Recruitment of labelled monocytes by experimental venous thrombi

Objective. Infusing monocytes that have been stimulated to produce fibrinol ytic activators and factors that regulate cell proliferation, migration and maturation, might enhance venous thrombus resolution. The aim of this stud y was to determine the time course of infused monocyte recruitment into ven ous thrombus in an appropriate model of this di;ease. Design and Methods. T hrombus was induced in the inferior vena cava of male Wistar rats using red uced flow (80-90% stenosis). The vessel wall was examined at 1hr by scannin g electron microscopy. Resolving thrombi with surrounding vena cava were ob tained at 1, 7, 14 and 21 days after induction (n = 8). Sections, taken at 0.5 mm intervals (10-15 sections per thrombus), were stained using haematox ylin, Martius Scarlet Blue and antibodies against monocytes, platelets and fibrin. Sections from human venous thrombi (n = 4) were similarly stained. The area occupied by monocytes tin relative pixel units. RPU) was determine d using computer aided image analysis, Peripheral rat blood monocytes were extracted. fluorescently labelled and injected intravenously into 7 rats pr ior to thrombus induction, Vena cava with thrombus was harvested 1 h, 2, 3, 4, 7, 14 and 25 days after induction and their fluorescence measured. The fluorescent content of the caval wall and thrombus was analysed in greater detail at 2 and 25 days after thrombus induction (n = 4 at each time interv al), Results. Experimental thrombi were structurally similar to human throm bus and resolved within 14-21 days. Scanning electron microscopy showed min imal endothelial damage at I h with signs of early thrombus formation (plat elet, red cell leukocyte and fibrin deposition), Neutrophils were the predo minant leukocyte in the thrombus at 1 day, with monocytes making up only 0. 3% (0.04% sem) of the area of the thrombus. There was a steady increase in thrombus monocyte content and by 21 days the percentage area of thrombus co vered by monocytes had increased by over 35 fold to 11.5% (2.3% sam) (p < 0 .001). Initially, monocytes appeared around the edge of the thrombus and be came more evenly distributed through the thrombus as resolution progressed. Labelled monocytes could be found in the circulation up to I week after in fusion. The fluorescent content (RPU) of the thrombus increased over 25 day s (mean RPU At 2 days 0.012. sem 0.005: mean RPU at 25 days 1.062. sem 0.25 2, p = 0.008). The number of labelled monocytes in the vessel wall peaked a t 2 days and decreased thereafter. Conclusion. The structure of thrombi pro duced by this model was comparable to that of human venous thrombi. Endogen ous and injected monocytes migrated into the thrombus during natural resolu tion, possibly via the vein wall. Monocyte targeting could therefore be use d to develop novel treatments for venous thrombosis. with the aim of reduci ng post-thrombotic complications.