Endothelium cell medium

Mouse lung microvascular endothelial cells (MLECs) were isolated from C57BL/6 mice. Briefly, the lung was perfused, lavaged and cut into small pieces which were in turn digested with the enzymes dispase and collagenase A (Sigma) for 60 min at 37 °C. Following digestion, single-cell suspensions were passed through a 70-μm filter to remove debris. Endothelial cells were isolated by positive selection using Microbeads binding to CD31. Flow cytometry confirmed that 90% of cells in the final suspension are CD31-positive. Primary MLECs were maintained in endothelium cell medium (Sciencell). For HDL treatment experiments, endothelial cells were cultured in endothelium cell medium (containing 1% FBS) with HDL (50 μg/ml) or human albumins (sigma).

SiHa, HeLa, and HUVEC cell lines were obtained from American Type Culture Collection. SiHa and HeLa cells were grown in minimum essential medium and RPMI‐1640 supplemented with 10% fetal bovine serum (FBS) (Gibco, Thermo Fisher Scientific), 100 U mL⁻¹ of penicillin, 100 mg mL⁻¹ of streptomycin, and 250 ng of amphotericin B (NCM Biotech). HUVECs were cultured in an endothelial cell growth medium (Endothelium Cell Medium, ScienCell) supplemented with 5% FBS (Gibco). Primary CAFs were isolated as described previously^[46] and cultured in the Dulbecco Modified Eagle Medium/F12 (Gibco) supplemented with 10% FBS. All cells were cultured at 37 °C in an atmosphere of 5% carbon dioxide. CS Well 600 chambers (JIYAN, China), which contained 261 wells per chamber, were used to fabricate the 3D organoids. A total of 1000 SiHa/HeLa cells, HUVECs and CAFs per well were mixed in a proportion of approximately 9:3:1, seeded into the chambers and cultured for 48 h to generate organoids.