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. Author manuscript; available in PMC: 2014 Oct 17.
Published in final edited form as: Oncogene. 2013 May 13;33(16):2110–2122. doi: 10.1038/onc.2013.160

Figure 3. YKL-40 mediates the interaction of VE-cadherin/β-catenin/actin and cell-cell adhesion in HMVECs.

Figure 3

A. Western blot analysis of VE-cad (VE-cad), N-cad, β-cate, and actin protein expression from the lysates of HMVECs treated with either control or YKL-40 shRNA GSDC-conditioned media for 24 hours. B. Immunoprecipitation of VE-cad from the HMVEC lysates treated for 24 hours with conditioned media from control, YKL-40 shRNA, or YKL-40 shRNA cells plus an anti-VEGF Ab (100 ng/ml) followed by immunoblotting of β-cate. IgG levels were used as a control. Immunoprecipitation of β-cate followed by immunoblotting against actin was similarly performed in HMVEC lysates. *P<0.05 compared with control. n=3. C. Double staining of VE-cad (red) with β-cate (green) in HMVECs treated with conditioned media from control, YKL-40 shRNA, or YKL-40 shRNA cells plus an anti-VEGF Ab to determine the extent of co-localization (yellow) indicated by arrows. Nuclei (blue) were stained by DAPI. A bar: 20 μm. D. Quantification of the immunocytochemistry images in part C, normalized to cell number. N=3, *P≤0.05 compared to control. +P≤0.05 compared to control and YKL-40 shRNA. E. HMVEC aggregation was measured and quantified in the presence of GSDC control or YKL-40 shRNA media with an anti-VE-cad Ab (50 μg/ml). N=3, *P≤0.05 compared to control. F. HMVECs were treated with GSDC control medium in the presence of mAY or mIgG (10 μg/ml) overnight. Cell lystates were subjected to immunoprecipitation with an anti-VE-cadherin antibody followed by immunoblotting against β-cate. G. HMVECs treated with GSDC control medium in the presence of mAY or mIgG (10 μg/ml) were measured for cell aggregation. N=3, *P≤0.05 compared to control.