Mark Fisher

Background and Purpose Endothelial cells regulate hemostasis in part via expression of thrombomodulin, a potent anticoagulant protein. The purpose of this study was to analyze brain capillary endothelial cell expression of thrombomodulin mRNA. Methods Bovine brain capillary endothelial cells were grown in a blood-brain barrier model in which endothelial cells form capillary-like structures. In situ hybridization and polymerase chain reaction (PCR) were used to examine thrombomodulin expression. Endothelial cells were then cocultured with astrocytes. We examined both coculture and monoculture preparations for γ-glutamyl transpeptidase (GGTP), a marker of the blood-brain barrier. We then used quantitative-competitive PCR to compare thrombomodulin expression in endothelial monocultures and astrocyte-endothelial cocultures after 1 and 7 days of culture. Results Both in situ hybridization and PCR studies demonstrated thrombomodulin mRNA expression by endothelial cells. During 1 week of astrocyte-endothelial coculture, there was (1) progressive association of astrocytes with capillary-like structures and (2) expression of GGTP; endothelial monocultures did not express GGTP. There was no significant difference in thrombomodulin mRNA expression for cocultures versus monocultures after 1 day. After 1 week, however, astrocyte-endothelial cocultures had markedly decreased thrombomodulin mRNA compared with monocultures (9±2 versus 189±62 pg/mL; P<.025). This thrombomodulin mRNA decrease thus occurred when elements of the blood-brain barrier phenotype were demonstrable, ie, when astrocyte association with capillary-like structures was maximal and when GGTP was expressed in cocultures. Conclusions These findings indicate astrocyte regulation of thrombomodulin mRNA expression in vitro and suggest an important role for the blood-brain barrier in the regulation of thrombomodulin.

Background and Purpose Infection/inflammation appears to be an important predisposing risk factor for brain infarction, but little is known regarding underlying molecular mechanisms. We examined the hypothesis that patients with brain infarction preceded by infection/inflammation within 1 week could be identified by a distinctive procoagulant laboratory profile characterized by abnormalities in the protein C system and endogenous fibrinolysis. Methods We performed a case-control study examining the relationship between preceding systemic infectious/inflammatory syndromes and selected immunohematologic variables in 36 patients with acute brain infarction and 81 control subjects (community control subjects [n=47] and hospitalized nonstroke neurological patient controls [n=34]). Results The stroke group had a lower mean level of the circulating antithrombotic enzyme activated protein C (APC) (4.33±0.34% [log-transformed percentage of control value, mean±SD]) than community control subjects (4.51±0.27%, P <.02) or hospitalized neurological patient controls (4.57±0.31%, P <.005). The lowest circulating APC levels were found in the stroke group with antecedent infection/inflammation within 1 week preceding index brain infarction (4.23±0.4%, n=12). Within the stroke group, circulating APC levels were inversely related to IgG isotype anticardiolipin antibody titers ( r =−.55, P <.001). Only the stroke group with infection/inflammation within 1 week had elevated plasma C4b binding protein compared with control subjects (141±61% versus 112±44%, P <.05). Stroke patients with antecedent infection/inflammation had a distinctively lower ratio of active tissue plasminogen activator to plasminogen activator inhibitor (0.11±0.04, n=9) than other stroke patients (0.19±0.06, n=9, P <.01) and control subjects (0.22±0.16, n=17, P <.02). Conclusions Impairments in the protein C pathway and endogenous fibrinolysis may contribute to the increased risk for brain infarction after recent (≤1 week) infection/inflammation. A decrease in the circulating anticoagulant APC may be related to elevated antiphospholipid antibody titers.

Background and Purpose Antecedent febrile infection and psychological stress are described as predisposing risk factors for brain infarction. We examined the temporal relationship between preceding infection/inflammation and stroke onset as well as the role of recent psychological stress as a potential precipitant for brain infarction. Methods In this case-control study, a standardized evaluation including a signs/symptoms-based questionnaire was used to characterize the prevalence and timing of recent prior (Results The prevalence of infection/inflammation was significantly higher in the stroke group only within the preceding 1 week compared with either community control subjects (13/37 versus 6/47, P <.02) or hospitalized neurological patient controls (3/34, P <.02). Upper respiratory tract infections constituted the most common type of infection. A substantial proportion of stroke patients with preceding (Conclusions Our data suggest that both febrile and nonfebrile infectious/inflammatory syndromes may be a common predisposing risk factor for brain infarction and that the period of increased risk is confined within a brief temporal window of less than 1 week. Results of this study argue against a role for recent psychological stress as a precipitant for cerebral infarction.

Background and Purpose Resistance to activated protein C (APC), a natural plasma anticoagulant, is the most common identifiable risk factor for venous thromboembolic disease. One point mutation in coagulation factor V that renders it APC-resistant is found in >90% of APC-resistant venous thrombosis patients. To determine the prevalence of APC resistance and of this factor V mutation in stroke, we screened a group of ischemic stroke patients. Methods Hispanic ischemic stroke patients were screened using two different activated partial thromboplastin time-based assays. One assay using neat patient plasma determined APC resistance, and the other assay using patient plasma diluted into factor V-deficient plasma determined APC-resistant factor V, including the Arg ⁵⁰⁶ →Gln mutation. Results were compared with those in 31 Hispanic control subjects of similar ages. Results Six of 63 (9.5%) stroke patients had APC resistance compared with none of 31 (0%) control subjects. No patient or control subject had APC-resistant factor V, ie, the factor V Arg ⁵⁰⁶ →Gln mutation. Conclusions In Hispanic patients with ischemic stroke, the incidence (approximately 10%) of APC resistance is not caused by the factor V Arg ⁵⁰⁶ →Gln mutation. APC resistance not caused by this factor V mutation may be a risk factor for ischemic stroke in this population.

Atherosclerotic plaques may not be demonstrated adequately by standard anteroposterior (AP)-lateral projections of carotid arteriography. We evaluated selective carotid arteriography in oblique projections for 60 individual carotid arteries. Two neuroradiologists evaluated arteriograms in four separate sessions devoted to AP-lateral views alone and in combination with oblique projections. The lumen surface was characterized as “smooth,” “irregular,” or “ulcerated.” When oblique projections were added to AP-lateral views, there was a significant shift of radiologic diagnosis from smooth to irregular or ulcerated. Addition of oblique projections to standard AP-iateral views increases the radio-graphic diagnosis of carotid artery lesions that may be the source of emboli.