FIG. 1.
Experimental setup of the 3D SW-CC. (A) Day 1, the surface of an Ibidi μ-Slide 4 Well Ph+ was coated with rat tail collagen I. Then, 105 SMC/cm2 in SMC growth medium were seeded into each well, allowed to adhere, covered with a second coating of collagen I, and cultivated in SMC growth medium for 48 h at 37°C. Then, the growth medium was replaced with serum-free quiescence medium. (B) Day 4, following 24 h of serum starvation, SMC acquired a contractile phenotype. (C) Day 5, 1.5 × 105 EC in growth medium were seeded on top of SMC-collagen I sandwich and incubated for 48 h at 37°C. (D) Day 7, EC growth medium was replaced by coculture medium with 3.3% FCS. The 3D SW-CC can now be cultured for up to 2 weeks. 3D SW-CC, three-dimensional sandwich coculture; EC, endothelial cells; FCS, fetal calf serum; SMC, smooth muscle cells.