Diagnosis and management of patients with the antiphospholipid syndrome.

The antiphospholipid syndrome is an autoimmune disorder characterized by venous or arterial thrombosis, recurrent pregnancy loss, and thrombocytopenia combined with laboratory tests that indicate the presence of antibodies against phospholipid-binding proteins. The antibodies are directed against a complex of phospholipid with a protein such as beta 2-glycoprotein I (beta 2-GPI) or prothrombin and are detected by means of phospholipid-dependent coagulation assays (known as assays for lupus anticoagulants) and by ELISAs that contain beta 2-GPI and a phospholipid (e.g., cardiolipin). The antiphospholipid syndrome can be associated with other connective tissue disorders such as systemic lupus erythematosus or may be the only manifestation of an autoimmune disorder. Management of patients with this disorder usually includes anticoagulation, which has been found to reduce the rate of recurrence of venous and arterial thrombosis as well as the rate of fetal loss.

Parvovirus B19: implications for transfusion medicine. Summary of a workshop.

This 1-day workshop showed that the infectivity of B19 DNA in donor blood and the neutralizing action of different antibodies present in the donated blood are not yet fully understood. It is possible that B19-induced anemia and reticulocytopenia are not being recognized in transfused recipients other than those in specific risk groups. The testing of blood components for any infectious agent is usually clinically driven, and, if B19 NAT were recommended at the present time in other than plasma products, a CMV-like model might prove appropriate; that is, virus screening would be performed on blood components destined for high-risk groups only. Currently, there is insufficient evidence to recommend universal testing, especially for single units. Workshop participants recommended that basic research continue in the scientific areas addressed. If clinical trials were to be developed, participants recommended that they include special risk groups such as seronegative pregnant women and children with malignancies who are receiving chemotherapy.

Fibrin sealant in preclinical and clinical studies.

Fibrin sealant, now commercially available in the United States, is a virally inactivated preparation of highly purified human fibrinogen and human thrombin that includes aprotinin to reduce fibrinolysis. Although the product is relatively expensive, cost can be justified when the sealant is used to produce localized hemostasis in surgery in which bleeding cannot be controlled by sutures. Fibrin sealant can also be justified as an alternative to factor concentrates in patients with coagulopathies who have a localized site of bleeding. Newer formulations of fibrinogen and thrombin in a freeze-dried form applied as a bandage may be useful in immediate, on-site treatment of trauma victims in either a civilian or military setting.

Hemostatic efficacy of fibrin sealant (human) on expanded poly-tetrafluoroethylene carotid patch angioplasty: a randomized clinical trial.

PURPOSE: The efficacy of solvent-detergent-treated fibrin sealant (human [FSH]) for controlling anastomotic bleeding from expanded polytetrafluoroethylene (ePTFE) patch angioplasty during carotid endarterectomy was evaluated, and FSH was compared with thrombin-soaked gelatin sponge (Gelfoam; TSG). METHODS: The study was of a randomized, open-label, single-site, single-treatment, parallel design that took place in a referral center with hospitalized patients. Forty-seven adult patients (33 men, 14 women) underwent elective carotid endarterectomy. Patients were randomized to receive either FSH (N = 24) or TSG (N = 23). FSH was obtained as an investigational new drug. FSH was applied as a liquid by means of a dual-syringe technique. Heparin anticoagulation, patch thickness, and suture type were standardized. Two different needle sizes were used (CV-6, PT-13: N = 21 [FSH: N = 10, TSG: N = 11]; CV-6, PT-9: N = 26 [FSH: N = 14, TSG: N = 13]). The FSH or TSG was applied to the ePTFE patch, and then blood flow was restored through the carotid artery. Degree of anticoagulation was assessed by anti-factor Xa activity. The time from restoration of carotid blood flow until achieving hemostasis was recorded. The blood loss from patch suture hole bleeding was measured. Completion intraoperative duplex ultrasound scanning was performed in all cases. Heparin was reversed with protamine sulfate. The primary end point was successful hemostasis within 15 minutes of restoration of carotid blood flow. The secondary end points were the amount of blood loss caused by suture line bleeding and the time to achieve hemostasis. RESULTS: There was no difference in the number of patients with complete hemostasis at 15 minutes (TSG, 13 of 23; FSH, 12 of 24; P =.77). The measured blood loss was 99.0 +/- 119.9 (SD) mL for TSG, and 105.0 +/- 107.9 mL for FSH (P =.86). The time to hemostasis was the same for both groups (TSG, 16.5 +/- 16.5 minutes; FSH, 16.6 +/- 14.2 minutes; P =.97). Within both treatment groups, the use of larger needles (PT-13) was associated with greater blood loss (FSH, 169.7 +/- 124.2 mL; TSG, 172.7 +/- 151.5 mL) than was the use of smaller needles (PT-9; FSH, 58.8 +/- 66.3 mL; TSG, 34.1 +/- 25.6 mL; P =.036, P =.001, respectively). There were no postoperative strokes or bleeding complications in either group. No abnormalities were shown in either group by means of completion carotid duplex ultrasound scanning. CONCLUSION: FSH was equivalent, but not superior to, TSG in achieving hemostasis during carotid endarterectomy performed with ePTFE patch angioplasty. Adhesion properties of FSH to ePTFE are possibly different than those to native tissue and warrant additional investigation.

How to improve transfusion medicine. A treating physician's perspective.

As transfusion medicine becomes more complex, cooperative strategies are gaining increasing importance in relaying information to the treating physician and in incorporating the treating physician into the education and quality control processes. The broad domain of transfusion medicine is illustrated by the variety of disciplines involved in defining the use of products such as fresh frozen plasma and the newly released solvent-detergent-treated plasma, fibrin glue and highly purified fibrin sealant, and leukoreduced and irradiated blood products. Cooperative efforts among physicians and other personnel of multiple disciplines are essential to ensure appropriate use and continuous evaluation of blood products.

Near-site monitoring of the antiplatelet drug abciximab using the Hemodyne analyzer and modified thrombelastograph.

OBJECTIVE: This investigation examines the hypothesis that the antiplatelet effect of abciximab and its reversal can be monitored using the Hemodyne (Hemodyne, Inc, Midlothian, VA) analyzer and modified Thrombelastograph (Haemoscope, Skokie, IL). DESIGN: In vitro dose-response and reversal study. SETTING: Anesthesia Research (Dallas, TX) and Special Studies Coagulation Laboratories (Washington, DC). PARTICIPANTS: Nine healthy volunteers. INTERVENTIONS: The addition of increasing concentrations of abciximab, 0 to 10 microg/mL, and purified fibrinogen, 50 to 400 mg/dL. The reversal of abciximab, 4 microg/mL, with the addition of fresh platelet-rich plasma (PRP) sufficient to increase the platelet concentration by approximately 10%. MEASUREMENTS AND MAIN RESULTS: Platelet aggregation and platelet contractile force using the Hemodyne analyzer were used as platelet-specific measurements. The Thrombelastograph maximum amplitude (MA) for platelets (MA(PLT)) was calculated by subtracting the MA from a platelet-poor plasma (PPP) sample (MA(ppp)) determined in one thromboelastography well from that of whole-blood MA (MA(WB)) run simultaneously in the second thromboelastography well. The addition of abciximab, 0 to 10 microg/mL, resulted in significant concentration-dependent reductions in platelet aggregation (p < 0.001), platelet contractile force (p < 0.001), and MA(PLT) (p < 0.001). Platelet contractile force (p < 0.03) and MA(PLT) (p < 0.05) were significantly more responsive than MA(WB) to the effect of abciximab, 4 microg/mL, and its reversal with the addition of fresh PRP. Purified fibrinogen concentration directly correlated with thromboelastography MA (r(s) = 0.97; p < 0.001), yet had no effect on platelet contractile force. The addition of abciximab had no measurable influence on the MA(ppp). CONCLUSION: This in vitro study suggests that the Hemodyne analyzer and modified Thrombelastograph might be clinically useful methods to monitor the platelet inhibitory effects of agents such as abciximab.

Physiologic responses of cross-linked hemoglobin in endotoxin-treated rats.

Purified human cross-linked hemoglobin (alpha alpha Hb) as well as recombinant human hemoglobin is undergoing clinical trials in the setting of acute blood loss and perioperative hemodilution. We have previously demonstrated that in rabbits with circulating plasma Hb, such as alpha alpha Hb, infusion of endotoxin (LPS) impairs myocardial contractility which results in hypotension, tissue hypoperfusion and increased mortality. The untoward cardiovascular effects occurring after the combined infusion of LPS and alpha alpha Hb in this model are similar to those reported for other agents that inhibit nitric oxide (NO) availability. To determine if the deleterious effects of alpha alpha Hb and LPS were species specific, we performed similar studies in rats. Anesthetized Sprague-Dawley rats received LPS (4 mg/kg or 40 mg/kg) alone or in combination with alpha alpha Hb (0.7 g/kg). Mean arterial blood pressures (MAP) increased in the group that received alpha alpha Hb alone (105 +/- 8 to 120 +/- 7 mm Hg, p = 0.2) and a decrease was noted in the groups that received low dose LPS (4 mg/kg, p = 0.5) and high dose LPS (40 mg/kg, p = 0.016). MAP in rats treated with the LPS at either dose combined with alpha alpha Hb remained unchanged. Levels of urine nitrite, which was measured as a surrogate marker for plasma NO, were significantly decreased at 2 hr in groups that received the combination of alpha alpha Hb and LPS at 4 mg/kg (p = 0.022) and 40 mg/kg (p = 0.003). No significant decrease was observed in animals treated only with alpha alpha Hb (p = 0.21) or LPS (4 mg/kg; p = 0.78 and 40 mg/kg; p = 0.65). Survival was evaluated during 72 hr in animals that were infused with high dose LPS (40 mg/kg) alone or in combination with alpha alpha Hb and then allowed to recover. The survival of rats treated with LPS alone or the combination was 29% at the end of 24 hr and was 100% for rats receiving only alpha alpha Hb. The data suggest that the toxicity of alpha alpha Hb appears to be a species specific phenomenon.

Women with thrombophilia: assessing the risks for thrombosis with oral contraceptives or hormone replacement therapy.

Women with inherited or acquired thrombophilia are at increased risk for venous thromboembolism (VTE) when they use oral contraceptives (OCs) of either the second or third generation. For women who are heterozygous for Factor V Leiden, the risk is probably 28 to 50 of 10,000 women-years compared to 2 to 5 of 10,000 years for those not known to have thrombophilia. The thrombotic risk is highest during the first year that OCs are used. Whether women with thrombophilia are at increased risk for VTE when they use hormone replacement therapy (HRT) has not been assessed in any study. For women without thrombophilia, the risk for VTE associated with HRT is probably 2 to 3 of 10,000 years. The benefits of HRT include reduced risk for myocardial infarction and Alzheimer's disease, and increased bone density. The physiological changes induced by HRT are not the same as those induced by OCs. Small studies have suggested that for women who have additional risks of thrombosis (i.e., perioperative setting, underlying systemic lupus erythematosus), HRT does not confer the same increased risk of thrombosis, as does the use of OCs. Until data are available to address the magnitude of any increase in thrombotic risk induced by HRT for women with thrombophilia, physicians probably serve their patients best by providing information about the benefits of HRT, emphasizing that the risk of VTE is unknown, and encouraging patients to take an active role in decisions about their healthcare.

Hemostatic efficacy of a fibrin sealant dressing in an animal model of kidney injury.

OBJECTIVE: The purpose of this study is to determine the hemostatic efficacy of a fibrin sealant dressing compared with a standard collagen control dressing in an animal model of kidney injury. METHODS: Twenty adult male Sprague-Dawley rats were administered general anesthesia and underwent partial nephrectomy with heparin anticoagulation (300 U/kg intravenous). Treatment of the cut surface of the kidney was randomized to three groups: group I, no hemostatic agent; group II, collagen dressing; and group III, fibrin sealant dressing. RESULTS: Blood loss was significantly less in group III (3.39+/-0.63 mL) than in group I (8.64+/-2.26 mL) and group II (8.63+/-1.72 mL; p < 0.001). The percentage decrease in the mean arterial pressure was significantly less in group III (34.09+/-15.58%) than in group I (59.66+/-16.19%) and group II (60.35+/-15.66%; p=0.015). CONCLUSION: Fibrin sealant dressings provide effective hemostasis and are superior to collagen dressings in an animal model of kidney injury. Additional development of fibrin sealant dressings for potential clinical use is warranted.

The incidence of heparin-induced antibodies in patients undergoing vascular surgery: a prospective study.

PURPOSE: This study prospectively assessed the incidence of heparin-induced antibodies in patients undergoing peripheral vascular surgery and determined whether the incidence is influenced by previous heparin exposure. METHODS: Fifty-four hospitalized patients (36 men and 18 women) undergoing peripheral vascular surgery and receiving intraoperative heparin anticoagulation were studied. Unfractionated porcine heparin was given for intraoperative anticoagulation and was not continued postoperatively. Carotid endarterectomy was performed in 36 patients, aortic reconstruction in 11 patients, and infrainguinal bypass in 7 patients. Plasma was tested before and after (14 +/- 7.5 [SD] days) surgery for IgG antibodies to the complex of heparin/platelet factor 4, using a standardized, validated enzyme-linked immunosorbent assay (ELISA). Results are expressed as an optical density ratio (ODR) of patient plasma to normal plasma, with the threshold for a positive result of > or = 1.8. Platelet counts and clinical outcomes were also assessed. RESULTS: The mean patient age was 67.2 +/- 9.7 years. A prior exposure to heparin was documented in 41% of patients. The mean intraoperative heparin dose was 9089 +/- 3607 units. Only 1 patient converted from a negative antibody status to a positive status (1.9%, 95% CI = 0.10%-11.18%). The change in the ELISA ODR after surgery was not significantly different for patients with (+0.042 +/- 0.272) and without (-0.022 +/- 0.299, P = 0.57) prior heparin exposure. Postoperatively, the platelet counts dropped from 227,620 +/- 78,308 microL, to 185,706 +/- 80,842 microL (P < .001). The decrease in platelet count was the same in patients with prior heparin exposure (-23.0 +/- 18.0%) and without (-18.0 +/- 14.0%, P = .46). One thrombotic complication occurred, a femorotibial bypass graft occlusion in a patient who tested negative for antibodies. CONCLUSION: Heparin-induced antibodies occur infrequently after peripheral vascular surgery. The commonly observed, mild degree of postoperative thrombocytopenia does not appear to be caused by heparin-induced antibodies. These results indicate that a standard dose of heparin for intraoperative anticoagulation during vascular surgery is not associated with a significant risk of heparin-induced thrombocytopenia and thrombosis.

Update on recognition and management of patients with acquired or inherited hypercoagulability.

As many as 20% of individuals who develop venous thromboembolism (VTE) in an outpatient setting may have a mutation in factor V (factor V Leiden) or the prothrombin gene. Through testing for the inherited and acquired conditions for VTE, the clinician can develop strategies to prevent recurrences.

A method for the quantitative assessment of platelet-induced clot retraction and clot strength in fresh and stored platelets.

BACKGROUND AND OBJECTIVES: The changes that occur in platelets as they undergo storage have been documented by aggregometry as well as by flow cytometry. However, one of the most essential platelet functions, the induction of clot retraction, has not been quantitatively assessed in stored platelets. We describe two potentially useful methods, platelet-induced clot retraction and clot strength, to assess effect of storage of platelets in blood banks or of platelet preparations subjected to freezing or freeze-drying. These methods have previously been developed for bedside monitoring of patients receiving c7E3 (Reopro(R)). MATERIALS AND METHODS: Platelet-induced clot retraction (PICR) and clot strength were measured with the Hemodyne and Thromboelastograph, respectively. Paired Study: Fresh platelet concentrates (n = 3) were obtained from leukapheresis donors and divided into two equal units; one unit was tested within 4 h of collection and the other stored for 5 days at 22 degrees C in a platelet incubator and tested. Unpaired Study: Fresh platelet concentrates (n = 15) were obtained from leukapheresis donors and tested within 4 h of collection and compared to outdated platelets (n = 30; random or single donor) that had been stored for 5 days at 22 degrees C in a platelet incubator. Alternative Preservation Methods: Lyophilized platelets, platelets chilled to 4 degrees C, platelets frozen at -70 degrees C in 5% dimethyl sulfoxide (DMSO) or in the absence of a cryoprotectant. RESULTS: Paired Study: Stored platelets demonstrated an increase in PICR; the difference was not significant (p = 0.55). There was no difference in clot strength between fresh and outdated platelets (p = 0.90). Unpaired Study: When compared to fresh platelets, stored platelets demonstrated a 2-fold higher PICR (p = 0.0011). On the other hand, there was no difference in the time to onset of PICR (p = 0.08) and there was no difference in clot strength between fresh and outdated platelets (p = 0.14). Alternate Preservation Methods: In contrast, PICR and clot strength were reduced in platelets frozen at -70 degrees C in 5% DMSO and absent in lyophilized platelets, in platelets frozen at -70 degrees C in the absence of cryoprotectants or stored at 4 degrees C. CONCLUSION: The data indicate that the ability of platelets to induce clot retraction and to enhance clot strength is not altered by storage, despite functional abnormalities in aggregation and agglutination. These data suggest that quantitative measurements of PICR and clot strength may be simple, useful tools for assessing the function of stored platelet concentrates, platelets that have undergone freezing or exposure to alternative buffers and for evaluating platelet functions relevant to PICR.

Cardiovascular toxicity of human cross-linked hemoglobin in a rabbit endotoxemia model.

OBJECTIVE: To determine the possible adverse effects of human cross-linked hemoglobin in endotoxemia. DESIGN: Prospective, controlled, laboratory trial. SETTING: Animal research laboratory. SUBJECTS: New Zealand white rabbits. INTERVENTIONS: Conscious rabbits received intravenous infusions of either lipopolysaccharide (LPS) alone (10 micrograms/kg, Escherichia coli 0111:B4), human hemoglobin cross-linked between the alpha chains (alpha alpha Hb, 0.7 g/kg), or both LPS and alpha alpha Hb. The cardiovascular effects of alpha alpha Hb and LPS as single agents or administered together were then studied in anesthetized rabbits. MEASUREMENTS AND MAIN RESULTS: Mortality in conscious animals that received alpha alpha Hb followed by LPS 4 hrs later (n = 5), or LPS and alpha alpha Hb at the same time (n = 6) was 60% and 67%, respectively. In anesthetized animals, infusion of both LPS and alpha alpha Hb (n = 6) resulted in hypoxia, lactic acidosis, ventricular arrhythmias, and decreased myocardial contractility and left ventricular pressure. In contrast, anesthetized rabbits that received alpha alpha Hb (n = 5) or LPS (n = 5) alone did not develop hypoxia, acidosis, alteration in myocardial contractility, or arrhythmias. Furthermore, death did not occur in any of the conscious animals that received either LPS (n = 7) or alpha alpha Hb (n = 4) as single agents. CONCLUSIONS: In an animal model of nonlethal endotoxemia, infusion of alpha alpha Hb significantly increases mortality. Our data suggest that mortality may be due to the acute increased cardiopulmonary toxicity of alpha alpha Hb in animals with underlying endotoxemia.

Hemostatic efficacy of a fibrin sealant-based topical agent in a femoral artery injury model: a randomized, blinded, placebo-controlled study.

PURPOSE: The efficacy of currently available topical hemostatic agents requires the formation of fibrin generated from circulating blood. Fibrin sealant, which is prepared from high concentrations of thrombin and fibrinogen, has been used in liquid form to promote hemostasis during vascular surgery. In a blinded, randomized, placebo-controlled fashion, we evaluated a dry dressing of purified, viral-inactivated human fibrinogen and human thrombin in a large animal model of arterial injury. METHODS: Dressings were prepared by application of a layer of lyophilized human fibrin sealant or immunoglobulin G (IgG, control) to a silicone backing material. Six anesthetized female Yorkshire pigs (16 to 27 kg) received bilateral, 4 mm longitudinal femoral arteriotomies after surgical exposure of the arteries. The arteriotomies were not closed. In each animal a fibrin sealant dressing was applied to one artery and a control dressing to the other. Each dressing was secured on the arteriotomy by a mechanical device. After application of the dressings, blood flow was restored to each limb for 1 hour. The compressive device was released for 5 seconds at intervals of 15 minutes to assess hemostasis. Blood flow was measured distal to each arteriotomy with a dual-channel flowmeter to adjust equal bilateral compression. RESULTS: Blood loss (mean +/- SEM) was significantly less from the arteriotomy treated with the fibrin-based dressing compared with the control dressing (4.9 +/- 4.0 ml versus 82.3 +/- 11.1 ml; p = 0.0005). Complete hemostasis was achieved at the first 15-minute interval in five of six arteriotomies treated with fibrin sealant and in none of the six control arteriotomies during 1 hour of assessment (p = 0.03). Blood flow through each femoral artery at baseline was the same in both treatment and control arteries (fibrin sealant, 114.2 +/- 17.4 ml/min; control, 106.7 +/- 16.5 ml/min; p = 0.24) and was not significantly different throughout the experiment. CONCLUSIONS: Fibrin-based dressings provide effective hemostasis in a large animal model of arterial injury. Further development of these dressings will address optimal formulation and configuration for clinical use. Our results suggest that fibrin-based dressings will be effective in promotion of hemostasis in arterial bleeding, without compromising blood flow.

Diagnosis of heparin-induced thrombocytopenia in the vascular surgery patient.

BACKGROUND: Recent studies have shown that patients with heparin-induced thrombocytopenia (HIT) form immunoglobulin G (IgG) and/or IgM antibodies directed against a complex of platelet factor 4 (PF4) and heparin. This recognition has resulted in the development of enzyme-linked immunosorbent assays (ELISAs) that use the heparin/PF4 complex as the antigen. This study describes the use of a standardized ELISA to assess antibody formation in five patients suspected of having HIT. METHODS: Five patients received heparin for treatment of arterial or venous thrombotic disorders. All patients had the ELISA performed to detect IgG or IgM antibodies directed against heparin-PF4, as well as the 14C serotonin release assay, when HIT was clinically suspected. RESULTS: HIT was diagnosed in four patients and ruled out in a fifth by using the ELISA. All patients had a 40% decrease in platelet count that returned to normal after heparin cessation. Only one of the four patients who tested positive by ELISA for IgG antibodies also tested positive by the 14C serotonin release assay. Treatment was significantly altered by the ELISA results in all five patients. CONCLUSIONS: It is likely that the ELISA is more sensitive in the diagnosis of HIT than the more traditional aggregation tests, and it may emerge as a new gold standard. Prospective studies in which serial laboratory testing is combined with measurement of clinical outcomes are needed and will eventually provide a greater understanding of the full spectrum of HIT and the clinical settings that precipitate thrombosis in the vascular surgery patient.