Generation and characterisation of scalable and stable human pluripotent stem cell-derived microvascular-like endothelial cells for cardiac applications

DOI: 10.1007/s10456-024-09929-5

Authors: Qasim A. Majid; Bishwa R. Ghimire; Bela Merkely; Anna M. Randi; Sian E. Harding; Virpi Talman; Gábor Földes

Step 1: iPSC/ESC maintainance

day 0 ~ 1

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

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

Step 2: Primitive streak 1

day 1 ~ 2

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

Step 3: mesoderm 1

day 2 ~ 13

culture “adherent” human ESC in “Stemline II” for “11” days. For ECM, use “Matrigel” at “2” “%”. Medium should contain factors shown in the following table, and should be changed once in “11” days.

Step 4: endothelial 1

day 13 ~ 14 (duration missing in database; assumed 1 day)

culture “adherent” unknown species cell in “unspecified medium”.

Ideally, “EC” should express “PECAM1”, “CDH5”, “VWF”, “NRP1” at the end of culture.

Step 5: endothelial 2

day 14 ~ 16

culture “adherent” unknown species cell in “EGM2” for “2” days. For ECM, use “Collagen IV” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “2” days.

Ideally, “EC” should express “PECAM1”, “SOX7”, “VEGF”, “FLT1”, “KDR”, “FLT4”, “CD34”, “COL15A1”, “ANGPT2”, “PNP”, “ESM1”, “IGFBP2”, “RGCC”, “HEY1”, “ILK”, “LIMS1”, “PARVA”, “CTNNB1”, “CTNND”, “WNT7B”, “ADGRA2”, “FZD4”, “NRP1”, “SOX17”, “HCLS1”, “SOX18”, “YAP1”, “LYL1”, “HOXB7”, “HOXB3” at the end of culture.