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. 2005 Jul 18;202(2):271-81.
doi: 10.1084/jem.20050664. Epub 2005 Jul 11.

Defective thrombus formation in mice lacking coagulation factor XII

Thomas Renné  1 , Miroslava Pozgajová, Sabine Grüner, Kai Schuh, Hans-Ulrich Pauer, Peter Burfeind, David Gailani, Bernhard Nieswandt
Affiliations

Defective thrombus formation in mice lacking coagulation factor XII

Thomas Renné et al. J Exp Med. .

Abstract

Blood coagulation is thought to be initiated by plasma protease factor VIIa in complex with the membrane protein tissue factor. In contrast, coagulation factor XII (FXII)-mediated fibrin formation is not believed to play an important role for coagulation in vivo. We used FXII-deficient mice to study the contributions of FXII to thrombus formation in vivo. Intravital fluorescence microscopy and blood flow measurements in three distinct arterial beds revealed a severe defect in the formation and stabilization of platelet-rich occlusive thrombi. Although FXII-deficient mice do not experience spontaneous or excessive injury-related bleeding, they are protected against collagen- and epinephrine-induced thromboembolism VSports手机版. Infusion of human FXII into FXII-null mice restored injury-induced thrombus formation. These unexpected findings change the long-standing concept that the FXII-induced intrinsic coagulation pathway is not important for clotting in vivo. The results establish FXII as essential for thrombus formation, and identify FXII as a novel target for antithrombotic therapy. .

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Figures (V体育官网)

Figure 1.
Figure 1.
Hemostasis and coagulation in FXII deficient mice. (A) Tail bleeding times for wild-type (n = 12) and FXII−/− (n = 11) mice. Each symbol represents one individual. (B) Peripheral blood counts in thousands/microliter and coagulation parameters for FXII−/− and wild-type mice. Values are mean ± SD for 10 mice of each genotype. WBC, white blood cells; aPTT, activated partial thromboplastin time; PT, prothrombin time. (C) Western blots of wild-type or FXII deficient plasma (0.3 μl per lane) using primary antibodies recognizing the contact proteins FXII (αFXII), plasma kallikrein (αPK), and high molecular mass kininogen (αHK). The arrowheads point to FXII (left), plasma kallikrein (middle), and high molecular mass kininogen (right). Numbers indicate molecular mass standards. (D) Wild-type PFP or FXII-deficient plasma was reconstituted with platelets stimulated with calcium ionophore A23187 (+) or buffer (−), and recalcification clotting times were determined. Reduction of the clotting time is given relative to PFP substituted with buffer (n = 6). (E) Recalcification clotting times were determined in PRP (left) and PFP (right) from wild-type (129Sv and C57BL/6), FXII−/−, and FcRγ−/− mice after activation with kaolin (black bars) or collagen (white bars). Mean ± SD from six experiments is given. (F) Thrombin generation in PRP (white bars) and PFP (black bars) from wild-type and FXII−/− mice initiated by the addition of 30 μg/ml of collagen. Thrombin formation was analyzed using the chomogenic substrate S-2238, and absorbance was detected at a 405-nm wavelength after incubation for 20 min. Data shown are mean ± SD (n = 6).
Figure 2.
Figure 2.
Collagen-induced pulmonary embolism model. (A) Mortality associated with i.v. injection of 0.8 mg/kg of collagen and 60 μg/kg of epinephrine. All wild-type mice died within 5 min. Animals alive 30 min after challenge were considered survivors. (B) Platelet counts in control (n = 19), FXII−/− (n = 14), and FcRγ−/− (n = 5) mice 2 min after infusion of collagen/epinephrine. (C) Heparinized PRP from wild-type and FXII−/− mice was stimulated with 10 μg/ml of collagen or 5 μM ADP, and light transmission was recorded in a standard aggregometer. Representative results for each genotype are shown (n = 6). (D) Hematoxylin and eosin–stained sections from lungs of mice 2 min after collagen/epinephrine infusion. Thrombi per visual field were counted at 20×. Mean ± SD for 100 fields is shown. Bar, 200 μm.
Figure 3.
Figure 3.
Defective thrombus formation in FeCl3-injured mesenteric vessels in FXII−/− and factor XI−/− mice. Thrombus formation in vivo was monitored on mesenteric arterioles after topical application of 20% FeCl3. (A) Mean ± SD of platelets adherent at 5 min (129Sv wild type, n = 14; FXII+/−, n = 14; FXII−/−, n = 11; FXI−/−, n = 9). (B) The number of vessels in which one or more thrombi >20 μm in diameter formed during the 40-min observation period. (C) Time to complete occlusion after injury. Each symbol represents one monitored arteriole. (D) Representative images of one experiment.
Figure 4.
Figure 4.
Defective thrombus formation in the injured aorta and carotid artery in FXII−/− and factor XI−/− mice. (A) Thrombosis was induced in the aorta of wild-type (n = 10), FXII−/− (n = 10), and FXI−/− (n = 11) mice by one firm compression with a forceps. Blood flow was monitored with a perivascular ultrasonic flow probe until complete occlusion. The experiment was stopped after 40 min. Each symbol represents one individual. (B) Mechanical injury to the carotid artery was induced by ligation with a surgical filament. 5 min after removal of the filament, thrombus areas (μm2) in wild-type and FXII−/− (n = 10) mice were measured and are expressed as mean ± SD. (C) The photomicrographs show representative images 5 min after injury.
Figure 5.
Figure 5.
Thrombus formation in FXII−/− mice reconstituted with exogenous FXII. FXII-deficient animals received human FXII (hFXII, 2 mg/kg of body weight), and thrombus formation after FeCl3-induced injury was analyzed. (A) Time to complete occlusion after injury. Each symbol represents one monitored arteriole. (B) Representative images from one experiment. (C) FXII−/− mice received 2 mg/kg hFXII, and aortic thrombosis was induced by compression with a forceps. Blood flow was monitored with a perivascular ultrasonic flow probe. Each symbol represents one individual.
Figure 6.
Figure 6.
A model of arterial thrombus formation. (A) Initiation of thrombus formation at sites of vascular lesions is predominantly caused by TF and collagens exposed in the subendothelial matrix. TF in complex with FVII initiates thrombin formation, which promotes fibrin formation and platelet activation. The contribution of FXII in this early phase of thrombus formation is minor. (B) FXII activity on the exposed surface of the thrombus contributes to thrombin generation and additional platelet activation, propagating thrombus growth. Accordingly, FXII, as well as factor XI, deficiency severely impairs thrombus formation.
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Comment in

V体育官网入口 - References

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