Platelet Activation Modulation by Drugs

After treatment (in vitro: metformin: 1 mM, 6 h, 37 °C; aspirin: 0.1 mM, 6 h, 37 °C; mtDNA: 40 ng/μL, 6 h, 37 °C. In vivo: metformin: 200 mg/kg twice a day for 7 days; aspirin: 15 mg/kg twice a day for 7 days; mtDNA: 25 μg/kg twice a day for 7 days), platelets were incubated with PE-CD62P (Sigma, USA) or PAC-1-FITC (Sigma, USA) for 30 min in the dark, and reaction was stopped by adding ice-cold PBS. The P-selectin and integrin-αIIbβ3 expression on the platelets was measured at 585 nm (FL2) and 530 nm (FL1) by using flow cytometer (Becton - Dickinson, San Jose, CA, USA). In selected experiments platelets were pre-incubated for 0.5 h at 37 °C with neutralizing antibodies against DC-SIGN (120507; R&D Systems, USA, 25 μg/mL) or DNase I (Genentech, USA, 20 μg/mL)33 (link)45 (link).

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Xin G., Wei Z., Ji C., Zheng H., Gu J., Ma L., Huang W., Morris-Natschke S.L., Yeh J.L., Zhang R., Qin C., Wen L., Xing Z., Cao Y., Xia Q., Lu Y., Li K., Niu H., Lee K.H, & Huang W. (2016). Metformin Uniquely Prevents Thrombosis by Inhibiting Platelet Activation and mtDNA Release. Scientific Reports, 6, 36222.

Publication 2016

PE-CD62P PAC-1-FITC DNase I

Aspirin Cold Dc sign Dnase i Fitc Integrin αiibβ3 Metformin Mtdna Neutralizing antibodies P selectin Platelets

Corresponding Organization :

Other organizations : Sichuan University, University of Electronic Science and Technology of China, West China Hospital of Sichuan University, University of North Carolina at Chapel Hill, Kaohsiung Medical University, China Medical University Hospital, China Medical University

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Variable analysis

independent variables

Metformin, 1 mM, 6 h, 37 °C, aspirin, 0.1 mM, 6 h, 37 °C, mtDNA, 40 ng/μL, 6 h, 37 °C, metformin, 200 mg/kg, twice a day, 7 days, aspirin, 15 mg/kg, twice a day, 7 days, mtDNA, 25 μg/kg, twice a day, 7 days

dependent variables

P-selectin expression on platelets
Integrin-αIIbβ3 expression on platelets

control variables

Platelet incubation with PE-CD62P or PAC-1-FITC for 30 min in the dark
Reaction stopped by adding ice-cold PBS
P-selectin and integrin-αIIbβ3 expression measured by flow cytometer

positive controls

Neutralizing antibodies against DC-SIGN, 25 μg/mL, 0.5 h, 37 °C
DNase I, 20 μg/mL, 0.5 h, 37 °C

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