Hinokitiol Modulates Constitutively Active AKT

B16F10 or 4T1 cells (1 × 10⁶ cells per well) were seeded in 6-well culture plates incubated for 24 h at 37°C, 5% CO2. Then, the cells were either transfected with 5 μg pCDNA 3.1 control plasmid or constitutively active AKT plasmid (Provided by Dr. Chiau-Yuang Tsai of the Department of Molecular Immunology, Osaka University, Japan) using Lipofectamine 2000 18 (link), 19 and then incubated for another 24 h at 37°C, 5% CO2. Afterwards, the cells were washed and treated with hinokitiol (Sigma Aldrich, St. Louis, MO, USA) for 16 h and prepared for Western blotting.

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Wu Y.J., Hsu W.J., Wu L.H., Liou H.P., Pangilinan C.R., Tyan Y.C, & Lee C.H. (2020). Hinokitiol reduces tumor metastasis by inhibiting heparanase via extracellular signal-regulated kinase and protein kinase B pathway. International Journal of Medical Sciences, 17(3), 403-413.

Publication 2020

hinokitiol

Cells Hinokitiol Lipofectamine 2000 Plasmid

Corresponding Organization : China Medical University Hospital

Other organizations : Kaohsiung Armed Forces General Hospital

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Variable analysis

independent variables

Transfection with pCDNA 3.1 control plasmid
Transfection with constitutively active AKT plasmid

dependent variables

Outcomes measured by Western blotting

control variables

Cell line (B16F10 or 4T1 cells)
Cell seeding density (1 × 10^6 cells per well)
Cell culture conditions (37°C, 5% CO2)
Incubation time (24 h before transfection, 24 h after transfection)
Transfection method (Lipofectamine 2000)
Treatment with hinokitiol (16 h)

positive control

Transfection with pCDNA 3.1 control plasmid

negative control

No transfection

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