GSK3β inhibition activates the CDX/HOX pathway and promotes hemogenic endothelial progenitor differentiation from human pluripotent stem cells

DOI: 10.1016/j.exphem.2015.09.007

Authors: Kenji Kitajima; Marino Nakajima; Mai Kanokoda; Michael Kyba; Abhijit Dandapat; Jakub Tolar; Megumu K. Saito; Masashi Toyoda; Akihiro Umezawa; Takahiko Hara

Step 1: iPSC/ESC maintainance

day 0 ~ 1

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

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

Step 2: Primitive streak 1

day 1 ~ 2

culture “adherent” human PSC in “STEMDiff APEL medium” for “1” days. For ECM, use “Matrigel” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “1” days.

Ideally, “PS” should express “EPCAM”, “NCAM” at the end of culture.

Step 3: mesoderm 1

day 2 ~ 3

culture “adherent” human PSC in “STEMDiff APEL medium” for “1” days. For ECM, use “Matrigel” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “1” days.

Ideally, “mesoderm” should express “CDX1”, “CDX2”, “CDX4”, “HOXB2”, “HOXA1”, “TBX3”, “HAND1”, “NKX2-5”, “LEF1” at the end of culture.

Step 4: mesoderm 2

day 3 ~ 5

culture “adherent” human PSC in “STEMDiff APEL medium” for “2” days. For ECM, use “Matrigel” at “N/A” “N/A”. 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 5: endothelial 1

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

culture “adherent” human HEP in “unspecified medium”. Medium should contain factors shown in the following table.

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