Thrombotic thrombocytopenic purpura and hemolytic uremic syndrome following allogeneic bone marrow transplantation.

We monitored the levels of various cytokines and chemokines, as well as an adhesion molecule and factors related to vascular endothelial damage, in three patients with thrombotic thrombocytopenic purpura and hemolytic uremic syndrome after bone marrow transplantation. Measurements were done at the onset of this condition and during plasma exchange for treatment. At the onset of thrombotic thrombocytopenic purpura and hemolytic uremic syndrome, the levels of interleukin-8, thrombomodulin, and plasminogen activator inhibitor-1 were all markedly increased. A close relationship was observed between improvement in symptoms by plasma exchange and a decrease in interleukin-8 level, suggesting that this chemokine may be related to the development of thrombotic thrombocytopenic purpura and hemolytic uremic syndrome after bone marrow transplantation.

Enhancement by cyclosporine A and tacrolimus of serotonin-induced formation of small platelet aggregation.

Cyclosporine A (CsA) may increase the incidence of thrombotic events, but whether tacrolimus (Tc) has such effects is still unclear. The serotonergic system has been linked to the thrombotic effects of CsA, but a direct effect of CsA on serotonin-induced platelet aggregation has not been demonstrated because of methodological difficulties. We measured the effects of CsA and Tc on serotonin-induced platelet aggregate formation by particle counting using light scattering. CsA and Tc both enhanced serotonin-induced formation of small platelet aggregates, however, neither CsA nor Tc affected aggregation induced by high or low concentrations of ADP, with or without addition of a serotonin receptor antagonist. Both CsA and Tc enhance platelet aggregation induced via the serotonin pathway.

Daunorubicin inhibits gene expression of cyclooxygenase-2 in vascular smooth muscle cells.

Daunorubicin (0.1-1 microM) concentration-dependently inhibited prostacyclin production induced by interleukin-1beta (IL-1beta, 2.5 ng/ml) in cultured aortic smooth muscle cells isolated from rats. IL-1beta stimulation caused activation of nuclear factor-kappaB (NF-kappaB) and expression of cyclooxygenase-2 (COX-2) mRNA and protein, which were inhibited by daunorubicin. However, COX activity, evaluated by conversion of exogenous arachidonic acid to prostacyclin, was not affected by daunorubicin (0.1-1 microM). Protein expression of COX-1 and NF-kappaB was not affected by daunorubicin. Daunorubicin also inhibited nitric oxide (NO) production induced by IL-1beta. These results suggest that daunorubicin attenuated prostacyclin synthesis through inhibiting expression of COX-2 mRNA, which could be explained by perturbation of NF-kappaB activation.

Oral eicosapentaenoic acid for complications of bone marrow transplantation.

The 'systemic inflammatory response syndrome' (SIRS) may represent the underlying cause of complications after bone marrow transplantation (BMT). This study was conducted to determine whether blocking the etiologic factors of SIRS could improve the complications of BMT. Sixteen consecutive patients with unrelated donors were allocated alternately to two groups. Seven patients received 1.8 g/day of eicosapentaenoic acid (EPA) orally from 3 weeks before to about 180 days after transplantation, while nine patients did not. These two groups were compared with respect to complications, survival, and various cytokines and factors causing vascular endothelial damage. All seven patients receiving EPA survived and only two had grade III graft-versus-host disease (GVHD). Among the nine patients not receiving EPA, three had grade III or IV GVHD. In addition, thrombotic microangiopathy developed in four patients and cytomegalovirus disease occurred in four. Five patients died in this group. The levels of leukotriene B(4), thromboxane A(2), and prostaglandin I(2) were significantly lower in patients receiving EPA than in those not receiving it (all P < 0.01). Cytokines such as tumor necrosis factor-alpha, interferon-gamma, and interleukin-10 were also significantly decreased by EPA (P < 0.05), as were factors causing vascular endothelial damage such as thrombomodulin and plasminogen activator inhibitor-1 (P < 0.05). The survival rate was significantly higher in the group given EPA (P < 0.01). EPA significantly reduced the complications of BMT, indicating that these complications may be manifestations of the systemic inflammatory response syndrome.

Complications after bone marrow transplantation are manifestations of systemic inflammatory response syndrome.

Bone marrow transplantation has been established as a useful treatment for various hematological disorders and is now performed widely, but the mortality rate is still high due to various complications. A clear therapeutic policy for such complications has not yet been established because of their complex nature. We investigated whether the major complications occurring after bone marrow transplantation could be classified as aspects of the systemic inflammatory response syndrome. Subjects were 10 patients who developed severe complications after bone marrow transplantation (graft-versus-host disease, thrombotic microangiopathy, respiratory disorders, and cytomegalovirus interstitial pneumonitis) and 16 patients without complications. Their symptoms, serum cytokines, and factors related to vascular endothelial damage were compared before and after transplantation. Whereas all 10 patients who developed complications had fever in the aplastic phase after transplantation, 15 of the 16 patients without complications remained afebrile (P < 0.001, t-test). When compared with the patients who did not develop complications, the patients with complications also showed significantly higher cytokine levels during the recovery phase after transplantation (P < 0.0001, t-test). Thus, the patients with complications developed fever in the aplastic phase and showed an increase of cytokines during the recovery phase, which triggered the occurrence of vascular endothelial damage shown by factors such as the thrombomodulin and plasminogen activator inhibitor type 1. This sequence of events corresponds with that occurring during systemic inflammatory response syndrome, so many of the complications of bone marrow transplantation can be considered as manifestations of this syndrome.

Platelet aggregating activity of plasma in patients with thrombotic thrombocytopenic purpura evaluated by a particle counting method using light scattering.

The platelet aggregating activity of plasma obtained from five patients with thrombotic thrombocytopenic purpura (TTP) was evaluated by a particle counting method using light scattering. When normal platelets were suspended in the plasma obtained from TTP patients, small aggregate formation was observed after stirring at 1000 rpm without the addition of platelet aggregating agents; no aggregate was observed, however, in the plasma obtained from healthy donors. Since the inhibitory effect of the addition of normal plasma to TTP plasma on this reaction was dose-dependent and not additive, the efficacy of plasma therapy was not confirmed. Small aggregates were formed in the high molecular weight fraction (HMWF) of TTP plasma but not in the low molecular weight fraction (LMWF), suggesting that the platelet aggregating activity existed in HMWF. Among the antiplatelet agents usually used for TTP, dipyridamole was more effective for the inhibition of this reaction than aspirin. This spontaneous platelet aggregation reaction by a particle counting method using light scattering could be useful for evaluating the platelet aggregating activity in patients with TTP.

Effects of antitumor agents on inducible prostacyclin production in vascular smooth muscle.

The effects of various antitumor agents were examined on prostacyclin production induced by interleukin-1beta in rat aortic smooth muscle cells. Stimulation of the cells with interleukin-1beta (2.5 ng/ml) resulted in a great increase of prostacyclin production, which was abolished by indomethacin (1 microM) or cycloheximide (2 microM). Daunorubicin at 0.1-1 microM inhibited the inducible prostacyclin production in a concentration-dependent manner. However, other antitumor agents (cyclophosphamide at 1-100 microM, 5-fluorouracil at 1-100 microM and vincristine at 1-100 nM) tested did not significantly affect it. Protein expression of cyclooxygenase-2 induced by interleukin-1beta was inhibited by daunorubicin at 0.1-1 microM, but was not affected by other antitumor agents. These results suggest that daunorubicin inhibits induction of cyclooxygenase-2 and subsequent prostacyclin production in rat aortic smooth muscle cells.

Age related elevation of serum macrophage colony stimulating factor (M-CSF) level.

We recently reported that the serum level of macrophage colony-stimulating factor (M-CSF) was elevated in patients with cerebral infarction. In the present study, we measured serum M-CSF level, as well as coagulo-fibrinolytic markers and general laboratory tests in adult healthy subjects of various ages, and investigated the relationship between age and M-CSF level. M-CSF in aged subjects (>or=65 years of age) was significantly higher than that in the younger subjects (<65 years of age), and a significant positive correlation between age and M-CSF was found. Significant positive correlations between M-CSF, and plasma levels of thrombomodulin (TM), von Willebrand factor antigen (vWF), thrombin-antithrombin III complex (TAT), prothrombin fragment 1+2 (F1+2), d-dimer products cross-linked fibrin degradation products (d-dimer) and plasmin-antiplasmin complex (PAP) were also found. Among the general laboratory tests, there was only a significant correlation between M-CSF and serum creatinine; however, no significant correlation was found between M-CSF and other tests including blood cell counts. From these results, age-related elevation of serum M-CSF level was confirmed, and was suggested not to indicate the alteration of hemopoietic condition in aged subjects but to be related to thrombotic state or systemic damaged blood vessel in the apparently healthy aged people.

The levels of soluble P-selectin, von Willebrand factor and thrombomodulin in patients with neurological complications after allogeneic bone marrow transplantation.

We encountered two patients with uncommon neurological manifestations after allogeneic bone marrow transplantation (BMT), which occurred with rapid elevation of the leukocyte count at engraftment. Both patients then developed severe acute graft-versus-host disease (GVHD). To investigate the pathogenesis, we measured the levels of soluble P-selectin, von Willebrand factor (vWF) and thrombomodulin (TM), which reflect endothelial damage. The P-selectin, vWF and TM levels in the patients with (n=2) and without (n=5) neurotoxicity were, respectively, 168.5+/-52.5 ng/ml vs 27.7+/-3.9 ng/ml, 6.7+/-0.15 FU/ml vs 3.42+/-0.41 FU/ml and 459+/-37% vs 189.4+/-32.4% (mean+/-s.d.). All three parameters were much higher in the patients with neurological complications. These results suggest that neurotoxicity after BMT may be related to endothelial damage.

Age-related augmentation of stroke tendency evaluated using plasma P-selectin levels.

Plasma soluble P-selectin is thought to be a useful marker for thrombotic diseases. To evaluate the thrombotic state and risk of stroke in aged healthy subjects, we investigated plasma P-selectin levels in healthy subjects and ischemic stroke patients. Plasma P-selectin was measured in 67 healthy subjects and 35 aged (>or= 65 years of age) patients with chronic ischemic stroke using a sandwich enzyme-linked immunosorbent assay (ELISA). Plasma P-selectin was significantly higher in aged (>or= 65 years of age) healthy subjects than in young (< 65 years of age) healthy subjects. Significant difference did not exist between aged healthy subjects and aged stroke patients who were not receiving anti-platelet agents. Anti-platelet agent had no significant effect on plasma P-selectin levels in aged stroke patients. The amounts of P-selectin released from platelets into the plasma after stimulation with adenosin diphosphate in young and aged healthy subjects were not significantly different. Elevated levels of P-selectin in aged healthy subjects suggests the existence of a subclinical thrombotic state which can result in a stroke. Elevated P-selectin levels are not thought to be due to platelet hyperfunction.

Effects of perfluorocarbon exchange transfusion on reducing myocardial infarct size in a primate model of ischemia-reperfusion injury: a prospective, randomized study.

BACKGROUND: This study was undertaken to test the hypothesis that perfluorocarbons were able to reduce myocardial infarct size in a baboon model of ischemia-reperfusion injury. Exchange transfusion of perfluorocarbons has been shown to reduce myocardial infarct size in the dog, who, unlike the baboon, has an extensive collateral circulation. METHODS: After 15 minutes of occlusion of the left anterior descending coronary artery, 14 baboons were bled to attain a hematocrit of 24% to 26% and were simultaneously transfused, six with Fluosol-DA 20% emulsion and eight with FC-43 emulsion. After 2 hours of ligation, the coronary arteries were reperfused. Baboons were killed 24 hours after ligation, and the hearts were excised. Microvascular dye was infused into the coronary artery to delineate its perfusion bed. Ratios of the mean volume of infarct to the mean volume of perfusion bed were calculated and compared by use of planimetry. A similar protocol was followed in two other groups of baboons except that lactated Ringer's solution was infused into six of them, whereas eight had no exchange transfusions. RESULTS: The ratios of the mean volume of infarct to the mean volume of perfusion bed of the four groups were as follows: Fluosol-DA, 38.1% +/- 7.5%; FC-43, 37.7% +/- 8.3%; lactated Ringer's, 46.9% +/- 10.5%; controls, 65.6% +/- 6.9%. Statistical significance was reached when comparing both perfluorocarbon-treated groups with the controls (p < 0.05 for both groups) but not significant when comparing them with the Ringer's lactate-treated group. CONCLUSIONS: Results suggest that the beneficial effects of exchange transfusion with the perfluorocarbons may be primarily due to hemodilution.

Influence of atrial fibrillation on coagulo-fibrinolytic markers in patients with cerebral infarction.

To examine the influence of atrial fibrillation (Af) on stroke onset, we measured the plasma D-dimer level, thrombin antithrombin III complex and plasmin alpha 2 antiplasmin complex (PAP) in 46 stroke patients with Af and 87 stroke patients without Af. These marker levels were significantly higher in Af patients with stroke than in those without stroke (n = 16), and thus do not seem to be affected by Af alone. Abnormal values were also more frequent in acute Af stroke patients with visible occlusion of the major cerebral artery than in those without Af. The D-dimer and PAP levels in all Af stroke patients in the younger-aged patients (< or = 64 years) were significantly higher than those without Af, but not noted in the older-aged group (> or = 75 years). These results suggest that the D-dimer and PAP levels in younger-aged patients with Af indicate the existence of cerebral emboli due to Af.

Vasorelaxant effect of trapidil on human basilar artery.

We have investigated the vasorelaxant effect of trapidil on human isolated basilar artery. Trapidil (10(-5)-10(-4) M) dose-dependently caused relaxation in vascular strips with or without endothelium, with no significant difference between the two types of strips. The relaxation responses were not inhibited by atropine, propranolol or methylene blue. Trapidil increased the concentration of 6-keto-PGF1 alpha, a prostacyclin degradation product, released from an artery ring in the incubation medium, but trapidil-induced relaxation was not inhibited by indomethacin. Pretreatment of vascular strips with 10(-5) M trapidil increased the relaxation responses to forskolin and dibutyryladenosine cyclic monophosphate but not to sodium nitroprusside or 8-bromoguanosine cyclic monophosphate. Trapidil induced a significant increase in the cAMP concentration but not in the cGMP concentration in artery strips. These results suggest that the relaxation response to trapidil is not caused by prostacyclin release or an increase in cGMP in the smooth muscle, but possibly by an increase in the cAMP levels, probably via an inhibitory effect on cAMP phosphodiesterase.

Evaluation of platelet function by kinetic analysis of platelet aggregation.

Platelet aggregation is usually measured by a spectroscopic method using an aggregometer. However, when the process of platelet aggregation is followed by the changes in light transmission of platelet-rich plasma (PRP), factors such as platelet deformation or changes in plasma permeability must be taken into consideration. Thus, a better method was developed in our laboratory for measuring platelet aggregation by chronologically counting non-aggregated single platelets, one of the simplest parameters of the aggregation process, using PRP to which adenosine diphosphate (ADP) was added.(1,2) In the present study, we attempted to further evaluate platelet aggregability by measuring the velocity of platelet aggregation in different subjects using our single platelet counting method.

[Clinical usefulness of the measurement of plasma D-dimer levels].

To evaluate the clinical usefulness of D-dimer, various effects on the measurement of D-dimer were examined. Although both fibrinolytic and fibrinogenolytic products were detected by the measurement of FDP, only fibrinolytic products were detected by the measurement of D-dimer. In patients with DIC and other thrombo-embolic diseases, plasma D-dimer levels were significantly higher than in normal persons. A significant positive correlation between plasma D-dimer and serum FDP was found in DIC patients. In patients with DIC associated with acute promyelocytic leukemia, which is thought to be an increased fibrinogenolysis state, serum FDP was higher than the plasma D-dimer which suggests that increased fibrinogenolysis affects the result of serum FDP measurement. Plasma D-dimer significantly increased 5 minutes after endoscopic embolization with thrombin in the patients with esophageal varices. However serum FDP increased 30 minutes after the treatment, which suggests that the D-dimer is more useful for rapid detection of coagulo-fibrinolytic change than serum FDP. Plasma D-dimer was significantly higher in patients with cerebral infarction and increased with age. These finding suggest the usefulness of plasma D-dimer measurement for the specific and rapid evaluation of coagulo-fibrinolytic activation and thrombo-embolic state.