FIGURE 3.

Src is activated after LMW-HA treatment of HMVEC and regulates tubule formation. Panel A, siControl and siCD44 HLMVE cells were serum-starved for 6 h before incubation for 0, 5, 15, or 30 min with LMW-HA (100 nm). Cells lysates were then prepared and immunoblotted (IB) with phospho-Src (Tyr⁴¹⁸), total Src, CD44, and actin antibodies. Panel B, Src- and control-silenced cells were seeded on Matrigel-coated plates in the presence or absence of LMW-HA (100 nm), VEGF (200 pg/ml), or HMW-HA (100 nm). Images were captured after a 6-h incubation, and tubule formation was quantified using ImageJ image analysis software to measure tubule length and graphically depicted. The asterisks (*) indicate a statistically significant difference (p < 0.05) from control. No significant difference was detected between siControl and siSrc or siSrc and siSrc LMW-HA. Each treatment was performed in triplicate, and experiments were repeated three times. Results are expressed as tubule length per treatment. Panel C, immunoblot analysis of HLMVE cells treated with Src siRNA or control siRNA. Cell lysates were blotted using Src and actin antibodies to confirm Src silencing.