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Supplemental Materials and Methods
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Cell culture
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HUVECs were obtained from the American Type Culture Collection (CRL-1730:
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Manassas, VA, USA). The cells were plated in gelatin-coated tissue culture wells and
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grown in HuMedia-EB2 medium (KE-2350S: Kurabo Industries Ltd., Osaka, Japan)
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supplemented with 10% fetal bovine serum at 95% O2 and 5% CO2. HUVECs at
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passages 2 or 3 were used for all experiments.
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Measurement of the nitric oxide concentrations in the HUVECs
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HUVECs were incubated in phenol red-free medium without serum for 12
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hours and in the presence or absence of 200 μmol/l of bezafibrate (B7273:
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Sigma-Aldrich, St. Louis, MO, USA) for two hours. The cells were then cultured in the
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presence or absence of 100 μmol/l of carboplatin (C2538: Sigma-Aldrich) or 10 nmol/l
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of paclitaxel (T7402: Sigma-Aldrich) for two hours prior to stimulation with 1 mol/l of
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A23187 (A7522: Sigma-Aldrich) for 10 minutes. The cell culture medium was collected
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and used to determine the level of nitric oxide (NO) production using the Nitric Oxide
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Quantitation Kit (#40020: Active Motif, Carlsbad, CA, USA) according to the
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manufacturer’s instructions.
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Measurement of the PPARα DNA binding activity in the HUVECs
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HUVECs were incubated in phenol red-free medium without serum for 12 hours, and in
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the presence or absence of 100 μmol/l of bezafibrate for 24 hours. The cells were then
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cultured in the presence or absence of 100 μmol/l of carboplatin (CBDCA) or 10 nmol/l
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of paclitaxel (PTX) for 24 hours. The cell nuclear fraction was collected using a Nuclear
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Extraction Kit according to the manufacturer’s protocol (#10009277: Cayman Chemical
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Company, Michigan, USA) and was employed to determine the PPARα DNA binding
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activity using a PPARα Transcription Factor Assay Kit (#10006915: Cayman Chemical
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Company) according to the manufacturer’s instructions.
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