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Background. Therapy for deep vein thrombosis (DVT) resolution in those patients in whom a complication or contraindication to anticoagulation occurs is limited. As prior work suggests that thrombus maturation involves early influx of neutrophils (PMN) and neovascularization, we hypothesized that administering the proinflammatory/proangiogenic chemokine interleukin (IL)-8 might accelerate thrombus resolution. Materials and methods. An established rodent model of DVT (inferior vena cava [IVC] ligation) was used whereby daily intravenous recombinant human IL-8 (1 μg) or vehicle control was administered, with sacrifice at 4 and 8 days. Prior to sacrifice and at harvest, duplex ultrasound of the DVT and femoral venous pressure measurements were performed. Thrombi were analyzed by immunohistochemical techniques for PMN, monocytes, and neovascularization; for chemokines, by enzyme-linked immunoassay; and fibrosis, by hydroxyproline assay and trichrome staining. Results. IL-8 accelerated thrombus dissolution 4 days after IVC ligation, with 6-fold increased thrombus blood flow by duplex ultrasound and a 23% increased absolute femoral venous pressure compared with controls (both P < 0.05). These findings may be partially explained by the fact that animals receiving IL-8, as compared with controls, had 2.5-fold greater thrombus neovascularization (with a trend continuing to 8 days) and increased PMN at 4 days. Thrombus vascular endothelial growth factor was significantly reduced at 8 days postligation, while monocyte chemotactic protein-1 and macrophage inflammatory protein-1α were not altered by IL-8 administration. At 8 days post-IVC-ligation, fibrosis was 12-fold greater with IL-8 treatment compared with controls. Conclusions. A proinflammatory/proangiogenic thrombus milieu, as conferred by IL-8, enhances thrombus resolution and underscores the important relationship between neovascularity and inflammation.

Purpose: The purpose of this study was to evaluate the clinical presentation, diagnosis, and endovascular treatment of iliocaval compression syndrome (ICS). Patients and Methods: During a 3-year period, 18 patients (17 women, 1 man; mean age, 42 years) presented with clinical and imaging findings consistent with ICS. All patients were evaluated with venography and Doppler ultrasound (DUS), 13 of 18 with intravascular pressure measurements, 12 of 18 with intravascular ultrasound, 9 of 18 with air plethysmography (APG), and 4 of 18 with magnetic resonance venography. Seventeen patients were treated with endovascular stenting, one was treated with angioplasty alone, and six received adjunct thrombolysis. Results: Despite the presence of stenosis or occlusion in all cases, APG indicated no iliac vein obstruction (outflow fraction ≥ 40%) in nine patients. DUS revealed acute (6) or chronic (7) unilateral iliofemoral deep venous thrombosis in 13 of 18 patients, whereas the results of five of 18 DUS studies were normal. Recanalization and stent placement (n = 17) or angioplasty (n = 1) was achieved in all patients. The average pressure gradient was 5.6 mm Hg preprocedure and 0.6 mm Hg postprocedure. The primary patency rate demonstrated with DUS (n = 17) and venography (n = 7) at 6 months was 89%. The primary patency rate at 12 months was 79%. Conclusions: ICS often presents as sudden unilateral left lower extremity pain and swelling in young to middle-aged female patients after pregnancy, surgery, or a period of inactivity. Venography, intravascular ultrasound, and magnetic resonance venography demonstrate high sensitivity, whereas APG–outflow fraction demonstrates low sensitivity in the diagnosis of ICS. Endovascular stenting and angioplasty provide safe and effective early and intermediate-term treatment of symptomatic ICS. (J Vasc Surg 2001;34:106-13.)

Objective: Limb length discrepancies (LLDs) in growing children may accompany extremity arterial occlusions. Revascularization with vein grafts has been questioned because of degenerative graft changes observed at other sites. This study was undertaken to define vein graft durability and efficacy in lower extremity revascularizations in preadolescent children. Study Design: Fourteen children (10 boys, 4 girls) with a mean age of 7.3 years (range, 2-11 years) who underwent 16 lower extremity revascularizations with greater saphenous vein grafts were subjected to follow-up with graft ultrasonography, ankle/brachial indices (ABIs) with and without exercise, and limb length determinations. A mean of 5.7 years elapsed between the onset of ischemia and operation. Arterial occlusions resulted from cardiac catheterizations (11), arteritis (1), dialysis cannulation (1), and penetrating trauma (1). Indications for operation included LLD (6), claudication (4), both LLD and claudication (3), markedly diminished ABIs with a potential for LLD (2), and a traumatic transection with hemorrhage (1). The reconstructions with 15 reversed and one in situ vein grafts included iliofemoral (11), femorofemoral (1), aortofemoral (1), femoropopliteal (1), popliteal-popliteal (1), and popliteal-posterior tibial (1) arterial bypass grafts. Results: Among patent grafts available for follow-up, 36% (5 of 14) remained unchanged, 50% (7 of 14) developed nonaneurysmal dilatation, and 14% (2 of 14) exhibited nonprogressive aneurysmal expansion. One graft became occluded, and one graft was lost to follow-up. Collectively, the grafts manifest an 11.2% expansion at an average of 10.7 years postoperatively. ABIs increased from 0.75 preoperatively to 0.97, at an average of 11.0 years postoperatively. LLDs were reduced from 1.66 to 1.24 cm, at an average of 11.4 years postoperatively. Conclusion: Vein graft reconstructions of lower extremity arteries in preadolescent children are durable. They provide an efficacious means of restoring normal blood flow, and in 70% of children their preexisting LLDs were reduced. (J Vasc Surg 2001;34:34-40.)

Background : Selectins, among other adhesion-mediated functions, facilitate and augment thrombosis; standard anticoagulants address thrombosis but also increase bleeding risk. Previous work in animal models showed inhibiting E-selectin decreases venous thrombosis (VTE) and vein wall inflammation without adverse bleeding events, making E-selectin inhibition a favorable therapeutic candidate for VTE. GMI-1271 is a potent, specific E-selectin antagonist. Here we report final analysis of safety, PK, biomarker and bleeding risk profile for GMI-1271 in 2 phase 1 studies of healthy subjects. Methods: The first study was a blinded single ascending dose (SAD) evaluation of 2, 5, 20, or 40mg/kg of GMI-1271 (n=4/cohort), vs placebo control saline (n=4) or active control Lovenox 1mg/kg (n=4). The second was an open-label multiple ascending dose (MAD) study of 10 (n=3) or 20mg/kg (n=3) of GMI-1271 QD d 1-5 vs Lovenox 1mg/kg (n=2) d 1-5. Assessments included safety (adverse events [AEs], clinical labs, bleeding time, PT/PTT, vitals, exam); PK (plasma and urine); and biomarkers. Biomarkers included ELISAs (CRP, D-dimer, IL-10, MPO, Prothrombin fragment 1.2, soluble E-selectin (sEsel), soluble P-selectin (sPsel), sICAM, Thrombin-antithrombin complex (TAT), Tissue Factor (TF), TNFα, VWF activity, and sCD40L); PicoGreen Assay for circulating DNA; flow cytometry (Platelet Monocyte Aggregates (PMA), Mac-1, LFA-1, and CD44) and Thromboelastography (TEG). See Table 1 for functional description. SAD remains blinded to GMI-1271 or placebo (GMI-1271/p). In SAD, we measured biomarker values at baseline, 8 and 24 h after dosing. Analysis was performed of biomarkers at each dose level and then pooled by GMI-1271/p vs Lovenox. In MAD, we measured biomarker values at baseline and day 4. Comparisons were made using unpaired t-test. Results: In total, 32 subjects enrolled and received GMI-1271/placebo (GMI-1271/p; 20), GMI-1271 (6) or Lovenox (L; 6). Safety: All subjects completed dosing uneventfully. Safety findings for GMI-1271/p were unremarkable. No moderate or serious AE were seen. AE overall were as expected in healthy volunteers. In SAD, only 1/20 GMI-1271/p subjects experienced an AE possibly related to study drug (mild transient headache); 0/6 in MAD. In the L group we saw expected mild transaminitis and injection site bruising. In all studies, GMI-1271 had no effect on bleeding time, PT, or PTT. PK: Plasma levels, Cmax, and AUC increased in a linear manner. Cl, Vz, and t ½ were not dose dependent; no accumulation was seen with multiple dosing (Fig 1 and 2). TEG: In SAD, there was a tendency to increase R, K, and decrease A and MA with no change in % lysis in L vs GMI-1271/p. In MAD, we saw a similar trend as SAD except for an increase in % lysis in L. In the pooled SAD analysis, there was a statistically significant difference between GMI-1271/p and L (higher values) for R (p sEsel: In SAD, there was a trend for sEsel to decrease with treatment in all cohorts (combined GMI-1271/p vs L p= 0.017). In MAD, there was non-significant trend for sEsel to decrease between Day 0 and Day 4 in GMI-1271; sEsel levels trended upward with L in comparison. MPO: In SAD, there was a trend for increased values in L vs GMI-1271/p, except for the 40mg/kg cohort. When pooled, there was a significant difference between GMI-1271/p vs L (higher levels) p MAC-1 In SAD, there was no change. In MAD, there was a significant decrease in MAC-1 at the 10mg/kg dose (p No other notable trend was seen in the other biomarkers measured. Conclusion: We report a favorable safety, biomarker and bleeding profile for the E-selectin antagonist GMI-1271 in healthy subjects. There is no signal to suggest GMI-1271 increases bleeding potential based on adverse events, PT, PTT, bleeding time, and TEG values, unlike traditional anticoagulants. An additional unblinded analysis of the biomarkers will be presented at ASH. We note a trend for sE-sel to decrease with GMI-1271 treatment even in this healthy volunteer population, consistent with previous experience and as expected based on mechanism of action. A phase 1 study of GMI-1271 is currently ongoing to evaluate the safety and efficacy of E-selectin antagonism for the treatment of patients with calf vein DVT. PK Figures: Figure 1: SAD; Figure 2: MAD. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures Hemmer: GlycoMimetics, Inc.: Employment, Equity Ownership. Flanner:GlycoMimetics, Inc.: Employment. Parker:GlycoMimetics, Inc.: Employment. Li:GlycoMimetics, Inc.: Employment, Equity Ownership. Froehlich:Novartis: Consultancy; Janssen: Consultancy; Pfizer: Membership on an entity's Board of Directors or advisory committees; Boehringer-Ingelheim: Membership on an entity's Board of Directors or advisory committees; Fibromuscular Disease Society of America: Research Funding; Blue Cross/Blue Shield of Michigan: Research Funding; Merck: Consultancy. Magnani:GlycoMimetics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Thackray:GlycoMimetics: Employment, Equity Ownership. Sood:Bayer: Research Funding.

Introduction: Despite efforts toward prevention, thousands of patients suffer from venous thromboembolic disease (VTE), consisting of deep vein thrombosis (DVT) and pulmonary embolism (PE), each year; this results in significant morbidity and mortality. Clinical prediction rules, such as the Wells score, in conjunction with biomarkers like the D-dimer, are suggested by national guidelines to aide in evaluating for DVT. Substantial interest exists in developing improved diagnostic testing strategies and algorithms to limit the need for more costly, time consuming, and/or invasive testing. Novel biomarkers, like the soluble P selectin (sP-Sel), a cell adhesion molecule that functions to mediate thrombus formation and amplification, have shown promise in this area. While cancer patients suffer from a high burden of VTE, the D-dimer and Wells score are less helpful in this group. This is due to non-specific D-dimer elevations in these patients, and cancer being part of the Wells risk stratification schema. We sought to identify a more specific combination of biomarkers and Wells score, for the diagnosis of DVT that would apply to cancer patients as well. Methods: We previously performed a prospective cohort study of adults presenting with symptoms of DVT afflicting the upper or lower extremities (Vandy et al. J Vasc Surg Venous Lymphat Disord., 2013). Patients were enrolled from December 2008 to July 2013. Those with isolated calf DVT, superficial venous thrombosis, pregnant or breastfeeding, on therapeutic anticoagulation, or with symptoms of simultaneous upper and lower DVT were excluded. After informed consent was obtained, clinical characteristics and biomarkers (D-dimer, C-reactive protein (CRP), Von Willebrand Factor activity (VWF), and sP-Sel) were collected, and duplex ultrasonography was performed to determine if DVT was present. In this subset analysis, cancer patients were compared to non-cancer patients with regards to thrombotic risk factors, biomarker values, and to assess the test characteristics of various combinations of biomarkers and Wells score. Results: A total of 373 patients were enrolled, 151 (40%) in the cancer group, compared to 222 (60%) in the non-cancer group. Cancer patients were more likely to have a DVT on confirmatory testing (58.9% vs. 43.2%). Cancer patients tended to be older, male, have a lower BMI, more recent acute illness, and more central lines, relative to the non-cancer group. Non-cancer patients were more likely to have other potential risk factors for thrombosis, i.e. recent surgery, oral contraceptive use, and a family history of thrombosis. Biomarker values for CRP and sP-Sel were similar, however VWF and D-dimer values were significantly higher in the cancer patients (see table). D-Dimer > 500 with Wells score ≥ 2 was less specific for the diagnosis of DVT among cancer patients compared to non-cancer patients (p=0.003). However, D-Dimer ≥ 500 with sP-Sel ≥ 90 showed not only high specificity, but that specificity was not different between the cancer and non-cancer populations (p=0.88). sP-Sel ≥ 90 and Wells ≥ 2 had similar performance characteristics in both groups as well (p=0.54 for specificity, p=0.14 for positive predictive value (PPV)). Conclusion: Co-morbid risk factors for thrombosis among cancer patients included an older age, male gender, and an increased use of central lines. This group was less likely to have other potential thrombotic risk factors like family history or recent surgery. Our results concur with the finding that the D-dimer, combined with a clinical prediction rule, is not as helpful for DVT in cancer patients. Moreover, this study further supports sP-Sel as a specific test for DVT, that can be combined with clinical information (Wells score) or other laboratory data (D-dimer) to reflect the presence of DVT and potentially rule in clot; the test seems equally useful for both cancer and non-cancer populations. This could potentially be used to guide initial patient management in scenarios where imaging or further testing is not immediately available, such as in an outpatient clinic or rural area. Further exploration of the optimal strategy for utilizing biomarkers and clinical prediction rules in the diagnosis of DVT in cancer is needed. Table Table. Disclosures Sood: Bayer: Research Funding.

SummaryInhibition of P-selectin by antibody or selectin antagonist decreases inflammation and thrombosis. This study evaluates the dose-response relationship using a selectin receptor antagonist. Eight male baboons (Papio anubis) underwent inferior vena caval thrombosis using a 6 h balloon occlusion model. Three animals received 500 μg/kg P-selectin antagonist (rPSGL-Ig) and five 1 mg/kg rPSGL-Ig with or without a non-anticoagulant dose of Dalteparin. These animals were compared to our published results in this model with 4 saline controls and 8 animals that received 4 mg/kg rPSGL-Ig. A statistically significant dose-response relationship existed between rPSGL-Ig dose and thrombosis (p < 0.01), and between rPSGL-Ig dose and spontaneous recanalization (p < 0.05). Inflammatory assessment revealed decreased gadolinium enhancement in all rPSGL-Ig groups compared to previously reported control, despite no significant differences in inflammatory cell extra-vasation. No dose of rPSGL-Ig caused anticoagulation. Selectin antagonism results in a dose-dependent decrease in thrombosis and increase in spontaneous recanalization.

Objective: The purpose of this study was to determine the incidence of femoral and popliteal aneurysms in men and women who have abdominal aortic aneurysms (AAAs) and to assess potential etiologic differences in patients with and without these lower extremity aneurysms. Methods: We studied 313 consecutive patients with AAAs encountered from 1995 to 1998 who underwent prospective ultrasound scanning to detect the presence or absence of femoral and popliteal aneurysms. Patients with and without these extremity aneurysms were compared for differences in potential etiologic risk factors with each other and with a statewide population of patients with AAAs. Results: A total of 51 femoral and popliteal aneurysms were encountered, all occurring in male patients. Among the 251 men with AAAs, the incidence of femoral or popliteal aneurysms was 14%, compared with 0% among the 62 women with AAAs (P

Objective: This study was performed to determine the effectiveness of recombinant P-selectin glycoprotein ligand Ig (rPSGL-Ig) pretreatment to decrease thrombosis and inflammation in experimental venous thrombosis. rPSGL-Ig, a unique mucin-like glycoprotein, has a high affinity for P-selectin. Methods: Twelve juvenile baboons underwent inferior vena cava (IVC) thrombosis with temporary 6-hour IVC balloon occlusion. Before balloon placement, the animals received rPSGL-Ig (4 mg/kg; n = 8) or saline solution for control (n = 4). The animals underwent evaluation with duplex ultrasound scan imaging, magnetic resonance venography (MRV), phlebography, coagulation profile, and tissue analysis at death for cytokines and vein wall leukocyte morphometrics. With the MRV results, thrombus development, thrombus resolution, and inflammation (gadolinium; square millimeters of enhancement) were assessed. Results: Each animal provided two time points for evaluation (days 2 and 6 after balloon occlusion). A significant decrease in IVC thrombosis between balloons was found in the rPSGL-Ig animals (1 of 16) versus the control animals (5 of 8; P

Deep venous thrombosis (DVT) and pulmonary embolism (PE) remain significant heath care problems. Although standard unfractionated heparin effectively treats DVT and PE, low molecular weight heparins (LMWHs) have many theoretical advantages, including less bleeding potential, less thrombocytopenia, improved bioavailability, and more even pharmacokinetics. Additionally, LMWHs are given subcutaneously based on patient weight and do not need to be monitored by blood tests. Multiple studies have compared LMWH to standard heparin for treatment of DVT/PE. LMWHs have a lower risk for major bleeding, recurrent thromboembolic disease, and mortality rate for the treatment of DVT, whereas for PE treatment they are at least equivalent to standard heparin and more convenient. However, not all patients are candidates for such outpatient therapy. The ability to give LMWH subcutaneously and the lack of need for intensive blood monitoring allows for outpatient therapy.

Purpose: A phase I feasibility study was conducted to determine whether cryopreserved venous valved segments would remain patent/competent in a short-term period (6 months). Methods: The target group consisted of 10 patients (C 4-6 , E, A D , P R ). The exclusion criteria included untreated superficial/perforator venous disease, significant venous or arterial obstruction, hypercoagulability or coagulopathy, and significant preexisting medical conditions. Required preoperative tests were venous duplex, ascending/descending venography, and a physiologic study (eg, APG, blood typing, an ankle/brachial index, and if post-thrombotic, a hypercoagulability work-up). A single-valve transplant was placed below all reflux, aided by anticoagulation with or without a distal arteriovenous fistula. Postoperative assessment included duplex scanning/clinical examination (at 1, 3, and 6 months), descending venogram (at 1 month), and physiologic study (at 1 and 6 months). The primary end point was valve patency/competence, with clinical outcome as a secondary end point. Adverse events were recorded. Results: After eliminating protocol violations, nine patients with superficial femoral (5) or popliteal (4) vein valve transplants were studied. Six-month actuarial results show a patency rate of 67% ± 16% and 78% ± 13%, respectively, a primary and secondary competency rate of 56% ± 17% and 67% ± 16%, respectively, and a 100% patient survival rate. Clinical outcome averaged 1.1, with healing and/or freedom from ulcer recurrence, in six of nine patients. A postoperative risk of seroma formation (3) and cellulitis (1) exists. Conclusion: In patients with few remaining therapeutic options, one can achieve a 6-month assisted patency and competency rate of 78% and 67%, respectively, with an improved clinical outcome. (J Vasc Surg 1999;30:854-66.)

Purpose: This review examines the many techniques that have been used for the non-invasive diagnosis of acute and chronic venous disease and was conducted by members of the Committee on Research of the American Venous Forum. It proposes to identify those techniques with the greatest clinical potential, to suggest algorithms for the clinical application of non-invasive techniques in the identification of acute deep venous thrombosis and chronic venous insufficiency, and to identify areas of deficient knowledge and potential areas for future research initiatives. Methods: Review of pertinent clinical and research material. Results: Impedance plethysmography and ultrasonic imaging are the primary non-invasive tools used in the diagnosis of acute deep venous thrombosis. At present, ultrasonic imaging techniques are recommended on the basis of greater diagnostic accuracy in recent comparative clinical trials. Data would suggest that serial evaluation should probably be viewed as the preferred option for symptomatic patients with a negative initial examination and the presence of risk factors or physical findings suggesting a proximal deep venous obstruction/thrombosis. Chronic venous disease is the result of valvular incompetence, with or without associated venous obstruction. Duplex imaging can be used to determine the location and extent of reflux; however, there are reported procedural variations in the performance and interpretation of such studies. Recent innovations in air plethysmography may provide a means of quantifying volume changes, and permit an objective characterization of venous reflux and calf pump efficiency. Conclusions: There are still significant questions that need to be answered by well-designed research initiatives. Research applications that incorporate non-invasive diagnostic techniques may involve the diagnosis, treatment and natural history of acute deep venous obstruction/thrombosis and chronic venous insufficiency, assessment prior to and following venous reconstruction, and the basic science aspects of acute and chronic venous disease. At present, a lack of common standards is, by far, the greatest impediment to an organized research approach to venous disease.

A polycation-sensitive membrane electrode based on the ion-exchanger dinonylnaphthalene sulfonate has previously been developed and used as an end-point detector for the determination of unfractionated heparin in whole blood samples via simple potentiometric titration with protamine. Herein, we report the application of the same methodology for the quantitation of a commercial low-molecular-weight heparin (LMWH) preparation (Fragmin) in whole blood samples at concentrations up to 2 U/ml. Further, an analogous polyanion (heparin)-sensitive electrode is used to estimate the binding constants between protamine and various LMWH preparations. The equilibrium constants (Keq) and the number of binding sites per mole of heparin (n) are determined by recasting the data in the form of a Scatchard plot. Results show that the average molecular weight and molecular weight distribution of the LMWH preparation are important parameters affecting their binding with protamine. Comparable binding constants are obtained for the same LMWH preparations titrated with a synthetic protamine analog, [+18RGD] [acetyl-EA(R2A2R2A)4R2GRGDSPA-NH2].

ch Acute and chronic venous disorders are being increasingly recognized as a healthcare priority in the Western world. The ageand sex-adjusted incidence of venous thromboembolism in the United States has been estim to be 77.6 per 100,000, corresponding to over 275,000 new cases per year. 1 Varicose veins are the most comm clinical manifestation of chronic venous disease, occurring in one-quarter to one-third of Western adult populations, and severe chronic venous insufficiency, with changes and ulceration, may be present in 2% to 5% of Western populations. 4

The purpose of this review is to define animal models of chronic venous disease and to demonstrate how animal studies can impact our understanding and treatment of this disorder. To this end an extensive literature search was conducted highlighting potential animal models of chronic lower extremity venous disease. Scientific investigations using animals to study particular aspects of this disease are also reviewed. This review was conducted by members of the Committee on Research of the American Venous Forum to help provide direction for future venous research endeavors. Useful models of chronic venous occlusive disease involve controlled ligation of a major lower limb vein and multiple tributaries. Such a model can provide sustained venous hypertension and studies using this model have confirmed that an isodiametric graft can provide early hemodynamic relief. Models of primary, postphlebitic, and isolated chronic deep venous insufficiency are available for study. Valve repair or transplantation can positively impact the insufficiency observed in these models. Investigations into valve substitutes have generally been disappointing or are undergoing early evaluation. In conclusion, animal models for the study of some aspects of chronic venous disease do exist and have already affected our clinical approach to patients. The scientific study of basic pathophysiology, diagnostics, end-organ response, and long-term surgical treatments of this disorder in well-controlled animal experiments have not been conducted. (Ann Vasc Surg 1998;12:487–494.)

Vein wall inflammation associated with venous thrombosis is mediated by an imbalance in proinflammatory as compared with antiinflammatory molecules. We hypothesize that IL-10 is an important antiinflammatory cytokine that influences vein wall inflammation and thrombus propagation during venous thrombosis. To test this hypothesis a model of inferior vena caval thrombosis was used. Studies were performed at sacrifice 2 days after thrombus induction and included leukocyte morphometrics, myeloperoxidase activity, vein wall permeability, thrombus weight, and IL-10 ELISA analysis from the vein wall. IL-10 was elevated in the vein wall during venous thrombosis. Neutralization of IL-10 increased inflammation, while supplementation with rIL-10 demonstrated a dose- and time-dependent decrease in inflammation. Interestingly, a low 2.5-μg rIL-10 dose given at time of initiation of thrombosis most significantly decreased inflammation. Thrombus weight was importantly diminished by reconstitution of IL-10. These studies support an important role for IL-10 in the regulation of thrombus-associated inflammation and thrombosis and suggest that IL-10 could be used as a therapeutic agent in the treatment of venous thrombosis.

Purpose: Protamine reversal of heparin anticoagulation during cardiovascular surgery may cause severe hypotension and pulmonary hypertension. A novel protamine variant, [+18RGD], has been developed that effectively reverses heparin anticoagulation without toxicity in canine experiments. Heretofore, human studies have not been undertaken. This investigation hypothesized that [+18RGD] would effectively reverse heparin anticoagulation of human blood in vitro. Methods: Fifty patients who underwent anticoagulation therapy during vascular surgery had blood sampled at baseline and 30 minutes after receiving heparin (150 IU/kg). Activated clotting times were used to define specific quantities of [+18RGD] or protamine necessary to completely reverse heparin anticoagulation in the blood sample of each patient. These defined amounts of [+18RGD] or protamine were then administered to the heparinized blood samples, and percent reversals of activated partial thromboplastin time, thrombin clotting time, and antifactor Xa/IIa levels were determined. In addition, platelet aggregation assays, as well as platelet and white blood cell counts were performed. Results: [+18RGD] and protamine were equivalent in reversing heparin as assessed by thrombin clotting time, antifactor Xa, antifactor IIa levels, and white blood cell changes. [+18RGD], when compared with protamine, was superior in this regard, as assessed by activated partial thromboplastin time (94.5 ± 1.0 vs 86.5 ± 1.3%δ, respectively; p < 0.001) and platelet declines (–3.9 ± 2.9 vs –12.8 ± 3.4 per mm3, respectively; p = 0.048). Platelet aggregation was also decreased for [+18RGD] compared with protamine (23.6 ± 1.5 vs 28.5 ± 1.9%, respectively; p = 0.048). Conclusions: [+18RGD] was as effective as protamine for in vitro reversal of heparin anticoagulation by most coagulation assays, was statistically more effective at reversal than protamine by aPTT assay, and was associated with lesser platelet reductions than protamine. [+18RGD], if less toxic than protamine in human beings, would allow for effective clinical reversal of heparin anticoagulation. (J Vasc Surg 1997;26:1043-8.)

The purpose of this study was to summarize the current issues in chronic venous disease by reviewing of the literature relating to the condition. The review was conducted by members of the Committee on Research of the American Venous Forum and includes the Committee's venous disease and current/future directions. Progress in the understanding and management of chronic venous problems has lagged behind that in arterial disease, despite the large number of patients affected. The complex pathophysiology of venous problems, lack of accepted evaluation standards and lack of prosthetic conduits are some of the factors which contribute to this. New information in these areas has laid the foundation for advances in both operations and non-operative therapy. In conclusion, many opportunities for clinical and basic research in the area of chronic venous disease are available. Application of basic science techniques, including those of molecular biology, will lead to new insights into pathophysiology of chronic venous syndrome. Developments in technology, classification and basic science suggest multiple new potentials therapeutic approaches in the next decade.

Purpose: Venous thrombosis and inflammation are interrelated. P-selectin contributes to activation of leukocyte-mediated inflammation. Therefore, we hypothesized that the neutralization of P-selectin would decrease vein wall inflammation and thrombosis.Methods: Twelve baboons underwent infrarenal inferior vena caval balloon occlusion to induce thrombosis. Two groups of four baboons received neutralizing intravenous anti-P-selectin antibody (PSab) GA6 or CY1748 before occlusion and at days 2 and 4. Four baboons received saline control injections. One baboon per group was killed at days 2, 6, and 13, and at 2 months. Analysis included phlebography, ultrasound, gadolinium (Gd)-enhanced magnetic resonance venography (reflecting vein wall inflammation), and histologic, morphometric, and protein evaluation of the vein wall. Thrombus presence or absence was assessed.Results: By day 2 in PSab baboons, vein wall Gd enhancement was decreased in the mid–inferior vena cava and the right iliac vein (p < 0.05; GA6 vs control baboons), normalizing by 2 months. The mid–inferior vena cava revealed fewer neutrophils and total leukocytes in PSab baboons; however, for GA6 in the right iliac vein these decreases were not present despite the absence of Gd enhancement; they were decreased with CY1748. PSab baboons demonstrated significantly less thrombus than control baboons (p < 0.01, GA6 and CY1748 vs control baboons).Conclusions: Anti-P-selectin antibody decreases vein wall inflammation and thrombus formation. Inhibition of P-selectin may be useful in venous thrombosis prophylaxis. (J Vasc Surg 1997;25:816-28.)

Although duplex ultrasound is the standard for the diagnosis of lower extremity deep venous thrombosis (LE-DVT), imaging is not always available. The use of D-dimer can exclude (high-sensitivity), but not rule in (low-specificity) LE-DVT. Previously, we demonstrated that soluble P-selectin (sP-sel) in combination with the Wells score, establishes the diagnosis of LE-DVT with a specificity of 96% and a positive predictive value of 100%. In order to validate our previous results, we applied the model to a separate but similar patient cohort. Additionally, we analyzed the role of biomarkers for diagnosing upper extremity DVT (UE-DVT).Between April 2009 and March 2012, all patients presenting for a duplex ultrasound exam with concern of DVT were screened. Demographics, clinical data, D-dimer, sP-sel, C-reactive protein, a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13, and von Willebrand factor levels were prospectively collected in 279 patients (234 LE-DVT, 45 UE-DVT). Continuous and categorical variables among patients with DVT were compared with patients without DVT. The diagnostic sensitivity, specificity, positive predictive value, and negative predictive value were then calculated using our previously derived cut points to rule in or exclude DVT.Among 234 patients evaluated for LE-DVT, 112 (48%) patients had a confirmed LE-DVT with significant differences in all biomarkers. When Wells score ≥2, sP-sel could rule in LE-DVT with a specificity of 97.5% and a positive predictive value of 91%, which was more accurate than Wells score ≥2 and D-dimer (specificity, 65%; positive predictive value, 69%). When Wells score was2, D-dimer was superior to sP-sel for excluding the diagnosis of LE-DVT (sensitivity, 98%; negative predictive value, 95% vs sensitivity, 91%; negative predictive value, 79%). The use of additional biomarkers did not increase accuracy. Had imaging not been available, we could have correctly ruled in or ruled out LE-DVT in 29% (67/234) of patients. The use of sP-sel in UE-DVT was nondiagnostic.We demonstrate that when Wells score ≥2, sP-sel is an excellent biomarker to rule in LE-DVT. Different from our previous study, D-dimer and a Wells score2 was most sensitive at excluding a diagnosis of LE-DVT. Combined, Wells score, sP-sel, and D-dimer can both rule in and exclude LE-DVT in approximately one-third of patients.

The use of thrombolytic agents has greatly improved patient outcomes, but the prothrombotic response to these drugs in vivo is unknown. Approximately 24 h after we induced thrombosis in male Sprague–Dawley rats, we placed an infusion line in the inferior vena cava and administered either saline or a thrombolytic agent (tissue plasminogen activator [tPA] or plasmin) for 30 min. Blood was drawn immediately after infusion; rats were euthanized 24 h after infusion for collection of blood and tissue (inferior vena cava and thrombus). Thrombus size was decreased in the tPA-treated rats but not in those that received saline or plasmin; this change correlated with the significant rise in D-dimer levels noted immediately after infusion in the tPA-treated rats. Plasma soluble P-selectin, a prothrombotic marker, was elevated at 24 h in the plasmin group compared with the other treatment groups. There were no significant differences in plasma C3a, C5a, or C5b9 levels or in thrombus C3 levels between groups. According to ultrastructural analysis, thrombus structure and vein wall effects did not differ between groups. Local tPA did not induce a prothrombotic state during acute DVT or after thrombolytic therapy in a rodent model of venous thrombolysis. Conversely, levels of the prothrombotic marker plasma soluble P-selectin increased when plasmin was administered.

OBJECTIVE To investigate if plasma DNA is elevated in patients with deep vein thrombosis (DVT) and to determine whether there is a correlation with other biomarkers of DVT. BACKGROUND Leukocytes release DNA to form extracellular traps (ETs), which have recently been linked to experimental DVT. In baboons and mice, extracellular DNA co-localized with von Willebrand factor (VWF) in the thrombus and DNA appeared in circulation at the time of thrombus formation. ETs have not been associated with clinical DVT. SETTING From December 2008 to August 2010, patients were screened through the University of Michigan Diagnostic Vascular Unit and were divided into three distinct groups: 1) the DVT positive group, consisting of patients who were symptomatic for DVT, which was confirmed by compression duplex ultrasound (n=47); 2) the DVT negative group, consisting of patients that present with swelling and leg pain but had a negative compression duplex ultrasound, (n=28); and 3) a control group of healthy non-pregnant volunteers without signs or symptoms of active or previous DVT (n=19). Patients were excluded if they were less than 18 years of age, unwillingness to consent, pregnant, on an anticoagulant therapy, or diagnosed with isolated calf vein thrombosis. METHODS Blood was collected for circulating DNA, CRP, D-dimer, VWF activity, myeloperoxidase (MPO), ADAMTS13 and VWF. The Wells score for a patient's risk of DVT was assessed. The Receiver Operating Characteristic (ROC) curve was generated to determine the strength of the relationship between circulating DNA levels and the presence of DVT. A Spearman correlation was performed to determine the relationship between the DNA levels and the biomarkers and the Wells score. Additionally the ratio of ADAMTS13/VWF was assessed. RESULTS Our results showed that circulating DNA (a surrogate marker for NETs) was significantly elevated in DVT patients, compared to both DVT negative patients (57.7±6.3 vs. 17.9±3.5ng/mL, P

Venous thromboembolic disease (VTE) remains a significant source of morbidity and mortality. As non-specific subjective complaints and a paucity of objective clinical examination findings complicate the diagnosis of both deep venous thrombosis (DVT) and pulmonary embolism, diagnostic modalities remain essential. Compression ultrasound remains the gold standard for DVT diagnosis. Reliable imaging is not always available making a serologic diagnosis, or biomarker, highly desirable. While D-dimer, a highly sensitive biomarker, is useful for excluding acute VTE, it lacks the specificity necessary for diagnostic confirmation. As such, ongoing research efforts target and support the utility of alternative plasma biomarkers to aid in the diagnosis of VTE including selectins, microparticles, IL-10 and other inflammatory markers. These molecular markers may also predict recurrence risk, guide length and modality of treatment, and predict which thrombi will resolve spontaneously or recanalize, thus potentially identifying patients who would benefit from more aggressive therapies than standard anticoagulation.

Protamine sulfate reversal of heparin anticoagulation may be associated with adverse cardiovascular side effects. The purpose of this study was to determine whether diminished systemic oxygen consumption and hemodynamic changes were more likely to accompany rapid versus slow protamine administration.Fifteen patients undergoing abdominal aortic aneurysm resection in a prospective randomized double-blinded study received intravenous protamine (1.5 mg/kg) rapidly during a 3-minute period (group I, n = 7) or slowly during a 15-minute period (group II, n = 8). Systemic oxygen consumption (VO2) and hemodynamic parameters were assessed for up to 20 minutes after protamine administration began.Blood pressure declines (millimeters of mercury) were greatest in group I with rapid protamine administration (-19 systolic and -9 diastolic) compared with group II with slow protamine administration (-12 systolic and -1 diastolic). Heart rate fell markedly in both groups I and II. Cardiac output (CO) declined in group I at virtually all time periods. Similar CO declines in group II occurred 10 minutes after protamine infusion had begun and persisted for 3 minutes after protamine administration was complete. Maximum VO2 decreases were -16% (60 seconds into protamine infusion) and -13% (1.5 minutes after protamine infusion) in groups I and II, respectively, with statistically significant declines (p0.05) occurring only in group I compared with baseline values. Statistically significant differences (p0.01), however, were found when mean declines during and after protamine infusion were compared with controls for both CO and VO2 in both groups.Significant declines in systemic VO2 and hemodynamic perturbations accompany protamine reversal of heparin anticoagulation during aortic surgery. Rapid protamine administration increases the magnitude of these adverse responses.

Background Administration of the arginine-rich, highly charged protamine (PROT) molecule has been associated with episodes of acute left ventricular (LV) dysfunction. The objective of the present study was to test the hypothesis that PROT has direct effects on isolated LV myocyte contractile processes and sarcolemmal transduction systems. Methods and Results Exposure of porcine LV myocytes (n=305) to 40 μg/mL PROT (reflecting a dose of 2.5 mg/kg) decreased basal contractile function and β-adrenergic responsiveness. For example, myocyte percent shortening was 4.3±0.1% in control myocytes and decreased to 2.8±0.2% in the presence of 40 μg/mL PROT ( P P 40% from forskolin. In addition, PROT abolished the inotropic effects of ouabain on myocyte contractile function. To determine contributory mechanisms for the effects of PROT on myocyte sarcolemmal systems, β-receptor– and cardiac glycoside–binding characteristics were determined in sarcolemmal preparations. β-receptor binding was 175±10 fmol/mg and was reduced to 140±6 fmol/mg in the presence of PROT ( P Conclusions The unique findings from the present study suggest that a fundamental contributory mechanism for the changes in LV function observed after protamine administration may be the direct effect of unbound protamine on myocyte contractile processes.

This investigation assessed protamine reversal of heparin anticoagulation by formation of a protamine-heparin alpha-helix by use of a new designer-variant protamine [+18BE] that was made from an existing protamine variant [+18B] whose non-alpha-helix-forming amino acid proline (P) was replaced by an alpha-helix-forming glutamic acid (E). The rate of administration of the new [+18BE] variant protamine on efficacy and toxicity in comparison to that of [+21] standard protamine and [+18B] was also studied.Acetyl-EAA(K2A2K2A)4K2-Amide [+18BE] was administered intravenously in a 1:1 dose to low-molecular-weight heparin (LMWH)-anticoagulated (intravenous 150 IU antifactor Xa/kg) dogs over 10 seconds or 3 minutes (n = 7, each group). Reversal efficacy was documented by measuring activated clotting time, thrombin clotting time, antifactor Xa, and antifactor IIa. Toxicity was defined by measuring systemic blood pressure, heart rate, cardiac output, pulmonary artery pressure, and oxygen consumption. Measurements were made at baseline, after administration of LMWH, before its reversal, and for 30 minutes thereafter. Results were compared with those after LMWH reversal with [+21] standard protamine and the [+18B] variant. A total toxicity score (TTS) was calculated for each compound from maximal declines in blood pressure, heart rate, cardiac output, and oxygen consumption.LMWH anticoagulation reversal was significantly (p0.01) less toxic over 10 seconds and 3 minutes with the [+18BE] designer variant (TTS -2.3, -2.2) compared with the [+21] standard protamine (TTS -6.4, -7.2). Percent LMWH reversal at 3 minutes revealed [+18BE] to have antifactor Xa activity as high as 91%, compared with 68% for protamine [+21], when given over 3 minutes (p0.05).This investigation documents that a new designer variant of protamine [+18BE] has superior efficacy compared with [+21] standard protamine for reversal of LMWH anticoagulation and that this occurs with a highly favorable toxicity profile.

Objectives:To define current practice regarding the use of pharmacological prophylaxis to prevent postoperative graft occlusion.Design:Prospective open questionnaire.Materials and Methods:Questionnaires regarding this subject were sent to vascular surgeons throughout the world to analyse current practice.Results:651 questionnaires were returned with a response rate of 62% and form the basis for this report. Data from 100 334 vascular reconstructions were reported in this survey. Prophylaxis against postoperative graft occlusions was common. Treatment periods were usually greater than 1 year. Among carotid surgery patients, 82% received prophylaxis, consisting mainly of low-dose acetysalicylic acid (ASA). In Mid-Europe the use of oral anticoagulation was more common than in other regions (p < 0.001). Among aneurysm surgery patients, 38% received prophylaxis. For infrainguinal bypass, ASA in low dose was the most commonly used agent worldwide. However, oral anticoagulation was more frequent in Mid-Europe, in contrast to South America where the combination of ASA and dipyridamole was most common. Considerable geographical differences regarding patient selection, the frequency of specific procedures and operative techniques existed.Conclusions:Important world-wide differences exist regarding prophylaxis for postoperative graft occlusion.

Purpose: This study was undertaken to characterize the changing operative treatment of pediatric renovascular hypertension and subsequent outcomes in a 30-year experience at a single institution. Methods: Clinical data were analyzed on 57 pediatric patients, 24 girls and 33 boys, ranging in age from 10 months to 17 years, who underwent operations for renovascular hypertension from 1963 to 1993 at the University of Michigan. Renal artery disease included atypical medial - perimedial dysplasia, often with secondary intimal fibroplasia (88%), and inflammatory mural fibrosis (12%). Abdominal aortic narrowings affected 15 patients. Data were categorized into three chronologic eras (I:1963 - 1972, II:1973 - 1980, and III:1981 - 1993) to allow identification of therapeutic trends. Results: Primary surgical procedures were undertaken 74 times. Ex vivo reconstruction was necessary once. Primary operations included aortorenal bypass with autogenous vein grafts ( n = 26) or internal iliac artery grafts ( n = 7); iliorenal bypass with vein grafts ( n = 2); renal artery resection beyond the stenosis and reimplantation into the aorta ( n = 10), the main renal artery ( n = 2), an adjacent segmental renal artery ( n = 3), or the superior mesenteric artery ( n = 3); renal artery resection and reanastomosis ( n = 3); focal renal arterioplasty ( n = 2); operative dilation ( n = 7); splenorenal bypass ( n = 2); and primary nephrectomy ( n = 7). Among 23 primary operations performed in era I, 56.5% were aortorenal bypasses with vein grafts, but in era III this form of revascularization represented only 3% of 33 primary operations. No reimplantations were performed in era I, whereas reimplantations accounted for 51.5% of era III procedures. Thirteen patients underwent staged or concomitant aortic reconstructions with thoracoabdominal aortoaortic bypass grafts ( n = 5) or patch aortoplasty ( n = 8). Fourteen patients underwent a total of 20 secondary operations, including seven secondary nephrectomies. Operative therapy benefited 98% of these children: hypertension was cured in 45 (79%), improved in 11 (19%), and unchanged in one (2%). There were no operative deaths. Conclusions: Contemporary surgical management emphasizes direct reimplantation of main renal arteries into the aorta, reimplantation of segmental arteries into adjacent renal arteries, patch aortoplasty for associated abdominal aortic coarctations, and single-stage revascularizations. Pediatric patients with renovascular hypertension clearly benefit from carefully executed operative therapy. (J VASC SURG 1995;21:212-27.)

Abstract Thrombosis and inflammation are closely related. However, the response of the vein wall to venous thrombosis has been poorly documented. This study examines the hypothesis that venous thrombosis is associated with an inflammatory response in the vein wall. In a rat model of inferior vena caval thrombosis, vein wall was temporally examined for inflammation by assessment of histopathology, leukocyte morphometrics, and cytokine levels. Animals were killed 1 hour and 1, 3, and 6 days after thrombus induction. Our findings demonstrated an early (day 1) neutrophil infiltration into the vein wall followed by a later (days 3 and 6) monocyte/macrophage and lymphocyte response. Cytokines were elevated only under conditions of venous thrombosis. Levels of epithelial neutrophil activating protein–78 (ENA-78), tumor necrosis factor–α (TNF), interleukin-6, and JE/monocyte chemoattractant protein–1 (JE/MCP-1) increased over the 6-day period, while macrophage inflammatory protein-1α (MIP-1α) peaked at day 3 after thrombus induction. Additionally, rats were passively immunized with neutralizing antibodies to TNF, ENA-78, MIP-1α, JE/MCP-1, intercellular adhesion molecule–1 (ICAM-1), and CD18 compared with control antibodies. The most effective antibody early after thrombus induction for attenuating vein wall neutrophil extravasation was anti-TNF ( P P

Statins, particularly rosuvastatin, have recently become relevant in the setting of venous thrombosis. The objective of this study was to study the non-lipid lowering effects of rosuvastatin in venous thrombosis in mice with hyperlipidemia.An inferior vena cava ligation model of venous thrombosis in mice was utilized. Saline or 5mg/kg of rosuvastatin was administered by gavage 48hs previous to thrombosis. Blood, the inferior vena cava, thrombus, and liver were harvested 3, 6hours, and 2days post-thrombosis. Thrombus weight, inflammatory markers, and plasminogen activator inhibitor-1 expression and plasma levels were measured. Also, neutrophil migration to the IVC was assessed.Rosuvastatin significantly decreased thrombus weight, plasminogen activator inhibitor-1 expression and plasma levels, expression of molecules related to the interleukin-6 pathway, and neutrophil migration into the vein wall.This work supports the beneficial effects of rosuvastatin on venous thrombosis in mice with hyperlipidemia, due to its non-lipid lowering effects.

Introduction: Plasminogen activator inhibitor (PAI-1) is the primary regulator of urokinase (u-PA) and tissue (t-PA) plasminogen activators. Observations in mice with impaired fibrinolysis have demonstrated an alternate pathway of thrombus resolution by matrix metalloproteinase (MMP)- 9 and 2. TGF-β is a potent inducer of TIMP-1 expression and is found in both thrombus and vein wall post-thrombosis. Hypothesis: PAI-1 deficiency and resulting hyperfibrinolytic state will improve thrombus resolution and result in reciprocal downregulation of MMP-2 and MMP-9 via TGF-beta induction of TIMP-1, resulting in decreased vein wall fibrosis. Methods: Inferior vena cava (IVC) ligation was performed in wild-type (WT) mice (N=194) and PAI-1 deficent mice (N=164). IVC, thrombus and blood samples were collect at 2, 6 and 14 days post procedure. The IVC and thrombus were assessed for thrombus weight and samples collected for zymography (MMP-2 and MMP-9), ELISA (TIMP-1, TGF-β) and histology. Results: PAI-1 deficient mice had similar thrombus weight (TW) versus WT at 2 days, but smaller at days 6 (p Conclusions: (1) PAI-1 deficient mice exhibit a hyperfibrinolytic phenotype with normal thrombotic response to IVC ligation, but improved thrombus resolution when compared to WT mice. (2) MMP-9 and MMP-2 were suppressed, confirming a reciprocal relationship between plasmin and MMP activation as mediators of thrombus resolution. (3) TIMP-1 production was elevated in vein wall independent of TGF-beta. (4) Decreased neutrophils at day 2 correlated with decreased MMP-9 and decreased monocytes at day 14 correlated with decreased MMP-2, suggesting a plasmin-induced inhibition of leukocyte recruitment. (5) Final histology showed no difference in vein wall fibrosis or thickness at chronic time points, indicating that improved thrombus resolution does not lessen vein wall fibrosis.

Galectin-3 binding protein (Gal-3BP) was detected in procoagulant microparticles from mice and patients with venous thrombosis (VT), in prior studies from our laboratory. However, the mechanistic role of Gal-3BP and Galactin-3 (Gal-3) in the pathophysiology of thrombosis is unknown. Our hypothesis is that Gal-3BP and Gal-3 are critical to thrombus formation. Methods: Mice : Using our inferior vena cava ligation mouse model of VT, microparticle samples from thrombosed and non-thrombosed wild type mice were analyzed for Gal-3BP. We then either neutralized Gal-3BP, using an antibody (Anti-M-Gal-3BP), or utilized Gal-3 knock-out (Gal-3 KO) mice, compared to mice given saline as controls (NaCl). We evaluated thrombus weight (TW), inflammatory cell counts, and the cell source of the molecules Gal-3BP and Gal-3 using western blot. Patients: Microparticles and plasma samples from patients who were positive or negative for VT were tested for Gal-3BP using ELISA. Results: Mice : Two days after thrombosis, blocking Gal-3BP significantly reduced both vein wall inflammatory cells ( p =0.0369) and TW ( p =0.0024), compared to NaCl. In Gal-3 KO mice, TW was significantly smaller versus controls ( p =0.0061). We found Gal-3BP on microparticles and platelets while Gal-3 was on monocytes. Patients : Gal-3BP was significantly elevated in microparticles ( p =0.0495) and in plasma ( p =0.0400), versus patients negative for VT. Conclusions: We demonstrated a clear link between Gal-3BP and VT, in both mice and patients. This is the first time that Gal-3BP has been shown to be involved with thrombus formation, making it a promising biomarker and potential target for therapeutic interventions of VT.

Deep vein thrombosis (DVT) and pulmonary embolism are common mortal/morbid diseases worldwide, and improvements in prophylaxis and therapy have been elusive. The basic pathophysiology of how a thrombus forms in vivo and how it resolves are now better understood through experiments with animal models. Inflammation can both cause and incite venous thrombosis. Leukocytes, platelets, and coagulation factors coalesce locally after a thrombogenic stimulus occurs. Cell adhesion molecules, such as P-selectin, bridge the interface of thrombosis and inflammation, and are both biomarkers for, and pathogenic in DVT. Once DVT forms, natural thrombolysis occurs primarily via the urokinase-type plasminogen activator (uPA)–plasmin system. Alternative pathways also exist for venous thrombus resolution, including the matrix metalloproteinases. A new concept in DVT resolution related to sterile inflammation is clearance of procoagulant necrotic leukocytes and platelets, in part mediated by TLR9 signaling. A consequence of DVT is the proximate vein wall injury, which is dependent on the mechanism and duration of the thrombus-vein wall contact. Both vascular smooth muscle cells and endothelial cells maintain normal vein homeostasis, but adopt an injury response after thrombosis that may lead to vein fibrosis. In humans, multiple biomarkers are now available, but increased specificity for venous thrombosis is still elusive. Some of these biomarkers such as P-selectin may also play a causative role, and be attractive therapeutic targets for nonanticoagulant DVT prophylaxis and treatment.

Background: GMI-1271 is a potent, specific E-selectin antagonist which targets interactions between cancer cells and the bone marrow niche thereby increasing tumor sensitivity to chemotherapy; avoids HSC mobilization; and inhibits thrombosis without increasing bleeding risk. Delayed inflammatory cell recruitment into the vein wall post thrombus induction indicates a possible decrease in leukocyte activation, which may be crucial in treating both acute and cancer associated thrombosis. Here we report first-in-human evaluation of GMI-1271 for safety (including bleeding), PK, and biomarker data for effects on adhesion and mobilization. Methods: Two Phase I single dose escalation DBRCT in healthy subjects evaluated 2, 5, 10, and 20 mg/kg IV doses of GMI-1271. Study 1 is complete and unblinded (GMI-1271 vs placebo). Study 2 is ongoing, remains blinded (GMI-1271 vs placebo) with open-label positive control (Lovenox), and is reported in aggregate. In both studies, assessments included safety (adverse events [AEs], clinical labs, bleeding time, PT/PTT, vitals, and exam); PK (drug levels in plasma and urine); and biomarkers (plasma soluble E-selectin [sEsel], sPsel, sICAM-1 levels; and blood CD34+ counts). Only sEsel and CD34+ are reported for Study 1; sEsel, sPsel, sICAM-1 for Study 2. Comparisons to baseline were made by ANOVA and paired t-test models; PK analyses were for total clearance (CL), volume of distribution (Vz), elimination half-life (t½), fraction excreted (Fe), and renal clearance (CLr). Results: Forty-six subjects enrolled; 30 received GMI-1271 (10 each at 2 and 5 mg/kg; 6 at 10 mg/kg; and 4 at 20 mg/kg) and 16 received control (13 placebo, 3 Lovenox). Safety: All subjects completed the studies; labs, vitals, and exam were unremarkable. Bleeding times and PT/PTT were unaffected by E-selectin inhibition. No serious AEs were seen. For Study 1, AEs were seen in 12/18 (66.7%) of those on GMI-1271 and 5/10 (50%) on placebo. AEs were mostly mild events at the infusion site (i.e. tenderness or erythema), and occurred in both groups. One moderate AE occurred (vasovagal reaction) in the 2 mg/kg GMI-1271 cohort. Three AEs were possibly related to drug: 2 in the 2 mg/kg cohort (infusion site bruise, lightheaded), and 1 in the 5 mg/kg cohort (metallic taste). For Study 2, AEs were seen in 12/15 (80%) of those on GMI-1271/placebo and 3/3 (100%) on Lovenox. All AEs were mild. In GMI-1271/placebo group, all AEs were considered unrelated to study drug; 3 were infusion site bruise (1 per dose level). In the Lovenox group, AEs were bruise at injection site (related) and blood in urine (possibly related). PK: Plasma levels, Cmax, and AUC increased in a consistent dose-related manner in both studies (figure). CL, Vz, and t½ were not dose dependent; the latter averaged ~2.3 hours. ~66% of the dose was excreted unchanged in the urine independent of dose level, and CLr averaged 86 mL/min, less than estimated CrCl, suggesting tubular reabsorption is one component of CLr. Biomarkers: In Study 1, the absolute CD34+ and %CD34+ cell counts were normal across all cohorts. There was no dose-response for changes in peripheral CD34+ counts after administration of GMI-1271 (doses 2-10 mg/kg). Plasma concentrations of sEsel were relatively constant in the placebo and GMI-1271 2 mg/kg cohorts, with significant reduction (vs baseline, p=0.012) at 48hrs post-2mg/kg dose only. In the 5 and 10 mg/kg cohorts sEsel decreased significantly post-dose (p Conclusion: First-in-human experience with the potent E-selectin antagonist GMI-1271 demonstrated favorable safety and PK at single doses up to 20 mg/kg. These findings are consistent with the glycomimetic class for safety at anticipated therapeutic levels. Biomarkers of physiologic effect demonstrated clear reductions in sEsel/sPsel; decreased leukocyte adhesion with lower sICAM-1 levels; and no CD34+ rise indicating no mobilization of HSCs. Studies of GMI-1271 are ongoing to evaluate activity in hematologic malignancies and thrombosis. Figure 1. Plasma concentrations and Cmax are consistent for GMI-1271 between Phase 1 studies. Figure 1. Plasma concentrations and Cmax are consistent for GMI-1271 between Phase 1 studies. Disclosures Sood: Bayer: Research Funding. Hemmer:GlycoMimetics: Employment, Equity Ownership. Flanner:GlycoMimetics: Employment, Equity Ownership. Kramer:GlycoMimetics: Consultancy. Nietubicz:GlycoMimetics: Employment, Equity Ownership. Myers:GlycoMimetics: Research Funding. Wakefield:GlycoMimetics: Research Funding. Magnani:GlycoMimetics: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Thackray:GlycoMimetics: Employment, Equity Ownership.

Heparin anticoagulation during cardiovascular surgical procedures remains poorly investigated and understood. The objective of this investigation was to assess the effectiveness of three methods of heparin administration. Heparin sulfate (75 IU/kg) administered to patients undergoing aortoiliac surgery was randomised to one of three methods: Group I (n = 9) heparin was injected into a central venous line 5 minutes before infrarenal aortic clamping; Group II (n = 9) heparin was injected into the distal aneurysm immediately after infrarenal aortic clamping; and Group III (n = 8) heparin was injected into a central venous line immediately after infrarenal aortic clamping. Blood samples were analysed for anticoagulant activity from both the upper and lower extremities at 5, 15, 30, 60, and 120 minutes after heparin administration. Anticoagulation, as measured by aPTT, antifactor Xa levels, and ACT, was achieved in all three groups by 5 minutes, but initially with lower heparin activity (measured as antifactor Xa) in the upper extremity (Group II) and lower extremity (Group III), respectively. These differences were also evident in ACT and aPTT determinations. Intravenous heparin administration prior to aortic cross-clamping achieves excellent anticoagulation (anti-factor Xa approximately 1 U/ml) in both upper and lower extremities after 5 minutes. With regional administration, rapid heparin redistribution occurs, but it takes longer to achieve the same level of anticoagulation distant from the site of administration. Nevertheless, from a practical perspective the method of administration does not appear to have a great influence on the eventual achievement of adequate anticoagulation.

Reperfusion brain edema occurs infrequently after carotid endarterectomy and has been reported only ipsilateral to the side of surgery. We report a 51-year-old woman who presented with transient right arm weakness followed by left hemiparesis. Angiography showed occlusion of the left internal carotid artery, 90% stenosis of the right internal carotid artery, filling of the left anterior cerebral artery from the right carotid circulation only, and filling of the left middle cerebral artery branches by pial collaterals from the left anterior cerebral and posterior cerebral arteries. The patient had a right carotid endarterectomy and 1 day postoperatively developed a severe headache but had no new focal neurologic findings. Computed tomography showed effacement of sulci and scattered areas of high attenuation in the left parietal lobe, consistent with cerebral edema and petechial hemorrhage or vascular engorgement. Magnetic resonance imaging 2 days later did not show a lesion in the left hemisphere, suggesting resolution of the edema. This report suggests that reperfusion brain edema can occur contralateral to carotid endarterectomy.

Vessel wall endothelial damage initiates a local inflammatory response, which promotes a prothrombotic state driven by tissue factor, adhesion molecules, and pro-inflammatory cytokines. Understanding how natural inflammatory mechanisms promote a procoagulant state, may lead to the development of new pharmacological interventions targeted at thrombosis.

BackgroundThe anticoagulant treatment of acute deep venous thrombosis (DVT) has been historically directed toward the prevention of recurrent venous thromboembolism. However, such treatment imperfectly protects against late manifestations of the postthrombotic syndrome. By restoring venous patency and preserving valvular function, early thrombus removal strategies can potentially decrease postthrombotic morbidity.ObjectiveA committee of experts in venous disease was charged by the Society for Vascular Surgery and the American Venous Forum to develop evidence-based practice guidelines for early thrombus removal strategies, including catheter-directed pharmacologic thrombolysis, pharmacomechanical thrombolysis, and surgical thrombectomy.MethodsEvidence-based recommendations are based on a systematic review and meta-analysis of the relevant literature, supplemented when necessary by less rigorous data. Recommendations are made according to the Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology, incorporating the strength of the recommendation (strong: 1; weak: 2) and an evaluation of the level of the evidence (A to C).ResultsOn the basis of the best evidence currently available, we recommend against routine use of the term “proximal venous thrombosis” in favor of more precise characterization of thrombi as involving the iliofemoral or femoropopliteal venous segments (Grade 1A). We further suggest the use of early thrombus removal strategies in ambulatory patients with good functional capacity and a first episode of iliofemoral DVT of

Tissue factor (TF) is a potent initiator of the extrinsic coagulation cascade. The role and source of TF in venous thrombotic disease is not clearly defined. Our study objective was to identify the contribution of myeloid cell TF to venous thrombogenesis in mice.The mouse electrolytic inferior vena cava model was used to induce thrombosis. The following groups of mice were used (1) TF(flox/flox)LysMCre(+) mice that have reduced TF expression in myeloid cells, (2) TF(flox/flox)LysMCre(-) littermate controls, (3) Wild type mice given a monoclonal anti-mouse TF antibody (1H1) to inhibit TF activity, and (4) Wild type mice given rat IgG. Evaluations at baseline, day 2, and day 6 post thrombosis included thrombus weight, vein wall inflammatory cell migration, vein wall TF mRNA, and plasma D-dimer levels.Inhibition of TF significantly decreased thrombus weight 2days post venous thrombosis. In contrast, TF(flox/flox)LysMCre(+) had no change in thrombus weight when compared to littermate controls. The absence of myeloid cell TF did not affect infiltration of neutrophils or monocytes into the vein wall. TF mRNA expression in the vein wall decreased at 2days but then returned to baseline levels by 6days post thrombosis. D-dimer levels peaked at 2days post thrombosis in mice with or without myeloid cell TF.TF is important in the formation of venous thrombi in the macrovasculature. However, TF expression by myeloid cells does not significantly contribute to venous thrombogenesis in this model.

Animal models serve a vital role in deep venous thrombosis (DVT) research in order to study thrombus formation, thrombus resolution and to test potential therapeutic compounds (1). New compounds to be utilized in the treatment and prevention of DVT are currently being developed. The delivery of potential therapeutic antagonist compounds to an affected thrombosed vein has been problematic. In the context of therapeutic applications, a model that uses partial stasis and consistently generates thrombi within a major vein has been recently established. The Electrolytic Inferior vena cava Model (EIM) is mouse model of DVT that permits thrombus formation in the presence of continuous blood flow. This model allows therapeutic agents to be in contact with the thrombus in a dynamic fashion, and is more sensitive than other models of DVT (1). In addition, this thrombosis model closely simulates clinical situations of thrombus formation and is ideal to study venous endothelial cell activation, leukocyte migration, venous thrombogenesis, and to test therapeutic applications (1). The EIM model is technically simple, easily reproducible, creates consistent thrombi sizes and allows for a large sample (i.e. thrombus and vein wall) which is required for analytical purposes.

Venous thromboembolism (VTE) includes both deep vein thrombosis (DVT) and pulmonary embolism (PE). In the United States (U.S.), the high morbidity and mortality rates make VTE a serious health concern (1-2). After heart disease and stroke, VTE is the third most common vascular disease (3). In the U.S. alone, there is an estimated 900,000 people affected each year, with 300,000 deaths occurring annually (3). A reliable in vivo animal model to study the mechanisms of this disease is necessary. The advantages of using the mouse complete stasis model of inferior vena cava thrombosis are several. The mouse model allows for the administration of very small volumes of limited availability test agents, reducing costs dramatically. Most promising is the potential for mice with gene knockouts that allow specific inflammatory and coagulation factor functions to be delineated. Current molecular assays allow for the quantitation of vein wall, thrombus, whole blood, and plasma for assays. However, a major concern involving this model is the operative size constraints and the friability of the vessels. Also, due to the small IVC sample weight (mean 0.005 grams) it is necessary to increase animal numbers for accurate statistical analysis for tissue, thrombus, and blood assays such as real-time polymerase chain reaction (RT-PCR), western blot, enzyme-linked immunosorbent (ELISA), zymography, vein wall and thrombus cellular analysis, and whole blood and plasma assays (4-8). The major disadvantage with the stasis model is that the lack of blood flow inhibits the maximal effect of administered systemic therapeutic agents on the thrombus and vein wall.

Objectives: The relationship between CEAP class and Venous Clinical Severity Score (VCSS) change following great saphenous vein ablation (GSVa) is under reported. We further defined this relationship in patients including those with severe chronic venous disease (CVD) and the obese. Methods: We reviewed our prospectively collected venous procedural database from January 2007 to October 2010 for patients who had undergone GSVa ( phlebectomies). A CEAP class 5 or 6 designated severe CVD. VCSS were obtained preoperatively and at the 1st, 2nd, and 3rd follow up visits.

Objective: The combination of D-dimer and Wells score can exclude, but not confirm, the diagnosis of deep venous thrombosis (DVT). Since thrombosis and inflammation are interrelated, we evaluated the combination of soluble P-selectin (sPsel) with other inflammatory biomarkers for the diagnosis of DVT. Methods: Sixty-two positive and one hundred and sixteen patients with negative DVT, by duplex scan, were prospectively evaluated for sPsel, D-dimer, C-reactive protein (CRP), microparticles (MPs; total, leukocyte, and platelet-derived and tissue factor positive microparticles), and clinical Wells score. Results: Biomarkers and clinical scores that differentiated DVT positives from negatives were sPsel (87.3 vs 53.4 ng/mL, P < .0001), D-dimer (5.8 vs 2.1 mg/ L, P < .0001), CRP (2.1 vs 0.8 μg/mL, P < .0005), and Wells score (3.2 vs 2.0, P < .0001). For MP analysis, platelet-derived MPs were found to differentiate DVT from negatives. Using multivariable logistic regression, a combination of sPsel and Wells score could establish the diagnosis of DVT (cut point ≥90 ng/mL + Wells ≥2), with a specificity of 96% and positive predictive value (PPV) of 100%, and could exclude DVT diagnosis (cut point ≤60 ng/mL and Wells

Traditional therapeutic oral anticoagulation strategies often require invasive dosing or monitoring. Vitamin K antagonists (VKAs) have a large number of interactions, delayed onset requires frequent dose monitoring, and they have a small margin between therapeutic dose and bleeding complications. Novel oral anticoagulants, such as dabigatran, rivaroxaban and apixaban, are being developed to prevent those VKAs drawbacks. Besides oral bioavailability, those compounds are designed to require minimal to no monitoring and have a favourable safety profile. This review reports efficacy and safety data of these compounds throughout clinical development, as well as new approaches for oral pharmacological management of venous thromboembolism.

Diagnostic approaches for thromboembolism include Doppler probe examination, impedance plethysmography, phleborheography, radio-labeled fibrinogen scanning, magnetic resonance imaging, duplex imaging, and venography. The two primary diagnostic approaches for pulmonary embolism include ventilation/ perfusion lung scanning and pulmonary arteriography. Surgical options available for venous thromboembolism are generally limited to those condition in which limb viability is threatened from acute iliofemoral thrombosis. Vena caval interruption, performed in patients with a contraindication to anticoagulation, a complication during anticoagulant therapy, and recurrent pulmonary embolism in the presence of adequate anticoagulation (among other indications) is best accomplished with the Greenfield filter. Surgical approaches for pulmonary embolism, limited to patients with massive embolism with hypotension, include catheter pulmonary embolectomy and open pulmonary embolectomy.