Manipulation of a VEGF-Notch signaling circuit drives formation of functional vascular endothelial progenitors from human pluripotent stem cells

DOI: 10.1038/cr.2014.59

Authors: Makoto Sahara; Emil M Hansson; Oliver Wernet; Kathy O Lui; Daniela Später; Kenneth R Chien

Step 1: iPSC/ESC maintainance

day 0 ~ 1

culture “adherent” human PSC in “mTeSR-1” for “1” days. For ECM, use “Matrigel” at “0.3” “g/l”. Medium should be changed once in “1” days.

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

Step 2: mesoderm 1

day 1 ~ 4

culture “adherent” human PSC in “N2/B27 medium” for “3” days. For ECM, use “Matrigel” at “0.3” “g/l”. Medium should contain factors shown in the following table, and should be changed once in “3” days.

Ideally, “mesoderm” should express “KDR” at the end of culture.

Step 3: endothelial 1

day 4 ~ 6

culture “adherent” human mesoderm in “StemPro” for “2” days. For ECM, use “Matrigel” at “0.3” “g/l”. Medium should contain factors shown in the following table, and should be changed once in “2” days.

Ideally, “EC” should express “VEC”, “PECAM1”, “KDR”, “CD34”, “ENG”, “NOS3” at the end of culture. Ideally, “EP” should express “KDR”, “FLT1”, “Id1”, “NRP”, “CD34”, “VEC”, “PECAM1”, “CD14” at the end of culture.

Step 4: endothelial 2

day 6 ~ 14

culture “adherent” human EP in “EBM-2” for “8” days. For ECM, use “FN1” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “8” days.

Ideally, “EC” should express “VEC”, “PECAM1”, “ENG”, “NOS3” at the end of culture.