Fig. 6

Mechanism of migrasome formation. A RPE were transfected with Lv-ctrl or Lv-TSPAN4. Cells were then lysed for western blotting to detect the expression of EVs-secretion related proteins. B RPE cells were transfected with Alix overexpression plasmids or control plasmids, and expression of TSPAN4 was detected by western blot. C Expression of TSPAN4 detected by western blot was quantified by Image J software. D RPE were transfected with Lv-TSPAN4-GFP and were then stained with Alix. Cell were then visualized by a confocal microscope. The white box emphasized the bubbling migrasome. E RPE cells were treated with TGF-β1 and LDN-19,311,892 (antagonist of Smad1/5/8), nsc23766 (antagonist of Wnt), XAV-939 (antagonist of Rac), SB431524 (antagonist of Smad2/3). TSPAN4 expression was measured by western blot and quantified below. F RPE cells were co-treated with SB431524 and TGF-β1. Smad2/3 and phospho-Smad2/3 were detected by western blot. The Smad2/3 phosphorylation was normalized to levels of total Smad2/3. Values represent Mean ± SEM. N.S, not sufficient