Animal model of left atrial thrombus in congestive heart failure in rats

The aim of the present study was to develop and study a new model of left atrial thrombus (LAT) in rat with congestive heart failure (CHF). CHF was induced by aortic banding for 2 mo, followed by ischemia-reperfusion (I/R) and subsequent aortic debanding for 1 mo. Cardiac function and the presence of LAT were assessed by echocardiography. Masson’s staining was performed for histological analysis. All CHF rats presented with significantly decreased cardiac function, fibrosis in remote myocardium, and pulmonary edema. The incidence rate of LAT was 18.8% in the rats. LAT was associated with severity of aortic constriction, aortic pressure gradient, aortic blood flow velocity, and pulmonary edema but not myocardial infarction or a degree of left ventricular depression. The progressive process of thrombogenesis was characterized by myocyte hypertrophy, fibrosis, and inflammation in the left atrial wall. Fibrin adhesion and clot formation were observed, whereas most LAT presented as a relatively hard “mass,” likely attributable to significant fibrosis in the middle and outer layers. Some LAT mass showed focal necrosis as well as fibrin bulging. Most LAT occurred at the upper anterior wall of the left atrial appendage. Aortic debanding had no significant impact on large LATs (>5 mm2) that had formed, whereas small LATs (<5 mm2) regressed 1 mo after aortic release. LAT is found in a rat model of aortic banding plus I/R followed by aortic debanding. The model provides a platform to study molecular mechanisms and potential new pathways for LAT treatment. NEW & NOTEWORTHY It is critically important to have a rodent model to study the molecular mechanism of thrombogenesis in the left atrium. Left atrial thrombus (LAT) is not a simple fibrin clot like those seen in peripheral veins or arteries. Rather, LAT is a cellular mass that likely develops in conjunction with blood clotting. Studying this phenomenon will help us understand congestive heart failure and promote new therapies for LAT.

A Cyclic Pentapeptide Derived from the Second EGF-Like Domain of Factor VII Is an Inhibitor of Tissue Factor Dependent Coagulation and Thrombus Formation

SummaryWe have previously reported the finding of a cyclic dodecapeptide representing loop I of the second EGF-like domain of FVII, which inhibited TF-dependent FX activation (Örning et al. 1997). The biological activity was localized to the tripeptide motif, Glu-Gln-Tyr. We have now synthesized a cyclic analog of this motif, Cys-Glu-Gln-Tyr-Cys (PN7051), evaluated its anticoagulant and antithrombotic properties and performed a detailed structural characterization of the peptide.PN7051 is a dose-dependent inhibitor of TF-dependent FX activation and coagulation of plasma with IC50 values of 10 ± 2 µM and 1.3 ± 0.2 mM, respectively. It shows inhibitory efficacy on acute thrombus formation in an ex vivo model of human thrombosis using native blood. Fibrin deposition, platelet-fibrin adhesion, platelet-thrombus formation, and thrombin-antithrombin complex formation were all inhibited by PN7051 at IC50 values between 0.3 and 0.7 mM. The cyclic peptide is a non-competitive inhibitor of FX activation with no significant activesite effects on FXa or FVIIa, indicating it affects FVII/TF/FX complex formation and function. Studies on the structure activity relationship revealed that Gln3-Tyr4, but not Glu2 were of importance for inhibition. In line with biological results, NMR measurements of PN7051 suggested that the Gln and Tyr residues configure a structural feature that contributes to the anticoagulant activity. Modeling of the Glu99Gln100Tyr101 motif in FVII and comparison with the solution structure of PN7051 suggest that the cyclic pentapeptide exerts its antithrombotic effect by interfering with the docking of Tyr101 into a hydrophobic pocket in the catalytic domain thereby disrupting an essential interaction between the second EGF-like and the catalytic domains of FVII.