Cyclosporin A toxicity on endothelial cells differentiated from induced pluripotent stem cells: Assembling an adverse outcome pathway

DOI: 10.1016/j.tiv.2024.105954

Authors: Zahra Mazidi; Matthias Wieser; Nicoleta Spinu; Adelheid Weidinger; Andrey V. Kozlov; Kristijan Vukovic; Sara Wellens; Cormac Murphy; Pranika Singh; Liadys Mora Lagares; Madhusudhan Reddy Bobbili; Lisa Liendl; Markus Schosserer; Andreas Diendorfer; Bruno Bettelheim; Wolf Eilenberg; Thomas Exner; Maxime Culot; Paul Jennings; Anja Wilmes; Marjana Novic; Emilio Benfenati; Regina Grillari-Voglauer; Johannes Grillari

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 “spheroid” human PSC in “RPMI 1640” for “1” days. Medium should contain factors shown in the following table, and should be changed once in “1” days.

Step 3: Primitive streak 2

day 2 ~ 3

culture “spheroid” human PSC in “RPMI 1640” for “1” days. Medium should contain factors shown in the following table, and should be changed once in “1” days.

Step 4: mesoderm 1

day 3 ~ 5

culture “spheroid” human PSC in “RPMI 1640” for “2” days. Medium should contain factors shown in the following table, and should be changed once in “2” days.

Step 5: endothelial 1

day 5 ~ 7

culture “spheroid” human PSC in “EGM2” for “2” days. Medium should contain factors shown in the following table, and should be changed once in “2” days.

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

Step 6: endothelial 2

day 7 ~ 8 (duration missing in database; assumed 1 day)

culture “adherent” human PSC in “EGM2”. For ECM, use “Collagen type I” at “0.1” “g/m2”. Medium should contain factors shown in the following table.

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