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. 2007;120(2):251–258. doi: 10.1016/j.thromres.2006.09.003

Figure 4.

Figure 4

αIIbβ3 outside-in regulation of MLC phosphorylation was inhibited by Src inhibitor or PLC inhibitor. 0.4 ml of human washed platelets at 3 × 108/ml pretreated with 10 μM indomethacin and 3 U/ml of apyrase followed by being incubated without (A) or with 50 μM PP2, PP3 (B), 10 μM U73122, or U73323 (C), and then they were seeded on the surfaces coated with BSA or fibrinogen (fib) for indicated times. Bound and unbound platelets were solubilized by 4× Laemmli sample buffer, followed by immediate sonication for three periods of 5 s. Platelet proteins were separated by SDS-PAGE on 15% gels, electrotransferred, and then phospho-MLC or total MLC were blotted using anti-phospho-MLC mAb or anti-MLC pAb. (D) 0.4 ml of washed platelets at 3 × 108/ml from PLCγ2+/+ and PLCγ2-/- mice were pretreated with 10 μM indomethacin and 3 U/ml of apyrase and then they were seeded on the surfaces coated with BSA or fibrinogen (fib) for indicated times. The following procedure is the same as that for human platelets. The data are representatives of at least two experiments.