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. 2012 Oct 29;7(10):e48393. doi: 10.1371/journal.pone.0048393

Figure 1. Insulin increases ROS production through a Nox4-based NADPH oxidase in HMVECs.

Figure 1

(A) Growth-arrested HMVECs were pre-incubated with DPI (5 μM) for 30 minutes, and then stimulated with insulin (100 nM) for 5 minutes. Measurement of ROS formation by DCF-DA method was as described in Methods. (B) mRNA expression of Nox2 and Nox4 in HMVECs was determined by real-time RT-PCR. (C) HMVECs were transfected with scrambled (control) or Nox4 siRNA. In 48 hours, the cells were harvested for detecting the Nox4 and Nox2 protein. (D) HMVECs were transfected with control siRNA, Nox4 siRNA, or Nox2 siRNA. In 24 hours the cells were growth-arrested, followed by stimulation with insulin (100 nM) for 5 minutes. ROS production was determined using DCF-DA as substrate. **P<0.01 vs. the control; ##P<0.01 vs. the insulin-treated group.