2015). The orexin 1 receptor modulates kappa opioid receptor function via a JNK-dependent mechanism. Cellular signalling, 27(7), 1449-1456.
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9

Ginsenoside Rg3 Kinase Inhibitor Protocol

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The ginsenoside Rg3 was purchased from NPC & BioTech (Daejon, Korea). The Rg3 was dissolved in dimethyl sulfoxide (DMSO, 5 mg/mL) and stored at −80 °C. Specific kinase inhibitors, PD98059, SB203580, and SP600125, were purchased from Thermo Fisher Scientific (Pittsburgh, PA, USA).
Kim J.H., Bae Y.C., Lee J.W., Choi J.S, & Bae S.H. (2019). Effects of ginsenoside Rg3 on apoptosis in A375.S2 melanoma cells. Translational Cancer Research, 8(2), 357-366.
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10

Biochemical Analyses of Cellular Signaling

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Catalase (CAT) was purchased from MP Biomedicals (Solon, OH). N-acetyl-L(+)-cysteine (NAC) and MitoSOX™ Red reagent were from Fisher (Fair Lawn, NJ). Trolox, Mito-TEMPO, and MMP-3 substrate β-casein were from SIGMA (Saint Louis, MO). TRIzol™ Reagent was purchased from Invitrogen (Carlbad, CA). SB203580 was from TOCRIS (Ellisville, MO), PD98059 from Cell Signaling Technology (Beverly, MA), and SP600125 from Thermo Fisher (Ward Hill, MA).
Antibodies against β-actin (cat.# 58169), total p38 (cat.# 9212), phospho-p38 (cat.# 9215), total ERK1/2 (cat.# 9107), phospho-ERK1/2 (cat.# 9101), total JNK2 (cat.# 9258), phospho-JNK (cat.# 4668), E-cadherin (cat.# 3195), fibronectin (cat.# 26836), and horseradish peroxidase (HRP)-conjugated horse anti-mouse IgG (cat.# 7076) and goat anti-rabbit IgG (cat.# 7074) were obtained from Cell Signaling Technology (Beverly, MA). Anti-MMP-3 antibody (cat.# 53015) was from abcam (Cambridge, MA), and anti-α-SMA (cat.# A5228) and anti-vimentin (cat.# V2258) antibodies from SIGMA (Saint Louis, MO). All other chemicals were purchased from Fisher Scientific (Fair Lawn, NJ) except when otherwise stated. All chemicals used were of analytical grade.
Zhang Y., Mo Y., Yuan J., Zhang Y., Mo L, & Zhang Q. (2022). MMP-3 activation is involved in copper oxide nanoparticle-induced epithelial-mesenchymal transition in human lung epithelial cells. Nanotoxicology, 15(10), 1380-1402.
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