Mechanism Of Activation And Aggregation Of Human Platelets By Bacterium, Streptococcus Sanguinis And Porphromonas Gingivalis

[Objective]To better understand the mechanism of platelet activation and aggregation by Streptococcus sanguinis (S. sanguinis) and Porphyromonas gingivalis (P. gingivalis). The oral bacterium S. sanguinis and P. gingivalis, are amongst the most common pathogens isolated from patients with periodontitis, and have the property of being able to activate platelets, simultaneously trigger intracellular signaling cascades leading to platelet aggregation and thrombus formation. Therefore, in the present study, the mechanism of platelet activation and aggregation by S.sanguinis and P.gingivalis was investigated.[Methods and results]The underlying mechanisms of platelet activation and aggregation stimulated by S.s, P.g and their mixed solution are investigated.1. Effects of Ss, Pg, Ss+Pg mixed solution on platelet aggregation. Washed human platelets were incubated for with same concentration of Ss, Pg, Ss+Pg mixed solution. Platelet aggregation rate of Ss+Pg group was 29.57%±11.24%, the rate of Ss group was 64.84%±4.27%(P＜0.01), and the rate of Pg was 85.31%±3.50% (P＜0.01).2. Flow cytometry assay- The same amount of Ss, Pg, Ss+ Pg mixed solution were selected for the flow cytometric assay with whole blood to test their effects on platelets activation and aggregation, which were determined by P-selectin (CD62P) expression and fibrinogen binding respectively. The results indicated that Ss, Pg could significantly induce platelet activation and aggregation while Ss+ Pg mixed solution has relatively inapparent effect. (p-selectin expression:negative control group was 2.36%±2.81%. Ss+Pg group was 11.63%±2.83%. Compared with Ss+ Pg group, Ss group was15.42%±2.77%(P>0.05), Pg group was upregulated to 21.87%±4.48%(P<0.05); Fibrinogen binding:negative control group: 3.55%±1.99%. Ss+Pg group was 10.54%±1.30%; compared with Ss+Pg group, Ss group was 21.92%±6.17%(P＜0.05), Pg group was upregulated to 21.78%±2.99%(P<0.01).3. Western blot analysis- The platelets were lysed, the total lysate was resolved by 10% sodium dodecylsulfate polyacrylamide gel electrophoresis, and probed with antigelsolin antibody. It was observed that S.s and P.g potently stimulated phosphoinositide 3-kinase (PI3K) and MAPK, as shown by induced Akt, p-38 and Erk phosphorylation. Our results suggest that higher levels of p-selectin expression and fibrinogen binding were found in Ss, Pg, Ss+ Pg mixed solution groups compared with rest group, and the Akt, p-38 and Erk phosphorylation was induced by previous agonist as well. But the stimulation effect of Ss+ Pg mixed solution was relatively lower than those of Ss and Pg.[Conclusion]Streptococcus sanguinis (S.sanguinis) and Porphyromonas gingivalis (P.gingivalis) induced platelet activation and aggregation that likely contribute to the thrombus formation. When Ss is mixed with Pg, the effect of their solution is relatively reduced, which means Ss may play as antagonist with Pg. Further investigations on the molecular mechanisms of human platelets activation and aggregation under stimulation of oral bacterium may still be warranted...