Proangiogenic property of HMCLs: effect of Tie2 blocking. The proangiogenic effect of HMCLs was tested with an angiogenic in vitro assay (Angio-kit). Endothelial-like cells were stimulated with CM (1:2) of XG-6 or U266 in the presence or absence of anti-Tie2 blocking polyclonal antibody (5 μg/mL) or irrelevant anti-IgG antibody. Every 3 days, the medium was replaced with a fresh one. At day 13, cells were fixed and stained using an anti-CD31 antibody. To measure the formation of the capillary network, the number of connections between 3 or more capillary-like structures was counted and quantified. (A) Mean percentage variation ± SD of capillary connections, tubules, and tubule length compared with control of 3 replicate wells of 2 independent experiments (^*P < .05; ^**P < .01). Immunostaining with anti-CD31 antibody to evaluate vessel formation at original magnifications × 4 (B) and × 10 (C) in the following conditions: control (i), VEGF (2 ng/mL) (ii), CM of XG-6 (iii), CM of U266 (iv), CM of XG-6 plus anti-Tie2 blocking antibody (v), and CM U266 plus anti-Tie2 blocking antibody (vi).