APLA/anti-β₂GPI–mediated EC activation is mediated by TLR4. HUVECs were incubated in medium alone, treated with control RNA of random sequence but identical nucleotide composition as specific siRNA, or with specific siRNA against TLR2, TLR4, or apoER2. Twenty-four hours later, cells were replated in 6- or 96-well plates, incubated for an additional 24 hours, then harvested and analyzed for expression of the targeted protein or assessed for cell-surface E-selectin expression in response to β₂GPI and anti-β₂GPI Abs or TNFα. (A) Cells treated with siRNA to TLR2 or TLR4. (B) Cells treated with siRNA to apoER2. Error bars represent the mean ± SD of triplicate points. P = .0016 by ANOVA for cells treated with TLR4 siRNA versus control or TLR2 siRNA. P = .48 for cells treated with apoER2 siRNA versus control RNA. The results shown are from 1 representative experiment of 3.