Differential Proteomic Analysis of Platelets Suggested Possible Signal Cascades Network in Platelets Treated with Salvianolic Acid B

Background: Salvianolic acid B (SB) is an active component isolated from Danshen, a traditional Chinese medicine widely used for the treatment of cardiovascular disorders. Previous study suggested that SB might inhibit adhesion as well as aggregation of platelets by a mechanism involving the integrin alpha 2 beta 1. But, the signal cascades in platelets after SB binding are still not clear. Methodology/Principal Findings: In the present study, a differential proteomic analysis (two-dimensional electrophoresis) was conducted to check the protein expression profiles of rat platelets with or without treatment of SB. Proteins altered in level after SB exposure were identified by MALDI-TOF MS/MS. Treatment of SB caused regulation of 20 proteins such as heat shock-related 70 kDa protein 2 (hsp70), LIM domain protein CLP-36, copine I, peroxiredoxin-2, coronin-1 B and cytoplasmic dynein intermediate chain 2C. The regulation of SB on protein levels was confirmed by Western blotting. The signal cascades network induced by SB after its binding with integrin alpha 2 beta 1 was predicted. To certify the predicted network, binding affinity of SB to integrin alpha 2 beta 1 was checked in vitro and ex vivo in platelets. Furthermore, the effects of SB on protein levels of hsp70, coronin-1B and intracellular levels of Ca(2+) and reactive oxygen species (ROS) were checked with or without pre-treatment of platelets using antibody against integrin alpha 2 beta 1. Electron microscopy study confirmed that SB affected cytoskeleton structure of platelets. Conclusions/Significance: Integrin alpha 2 beta 1 might be one of the direct target proteins of SB in platelets. The signal cascades network of SB after binding with integrin alpha 2 beta 1 might include regulation of intracellular Ca(2+) level, cytoskeleton-related proteins such as coronin-1B and cytoskeleton structure of platelets.