Fig. 2

Tie2 plays essential role in induction as well as sustenance of TGFβ-induced myofibroblastic differentiation of CAF. (A) (i) Representative images and quantification of myofibroblasts frequency in UT-CAF and TGF-CAF. Tie2-inhibitor was added 1 h before TGFβ induction (Tie2i > > TGF-CAF) or 48 h after TGFβ induction (TGF-CAF > > Tie2i). Cells were quantified using ImageJ. (ii-iii) Frequency of αSMA stress fibre-positive cells were plotted for three different patient derived CAF. (B) (i) Representative images of Tie2 and pTie2 (Y992) in UT-CAF and TGF-CAF after Tie2-inhibition for 1 h before (Tie2i > > TGF-CAF) or 6 h after TGFβ induction (TGF-CAF > > Tie2i). (ii-v) Bar graph showing quantification of total Tie2 protein and pTie2 (Y992) puncta, calculated using ImageJ software. (C) (i) Representative images of αSMA and pTie2 (Y992) in UT-CAF, TGF-CAF or with increasing doses of ANGPT2 (200 ng/ml, 400 ng/ml) in the presence of TGFβ. Arrowhead indicates pTie2 (Y992) puncta. (ii) Bar graph showing cell frequency with αSMA stress fiber-positive CAF and (iii) pTie2 (Y992) expression by CAF in given conditions. Scale bar = 20 µm. *P < 0.05, **P < 0.01, ***P < 0.001