Five hundred thousand VSMCs were seeded in a T75 flask overnight. The next day, siRNA duplexes for MCU (5′-CGACCUAGAGAAAUACAAUCAGCTC-3′ and 5′-GGGAAUAAAGGGAUCUUAAAUGCTG-3′ at a ratio of 1:1) or scramble control (5′-CGUUAAUCGCGUAUAAUACGCGUA-3′; Integrated DNA Technologies) were transfected (5 nM in 20 μl of DharmaFECT 4 reagent). Knockdown efficiency was determined after 72 hours by Western blotting for MCU, which was normalized to COXIV signal. The knockdown efficiency was consistently greater than 75%.

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