The miR-106a-5p mimics (5′-AAAAGUGCUUACAGUGCAGGUAG-3′), miR-106a-5p inhibitor (5′-UAUGGCUUUUUAUUCCUAUGUGA-3′) and scrambled mimic or inhibitor were designed and synthesized by Shanghai GenePharma Co., Ltd. si-STAT3 (5′-GCAGCAGCTGAACAACATG-3′) and si-Scramble (5′-UUCUCCGAACGUGUCACGUTT-3′) were also designed and purchased from Shanghai GenePharma Co., Ltd. miR-106a-5p mimics (50 nM), miR-106a-5p inhibitor (50 nM) or si-STAT3 (100 nM) were transfected into cells (2×104 cells) using Lipofectamine® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.), according to the manufacturer's protocols. After transfection for 48 h, cells were then exposed to ox-LDL for 48 h and collected for further experiments. Inhibitor experiments were performed using 30 µm STA-21 (Enzo Life Sciences, Inc.) as previously described (28 (link)). Briefly, HUVEC cells were co-treated with miR-106a-5p inhibitor and either 30 µm STA-21 or DMSO for 24 h and then exposed to ox-LDL for 48 h and collected for further experiments.
Sta 21
Manufactured by Enzo Life Sciences
Sourced in Italy, Switzerland
STA-21 is a laboratory instrument used for rheological analysis. It is designed to measure the viscoelastic properties of various materials, including polymers, gels, and suspensions. The core function of STA-21 is to provide accurate and reliable data on the flow and deformation behavior of these materials under controlled shear conditions.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions)
Sistat3, by Thermo Fisher Scientific (1 mentions)
Si scramble, by GenePharma (1 mentions)
Sistat3, by GenePharma (1 mentions)
Albumin bovine fraction 5, by Merck Group (1 mentions)
Recombinant human il 6, by Thermo Fisher Scientific (1 mentions)
Basic fibroblast growth factor (bfgf), by Thermo Fisher Scientific (1 mentions)
Bay11 7082, by Merck Group (1 mentions)
Dmem f12 medium, by Thermo Fisher Scientific (1 mentions)
Mk 2206, by Selleck Chemicals (1 mentions)
Epidermal growth factor (egf), by Thermo Fisher Scientific (1 mentions)
Insulin, by Merck Group (1 mentions)
Ly294002, by Merck Group (1 mentions)
Fk506, by Enzo Life Sciences (1 mentions)
3 protocols using sta 21
1
Modulating miR-106a-5p and STAT3 in HUVEC Cells
2
Lung Cancer Sphere Culture and Inhibition
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LCSs were obtained by plating 10,000 cells/ml in serum-free DMEM-F12 medium (Gibco-Invitrogen) containing 10 μg/ml insulin (Sigma-Aldrich, St. Luis, MO), 1% Albumin Bovine Fraction V (Sigma-Aldrich) 50 ng/ml EGF and 25 ng/ml bFGF (PeproTech, London, UK) (sphere medium) in ultra-low attachment flasks. LCSs were expanded by trypsinization and mechanical dissociation followed by re-plating of single cell suspensions (10,000 cells /ml) in fresh sphere medium. Self-renewal of LCSs was tested by assessing the capacity of primary LCSs to generate secondary LCSs after trypsin disaggregation. Secondary LCSs were assessed at day 7 for NCI-H460 and BEAS-2B cells and at day 12 for PEd/10 cells.
Anti-IL-6 mAb and recombinant human IL-6 were purchased from PeproTech. Cells were treated with LY294002 (Sigma-Aldrich), MK2206 (Selleckchem, Huston, TX), BAY11-7082 (Sigma-Aldrich), STA-21 (Enzo Life Science, Firenze, Italy) and STAT3 Inhibitor XIII (Calbiochem, Darmastadt, Germany). Drugs were added to cells every 4 days. The number of LCSs was assessed by counts under the microscope. LCSs diameters were determined using ImageJ software (NIH, Bethesda, MD).
Experiments were repeated at least three times and data are shown as mean ± SD of three independent experiments.
Anti-IL-6 mAb and recombinant human IL-6 were purchased from PeproTech. Cells were treated with LY294002 (Sigma-Aldrich), MK2206 (Selleckchem, Huston, TX), BAY11-7082 (Sigma-Aldrich), STA-21 (Enzo Life Science, Firenze, Italy) and STAT3 Inhibitor XIII (Calbiochem, Darmastadt, Germany). Drugs were added to cells every 4 days. The number of LCSs was assessed by counts under the microscope. LCSs diameters were determined using ImageJ software (NIH, Bethesda, MD).
Experiments were repeated at least three times and data are shown as mean ± SD of three independent experiments.
3
STAT3 Modulation in Glioma Cells
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We used U251 human neuroglioma cells (Cell Bank of Chinese Academy of Sciences, Shanghai, China) in the succeeding experiments. Cells incubated in serum-free media were exposed to 2% iso urane for 6 hr, performed similarly as our previous study [8] . The wild-type pcDNA3-6×Myc-mSTAT3 vector [10] (link) or an antisense oligonucleotide (ASO) of STAT3 (HSS186131, Invitrogen, USA) were transfected into cells using lipofectamine TM 2000 (Invitrogen, USA). An AxyPrep Endofree Plasmid Miniprep Kit (Axygen Biosciences, USA) was used to extract the plasmids from bacterial culture. After the transfection, cells were incubated at 37°C in a CO 2 incubator for 48 hr prior to iso urane exposure. The untransfected cells without iso urane exposure and the empty vector-transfected cells were set as the controls. In the interaction studies, cells were pretreated with a proteasome inhibitor MG-132 (30mM, Calbiochem, Germany) [11] (link), FK506 (1mM) or a STAT3 inhibitor STA21 (30mM, Enzo Life Sciences, Switzerland) [12] (link) 30 min before iso urane exposure.
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