The small-molecule inhibitors were all purchased from Targetmol, Wellesley Hills, MA, USA including Lipid metabolism compounds library (L2510), CB1 antagonists (AM251, T1915; AM281, T2264; JD5037, T4453; Otenabant, T3530; TM3837, T8511; CB1-IN-1, T5996), ferroptosis inducers (erastin, T1765; RSL3, T3646; ML-210, T8375; FIN56, T4066; CIL56, T4309), PI3K inhibitor (LY294002, T2008) and MEK inhibitor (PD98059, T2623).
Ly294002
Manufactured by Targetmol
Sourced in United States, China
LY294002 is a lab equipment product that functions as a phosphoinositide 3-kinase (PI3K) inhibitor. It is commonly used in research applications to study cellular signaling pathways and their regulation.
Automatically generated - may contain errors
Lab products found in correlation
Ml 210, by Targetmol (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (2 mentions)
Erastin, by Targetmol (1 mentions)
Am251, by Targetmol (1 mentions)
Fin56, by Targetmol (1 mentions)
Pd98059, by Targetmol (1 mentions)
Smc growth medium, by Cell Applications (1 mentions)
Cay10650, by MedChemExpress (1 mentions)
Nutrient mixture f12 ham kaighn s modification f12k, by Merck Group (1 mentions)
Low density lipoprotein cholesterol ldl c assay kits, by Nanjing Jiancheng (1 mentions)
Lipofectamine 3000 reagent, by Thermo Fisher Scientific (1 mentions)
Dimethyl sulphoxide dmso, by Avantor (1 mentions)
Opti mem, by Thermo Fisher Scientific (1 mentions)
Anti β actin, by Cell Signaling Technology (1 mentions)
Anti gapdh, by Cell Signaling Technology (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
3 4 5 dimethylthiazol 2 yl 5 3 carboxymethoxyphenyl 2 4 sulfophenyl 2h tetrazolium, by Promega (1 mentions)
Annexin 5 fitc apoptosis detection kit, by Dojindo Laboratories (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Minimum essential medium (mem), by Thermo Fisher Scientific (1 mentions)
8 protocols using ly294002
1
Screening Small-Molecule Inhibitors in Cell Lines
2
Umbilical Artery SMC Isolation and Inhibition
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Primary human umbilical artery SMCs isolated from artery of human umbilical cord were maintained in Nutrient Mixture F12 Ham Kaighn's Modification (F12K, Sigma Aldrich) supplemented with 20% (for cell maintaining) or 2% (for treatment) fetal bovine serum (FBS) (Gemini) and 10% SMC Growth Medium (Cell Applications). Cells at passage 3 to 7 were used in all of the experiments.
To inhibit cPLA2 activation, cells were incubated with CAY10650 (MedChemExpress) at 12 nmol L−1 for 1 h. PI3K inhibitor LY294002 (Targetmol) was used at 10 µmol L−1 for 24‐h‐treatment. AKT inhibitor GSK690693 (Targetmol) was used at 10 µmol L−1 for 24‐h‐treatment.
Rabbit polyclonal antibodies (pAbs) against DNMT1, DNMT3A, and DNMT3B were from ABclonal. Rabbit pAbs against SMA, SM22α, and MeCP2 were from Santa Cruz Biotechnology. Rabbit pAb against 5‐meC was from Active Motif. Rabbit pAbs against IgG and mouse mAb against ATP5a1, Lamin B1, IgG, IgM were from Proteintech. Anti‐mtDNA antibody was from PROGEN. Rabbit pAb against cPLA2, mouse mAb against AKT1, and mouse mAb against Myc‐tag were from Bioss. Mouse mAb against Dsred2 was from Origene. Rabbit pAb against phosphor‐AKT was from Cell Signaling Technology.
To inhibit cPLA2 activation, cells were incubated with CAY10650 (MedChemExpress) at 12 nmol L−1 for 1 h. PI3K inhibitor LY294002 (Targetmol) was used at 10 µmol L−1 for 24‐h‐treatment. AKT inhibitor GSK690693 (Targetmol) was used at 10 µmol L−1 for 24‐h‐treatment.
Rabbit polyclonal antibodies (pAbs) against DNMT1, DNMT3A, and DNMT3B were from ABclonal. Rabbit pAbs against SMA, SM22α, and MeCP2 were from Santa Cruz Biotechnology. Rabbit pAb against 5‐meC was from Active Motif. Rabbit pAbs against IgG and mouse mAb against ATP5a1, Lamin B1, IgG, IgM were from Proteintech. Anti‐mtDNA antibody was from PROGEN. Rabbit pAb against cPLA2, mouse mAb against AKT1, and mouse mAb against Myc‐tag were from Bioss. Mouse mAb against Dsred2 was from Origene. Rabbit pAb against phosphor‐AKT was from Cell Signaling Technology.
3
Lunasin Regulates LDL Cholesterol Metabolism
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Lunasin was prepared by using recombinant DNA technology as previously described [48 ], and dissolved in saline. Minimum essential medium (MEM) and Opti-MEM as well as FBS were purchased from Gibco (Grand Island, NY, USA). Lipofectamine 3000 reagent was obtained from Invitrogen (California, Carlsbad, USA). Antibodies against PCSK9 and LDLR were obtained from abcam (Cambridge, UK). Anti-GAPDH and Anti-β-actin were obtained from Cell Signaling Technology (Danvers, MA, USA). LY294002, an effective PI3K/Akt pathway chemical inhibitor, was purchased from TargetMol (Boston, MA, USA). LDL labeled with 1, 1′-dioctadecyl – 3, 3, 3′, 3′-tetramethyl- indocarbocyanine perchlorate (Dil-LDL), which is highly fluorescent lipophilic dyes that diffuse into the hydrophobic portion of the LDL complex without affecting the LDL-specific binding of the apoprotein. Dil-LDL was obtained from Yiyuan Biotechnologies (Guangzhou, China). Dimethyl Sulphoxide (DMSO) and other chemicals were purchased from Amresco (Solon, OH, USA). Total cholesterol (CHO) and low-density lipoprotein cholesterol (LDL-C) assay kits were obtained from Nanjing Jiancheng Bioengineering Institute (Nanjing, China).
4
Gastric Cancer Cell Signaling Pathway
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The study was conducted from April 2017 to January 2018 in Zhongshan Hospital. Gastric mucosal epithelial cells and gastric cancer cells were routinely subcultured after rapidly thawing in a 37°C water bath + resuscitation in complete culture solution for 5–8 hours, and after passage, gastric cancer cells were divided into three groups: gastric cancer cell group, LY294002 group and MK-2206 group. Cells in the LY294002 group were treated with 20 um of PI3K inhibitor LY294002 (TargetMol, China, cat no. T2008); cells in the MK-2206 group were treated with 4 nmol/L of AKT inhibitor MK-2206 (Wuhan Booute Biotechnology Co.,Ltd., cat no. orb322834), and the cells in the gastric cancer cell group were not treated. The levels of PI3K, Akt, phosphorylated PI3K (p-PI3K), phosphorylated Akt (p-Akt) and p-PRAS40-Thr246 in the four groups were measured, and the cell proliferation, invasion and apoptosis in the gastric cancer cell group, LY294002 group and MK-2206 group were detected.
5
DMSO and DON Induced Apoptosis Pathway
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Dimethylsulfoxide (DMSO)
and DON were kindly purchased from Sigma-Aldrich Corp (St. Louis,
MO, USA). 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) was obtained from Promega (Madison,
WI, USA). Annexin V-FITC Apoptosis Detection Kit was bought from DOJINDO
(Shanghai, China). The main antibodies against Bax, p62, LC3, AKT,
mTOR, and GAPDH and HRP-linked antibody were acquired from Servicebio
(Wuhan, China). Primary antibodies against p-AKT and p-mTOR were obtained
from Nanjing Jiancheng Bioengineering Institute (Jiangsu province,
China). 740Y-P was purchased from APEXBIO (Shanghai, China). LY294002
was purchased from TargetMol (Shanghai, China). 3-MA was purchased
from Selleckchem (Houston, TX). IPEC-J2 cells were procured from Biochemy
(Wuhan, China) and preserved in Dulbecco’s modified Eagle medium
(DMEM) incorporating 50 μg/mL streptomycin (Gibco Invitrogen),
50 μg/mL penicillin and 10% fetal bovine serum.
and DON were kindly purchased from Sigma-Aldrich Corp (St. Louis,
MO, USA). 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) was obtained from Promega (Madison,
WI, USA). Annexin V-FITC Apoptosis Detection Kit was bought from DOJINDO
(Shanghai, China). The main antibodies against Bax, p62, LC3, AKT,
mTOR, and GAPDH and HRP-linked antibody were acquired from Servicebio
(Wuhan, China). Primary antibodies against p-AKT and p-mTOR were obtained
from Nanjing Jiancheng Bioengineering Institute (Jiangsu province,
China). 740Y-P was purchased from APEXBIO (Shanghai, China). LY294002
was purchased from TargetMol (Shanghai, China). 3-MA was purchased
from Selleckchem (Houston, TX). IPEC-J2 cells were procured from Biochemy
(Wuhan, China) and preserved in Dulbecco’s modified Eagle medium
(DMEM) incorporating 50 μg/mL streptomycin (Gibco Invitrogen),
50 μg/mL penicillin and 10% fetal bovine serum.
6
Isolation and Treatment of Murine Myocardial Cells
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Myocardial cells were isolated from experimental mice and cultured in minimum essential medium (MEM) (Gibco; Thermo Fisher Scientific Inc., Waltham, MA, USA) supplemented with 5% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.). Myocardial cells were cultured at 37°C in a humidified atmosphere containing 5% CO2. Myocardial cells were treated with PI3K inhibitor LY294002 (1 mg/ml, TargetMol, Boston, MA, USA) or PBS for 12 h at 37°C for further analysis.
7
Comprehensive Pharmacological Toolkit for Research
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All small-molecule inhibitors were purchased from Targetmol (Wellesley Hills, MA, USA), including FAAH inhibitors (FAAH-IN-2, URB597, PF-3845, JNJ-42165279, BIA 10-2474, FAAH-IN-1, SA47, LY-2183240, FAAH inhibitor 1, JNJ-1661010, PF-04457845, 4-nonylphenylboronic acid, N-benzylpalmitamide, SA72, JZL184, JZL195), ferroptosis inducers (erastin, T1765; RSL3, T3646; ML-210, T8375; FIN56, T4066; CIL56, T4309; iFSP1; sorafenib; BAY 87-2243; sulfasalazine; ML162), PI3K inhibitor (LY294002, T2008), and MEK inhibitor (PD98059, T2623). These drugs were prepared as formulations for cell-based assays or the animal according the methods descripted in the previous publications [42 (link)–45 (link)].
8
Ligustrazine Cytoprotection for Retinal Cells
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RGC-5 cells in the logarithmic phase of growth were seeded in six-well plates and cultured in complete medium for 8 h until ~70% cell adhesion. I/R model cells were treated with 0, 5, 10, 20, 40, 80, 160, or 320 μg/mL ligustrazine (Baomanbio, D0188). Cultured I/R model cells were divided into I/R group [DMSO solution (Sigma, D2650)], ligustrazine group [10 μg/mL ligustrazine), ligustrazine+PI3K inhibitor group [10 μg/mL ligustrazine and 10 μM Ly294002 (TargetMol, T2008)], and ligustrazine+mTOR inhibitor group [10 μg/mL ligustrazine and 10 μM rapamycin (Solarbio, IR0010)]. In the normal (control) group, only an equivalent volume of DMSO was added to RGC-5 cells without hypoxia treatment. In addition to the RGC-5 cells in the normal group, the complete medium was replaced with low-glucose DMEM medium. RGC-5 cells were treated with ligustrazine and DMSO for 30 min, and the experiments were repeated five times.
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