Figure 5. Rescue of Ift88 leads to functional rescue of the cilia and prevents shear stress induced EndoMT in Tg737^orpk/orpk EC.

(A,B) Q-PCR and Western Blot analysis, respectively, showing Ift88 levels in WT, Tg737^orpk/orpk stably transfected with pEGFP-N1 (Tg737^o/o-eGFP, sham) and Ift88-mCherry/Ift88* (Tg737^o/o-Ift88*) expression constructs. Transfection with Ift88* results in a 2 fold increase in mRNA level of Ift88* in Tg737^orpk/orpk EC and normalizes protein levels to those of Ift88 in WT EC. (C) Confocal images with optical cross section of Tg737^orpk/orpk cells, immunostained for acetylated tubulin, showing the presence of cilia in rescued Tg737^orpk/orpk-Ift88* EC but not in non-rescued cells (inset). Arrowheads point towards primary cilia and scale bar represents 10μm. (D) Images of Tg737^orpk/orpk-Ift88* cells under static conditions and under 0.5 Pa shear stress. Arrows indicate the direction of flow and scale bars represent 25 μm. (E) Q-PCR showing relative mRNA expression of CD31, αSMA, Pai1, Snai1, Ncad, Klf2, and Klf4 in Tg737^orpk/orpk-Ift88* cells under 0.5 Pa shear stress. Expression is normalized to GAPDH and relative to static shams, as represented by the dashed line. The table shows the ratios of relative mRNA expression levels of Tg737^orpk/orpk–Ift88* and WT EC under shear stress.