American Society of Hematology

Figure 3.

$Release of GPVI from platelets is blocked by a broad-spectrum metalloproteinase inhibitor. Washed human platelets were resuspended in Tyrode buffer and treated with 0.5 μg/mL convulxin in the presence of (A) 100 μM GM6001 or DMSO, 5 mM EDTA, 0.1 mM pepstatin A, 0.1 mM leupeptin, or 0.1 mM soybean trypsin inhibitor (SBTI) or (B) 10 μM PP1, 10 μM PP2, 0.1 μM wortmannin, 30 μg/mL piceatannol, or DMSO. Platelets were incubated at room temperature for 1 hour, then pelleted and lysed in buffer containing proteinase inhibitors and Triton X-100 as described in “Materials and methods.” Aliquots of supernatants and platelet lysates were electrophoresed on SDS/acrylamide gels, transferred to nitrocellulose, and probed with anti-GPVI monoclonal antibody. An aliquot of resting platelet lysate is included on the left hand side of supernatant fractions for molecular weight comparison in panel A. Data are representative of at least 3 experiments with different donors.$

Release of GPVI from platelets is blocked by a broad-spectrum metalloproteinase inhibitor. Washed human platelets were resuspended in Tyrode buffer and treated with 0.5 μg/mL convulxin in the presence of (A) 100 μM GM6001 or DMSO, 5 mM EDTA, 0.1 mM pepstatin A, 0.1 mM leupeptin, or 0.1 mM soybean trypsin inhibitor (SBTI) or (B) 10 μM PP1, 10 μM PP2, 0.1 μM wortmannin, 30 μg/mL piceatannol, or DMSO. Platelets were incubated at room temperature for 1 hour, then pelleted and lysed in buffer containing proteinase inhibitors and Triton X-100 as described in “Materials and methods.” Aliquots of supernatants and platelet lysates were electrophoresed on SDS/acrylamide gels, transferred to nitrocellulose, and probed with anti-GPVI monoclonal antibody. An aliquot of resting platelet lysate is included on the left hand side of supernatant fractions for molecular weight comparison in panel A. Data are representative of at least 3 experiments with different donors.

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