Safety and efficacy of human ESC-derived corneal endothelial cells for corneal endothelial dysfunction
DOI: 10.1186/s13578-023-01145-w
Authors: Juan Yu; Nianye Yu; Yao Tian; Yifan Fang; Bin An; Guihai Feng; Jun Wu; Liu Wang; Jie Hao; Liqiang Wang; Qi Zhou; Wei Li; Yukai Wang; Baoyang Hu
Step 1: iPSC/ESC maintainance
day 0 ~ 1
culture “adherent” human ESC in “E8” for “1” days. For ECM, use “vitronectin” at “N/A” “N/A”. Medium should be changed once in “1” days.
Step 2: endothelial 1
day 1 ~ 2
culture “adherent” human NCC in “KnockOut DMEM/F-12” for “1” days. For ECM, use “vitronectin” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “1” days.
| concentration | unit | |
|---|---|---|
| SB431542 | 1.0e-5 | mol/l |
| DKK2 | 1.0e-5 | g/l |
| SU-5402 | 1.0e-7 | mol/l |
| Y27632 | 1.0e-5 | mol/l |
| KSR | — | — |
| GlutaMAX | 1 | % |
| NEAA | 1 | % |
| ITS | 1 | % |
| Ascorbic Acid | 0.1 | % |
Ideally, “CEC” should express “TJP1”, “ATP1A1”, “AQP1”, “CDH2”, “ALCAM” at the end of culture.
Step 3: endothelial 2
day 2 ~ 3
culture “adherent” human CEC in “KnockOut DMEM/F-12” for “1” days. For ECM, use “vitronectin” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “1” days.
| concentration | unit | |
|---|---|---|
| SB431542 | 1.0e-5 | mol/l |
| DKK2 | 1.0e-5 | g/l |
| SU-5402 | 1.0e-7 | mol/l |
| KSR | — | — |
| GlutaMAX | 1 | % |
| NEAA | 1 | % |
| ITS | 1 | % |
| Ascorbic Acid | 0.1 | % |
Ideally, “CEC” should express “COL8A2”, “AQP1”, “ALCAM”, “ZP4”, “SLC4A4”, “PCDH19”, “ATP1B2”, “TJP1”, “CDH2”, “ATP1A1”, “CLCN3” at the end of culture.