Complement anaphylatoxins C3a and C5a induce a failing regenerative program in cardiac resident cells. Evidence of a role for cardiac resident stem cells other than cardiomyocyte renewal
Authors: David Lara-Astiaso; Alberto Izarra; Juan Camilo Estrada; Carmen Albo; Isabel Moscoso; Enrique Samper; Javier Moncayo; Abelardo Solano; Antonio Bernad; Antonio Díez-Juan
Step 1: iPSC/ESC maintainance
day 0 ~ 1 (duration missing in database; assumed 1 day)
culture “adherent” mouse CPC in “DMEM/F12”. For ECM, use “Gelatin” at “0.1” “%”. Medium should contain factors shown in the following table.
| concentration | unit | |
|---|---|---|
| Neurobasal medium | 50 | % |
| PSA | 1 | % |
| GlutaMAX | 1 | % |
| Fungizone | 0.1 | % |
| Gentamicin | 0.1 | % |
| FBS | 10 | % |
| LIF | 1.0e-5 | g/l |
| ITS | 1 | % |
| FGF | 1.0e-5 | g/l |
| EGF | 2.0e-5 | g/l |
| N2 | 1 | % |
| B27 | 2 | % |
Ideally, “CPC” should express “Nkx2.5”, “GATA4”, “GJA1”, “Mef2c” at the end of culture.
Step 2: endothelial 1
day 1 ~ 8
culture “adherent” unknown species cell and mouse CPC in “EGM2” for “7” days. For ECM, use “Matrigel” at “N/A” “N/A”. Medium should be changed once in “7” days.
Ideally, “EC” should be obtained at the end of culture.