Coagulation Factor Activity Levels Research Articles

Variation in coagulation factor activity levels cause discrepancies between activated partial thromboplastin time and anti-Xa activity for heparin monitoring: a retrospective observational study

BackgroundUnfractionated heparin (UFH) is primarily monitored using activated partial thromboplastin time (APTT). However, the recent introduction of anti-activated factor X (anti-Xa) activity testing has provided a direct evaluation of Xa inhibition by anticoagulants. This study aimed to investigate discrepancies between APTT and anti-Xa activity during UFH monitoring in critically ill patients and explore their underlying causes.MethodsThis study analyzed 271 pairs of laboratory test results from blood samples of 99 critically ill patients receiving continuous intravenous UFH. Theoretical APTT values were calculated using fitted curve equations from spiked sample measurements with anti-Xa activity. Samples were categorized into three groups based on the measurement of the APTT/theoretical APTT ratio: the lower group (< 80%), the concordant group (80–120%), and the upper group (> 120%).ResultsThe overall concordance rate between APTT and anti-Xa activity was 45%, with a 55% discrepancy rate. The lower group frequently showed apparent heparin overdoses, while coagulation factor activities in the lower and upper groups were higher and lower, respectively, than those in the concordant group. Particularly, the lower group exhibited higher factor VIII activity levels than the upper and concordant groups.ConclusionsDiscrepancies between APTT and anti-Xa activity were frequently observed, influenced by changes in coagulation factors activity levels. The lower and upper groups were classified as pseudo-heparin-resistant and coagulopathy types, respectively. Accurate monitoring of heparin in critically ill patients is crucial, especially in cases of pseudo-heparin resistance, where APTT values may wrongly indicate inadequate heparin dosing despite sufficient anti-Xa activity. Understanding these discrepancies is important for managing heparin therapy in critically ill patients.Trial registration: Not applicable.

Potential utility of a multi-component coagulation factor panel to calculate MELD scores and assess the risk of portal vein thrombosis in chronic liver disease

BackgroundCurrent quantitative approaches to assess chronic liver disease (CLD) severity have limitations. Further, portal vein thrombosis (PVT) pre-liver transplant (LT) is a major contributor to morbidity in CLD; the means of detecting and/or predicting PVT are limited. We sought to explore whether plasma coagulation factor activity levels can serve as a substitute for prothrombin time/international normalized ratio (PT/INR) in the Model for End-stage Liver Disease (MELD), and/or help assess the risk of PVT.MethodsPlasma activity levels of Factor V (FV), Factor VIII (FVIII), Protein C (PC), and Protein S (PS) and the concentrations of D-dimer, sP-selectin, and asTF were assessed in two cohorts of CLD patients (ambulatory, n = 42; LT, n = 43).ResultsFV and PC activity levels strongly correlated with MELD scores, which enabled the development of a novel scoring system based on multiple linear regressions of the correlations of FV and PC activity with MELD-Na that substitutes PT/INR. Six-month and 1-year follow-up revealed that our novel approach was non-inferior to MELD-Na at predicting mortality. A significant inverse correlation between FVIII activity levels and PVT was found in the LT cohort (p = 0.010); FV and PS activity levels were in-trend (p = 0.069, p = 0.064). We developed a logistic regression-based compensation score to identify patients at risk of PVT.ConclusionsWe demonstrate that FV and PC activity levels may be used to replace PT/INR in MELD scoring. We also show the potential of using the combination of FV, FVIII, and PS activity levels to assess the risk of PVT in CLD.

P1658: DELAYED REVERSAL OF FACTOR V INHIBITOR ACTIVITY POST-IMMUNOSUPPRESSIVE THERAPY, FOLLOWED BY A SUCCESSFUL PREGNANCY: A CASE REPORT

Background: Acquired factor V inhibitor (AFVI) is a very rare autoimmune hemostatic disorder that leads to hemorrhagic manifestations of variable severity. Aims: We describe a case of a 35-year-old Caucasian female patient presenting with severe AFVI with life-threatening hemorrhages. The AFVI exhibited spontaneous reversal over several years after discontinuation of ineffective combination immunosuppressive therapies and subsequently a successful pregnancy. Methods: Diagnosis of AFVI was based on a marked prolongation of prothrombin time-international normalized ratio (INR) and activated partial thromboplastin time (APTI) not corrected by the addition of normal plasma (1:1 ratio), with FV activity reduced to <1.0%. Other coagulation factor activity levels were unchanged, and the presence of an FV inhibitor was revealed by the Bethesda inhibitor assay. Results: Over the course of 2.5 years, plasmapheresis plus immunoglobulins and different combination immunosuppressive regimens were adminstered (dexamethasone+rituximab; cyclophosphamide+dexamethasone + rituximab + cyclosporine; cyclosporine + sirolimus + cyclophosphamide + dexamethasone; bortezomib+sirolimus+methylprednisolone; sirolimus+mycophenolate mofetil). Coagulation could be effectively maintained using recombinant FVIIa. The combination immunosuppressive therapies resulted in intermittent reversal of AFVI over 2.5 years, which then became therapy-resistant. After discontinuation of the unsuccessful immunosuppressive therapies, the patient exhibited partial spontaneous remission: her FV activity gradually increased from ~1% to ~33% over the course of 5 years. The patient became pregnant and during her pregnancy, FV activity increased to 54%, and her coagulation parameters were normalized. She gave birth to a healthy child by Caesarean section without any bleeding complications. Summary/Conclusion: Patients with AFVI presenting with hemorrhagic manifestations should be treated aiming for hemostatic control as well as inhibitor eradication. The use of an activated bypassing agent for hemostatic control is effective in AFVI. The presented case is unique because the treatment regimens used included combinations of multiple immunosuppressive agents and also suggest that patients with AFVI resistant to combination immunosuppressive protocols receiving no therapy may undergo spontaneous remission. Long-term monitoring of the coagulation status and the anti-FV inhibitor levels is warranted.

Coagulation factor XII, XI, and VIII activity levels and secondary events after first ischemic stroke

BackgroundThough risk for recurrent vascular events is high following ischemic stroke, little knowledge about risk factors for secondary events post‐stroke exists. ObjectivesCoagulation factors XII, XI, and VIII (FXII, FXI, and FVIII) have been implicated in first thrombotic events, and our aim was to estimate their effects on vascular outcomes within 3 years after first stroke. Patients/MethodsIn the Prospective Cohort with Incident Stroke Berlin (PROSCIS‐B) study, we followed participants aged 18 and older for 3 years after first mild to moderate ischemic stroke event or until occurrence of recurrent stroke, myocardial infarction, or all‐cause mortality. We compared high coagulation factor activity levels to normal and low levels and also analyzed activities as continuous variables. We used Cox proportional hazards models adjusted for age, sex, and cardiovascular risk factors to estimate hazard ratios (HRs) for the combined endpoint. ResultsIn total, 94 events occurred in 576 included participants, resulting in an absolute rate of 6.6 events per 100 person‐years. After confounding adjustment, high FVIII activity showed the strongest relationship with the combined endpoint (HR = 2.05, 95% confidence interval [CI] 1.28–3.29). High FXI activity was also associated with a higher hazard (HR = 1.80, 95% CI 1.09–2.98), though high FXII activity was not (HR = 0.86, 95% CI 0.49–1.51). Continuous analyses yielded similar results. ConclusionsIn our study of mild to moderate ischemic stroke patients, high activity levels of FXI and FVIII but not FXII were associated with worse vascular outcomes in the 3‐year period after first ischemic stroke.

A gottingen minipig model of radiation-induced coagulopathy

Purpose Total body irradiation of the Gottingen minipig results in a characteristic hematopoietic response, including anemia, neutropenia, lymphocytopenia, and thrombocytopenia. Currently, there are no well-characterized large or small animal models for radiation-induced thrombocytopenia. The study described here using the Gottingen minipig was focused on understanding which aspects of the coagulation cascade leads to radiation-induced coagulopathy. In this study, multiple clinical pathology parameters were determined prior to and for 45-days following total body irradiation using a 6 MV photon linear accelerator. Materials and methods Following irradiation, frequent analyses of conventional hematology and coagulation parameters provided time-course information on the onset and recovery of thrombocytopenia. In addition, thromboelastography (TEG) was utilized to monitor coagulation dysfunction, namely clotting time, clot formation time, clot strength, and fibrinolysis. Coagulation factor activity levels were measured for factors II, V, VII, VIII, IX, X, XI, XII, XIII, Protein C, fibrin monomers, antiplasmin and D-dimer using a Siemen’s coagulation analyzer to provide time course information of changes in activity post irradiation exposure. Results These analyses revealed that in total body irradiated minipigs, TEG tracings demonstrate long R (time to initial clot formation) and K (time to achieve a certain clot strength) times, and low alpha-angle (rate of clot formation) and MA (overall stability of the clot) during onset of thrombocytopenia (typically post irradiation day 10–15). Low alpha-angle and MA directly correlated with decreased platelet counts. A long R time is suggestive of a deficiency in clotting factors and was compared to measured activity levels of individual coagulation factors. The data indicates that coagulation factors are significantly changed early after irradiation exposure prior to thrombocytopenia and factors VIII, XI, XII and XIII are markedly altered during the critical point of thrombocytopenia. Conclusion These data support the continued use of multiple approaches to evaluate the coagulation cascade in order to provide the most meaningful interpretation of the hematopoietic changes that occur post irradiation.

INCREASED SYSTEMIC HEPARANASE IN RETINAL VEIN OCCLUSION IS ASSOCIATED WITH ACTIVATION OF INFLAMMATION AND THROMBOPHILIA.

To investigate the levels of systemic heparanase, inflammatory markers, and coagulation factor activities in patients with retinal vein occlusion (RVO). This prospective study included 18 patients with central RVO, 22 patients with branch RVO, and 40 patients with age-related cataract as the control group. Serum heparanase protein levels and activities were measured by ELISA and a heparan degrading enzyme assay kit, respectively. Serum levels of MMP-2, MMP-9, TLR-2, and TLR-4 were measured by ELISA kits. The activities of coagulation factors (V, VII, VIII, and IX) were determined with an autoanalyzer. The Mann-Whitney U test was used to compare the above parameters between patients with RVO and control subjects. The relationship between two of the above parameters was analyzed by Spearman's correlation. Patients with RVO had higher levels of systemic heparanase protein, heparanase activities, coagulation factors' (V, VIII, and IX) activities, MMP-2, MMP-9, TLR-2, and TLR-4 compared with the control group. Systemic heparanase levels were correlated with serum levels of MMP-2, MMP-9, TLR-2, TLR-4, and activities of coagulation factors VIII and IX. Increase of systemic heparanase in RVO is associated with activation of systemic inflammation and blood hypercoagulability.

The Case Files

The Case Files

Hyperosmolar reconstituted lyophilized plasma is an effective low-volume hemostatic resuscitation fluid for trauma

We performed this study to optimize reconstituted lyophilized plasma (LP) into a minimal volume fluid that provides effective hemostatic resuscitation for trauma while minimizing logistical limitations. We performed a prospective, blinded animal study. Plasma was lyophilized following whole blood collection from anesthetized swine. The minimal volume needed for reconstitution was determined, and this solution was evaluated for safe infusion into the swine. Reconstituted LP was analyzed for electrolyte content, osmolarity, and coagulation factor activity. Twenty swine were anesthetized and subjected to a validated model of polytrauma and hemorrhagic shock (including a Grade V liver injury), then randomized to resuscitation with LP reconstituted to either 100% of the original plasma volume (100%LP) or the minimal volume LP fluid. Physiologic data were monitored, and blood loss and hematocrit were measured. Coagulation status was evaluated using thrombelastography. The minimal volume of reconstituted LP safe for infusion in swine was 50% of the original plasma volume (50%LP). The 50%LP had higher electrolyte concentrations, osmolarity, and increased coagulation factor activity levels by volume compared with 100%LP (p < 0.05). Blood loss, hematocrit, mean arterial pressure, and heart rate did not differ between animals receiving 100%LP (n = 10) or 50%LP (n = 10) at any time point (p > 0.05). International normalized ratio and thrombelastography parameters were not different between groups (R time, α angle, or maximal amplitude, p > 0.05). Resuscitation with 50%LP fluid was well tolerated and equally effective compared with 100%LP, with respect to physiologic and hemostatic properties. The smaller volume of fluid necessary to reconstitute hypertonic LP makes it logistically superior to 100%LP for first responders and may reduce adverse effects of large-volume resuscitation.

Quantitative evaluation of plasma after methylene blue and white light treatment in four Chinese blood centers

Pathogen reduction technology is an important process in the blood safety system, including solvent/detergent treatment, filtration and methylene blue-photochemical technology (MB-PCT). To investigate the quality of MB-PCT-treated plasma, plasma samples from four Chinese blood centers were analyzed over 12 months of storage to determine the recovery of activities and levels of various plasma proteins. Ten plasma units each from the Suzhou, Yancheng, Chongqing and Shandong blood centers were divided into two aliquots. One was subjected to treatment with one of two methylene blue-photochemical technology instruments and the other was used as control. The treated and untreated sample pairs were stored at -30°C. The recovery rates of coagulation factors, inhibitor proteins, total protein, immunoglobulins, and complement proteins were measured at different time points after storage. The mean recovery of most proteins exceeded 80% after MB treatment. The exceptions were factor XI and fibrinogen, of which only 71.3-74% and 69.0-92.3% were retained during storage. The two equipment types differed in terms of residual MB concentration in the plasma samples (0.18 μM and 0.29 μM, respectively). They had similar protein recovery rates at 0.5 month but differed at later time points. The four blood centers differed significantly with regard to factor II, V, VIII and fibrinogen activities. Only the samples from the Shandong blood center met the methylene blue treated fresh frozen plasma requirement. The major factor that influenced the quality of the MB-FFP samples was the time taken between blood collection and storage. Methylene blue treated plasma showed reduced coagulation factor (CF) activity and protein levels. The MB treatment-induced damage to the proteins was acceptable at the four Chinese blood centers, but the quality of the MB-treated plasma in general was not satisfactory. The main factor affecting plasma quality may be the duration of the collection and processing.

Acquired haemophilia A: a clinical case

AbstractBackground Acquired haemophilia A (AHA) is a rare medical emergency that may be characterised by severe and life‐threatening haemorrhage.Aims This case report highlights the need for a better awareness of this disorder in terms of diagnosis and urgent management.Materials and Methods This is a clinical presentation of a 77‐year‐old Caucasian man who presented to Accident and Emergency with profuse bleeding from his tongue, secondary to mechanical trauma. Local haemostatic measures were only temporarily effective as he returned on a further five occasions over a 3‐week period with recurrent bleeding from the same site. A coagulation screen revealed prolongation of the activated partial thromboplastin time that did not correct on a 50/50 mixing study with normal plasma. Subsequent analysis of intrinsic coagulation factor activity levels revealed severe factor VIII (FVIII) deficiency. A high‐titre FVIII inhibitor was identified in his plasma and the diagnosis of AHA was made. The patient was promptly transferred to the regional haemophilia comprehensive care centre for specialist management.Discussion AHA should be suspected in a patient with no personal or family history of a bleeding disorder who presents with soft tissue haemorrhage unresponsive to local measures.Conclusion This condition is particularly relevant to dental clinicians as patients may present with oral clinical signs of disease. In addition, affected individuals are also at risk of severe haemorrhage following accidental trauma or invasive oral surgery procedures.

Coagulation factor activity and clinical bleeding severity in rare bleeding disorders: results from the European Network of Rare Bleeding Disorders

The European Network of Rare Bleeding Disorders (EN-RBD) was established to bridge the gap between knowledge and practise in the care of patients with RBDs. To explore the relationship between coagulation factor activity level and bleeding severity in patients with RBDs. Cross-sectional study using data from 489 patients registered in the EN-RBD. Coagulation factor activity levels were retrieved. Clinical bleeding episodes were classified into four categories according to severity. The mean age of patients at data collection was 31 years (range, 7 months to 95 years), with an equal sex distribution. On linear regression analysis, there was a strong association between coagulation factor activity level and clinical bleeding severity for fibrinogen, factor (F) X, FXIII, and combined FV and FVIII deficiencies. A weaker association was present for FV and FVII deficiencies. There was no association between coagulation factor activity level and clinical bleeding severity for FXI. The coagulation factor activity levels that were necessary for patients to remain asymptomatic were: fibrinogen, > 100 mg dL(-1); FV, 12 U dL(-1); combined FV + VIII, 43 U dL(-1); FVII, 25 U dL(-1); FX, 56 U dL(-1) ; FXI, 26 U dL(-1); FXIII, 31 U dL(-1). Moreover, coagulation factor activity levels that corresponded with Grade III bleeding were: undetectable levels for fibrinogen, FV and FXIII, < 15 U dL(-1) for combined FV + VIII; < 8 U dL(-1) for FVI; < 10 U dL(-1) for FX; and < 25 U dL(-1) for FXI. There is a heterogeneous association between coagulation factor activity level and clinical bleeding severity in different RBDs. A strong association is only observed in fibrinogen, FX and FXIII deficiencies.

Coagulation Factor Activity Level and Clinical Bleeding Severity in Rare Bleeding Disorders: Results From the European Network of Rare Bleeding Disorders (EN-RBD),

Abstract 3312▪▪This icon denotes a clinically relevant abstractThe European Network of Rare Bleeding Disorders (EN-RBD) was established to bridge the gap between knowledge and practice in the care of patients with RBDs (rare bleeding disorders frequency <1/0.5-2M). The aim of this first report was to explore the relationship between the coagulation factor activity levels and clinical bleeding severity in patients with RBDs. A total of 592 records on patients with RBDs were cross-sectionally collected over a period of three years. Data on clinical bleeding episodes were available for 495 patients and were classified into four categories according to severity. The mean age of patients was 31 years (range, 7 months-95 years), with 51% being females. On linear regression analysis, there was a strong association between coagulation factor activity level and clinical bleeding severity for fibrinogen, factor (F)FX, FXIII, and combined FV and FVIII deficiencies. A weaker association was present for FV and FVII deficiencies. There was no association between coagulation factor activity level and clinical bleeding severity for FXI. It is of clinical relevance to identify levels that are not associated with spontaneous major bleeding, which was done by constructing receiver operating characteristic curves. These levels were highest for FXIII deficiency (25 U/dl), followed by FV+VII (15 U/dl), FVII (15 U/dl), FX (5 U/dl), and FV (1 U/dl) deficiencies. For fibrinogen deficiency, a factor activity level of 20 mg/dl is needed to ensure absence of major spontaneous bleeding. There is a clear relation between coagulation factor activity level and clinical bleeding severity in RBDs, which is different for the missing clotting factor. Disclosures:Peyvandi:NovoNordisk, CSL Behring: Honoraria. Bidlingmaier:CSL Behring, Bayer Schering: Research Funding; CSL Behring, NovoNordisk, Pfizer, Bayer Schering: Membership on an entity's Board of Directors or advisory committees; Baxter, Biotest, CSL Behring, NovoNordisk, Pfizer, Bayer Schering: Honoraria. Schved:LFB, CSL Behring, Novonordisk, Bayer Schering: Research Funding.

Interaction Between Oral Contraceptive Use and Coagulation Factor Levels in Deep Vein Thrombosis

Background: Since not all women using oral contraceptives (OC) develop deep vein thrombosis (DVT), the presence of additional risk factors in women with DVT and OC use is likely. High levels of coagulation factor activities are established risk factors of DVT. There are, however, limited data on the combined effect of OC use and high levels of procoagulant coagulation factors, a shortened aPTT or markers of inflammation like erythrocyte sedimentation rate (ESR) or CRP.Materials and Methods: 246 women with a first DVT (123 associated with OC use) and 246 normal women matched for age and date of birth were evaluated. Cut-off values for continuous variables were the 90% percentile determined in normal women. OC use influences the levels of many coagulation factors. Using the mean effect of OC use on coagulation factor levels (measured in the control subjects), we estimated the coagulation factor levels in patients prior to the index date, i.e. the time of the thrombosis.Results: Use of OC was associated with an increase in risk of 3.0. This risk increased due to interaction with factor I (FI) (Odds Ratio (OR) 14.2, p=0.0001), FVIII (OR 5.1, p=0.0001), FIX (OR 5.5, p=0.0001), FXI (OR 12.0, p=0.0001), vWF:Ag (OR 7.7, p=0.0001) and aPTT (OR 20.2, p=0.004). In contrast, no significant increase in risk was found when OC use was associated with high levels of ESR or CRP.Conclusion: The results of this study indicate that the joint effects of OC use and high coagulation factor levels increase the risk of DVT. This risk increase appears to be of clinical relevance particularly in individuals with shortened aPTT values or high levels of fibrinogen.

Coagulation Factors in Therapeutic Apheresis Plasma Held for 18 Hours at Ambient Temperature Prior to Pathogen Inactivation (INTERCEPT™).

Background. A photochemical treatment (PCT) using amotosalen HCl (S-59) and UVA light inactivates pathogens and leukocytes in therapeutic single donor apheresis fresh frozen plasma (INTERCEPT™, I-FFP) prepared within 8 hr of collection. Previous studies demonstrated a broad spectrum of pathogen inactivation (Transfusion 2006; 46:1168) and clinical efficacy of I-FFP for support of coagulopathies (Transfusion 2005; 45:1362; Blood 2006; 107:3753), and plasma exchange of TTP (Transfusion 2006;46). Preparation of therapeutic plasma up to 18 hr after collection would improve production logistics of frozen plasma provided sufficient levels of coagulation factors were retained.Aims. We measured coagulation factors in apheresis plasma stored for 18 hr at ambient temperature, processed with pathogen inactivation, and frozen.Methods. Fifteen jumbo plasma units (650 mL), were collected by apheresis with AB16 anticoagulant from group A, B, AB and O donors (MCS+. Haemonetics, Braintree, MA). Plasma collections were held at ambient blood bank temperature (20 – 24 °C) prior to further processing. After 18 hr, baseline samples for assay of coagulation factors were withdrawn before PCT. Plasma (635 mL plasma) was mixed with 15 mL of 6 mM amotosalen (150 uM: final concentration) and illuminated with a 3 J/cm2 UVA treatment. Following illumination (~ 8 min) and passage through a flow compound adsorption device (~20 min) to reduce levels of residual S-59, treated plasma units (650 mL) were divided into 3 equal storage units of ≥ 200 mL. Before freezing, post-treatment samples were withdrawn for factor assays. Treated plasma units were flash frozen at −80°C, and transferred to −30°C for 12-month storage. Plasma units were withdrawn to measure total protein, albumin, IgG, IgM, IgA, fibrinogen, factors II, V, VII, VIII, IX, X, XI, XII, VIII-vWF, Proteins C and S, AT III, plasminogen, alpha-2 antiplasmin, D-dimers, PT, and APTT.Results. Baseline coagulation factor levels (Mean ± SD) were in therapeutic ranges after 18 hr storage at ambient temperature. After PCT, all units had residual platelets < 1x109/L, WBC < 1x104/L, and RBC < 1 x 109/L. After PCT, total protein (59 ± 4 g/L), albumin (38 ± 2 g/L), IgG (9.0 ± 1.7g/L), IgA (1.6 ± 0.8 g/L) and IgM (0.9 ± 0.5 g/L) were unchanged from baseline. Mean values for fibrinogen (g/L), coagulation factors (IU/dL), coagulation inhibitors (IU/dL) were variably reduced from baseline, but within the ranges defined for therapeutic plasma (Table). Treated plasma showed no evidence of activation.Conclusions. Apheresis plasma held for 18 hr before processing with the INTERCEPT system for pathogen inactivation retained coagulation factor activity levels in conformance with French national standards for therapeutic frozen plasma (FP). Approximately 36 units (200 mL) could be prepared per hr with this system. A single UVA platform is compatible with the operational requirements of a regional blood center producing 12,000 doses (200 mL) of therapeutic FP and 12,000 doses of platelets per year. [Display omitted]

Coagulation Factor Activity in Therapeutic Plasma Prepared from Whole Blood with Pathogen Inactivation (INTERCEPT™) Treatment.

Background. A photochemical treatment (PCT) using amotosalen HCl (S-59) and UVA light was developed to inactivate pathogens and leukocytes in therapeutic plasma (INTERCEPT™, I-FFP) frozen within 8 hr of collection. Previous studies demonstrated a broad spectrum of pathogen inactivation (Transfusion 2006; 46:1168) and clinical efficacy of I-FFP for support of coagulopathies (Transfusion 2005; 45:1362; Blood 2006; 107:3753), and plasma exchange of TTP (Transfusion 2006;46). Preparation of therapeutic plasma from whole blood would complement blood center logistics and reduce the cost of therapeutic frozen plasma provided sufficient coagulation factors were retained.Aims. We measured coagulation factors in plasma isolated from whole blood held overnight at controlled temperature (21 ± 3°C), processed with pathogen inactivation, and frozen within 18 hr of blood collection.Methods. Whole blood units, approximately 460 mL, anticoagulated with CPD (Baxter, La Chatre, France) were drawn from group A, O, B and AB donors. Units were processed after 16 hr storage, and plasma was isolated by centrifugation. Two to 3 plasma units of matched blood group were pooled (n = 30: A = 14, O = 14, B = 1, AB =1) to a final volume of 635 mL. Baseline samples for assay of coagulation factors were withdrawn. Each of 30 pools was mixed with 15 mL of 6 mM amotosalen (150 uM: final concentration) and illuminated with a 3 J/cm2 UVA treatment. Following illumination (~ 8 min) and passage through a flow compound adsorption device (~20 min) to reduce levels of residual S-59, treated plasma units (650 mL) were divided into 3 equal storage units of ≥ 200 mL. Before freezing, post-treatment samples for assay of coagulation factors were withdrawn for assay of coagulation factors. Treated plasma units were flash frozen at -80°C, and transferred to −30°C for 12-month storage. Treated units were withdrawn after 1 month to measure total protein, albumin, IgG, IgM, IgA, fibrinogen, factors II, V, VII, VIII, IX, X, XI, XII, VIII-vWF, Proteins C and S, AT III, plasminogen, alpha-2 antiplasmin, D-dimers, PT, and APTT.Results. Baseline coagulation factor levels (Mean ± SD) were in suitable therapeutic ranges. After PCT, all units had residual platelets < 1×109/L, WBC < 1×104/L, and RBC < 1 × 109/L. After PCT and frozen storage for 1 month, total protein (59 ± 2 g/L), albumin (38 ± 1 g/L), IgG (8.9 ± 1.1g/L), IgA (1.8 ± 0.4 g/L) and IgM (0.9 ± 0.3 g/L) were unchanged from baseline. Mean values for fibrinogen (g/L), coagulation factors (IU/dL), coagulation inhibitors (IU/dL), were variably reduced from baseline, but within ranges defined as suitable for therapeutic plasma (Table). There was no evidence of plasma activation.Conclusions. Plasma prepared from whole blood after storage on cooling plates before processing with the INTERCEPT system for pathogen inactivation retained coagulation factor activity levels after frozen storage (−30°C) in conformance with French national standards for therapeutic frozen plasma (FP). Approximately 36 units (200 mL) could be prepared per hr of illumination time with this system. [Display omitted]

The INTERCEPT Blood System for Plasma: Process Validation Studies of Coagulation Factor Activity and Yield in Two European Blood Centers.

Introduction: INTERCEPT Plasma (I-FFP) is prepared as FFP for transfusion using photochemical treatment (PCT) with amotosalen HCl (S-59, 150 μM) and UVA light (3J/cm2) to inactivate a broad spectrum of blood-borne pathogens. Phase 3 clinical trials, supported with an inventory of > 19,000 I-FFP units processed in 200 mL/unit with a prototype set, demonstrated retention of coagulation factor activities and hemostatic function in I-FFP for support of patients with congenital and acquired coagulopathies or TTP. For commercial introduction, the clinical prototype I-FFP system has been improved to treat up to 650 mL of plasma in a single, less time-consuming, PCT process yielding up to three 200 mL I-FFP doses per PCT process. The effects of this modified process on coagulation factor activity and yield were evaluated in two European blood centers under routine operating conditions.Methods: A total of 60 plasma units (approximately 600 mL/unit) were collected using the Autopheresis C device (Baxter-Fenwall) at Haukeland University Hospital and the Institute of Immunology and Transfusion Medicine, University of Lübeck. I-FFP units were prepared at each center by blood bank personnel using the improved set. Baseline and I-FFP samples were collected, frozen (<−60 °C) and sent to Cerus for assay of: Fibrinogen (F I), and Factors II, V, VII, VIII, IX, X, XI, XIII, proteins C (PC) and S (PS), and antithrombin (AT).α-2 antiplasmin (AP) was assayed in a reference laboratory.Results: Fibrinogen (mg/dL) and coagulation factor activities (IU/dL) are expressed as the mean ±SD (Table). The post-PCT yield for each coagulation factor activity in I-FFP units prepared with the improved set (Improved) is expressed as a proportion (%) of total pre-treatment coagulation factor activity per FFP unit (Baseline). Comparative yield data for I-FFP used in Phase 3 studies (Prototype) were obtained by testing 275 plasma units randomly selected from 19,000 units prepared for use in clinical trials with the protype set at 6 U. S. blood centers.Conclusions: I-FFP prepared with the improved system retained 78–79% of baseline Fibrinogen and FVIII activity, and 82%–95% of baseline Factors II, V, VII, IX, X, XI, XIII, PC, PS, AT, and AP. Clinical trials have shown that I-FFP provided sufficient levels of coagulation factor activities for treatment of congenital and acquired coagulopathies and for therapeutic plasma exchange of TTP. The improved set, intended for commercialization, provides multiple I-FFP doses with a single PCT process. Using the improved processing set, coagulation factor activities and yields were similar to those for I-FFP used in Phase 3 clinical trials.Activity (IU)F IF IIF VF VIIF VIIIF IXBaseline300±54100±17135±23112±25142±4497±15I-FFP238±4898±16131±2291±20111±3883±13Yield (%)F IF IIF VF VIIF VIIIF IXImproved79±590±490±482±378±585±4Prototype78±690±595±482±577±788±5Activity (IU)F XFXIFXIIIPCPSATAPBaseline115±17107±20121±20120±21100±20100±11104±13I-FFP103±1696±18115±20104±1996±1996±1085±8Yield (%)F XFXIFXIIIPCPSATAPImproved90±389±595±586±395±595±382±8Prototype90±390±699±3100±8103±692±290±3F XIII, PC, PS, AT (n=34); AP (n=14)

Blood coagulation changes after hypoxemia: a fetal lamb model.

The effects of fetal hypoxemia on blood coagulation (platelet count, prothrombin time, partial thromboplastin time, fibrinogen, factors II, V, VI, VIII, IX, X, XI, XII, XIII and von Willebrands activities, fibrin degradation products, and fibrin monomer) were evaluated in nine chronically catheterized fetal lambs early in the third trimester of pregnancy (107-110 days gestation). Seven chronically catheterized fetal lambs of similar gestational ages served as controls. The hypoxemic episode (pO2 14 mm Hg) was maintained for 1 hr in the experimental group during which time there were only minimal changes in PCO2, arterial pressure, heart rate, and pH. Epinephrine and norepinephrine levels increased significantly in stressed animals--22 pg/ml pre- to 1025 pg/ml postepinephrine, and 475 pg/ml pre- to 2292 pg/ml postnorepinephrine. There were no significant changes in blood coagulation factor activities related to the hypoxic stress although, one fetus who experienced acidemia did develop a transient increase in fibrin monomer. Slight through significant increases in VIII coagulant activity activity (4.0%), von Willebrand activity (5.9%), and factor XII activity (4.3%) occurred in both the hypoxemic and control fetal lambs. These changes were associated with minimal increases in the white blood cell count (15%) and slight decreases in the mean arterial pressure (3.9 mm Hg) hemoglobin (1.2 g), and hematocrit (2.9%) and may have been related to the blood loss of 25% that occurred as a result of sampling in both groups. There were no differences between the hypoxic and control animals' levels of coagulation factor activities when measured during an 18-day follow-up period except for a slight increase in factor X activity (10%) in the control animals not apparent in the nine hypoxic animals. Thus an episode of severe fetal hypoxemia in the absence of hypotension, acidosis, and hypercarbia does not lead to acute or chronic alterations in blood coagulation factor activities in the fetal lamb.

Development of blood coagulation--a fetal lamb model.

To study the normal development of blood coagulation factor activities in a growing fetus while avoiding the effects of labor and delivery, a chronic fetal lamb model was developed in which serial blood samples from 10 fetuses were studied during the third trimester of pregnancy and 24 hr after birth. Under operating room conditions with sterile technique, a polyethylene catheter to which heparin had been bound to both internal and external surfaces was inserted into the femoral artery of the fetus. The catheter was brought out through a skin pouch to the side of the ewe and enclosed in a zip lock bag. Blood samples were withdrawn from the catheter three times each week for measurement of coagulation factor activities. Levels of coagulation factor activities at birth in noncatheterized animals were not different from those found in catheterized animals except for factor IX activity which was 12% higher in the catheterized animals (0.02 less than P less than 0.05). The patterns of development for each of the coagulation factors were similar in all 10 animals studied. Fibrinogen, prothrombin, and factor VII show a decrease in activity early in the last trimester of pregnancy. Both factors VIII and IX show a significant increase in activity (23% factor VIII and 12% factor IX) associated with the process of delivery. The levels of coagulation factor activities at birth in the lamb relative to adult sheep normals are similar to those found in humans with the exception of factor XIII. Factor XIII is at normal levels in the newborn lamb and is reported to be at levels approximately 50% of the adult level in human infants.

Diagnosis and treatment of coagulopathy in the newborn.

Appropriate management of the bleeding newborn is easily accomplished by first assessing the clinical circumstances under which the bleeding occurs. Having determined the clinical circumstances, knowledge of the pathophysiology of disseminated intravascular coagulation, liver failure, vitamin K deficiency, and hemophilia coupled with knowledge of the normal levels of coagulation factor activities at birth leads to selection of appropriate laboratory tests to confirm the etiology of the bleeding. Once the etiology is confirmed, treatment requires management of associated clinical conditions and replacement of vitamin K and/or deficient coagulation factors.