Extrinsic pathway of blood coagulation and thrombin in the cerebrospinal fluid after subarachnoid hemorrhage.

OBJECTIVE: The involvement of thrombin in the pathophysiology of subarachnoid hemorrhage (SAH) was investigated by comparing thrombin expression and extrinsic pathway activation in the cerebrospinal fluid (CSF) and blood of patients with SAH with the neurological grades, outcome, and presence of delayed cerebral vasospasm. METHODS: Blood and CSF samples were obtained from 38 patients with SAH on Days 3 through 5, 7 through 9, and 12 through 14 after the onset of SAH. CSF samples were also obtained from control patients. Thrombin-antithrombin III complex, prothrombin fragment F1 +2, tissue factor, and tissue factor pathway inhibitor were analyzed using enzyme-linked immunosorbent assay. RESULTS: No markers in the blood or CSF were correlated with neurological grades and outcome. Thrombin-antithrombin III complex and prothrombin fragment F1 +2 levels were significantly higher in the CSF of patients with SAH than in the blood or the CSF of control patients and were significantly higher in patients with vasospasm than in patients without vasospasm on Days 7 through 9. Tissue factor levels were significantly higher in the CSF of patients with SAH than in the blood, but the levels were close to those in the CSF of control patients. Tissue factor pathway inhibitor levels in the CSF of patients with SAH and control patients were under the detection limit. CONCLUSION: Thrombin in the blood may not reflect the pathophysiology of SAH. Imbalance between tissue factor and tissue factor pathway inhibitor in the CSF may tend to thrombin generation under normal physiological conditions and also after SAH. Thrombin in the CSF may be involved in the pathophysiology of vasospasm.

Cerebrospinal fluid tissue factor and thrombin-antithrombin III complex as indicators of tissue injury after subarachnoid hemorrhage.

BACKGROUND AND PURPOSE: No marker that reflects and predicts brain injury due to subarachnoid hemorrhage (SAH) and cerebral vasospasm has been reported. We hypothesized that membrane-bound tissue factor (mTF) and thrombin-antithrombin III complex (TAT) in the cerebrospinal fluid (CSF) of patients with SAH become markers indicating brain injury. To evaluate the hypothesis, we correlated levels of mTF and TAT in the CSF of patients with SAH with clinical severity, the degree of SAH, and outcome. METHODS: We assayed CSF mTF, TAT and myelin basic protein (MBP) in patients with SAH at intervals that included days 0 to 4 and days 5 to 9 after ictus. Classification of clinical severity of disease on admission was based on Hunt and Hess grade, degree of SAH on CT on Fisher's grading, and outcome 3 months after SAH on the Glasgow Outcome Scale. RESULTS: In the interval from days 0 to 4, mTF and TAT correlated with Hunt and Hess and Fisher grades, and occurrence of cerebral infarction due to vasospasm. Only mTF correlated significantly in this period with outcome. TAT, mTF, and MBP all correlated significantly with each other. From days 5 to 9, only mTF correlated with cerebral infarction, infarction volume, MBP levels, and outcome. CONCLUSIONS: Both mTF and TAT reflected brain injury from SAH and predicted vasospasm, though mTF was more sensitive and a better predictor of outcome. Unlike mTF, TAT did not correlate with vasospasm during the interval when it most commonly occurs, which raised doubt about thrombin activation as a cause.

Tissue factor antigen levels in various biological fluids.

Tissue factor (TF), a transmembrane surface protein, is known to initiate thrombogenesis through plasmatic and cellular activation processes. Besides complexing with factor VII, eventually leading to fibrin generation via the extrinsic pathway, TF can also activate factor IX, resulting in the intrinsic activation of coagulation. Other functions of TF are currently unknown, although various cells are believed to have TF receptors. Many of the post-surgical and post-interventional thrombotic events are due to the release of TF. Increased levels of TF are associated with several pathologic conditions such as cancer, sepsis and inflammation. Cellular necrosis also results in an increase of TF as the cells in the traumatized area lyse and release endogenous cell surface-bound TF. An ELISA method (American Diagnostica, Greenwich, CT) has been developed to assay TF antigen levels in various biological fluids. This ELISA employs a murine monoclonal antibody raised against native human TF for antigen capture. In this study, cerebrospinal fluid, peritoneal fluid, pleural effusion and urine from patients were assayed for their TF content using this ELISA method. Normal individual serum and plasma were also assayed as controls against which the levels of TF in the patients' body fluids could be compared. The amount of TF antigen in normal human plasma and serum was 165 +/- 139 pg/ml and 165 +/- 110 pg/ml, respectively. Concentrations of TF antigen in other fluids were: cerebrospinal fluid 868 +/- 721 pg/ml, peritoneal fluid 124 +/- 247 pg/ml, pleural effusion 385 +/- 569 pg/ml, synovial fluid 97 +/- 23 pg/ml, seminal plasma 11,485 +/- 875 pg/ml and urine 86 +/- 57 pg/ml.(ABSTRACT TRUNCATED AT 250 WORDS)

Thromboplastic activity of cerebrospinal fluid in neurological disease.

Estimates of thromboplastic activity in 1100 samples of cerebrospinal fluid indicate that an increased activity of this clotting factor is a nonspecific indicator of abnormality in the central nervous system, much like (e.g.) an increased count of mononuclear cells and an increased protein content. However, the proportion of abnormal results obtained by these three tests can differ markedly in different neurological disorders. Increased thromboplastic activity is about 14-fold more common in bacterial meningitis than in viral meningitis; thus the thromboplastin determination can be of value in discriminating between bacterial and viral meningitis.

Comparison of the spectrophotometric determination and the two-stage coagulation assay of tissue factor activity.

The tissue factor activity of human brain thromboplastin and 6 commercial thromboplastins was determined by a spectrophotometric method and a two-stage coagulation assay. The thromboplastins were incubated with an excess of Factor VII and Factor X, and the activation of Factor X was estimated from the rate of hydrolysis of the chromogenic substrate S-2222 and from the coagulation time of plasma enriched in phospholipids. The results obtained by the two methods were related linearly and showed a correlation coefficient of 0.89. The coefficient of variation was 11% in the spectrophotometric method and 25% in the two-stage coagulation assay.

Spectrophotometry of tissue thromboplastin in cerebrospinal fluid.

We describe a two-stage method for measuring thromboplastin in cerebrospinal fluid, based on the amidolytic determination of the generation rate of Factor Xa. We investigated the effect of variations in the duration of incubation, and in pH, temperature, and concentrations of calcium, buffer, and prothrombin complex (source of Factors VII and X). Optimal assay conditions are specified. The detection limit is 6 units/L and the results indicate an upper limit of normal of 14 units/L. The coefficients of variation were 7.7% within-day and 9.2--16.0% day-to-day, depending on the concentration.

Spinal-fluid procoagulant activity: a sensitive indicator of central-nervous-system damage.

Testing the procoagulant activity (P.C.A.) of cerebrospinal fluid (C.S.F.) by measuring its effect on plasma-recalcification time is a useful indicator of central-nervous-system damage. C.S.F. from 22 normal children and adolescents aged 6 months to 17 years had a mean +/- S.D. P.C.A. of 14 +/- 6%. P.C.A. in 13 children with C.N.S. infection was significantly increased to 59 +/- 13%. In 8/13 of these children activity remained high (42 +/- 11%) after therapy. In patients with acute lymphatic leukemia (A.L.L.) 12 aged greater than 2 1/2 years and diagnosis had a normal activity (17 +/- 8%) and 4 patients aged less than 2 1/2 years at diagnosis had an activity of 21 +/- 8%; during C.N.S. prophylaxis with intrathecal methotrexate and/or cranial irradiation, mean activity in these 16 patients rose significantly to 41 +/- 11%. More than 2 years after treatment P.C.A. decreased towards normal in the older children but remained high in the younger group. 5 children with neurological sequelae (including 3 with A.L.L. and the post-irradiation syndrome) had the highest activities. Ether extraction showed that the active material had lipid properties. P.C.A. did not correlate with protein, lactic dehydrogenase, or cell count in C.S.F.