FIGURE 4.

Human NGF (hNGF) inhibits metalloproteinase-dependent shedding of human platelet GPVI. Metalloproteinase-dependent GPVI shedding from washed human platelets was induced by CRP (2.5 μg/ml, final concentration) (A) or the monoclonal antibody, 14A2 (5 μg/ml, final concentration (B); it activates platelets via FcγRIIa) in the absence or presence of human NGF after 30 min at 22 °C. Supernatant fractions were analyzed by SDS-polyacrylamide gel electrophoresis (reducing conditions), immunoblotting with anti-GPVI antibody (6B12 or 1A5), and visualizing with horseradish peroxidase-conjugated secondary antibody and ECL. Platelet lysate containing intact GPVI is run as a blotting control.