Optimizing mesoderm progenitor selection and three-dimensional microniche culture allows highly efficient endothelial differentiation and ischemic tissue repair from human pluripotent stem cells

DOI: 10.1186/s13287-016-0455-4

Authors: Fengzhi Zhang; Lin Wang; Yaqian Li; Wei Liu; Fuyu Duan; Rujin Huang; Xi Chen; Sophia Chia-Ning Chang; Yanan Du; Jie Na

Step 1: iPSC/ESC maintainance

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

culture “adherent” human PSC in “E8”. For ECM, use “Gelatin” at “0.1” “%”.

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

Step 2: mesoderm 1

day 1 ~ 4

culture “adherent” human PSC in “DMEM/F12” for “3” 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 “3” days.

Ideally, “mesoderm” should express “MESP1”, “TBXT”, “GATA4”, “TBX5”, “NKX2.5”, “TAL1”, “GATA2”, “RUNX1”, “HOXB4”, “CXCR4”, “KDR”, “CD34”, “ETV2”, “HOXA9”, “FOXC1” at the end of culture.

Step 3: endothelial 1

day 4 ~ 5

culture “adherent” human mesoderm in “DMEM/F12” 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, “EC” should express “PECAM1”, “CDH5”, “CDH5”, “ID1”, “FLT1”, “FOXC1”, “TAL1”, “ETV2” at the end of culture.

Step 4: endothelial 2

day 5 ~ 8

culture “device” human mesoderm in “DMEM/F12” for “3” days. For ECM, use “gelatin microcryogel” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “3” days.

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