Serum starvation and adding serum

In this study, serum starvation was used as an in vitro model of ischemia, while adding serum after starvation was used as in vitro model of reperfusion.

Cardiac fibroblast culture was divided into four groups (four six-well plates). They were cultured in DMEM plus 10% FBS for 3 days. The first group was cultured with DMEM plus 10% FBS until day 8, while others were initially starved for 24 h by culturing in non-serum DMEM medium. After the starvation, one group of the cultures was collected for RNA collection, and the other two groups received serum with either 10% or 20% FBS and continued the culture for 4 days before RNA extraction.