Yasuhiko Ando

Ultrasonographic differentiation between tuberculous lymphadenitis and malignant lymph nodes

To assess the usefulness of ultrasonography in the differential diagnosis of cervical tuberculous lymphadenitis versus malignant lymph nodes.

Importance of fibrinogen and platelet membrane glycoprotein IIb/IIIa in shear-induced platelet aggregation

The mechanism of shear-induced platelet aggregation was investigated using a polycarbonate cone and plate viscometer. After exposed to shear stress of 54-90 dyne/cm2 for 2 min. at 37 degrees C, platelets aggregated without a significant amount of serotonin release and lactic dehydrogenase leakage from platelets. Under this conditions, platelets from 2 patients with thrombasthenia and a patient with congenital afibrinogenemia failed to aggregate. When fibrinogen was added to platelet rich plasma from a patient with afibrinogenemia, shear-induced platelet aggregation occurred at the same extent of aggregation as observed in normal platelets. Shear-induced platelet aggregation was inhibited by monoclonal antibody to GPIIb/IIIa (1 microgram/ml) and synthetic peptide, Arg-Gly-Asp-Ser (RGDS) (1 mM). Apyrase and hirudin showed no effect on this aggregation. Indomethacin (100 microM) and thromboxane A2 synthetase inhibitor, OKY-046 (100 microM) markedly inhibited aggregation, while thromboxane A2 competitive inhibitor, ONO-3708 (100 microM) exhibited only partial inhibition. These results indicate that fibrinogen and GPIIb/IIIa are important for shear-induced platelet aggregation and that the induction of fibrinogen receptor on GPIIb/IIIa may partially depend upon thromboxane A2 synthesis in platelets.

On the mechanism of heparin-induced potentiation of platelet aggregation.

The role of antithrombin III (AT III) in heparin-induced potentiation of platelet aggregation was investigated using purified AT III and AT III depleted plasma. When ADP or epinephrine was added to citrated platelet rich plasma (PRP) one minute after addition of heparin, marked enhancement of platelet aggregation was observed, compared with the degree of platelet aggregation in the absence of heparin. However, heparin exhibited no potentiating effect on ADP- or epinephrine-induced platelet aggregation when platelets were resuspended in AT III depleted plasma prepared by immunosorption using matrix-bound antibodies to AT III. When purified AT III was added to AT III depleted plasma at a concentration of 20 microgram/ml, potentiation of platelet aggregation by heparin was clearly demonstrated. These results suggest that the effect of heparin on platelet aggregation is also mediated by AT III.

Inhibition by endothelial cells of platelet aggregating activity of thrombin - role of thrombomodulin

Cultured human umbilical vein endothelial cells inhibited the platelet aggregating activity of thrombin in the absence of plasma. Using a new method in which thrombin-induced platelet aggregation was measured in the presence of endothelial cells, we showed that aspirin-treated endothelial cells inhibited platelet aggregating activity of thrombin in an incubation time- and cell number-dependent manner. This inhibitory effect of endothelial cells was partially blocked by the pretreatment of endothelial cells with monoclonal anti-thrombomodulin IgG (anti-TMIgG). It is suggested that (1) endothelial cells play a role in the clearance of thrombin by binding and inactivating this enzyme and that (2) thrombomodulin on endothelial cells, apart from its role for protein C activation, may be involved in this endothelial function.

Detection of hypercoagulability by the measurement of the dynamic loss modulus of clotting blood

The dynamic loss modulus of clotting whole blood was measured in thrombotic patients to characterize the physical properties of coagulation in the hypercoagulable state. The dynamic loss modulus was measured by a Sonoclot. Thrombotic patients consisted of 30 with deep vein thrombosis and 25 with arterial thrombosis. An accelerated increment rate of the dynamic loss modulus at the beginning of gelling was the characteristic of hypercoagulability. This characteristic occurred more frequently than other abnormalities in other tests for hypercoagulability (beta-thromboglobulin, antithrombin III and TEG). Only in deep vein thrombosis, a moderately positive correlation was noted between the increment rate of the dynamic loss modulus and the plasma fibrinogen level.

Quantitative Assay of Hepatitis C Virus RNA Using an Automated Extraction System for Specific Capture with Probes and Paramagnetic Particle Separation

ABSTRACT A commercially available automated specimen preparation instrument for specific probe capture and paramagnetic separation has been developed (AmpliCap/GT-12; Roche Molecular Systems). We evaluated assay performance of the AmpliCap/GT-12 in the quantitative assay for hepatitis C virus (HCV) RNA with the AMPLICOR HCV MONITOR Test (version 2.0). Assay linearity using serial dilutions from a serum panel was observed in the range of 500 to 850,000 IU/ml, with a slightly compromised slope in the higher viral titers. The overall within-run and between-run reproducibility of the entire detection process for 3 and 5 log10 (IU/ml) of HCV RNA in samples had a standard deviation of <0.2, which was comparable to a manual method based on organic extraction and isopropanol precipitation (Roche Molecular Systems). Comparison of the test results with those obtained by the manual method showed a good correlation (R2 = 0.972, n = 86). Using heparin (3, 6.5, and 13 U/ml), dextran sulfate (0.1, 1, and 5 mM), hemoglobin (1.13, 2.25, and 4.5 g/liter), conjugated or unconjugated bilirubin (7.5, 15, and 30 mg/dl), and ATP (1.25, 2.5, and 5.0 mM) as known inhibitors, inhibition was only detected at a dextran sulfate concentration of 1 mM with the manual method but not with the AmpliCap/GT-12 extraction. In summary, the AmpliCap/GT-12 system was shown to permit a stable extraction process and accurate results for the quantitative assay of HCV RNA, successfully eliminating the inhibitory effect of dextran sulfate. This automated extraction system provides reliable and reproducible test results and saves labor; thus, it is suitable for routine diagnostic PCR.

The role of calcium in platelet microtubule assembly-disassembly

Abstract The role of calcium in the equilibrium between assembled and disassembled microtubules was investigated in human platelets. The two pools of microtubule protein was “frozen” by addition of a glycerol-dimethyl sulfoxide-containing medium and were estimated by measuring the colchicine binding activities of total and polymerized tubulin. Addition of A23187 to platelet rich plasma produced a transient decrease in the pool of polymerized tubulin within 30 sec., followed by a return to base-line values within 2 min.. Total content of platelet tubulin remained unchanged during the course of aggregation. TMB-8, a known intracellular calcium antagonist, abolished this transient decrease in polymerized tubulin induced by A23187, while indomethacin did not. These findings strongly suggest the important role of intracellular calcium in microtubule assembly-disassembly.

Inhibition of Platelet Function by Sulbenicillin and Its Metabolite

The effect of sulbenicillin and its major metabolite, α-sulfobenzylpenicilloic acid, on platelet function was investigated. Sulbenicillin caused inhibition of platelet aggregation and release reaction in the same manner as carbenicillin. α-Sulfobenzylpenicilloic acid was found to cause much stronger inhibition of platelet function. The results indicate that the strong inhibitory action of α-sulfobenzylpenicilloic acid may also take part in impaired platelet functions following administration of sulbenicillin to humans.

Extensive Wall Thickening in Intestinal Burkitt Lymphoma

Objective. To evaluate intestinal lesions in Burkitt lymphoma. Methods. Ultrasonography was used in the initial evaluation of 6 Japanese patients with intestinal Burkitt lymphoma. Results. Ultrasonography revealed marked wall thickening of the colon from the cecum through either the ascending or the transverse colon, which led to a target sign (4 cases) or a pseudokidney sign (2 cases). The target sign histopathologically corresponded to invagination of an occult tumor of the cecum into the ascending colon. Wall thickening of the colon when associated with a target or pseudokidney sign corresponded to marked lymph edema or a diffuse infiltration of Burkitt lymphoma cells into the intramural layers, respectively. Conclusions. Ultrasonography provides useful information in the initial evaluation of intestinal lesions with distinctive histopathologic characteristics in Burkitt lymphoma.

Altered cytoskeletal structures of thrombasthenic platelets

The changes in cytoskeletal structures of thrombasthenic platelets were investigated following thrombin activation. Non-stimulated platelets from normal subjects and thrombasthenic patients yielded essentially the same amount of Triton-precipitated cytoskeletal proteins. When platelets were treated with thrombin at various concentrations, the amount of protein in Triton-precipitates was increased both in normal and thrombasthenic subjects; less degree in the latter. The Triton-insoluble materials from non-stimulated, normal and thrombasthenic platelets showed essentially similar polypeptide composition on SDSpolyacrylamide slab gel electrophoresis; three prominent bands of molecular weight 43,000, 200,000 and 250,000 respectively. A distinct difference was, however, observed in polypeptide composition of cytoskeletons between thrombinstimulated normal and thrombasthenic platelets. While polypeptides of molecular weight 43,000, 52,000, 56,000, 200,000 and 250,000 were increased after control platelets were stimulated with thrombin, cytoskeletal proteins of thrombinactivated thrombasthenic platelets showed no increase in the amount of polypeptides of molecular weight of 52, 000 and 56, 000. These findings may suggest the altered cytoskeletal structures of thrombasthenic platelets.