Jam-B and Jam-C are required for EC lumen and tube formation in 3D collagen matrices. (A) ECs were treated with the indicated siRNAs and were cultured in 3D collagen matrices for 24 hours before fixation, staining, and photography. Arrows indicate EC lumens. Bar equals 50 μm. SP indicates Smartpool siRNAs from Dharmacon; the other siRNAs are individual single siRNAs. Thus, we used 2 independent siRNAs for each target analyzed. In some cases, we mixed single siRNAs to assess the influence of more than 1 Jam family member. (B) EC lumenal areas were quantitated from the cultures in panel A were traced using Metamorph software. Data shown are normalized to the control sample and presents values of lumenal area per high-powered field (HPF) ± SD. n = 20, P < .01. (C) siRNA-transfected EC lysates were analyzed using Western blots to assess Jam-A, Jam-B, Jam-C, or β-actin (control) expression. (D) ECs were treated with the indicated siRNAs followed by treatment with the indicated adenoviruses. Cultures were established to assess the ability of the treated ECs to form lumens and tubes for 24 hours. Arrows indicate EC lumens. Bar equals 50 μm. (E) EC lumenal areas were quantitated from the cultures in panel D were traced using Metamorph software. Data shown are normalized to the control sample and presents values of lumenal area per HPF ± SD. n = 20, P < .01. Panels B and E: * indicates a significant decrease; +, a significant increase.