Real-time analysis of platelet aggregation and procoagulant activity during thrombus formation in vivo.
The exact mechanism of blood vessel thrombus formation remains to be defined. Here, we introduce a new approach to probe thrombus formation in blood vessels of living animals using intravital microscopy in green fluorescent protein (GFP)-transgenic mice to simultaneously monitor platelet aggregation and procoagulant activity. To this end, GFP-expressing platelets and annexin A5 labeled with a fluorescent dye were employed to visualize and analyze platelet aggregation and markers of procoagulant activity (platelet surface phosphatidylserine (PS)). Laser-induced thrombi increased and then decreased in size with time in vessels of living animals, whereas platelet surface PS initiated at the site of injury and then penetrated into the thrombus. PS-positive platelets were predominantly localized in the center of the thrombus, as was fibrin generation. The experimental system proposed here is a valuable tool not only for investigating mechanisms of thrombus formation but also to assess the efficacy of antithrombotic drugs within the vasculature.
Second Department of Surgery, Hamamatsu University School of Medicine, Hamamatsu, Japan.
Tadataka
T
Hayashi
Hideo
H
Mogami
Yusuke
Y
Murakami
Toshio
T
Nakamura
Naohiro
N
Kanayama
Hiroyuki
H
Konno
Tetsumei
T
Urano
D000818:Animals; D001241:Aspirin / Q000494:pharmacology; D001777:Blood Coagulation / Q000187:drug effects / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D002469:Cell Separation; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology; D049452:Green Fluorescent Proteins; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D015759:Ionomycin / Q000494:pharmacology; D008642:Mesenteric Veins / Q000166:cytology / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence; D009391:Nephelometry and Turbidimetry; D010718:Phosphatidylserines / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D013927:Thrombosis / Q000097:blood*
D010718:Phosphatidylserines; D010975:Platelet Aggregation Inhibitors; D049452:Green Fluorescent Proteins; D015759:Ionomycin; D013917:Thrombin; D001241:Aspirin; D002118:Calcium