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.
Hayashi
Tadataka
T
Second Department of Surgery, Hamamatsu University School of Medicine, Hamamatsu, Japan.
Mogami
Hideo
H
Murakami
Yusuke
Y
Nakamura
Toshio
T
Kanayama
Naohiro
N
Konno
Hiroyuki
H
Urano
Tetsumei
T
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