The horizontal migration ability of HUVECs was evaluated by using a wound healing assay. The cells were seeded into 24-well plates and grown to confluence. The confluent cells were scratched with a pipette tip, washed three times with PBS to remove the loose cells, and photographed. After being incubated for 18 h in different CM with 5% FBS, the cells were fixed, and pictures were taken in 3 random fields. The cell-free area between the confluent cells was measured by ImageJ (NIH, USA) and calculated by the following formula:

The cell-free clear distance (vertical scratch in the picture) = the clear area/height of the picture; the cell-free clear distance (horizontal scratch in the picture) = the clear area/width of the picture.

The migration distance = the cell-free clear distance at 0 h–the cell-free clear distance at 18 h.

The ratio of the migration distance between the experimental group and the control group was calculated as the relative ratio of the migration distance, which represents cell migration ability [31, 32].

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