Figure 1.

Schematic overview of experimental setup. (A) For analyses of effects of TGF-β1 treatment the cells were seeded (day 0) and cultivated to form spheroids for 5 days when the first medium refreshment occurred. TGF-β1 treatment started on day 7 and lasted for 3 days, after which the spheroids were harvested. When required, siRNA and transfection reagents were added during seeding at day 0 to achieve a high transfection efficiency. Following transfection, the cells were treated as above. (B) In the standard treatment schedule, the cells were seeded on day 0 and the first medium refreshment took place on day 5. All treatments of FFA, IL-6, CTGF, or angiotensin II (Ang-II), started on day 7 and lasted for 7 days. Medium and compounds were refreshed every 2 to 3 days, until the spheroids were harvested. In experiments, where cells were transfected, this took place on day 0 along with the seeding procedure. TGF-β1—transforming growth factor beta 1; FFA—free-fatty acids; IL-6—interleukin 6; CTGF—connective tissue growth factor; Ang-II—angiotensin II. Parts of the figure were created with BioRender.com.