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Objective— Deep vein thrombosis (VT) can result in vein wall injury, which clinically manifests as post-thrombotic syndrome. Postinjury fibrosis may be modulated in part through cellular cysteine-cysteine receptor 7 (CCR7)–mediated events. We tested the hypothesis that late vein wall fibrotic remodeling is dependent on CCR7. Approach and Results— CCR7 −/− and C57BL/6 wild-type mice had inferior vena cava VT induced by nonstasis or stasis mechanisms. In both models, VT size was largest at day 1 and trended down by day 21, and CCR7 + cells peaked at day 8 in wild-type mice. No significant differences in VT resolution were found in CCR7 −/− as compared with wild type in either model. In the nonstasis VT model, vein wall changes consistent with fibrotic injury were evidenced by significant increases in collagen I, III, matrix metalloproteinase 2, and transforming growth factor-β gene expression, increases in α-smooth muscle actin and fibroblast specific protein-1 antigen, and total collagen at 8 days. Correspondingly, SM22α and fibroblast specific protein-1, but not DDR2 + cells, were increased at 8 days. Early wild-type thrombus exposure inhibited profibrotic gene expression in CCR7 −/− in ex vivo vein wall culture. Bone marrow chimera experiments further showed that circulating CCR7 + leukocytes partially rescued midterm profibrotic changes in CCR7 −/− mice. In human histological sections of chronic thrombosed femoral veins, CCR7 + cells were present in the fibrotic areas. Conclusions— Post-thrombotic vein wall remodeling is impaired in CCR7 −/− mice, with a profibrotic phenotype, is dependent on the thrombotic mechanism, and is mediated by circulating CCR7 + cells. Unlike other postinjury fibrotic responses, CCR7 + signaling may be important for positive vein wall remodeling after VT.

Background: Venous thrombosis (VT) and the vein wall responses may be related to interleukin 6 (IL-6) signaling in humans and mice. Inflammatory and fibrotic effects of IL-6 largely signal through the trans pathway: IL-6/soluble IL-6Rα complex associates with ubiquitously expressed signal transducing protein gp130. We hypothesized that IL-6 signaling would differ depending upon thrombosis mechanism (complete v. partial stasis), and disruption of IL-6/IL-6Rα/gp130 axis would alter vein wall response to VT. Methods: Wild type C57BL/6 or IL-6-/- mice underwent induction of VT via inferior vena cava (IVC) stenosis (partial stasis) or ligation (complete stasis). Select WT mice received sgp130Fc chimera to inhibit trans pathway signaling. Vein wall, thrombus, and plasma were harvested at various time points and analyzed by PCR, western blot, ELISA or immunohistochemistry. Results: Compared to sham animals, those undergoing VT (complete or partial stasis) exhibited elevated vein wall IL-6 on post-VT day 4 (n=6-9, p Conclusions: Experimental VT stimulates IL-6 and downstream signaling processes and is dependent on the mechanism of thrombosis. Genetic deletion of IL-6 and selective disruption of trans-signaling pathway by sgp130 decreases vein wall inflammation and influences vein wall remodeling. Targeting of IL-6 trans-signaling pathway may represent a therapeutic option to treat vein wall inflammation post thrombosis.

Objective— Toll-like receptors (TLR) bridge innate immunity and host responses, including inflammation. Sterile inflammation such as a venous thrombus (V t ) may involve TLR signaling, including TLR9. Methods and Results— TLR9 signaling on thrombus resolution was investigated using a mouse model of stasis V t . V t were significantly larger in TLR9−/− mice compared with wild-type (WT) at 2 and 8 days, despite a 2-fold increase in thrombus polymorphonucleic neutrophils at 2 days and monocytes at 8 days, whereas thrombus collagen and neovascularization was 55% and 37% less, respectively, at 8 days. Coincidently, decreased fibrinogen and increased thrombin-antithrombin complex were observed in TLR9−/− mouse thrombi. Vein wall interferon-α, interleukin-1α, and interleukin-2 were significantly reduced in TLR9−/− mice compared with WT. Thrombus cell death pathway markers were not significantly altered at 2 days, but caspase-1 was reduced in TLR9−/− thrombi at 8 days. MyD88 confers TLR9 intracellular signaling, but MyD88−/− mice had V t resolution similar to that of WT. However, inhibition of the NOTCH ligand δ-like 4 was associated with larger V t . Finally, stimulation with a TLR9 agonist was associated with smaller V t . Conclusion— TLR9 signaling is integral for early and mid-V t resolution through modulation of sterile inflammation, maintaining a TH1 milieu, and effects on the thrombosis pathway.

Objective: Sepsis is a strong risk factor for the development of deep venous thrombosis (DVT). However, little is understood about the underlying pathobiology responsible for this phenomenon. This study was undertaken to evaluate the endothelial response of cellular adhesion molecules (CAM) involved in early thrombogenesis in a mouse model of sepsis. Methods: Wild-type mice underwent treatment with vehicle or LPS (10mg/kg), or followed by inferior vena cava (IVC) ligation to generate a venous thrombus (VT). Tissue and plasma were harvested at 2 hours post-intervention and were assessed for CAMs, thrombus size, and markers of extracellular DNA (NETs). Pre-treatment (12 hours prior and at time of thrombosis) with P/E selectin (sel) and ICAM exogenous inhibition was undertaken. Antibody depletion was accomplished in selected mice with anti-Ly6G antibody. To understand leukocyte trafficking to the vein wall under sepsis conditions, a separate group of animals underwent LPS or vehicle injection followed by intravital fluorescence microscopy of rhodamine-6G-labeled leukocytes. Results: Pre-thrombosis, IVC CAMs (P-sel p=0.15, E-sel, p Conclusions: In a mouse model of endotoxemia, CAM elevation and leukocyte trafficking occurs in the serum and local vein wall; and with stasis insult, larger VT form. Inhibitor experiments suggest that ICAM may be central to the sepsis related early VT, while PMNs and P/E sel may be less important. CAM inhibition may be a unique strategy to prevent sepsis-associated DVT.