FIGURE 1.

LPA activates Rho kinase and NF-κB pathways and induces expression of ICAM-1 and VCAM-1 protein in HUVECs. HUVECs were cultured in EGM-2 growth medium and serum-starved overnight in 0.5% serum prior to addition of 50 μm LPA. At the indicated times, cells were harvested and lysates were prepared for either Western blotting or ELISA as described under “Experimental Procedures.” Western blot analysis of Rho kinase substrates ppMLC and pMYPT-1 (A) and adhesion molecules ICAM-1 and VCAM-1 (B) at the indicated times after LPA addition. ELISAs of phospho NF-κB P65 and total NF-κB P65 in HUVEC lysates were performed at the indicated times after LPA addition (C). NF-κB activity is expressed as a ratio of these two measurements. Values are means (n = 3) ± S.E.