cANGPTL4-activated integrin β1 mediates vascular permeability via activated Rac/PAK signaling axis. (A-B) Immunodetection of indicated proteins in anti–Huts-21 (A) and anti-cANGPTL4 (B) immunoprecipitates from total protein lysates of rh-cANGPTL4–treated HMVECs (6 μg/mL). IgG immunoprecipitates serves as control. (C) Immunodetection of indicated proteins from anti–Huts-21 immunoprecipitates of HMVECs treated with either vehicle (PBS) or rh-cANGPTL4. Total integrin β1, Rac1, and PAK served as controls. Protein lysates were collected every 20 minutes (0-180 minutes). Values below each band represent the mean fold change in protein expression level from 3 independent experiments compared with the cognate zero time point. *P < .05; **P < .01. (D) In situ kinetic PLA detection of cANGPTL4: indicated binding partner complexes in cANGPTL4-treated HMVECs transfected with either constitutive-active Rac1 G12V or dominant-negative Rac1 T17N. Values (means ± SD) represent mean fold change in the number of interactions compared with the zero time point, as determined from n = 3 independent experiments (∼ 600 HMVECs) using BlobFinder. *P < .05; **P < .01.