Karnicki, Krzysztof

Infection increases the risk of thrombotic disease. Platelets contribute to defense against infection and to thrombosis. Gram negative bacteria produce lipopolysaccharides (LPS) that activate Toll‐like‐receptor‐4 (TLR4) in the host and initiate inflammatory responses to infection. Platelets express TLR4, but whether or how LPS affects platelet functions that contribute to thrombotic disease is not understood. To determine if TLR4 signaling influences platelet functions that contribute to thrombotic disease, we studied responses of platelets from normal mice (C57BL/10SnJ; n=10) and mice that do not express TLR4 (C57BL/10ScN; dTLR4, n=10). Prior to LPS exposure, the number of circulating platelets, platelet RNA content and expression of P‐selectin following stimulation with thrombin were diminished in mice that lack TLR4 compared to wild type mice, while platelet aggregation and secretion of ATP were unchanged. These results suggest that loss of TLR4 function affects platelet production and membrane binding characteristics of the circulating platelet pool. One week following a single intravenous injection of LPS (200μg/kg/iv), the time it takes for the circulating platelet pool to turnover, both wild‐type and dTLR4 mice had similar numbers of circulating platelets, and thrombin‐induced expression of P‐selectin and platelet aggregation to collagen increased comparably in both groups of mice. These results indicate that TLR4 function is required for normal platelet homeostasis. Exposure to LPS stimulates a TLR4‐independent shift of the platelet pool toward a reactive phenotype, which could contribute to increased thrombotic risk during or following infection.

Abstract Renal vein thrombosis (RVT), although uncommon, is associated with considerable morbidity and mortality. Neither the etiology nor the natural history of RVT has been adequately described in a large case series. Objective: To determine whether incident RVT is associated with an increased likelihood of recurrent local and systemic venous thromboembolic disease. Methods: Cases were identified by reviewing the Diagnostic Index for all patients with RVT at our institution between 1981 and 2000. Results: The study group consisted of 240 patients (34% women) with a mean age 55± 19 years. The diagnosis of RVT was made by CT (35%), Duplex Ultrasound (35%), MRI±gadolinium (10%) and either at surgery or autopsy in the remainder. The underlying etiology was deemed to be malignancy in 65% of cases. Of these, renal cell carcinoma (74%) and Wilms tumor (3%) made up the majority. Nephrotic syndrome primarily membranous glomerulonephritis was the second most common underlying etiology (23%). Of those tested, a definable thrombophilia was found in only 13 cases including lupus anticoagulant/antiphospholipid antibody (n=4); deficiencies of protein C or S, (n=2 each); heparin induced thrombocytopenia (n=2); antithrombin deficiency, dysfibrinogenemia and hyperhomocysteinemia (n=1 each). Eighty-one patients (34%) were treated with warfarin. Over a mean follow up of 3 years, the rate of recurrence of venous thrombosis at any site was 5%. The mortality rate over this time period was 49%. Conclusion: RVT represents a distinct clinical entity relative to venous thromboembolism of the lower extremity. The observed low rate of recurrence may be explained by the high mortality rate owing to a high percentage of renal malignancy. RVT should prompt an evaluation for an underlying renal malignancy.

Objective— The clinical use of venous stents is increasing dramatically. Although antiplatelet agents are required for arterial stent patency, optimal thrombo-prophylaxis after venous stenting remains undefined. To address this issue, PD0348292, a direct Factor Xa inhibitor, was compared with antiplatelet therapy in a porcine venous stent model. Methods and Results— Four hours before stent deployment, pigs (n=5 to 6 per group) received oral PD0348292 at 0.4, 0.9, 4.3 mg/kg, or 0.4 mg/kg plus aspirin (325 mg). Aspirin, clopidogrel (75 mg), aspirin plus clopidogrel, or vehicle (n=10) were administered daily for 2 days before the procedure. Two hours after stent placement, thrombi were quantified by autologous 111 In-platelet content and weights. Thrombus weight and platelet deposition were significantly reduced by PD0348292 at 0.4 (49±79 mg and 110±145×10 6 /cm 2 ), 0.9 (5±6 mg and 107±128×10 6 /cm 2 ), 4.3 mg/kg (0±0 mg and 87±125×10 6 /cm 2 ), and PD348292 plus aspirin (20±40 mg and 157±70×10 6 /cm 2 ) compared with vehicle (402±226 mg; 584±454×10 6 /cm 2 ). Despite prolonging bleeding times and inhibiting platelet aggregation, neither aspirin (567±683 mg and 533±622×10 6 /cm 2 ), clopidogrel (404±349 mg and 178±101×10 6 /cm 2 ), nor aspirin plus clopidogrel (247±261 mg and 231±266×10 6 /cm 2 ) significantly decreased stent thrombosis. Conclusions— PD0348292 completely inhibited thrombosis after venous stenting. Platelet accretion in these venous thrombi appear to involve pathways distinct from arachidonate metabolism or ADP P 2 Y 12 receptor activation.

Widespread applications of totally laparoscopic aortic reconstructions have been limited by the long cross-clamp time required to suture the aortic anastomosis despite improvement in instrumentation. The authors' hypothesis was that a “one-step anastomosis concept” using an intraluminal stapler would allow shorter cross-clamp time but similar patency and imperviousness as videoscopic suturing techniques. An intraluminal stapler (Endopath-ILS, Ethicon) with a modified anvil was used to perform videoscopic-assisted thoracic aorta-to-iliac artery bypass with a 21 mm by 8 mm polytetrafluoroethylene (PTFE) graft in 22 sheep through a minimally invasive approach using a 5 cm thoracotomy. The graft-to-iliac artery anastomoses were hand sutured through a flank incision. Twelve sheep were used to establish the technique and 10 subsequent animals constituted the study group. Aortic cross-clamp time, imperviousness, and need for additional sutures were recorded and compared to previously reported data using videoscopic suturing in pigs. Patency was assessed by comparing lower limb arterial pressures. Macroscopic and microscopic examinations of the anastomoses were performed at different time-points within the first 3 months. Videoscopic-assisted stapled anastomoses were also performed on atherosclerotic aortas of 3 human cadavers. Stapled anastomoses between the thoracic aorta and PTFE graft were completed in 8 of 10 animals. Two animals were euthanized after stapler failure and anastomotic bleeding. Sutures to strengthen the anastomosis had to be used in 4 cases. Mean aortic cross-clamp time in 8 successful cases was 4.3 ±2.9 minutes (range 2–11 minutes) and was significantly shorter than clamp time of videoscopic suturing technique (48.7 ±9.4 minutes, p<0.0001). Imperviousness was good or excellent in 4 animals and fair in 4 animals. All anastomoses were patent at the end of the procedure. Examination of the anastomosis of the 2 failed interventions showed medial aortic tear surrounding the anastomosis in 1 case and misfired staples in the other. No graft occlusion was noted during follow-up ranging from 0 to 12 weeks. At the time of harvest, no bleeding was noted after epinephrine and volume infusion to increase mean arterial pressure to 200 mm Hg for 15 minutes. Macroscopic examination of the anastomoses revealed adequate healing with circumferential stapling of the prosthesis to the aortic wall and no stenosis or thrombus except in 1 false aneurysm (1/7, 14%). Surface electron microscopy showed cells coverage of the anastomosis surface. When applied on human cadaver thoracic and abdominal aorta with atherosclerotic changes, clamping times of less than 5 minutes were achieved. However, imperviousness tested with saline was poor. An automatic stapling device allows performance of a graft-to-aorta anastomosis through a minimally invasive approach with shorter clamping time than a videoscopic suturing technique. However, the current technique of aortic stapling is unreliable and further improvements are needed.

Objective— Occurrence of arterial thrombosis secondary to vascular disease in an individual is not easily predicted. After establishing that this poor predictability arises at least in part from an intrinsic thrombosis propensity of the individual, we sought to determine whether the propensity for arterial thrombosis is governed by blood or arterial wall factors. Methods and Results— To evaluate the variability arising from the blood, autologous 111 In-labeled platelet deposition was measured after high-shear perfusion of compressed aortic strips, prepared from a single pig, with heparinized blood from 25 pigs. To evaluate the variability arising from the vessel wall, aortic strips from 8 pigs were superfused with blood from a single animal. Blood samples from 25 animals superfused over aortic substrate from a single source yielded a 24-fold range of platelet deposition. In contrast, when aortic substrates from 8 different animals were superfused with blood from a single animal, platelet deposition spanned a 3-fold range. Platelet deposition was significantly correlated with whole-blood lymphocyte counts and with platelet counts . Conclusions— Individual propensity for arterial thrombosis in pigs is more greatly influenced by blood components than by elements within the arterial wall.

Atherosclerosis manifests as a systemic disease with near global involvement of the named segments of the arterial tree. Acute thrombotic arterial occlusion, however, is not equally distributed. To evaluate intra-individual regional differences in arterial thrombogenicity, we compared 111 In-platelet deposition in porcine carotid and femoral arteries after a standardized crush injury. Within the unidirectional flow conditions of elastic carotid arteries, platelet deposition was more than 3-fold higher compared with predominantly muscular femoral arteries with triphasic arterial flow. To determine the influence of rheology on platelet deposition after crush injury, carotid arteries were transplanted into the femoral position and compared with the paired native carotid and femoral arteries. Similarly, femoral arteries transposed to the carotid position were compared with the paired native carotid artery. In each of these experiments, arterial transposition to a new anatomic location imparts a predilection for platelet deposition indigenous to the new location. In the controlled environment of two high-shear thrombin–independent and –dependent flow chambers, porcine carotid and femoral arterial substrates were indistinguishable from one another with respect to platelet deposition. Regional differences in arterial hemodynamics may account for substantial differences in thrombosis arising from deep arterial injury.