Sod kit

The content of Malondialdehyde (MDA) in cells was detected using the MDA kit (ab118970, Abcam, Cambridge, MA, United States). The level of reactive oxygen species (ROS) was detected using the ROS/Superoxide kit (ab139476, Abcam). The content of superoxide dismutase (SOD) was detected using the SOD kit (ab65354, Abcam).

VSMCs were seeded into the 96-well plates (5 × 10³ cells/well) and incubated at 37ºC for 24 hours for adherence before transfection. Two hours after the transfection, Ang II or PBS was added into medium to pretreat the transfected VSMCs for 24 hours. Then resuspended cells were incubated in pre-heated M231 containing 5 μM of 2,7-Dichlorofluorescein diacetate (H2DCFDA; Gibco, Carlsbad, California, USA) fluorescent probe at 37ºC for 30 minutes, and rinsed twice with PBS. The spectrophotometer (Agilent Technologies, Inc., Palo Alto, CA, USA) was utilized to measure the ROS level at the excitation wavelength of 485 nm and an emission wavelength of 530 nm using the ROS kit (ab113851, Abcam Inc., Cambridge, MA, USA). On the other hand, cell supernatant was collected to determine the MDA level with a thiobarbituric acid method using the MDA kit (ab118970, Abcam Inc., Cambridge, MA, USA) by checking the absorbance at the wavelength of 532 nm. SOD kit (ab65354, Abcam Inc., Cambridge, MA, USA) was used to determine the SOD content in cell supernatant via WST-1, by checking the absorbance at the wavelength of 450 nm.