Protocol to generate endothelial cells, pericytes, and fibroblasts in one differentiation round from human-induced pluripotent stem cells

DOI: 10.1016/j.xpro.2023.102292

Authors: Tea Soon Park; Rishabh Hirday; Amir Ali; Roba Megersa; Rafael Villasmil; Eric Nguyen; Kapil Bharti

Step 1: iPSC/ESC maintainance

day 0 ~ 1 (duration missing in database; assumed 1 day)

culture “adherent” human PSC in “E8”. For ECM, use “vitronectin” at “1” “%”. Medium should contain factors shown in the following table.

Ideally, “PSC” should be obtained at the end of culture.

Step 2: Primitive streak 1

day 1 ~ 3

culture “adherent” human PSC in “APEL2 medium” for “2” days. For ECM, use “vitronectin” at “1” “%”. Medium should contain factors shown in the following table, and should be changed once in “2” days.

Ideally, “PS” should be obtained at the end of culture.

Step 3: mesoderm 1

day 3 ~ 5

culture “adherent” human PSC in “APEL2 medium” for “2” days. For ECM, use “vitronectin” at “1” “%”. Medium should contain factors shown in the following table, and should be changed once in “2” days.

Ideally, “mesoderm” should be obtained at the end of culture.

Step 4: endothelial 1

day 5 ~ 6 (duration missing in database; assumed 1 day)

culture “adherent” human PSC in “EGM2”. For ECM, use “FN1” at “0.01” “g/l”. Medium should contain factors shown in the following table.

Ideally, “EC” should be obtained at the end of culture.

Step 5: endothelial 2

day 6 ~ 8

culture “adherent” human PSC in “VascuLife medium” for “2” days. For ECM, use “FN1” at “0.01” “g/l”. Medium should contain factors shown in the following table, and should be changed once in “2” days.

Ideally, “EC” should be obtained at the end of culture.