Ethyl acetate fraction from methanol extraction of Vitis thunbergii var. taiwaniana induced G₀/G₁ phase arrest via inhibition of cyclins D and E and induction of apoptosis through caspase‐dependent and ‐independent pathways in human prostate carcinoma DU145 cells

Abstract

contributed equally to this work. Funding informationThe authors gratefully acknowledged a research grant (Functional evaluation of extracts and pure compounds isolated from V. thunbergii var. taiwaniana on the improvements of metabolic syndromerelated diseases) from the National Science Council, Taiwan. AbstractVitis thunbergii var. taiwaniana (VTT) is a wild grape native to Taiwan, belonging to the Vitaceae family and Vitis genus, and widely used as folk herbal medicine. It is traditionally used for the treatment of diarrhea, hypertension, neuroprotection, jaundice, and arthritis. We used the wildcollected VTT and sterilized them to establish the plant tissue culture, and then took the leaves for DNA sequencing to determine its original base. We use methanol to extract VTT in four different solvents: 1-butanol, n-hexane, ethyl acetate, and water. These four preliminary extracts were used to treat human prostate cancer DU145 cells in vitro. We use the flow cytometry to check the cell survival situation. Finally, we found the ethyl acetate layer roughing product (referred VTEA) in human prostate cancer apoptotic effects of cell line DU-145. In the present studies, we use the crude extract of VTT to examine whether or not it can induce apoptosis of DU145 cells in vitro.Viability assays for extracts of VTT treatment showed that it had dose-dependent effect on human prostate cancer DU145 cells. We also found that the extract of VTT induces time-dependent mitochondrial and intrinsic-dependent apoptosis pathways. The in vitro cytotoxic effects were investigated by cell cycle analysis and the determination of apoptotic DNA fragmentation in

Ouabain induces apoptotic cell death in human prostate DU 145 cancer cells through DNA damage and TRAIL pathways

“…Human prostate cancer cell line (DU 145) was purchased from the Food Industry Research and Development Institute (Hsinchu, Taiwan). The cell line was cultured in MEM medium supplemented with 10% FBS, 2 mM l ‐glutamine and 1% penicillin‐streptomycin (100 Units/mL penicillin and 100 μg/mL streptomycin) at 37°C in a humidified incubator containing atmosphere of 5% CO 2 and 95% air …”

“…And then cells were collected, counted and stained with PI (5 μg/mL) for total cell viability determination by using flow cytometric assay (BD Biosciences, FACSCalibur, San Jose, California). Cells were treated with 10 μM of ouabain for 0, 6, 24, and 48 hours for cell cycle distribution assay by flow cytometry as previously described …”

“…Then, cells were harvested, resuspended in Annexin V binding buffer, followed incubated with Annexin V‐FITC/PI in the dark for 15 minutes. A 10,000 cells from each treatment were analyzed using BD FACSCalibur (BD Biosciences) for apoptotic cell number as described previously . Experiments were performed in triplicate.…”

Casticin inhibits human prostate cancer DU 145 cell migration and invasion via Ras/Akt/NF‐κB signaling pathways

“…Cells were grown in 90% RPMI 1640 medium, 10% FBS, 100 units/ml penicillin, and 100 μg/ml streptomycin. Cells were cultured in a humidified incubator at 37°C (an atmosphere of 5% CO 2 ) (Lin et al, ).…”

“…DU 145 cancer cells (1 × 10 5 cells/well) were subcultured in 12‐well plates overnight and treated with different concentrations of casticin (0, 1.25, 2.5, 5, 10, 20, 40, and 50 μM) or with 0.5% DMSO solution as vehicle control for 48 hr. After casticin treatment, cells were suspended, and then stained with PI solution (5 μg/ml) for measuring the viable cell number by flow cytometer (BD Biosciences, San Jose, CA, USA) according to our previous study (Lin et al, ).…”