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Biomedical subjects

M Kazama

Publications and source records attributed to M Kazama.

At least 37 records · Page 2Linked to original sources
PubMed1993

Nafamostat mesilate: a regional anticoagulant for hemodialysis in patients at high risk for bleeding.

107 hemodialysis patients at high risk for intradialytic bleeding due to previous surgery or active bleeding from other sites were treated with nafamostat mesilate (FUT-175; FUT) as hemodialysis anticoagulant for 2 weeks. In contrast to heparin. FUT prolonged clotting times only in the extracorporeal circuit. Clotting times were not prolonged even at the conclusion of the treatment, and bleeding from the puncture site after removal of the needle was shorter than with heparin. The exacerbation of bleeding by hemodialysis was noted in only 21 out of 573 hemodialysis procedures (3.7%), and 134 of 145 hemodialysis procedures (92.4%) with active bleeding were successfully completed without increasing the bleeding. Adverse effects of FUT were noted in only 6 cases (5.6%) or 1.2% of HD procedures. These results indicate that FUT is a very useful anticoagulant for HD, especially in patients with high risk of bleeding.

Aged↗
PubMed1992

Participation of calpain I activation in the ATP release reaction of platelets stimulated with thrombin.

Proteolytic activation of calpain (calcium-dependent neutral protease) I in thrombin-stimulated platelets was determined by following the production of the 76- and 78-kDa forms from the 80-kDa subunit of calpain I as measured by immunoblotting using monospecific antibody to human calpain I, and the correlation between the extents of calpain I activation and ATP release was investigated. When platelets were stimulated with thrombin in the range from 0.01 to 0.5 U/ml, the maximal 60% activation of calpain I was achieved within 15 s after the stimulation, and ATP release began after the maximal activation had been reached. The extent of ATP release decreased in parallel with the decrease in activation ratio of calpain I on treatment of platelets with EGTA or EST, a membrane-permeable inhibitor of calpain. Although pretreatment of platelets with EST did not affect the thrombin-dependent elevation of the cytosolic Ca2+ concentration, both the inhibition of calpain I activation and the reduction of ATP release were observed as a function of EST concentration. These results suggest that calpain I participates in one of the processes leading to the ATP release reaction of platelets stimulated with thrombin.

Adenosine Triphosphate↗
PubMed1992

Retinoic acid counteracts both the downregulation of thrombomodulin and the induction of tissue factor in cultured human endothelial cells exposed to tumor necrosis factor.

Inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) shift the hemostatic balance of endothelial cell surfaces in favor of prothrombotic properties by downregulating thrombomodulin (TM) and inducing tissue factor (TF) expression. We investigated the effects of retinoic acid (RA) on the prothrombotic properties of cultured umbilical vein endothelial cells exposed to TNF-alpha. The approximate 50% downregulation of TM antigen and cofactor activity induced by TNF-alpha (10 U/mL for 24 hours) was completely prevented when the cells were coincubated with both TNF-alpha and 10 mumol/L RA. In accordance with changes in cell surface TM antigen levels, the 70% decrease in TM messenger RNA (mRNA) induced by TNF-alpha was also prevented by 10 mumol/L RA. TNF-alpha induced TF activity of lysed cells (100-fold greater than untreated controls), an effect prevented when the cells were coincubated with both the TNF-alpha and 10 mumol/L RA. The 34-fold increase in TF mRNA levels induced by TNF-alpha (10 U/mL for 3 hours) was only two-fold in the presence of both TNF-alpha and RA. The effects of RA on the regulation of TM and TF expression in the cells exposed to TNF-alpha was dose-dependent from 0.01 to 10 mumol/L RA. The present results suggest that RA may affect on the mRNA level to alter TM and TF expression, effectively counteracting expression of prothrombotic properties of endothelial cells induced by inflammatory cytokines such as TNF-alpha.

Cells, Cultured↗
PubMed1992

Procalpain I in cytoplasm is translocated onto plasma and granule membranes during platelet stimulation with thrombin and then activated on the membranes.

Thrombin stimulation of platelets resulted in changes in the subcellular localization of calpain I, with a concomitant alteration of its molecular weight as measured by immunoblotting. Calpain I in resting platelets was distributed as procalpain I, an 80 kDa form which does not exhibit the enzyme activity, and 83% of the total antigen was localized in the cytosol fraction. When platelets were stimulated with thrombin, the total content of calpain I antigen was not significantly changed as compared with that of the resting platelets, though a decrease in the cytosolic distribution of 80 kDa form (from 83% to 47% of the total antigen) was observed with concomitant appearance of the active 76 kDa and intermediate 78 kDa forms of calpain I and increase in the 80 kDa form in the granule and membrane fractions. These results indicated that calpain I was translocated from the cytosol to both the plasma and granule membranes as procalpain I and then activated on the membranes during platelet stimulation with thrombin.

Blood Platelets↗
PubMed1992

Activation of calpain I in thrombin-stimulated platelets is regulated by the initial elevation of the cytosolic Ca2+ concentration.

The source and concentration of Ca2+ required to activate calpain I were investigated in thrombin-stimulated platelets. The concentration of cytosolic free Ca2+ ([Ca2+]i) was measured in platelets containing fura-2-AM, and exhibited a biphasic response after stimulation with 0.05, 0.1 or 0.5 NIH units of thrombin/ml. An initial transient elevation, which was predominantly dependent upon Ca2+ released from the internal stores into the cytosol, peaked at 15 s after stimulation, and a secondary sustained elevation, which was due to Ca2+ influx, was observed following the initial elevation. Calpain I was present at about 540 ng/10(8) unstimulated platelets, as measured by immunoblotting using rabbit anti-(human calpain I) IgG. Calpain I was activated 10 s after thrombin stimulation, as determined by the appearance of the 78 kDa and 76 kDa forms on immunoblots. The activation ratio of calpain I was calculated as the amount of the 78 + 76 kDa forms as a percentage of the total (80 + 78 + 76 kDa), and was influenced by the extent of the initial transient [Ca2+]i elevation after stimulation. An initial increase in [Ca2+]i of 300 nM was required to achieve the maximal activation (60%) of calpain I, and half-maximal activation occurred at 160 nM- Ca2+]i. These results suggest that the activation of calpain I in platelets is regulated by the initial elevation in Ca2+]i after thrombin stimulation, and does not necessarily require a Ca2+ influx.

Adult↗
PubMed1992

Changes in plasma thrombomodulin antigen in rabbit developing endotoxin-induced disseminated intravascular coagulation and the effect of heparin.

Soluble thrombomodulin (TM) antigen level was 1.64 +/- 0.64 microgram/ml (n = 18, mean +/- S.D.) in plasma of normal male rabbits as measured by enzyme immunoassay, and the antigen consisted of subspecies of 94, 83 and 51 kd. When disseminated intravascular coagulation (DIC) was induced by intravenous infusion of endotoxin into rabbits, the TM antigen level in plasma was elevated to about 1.5 times of the control value, and an increase in the 83 kd subspecies as well as the appearance of new subspecies of 76 and 48 kd was observed concomitantly with disappearance of the 94 kd subspecies in plasma. Elevation of the antigen level and disappearance of the 94 kd subspecies caused by infusion of endotoxin were reduced by simultaneous infusion of heparin. Addition of leukocytes stimulated with endotoxin plus FMLP to cultured endothelial cells induced release of TM antigen to the medium accompanying cell injury as measured by 51Cr release, which was prevented by treatment with heparin. It was suggested that the increase in plasma TM antigen level in parallel with the generation of DIC reflected endothelial injury of rabbits, and that the elevation of TM antigen and the endothelial cell injury were prevented by heparin treatment.

Animals↗
PubMed1992

Changes in thrombomodulin level in plasma of endotoxin-infused rabbits.

Changes in the plasma thrombomodulin (TM) level were examined in endotoxin-infused rabbits. The plasma TM level in normal rabbits was 143.8 +/- 8.4 ng/ml (n = 67) and the molecular weight of the major TM was about 55 kd. Endotoxin (lipopolysaccharide, LPS, E. Coli B8:0127) was intravenously infused. LPS infusion increased the plasma TM level dose-dependently between 0.2 mg/kg and 5 mg/kg. When 5 mg/kg LPS was infused, the plasma TM level started to increase immediately and was 2.3 times higher than the control value within 1 hr. The molecular weight of the major TM was about 75 kd. This rapid increase in TM occurred before the decrease in fibrinogen content and the prolongation of prothrombin time. To examine the effect of circulating leukocytes on the TM increase in endotoxin-infused rabbits, 5 mg/kg LPS was infused into rabbits with leukocytopenia induced by X-ray irradiation. The maximum plasma level of TM was significantly lower than in the untreated rabbits given LPS. These data suggest that the increase in plasma TM is caused by LPS-stimulated leukocyte's prior to hemostaseological changes. It is well known that endothelial cells can be injured by stimulated leukocytes, so this increase in plasma TM probably reflects the deterioration of endothelial cells. This deterioration decreases the ability of endothelial cells to inhibit thrombosis, which would, in turn, contribute to the development of disseminated intravascular coagulation in endotoxin-infused rabbits.

Animals↗
PubMed1992

Retinoic acid stimulates expression of thrombomodulin, a cell surface anticoagulant glycoprotein, on human endothelial cells. Differences between up-regulation of thrombomodulin by retinoic acid and cyclic AMP.

Thrombomodulin (TM) is a surface protein on endothelial cells, and represents one of the most valuable regulatory factors in the anticoagulant system. In this paper, we demonstrate that retinoic acid (RA) causes an increase in TM antigen on human umbilical vein endothelial cells (HUVECs) in vitro. The effect of RA on the surface TM level of HUVECs was dose-dependent in the range from 0.01 to 10 microM-RA. Antigen levels began to increase 3 h after addition of 10 microM-RA, and plateaued at a maximum level of approx. 2.5 times that of the untreated control at 24 h. TM levels remained at a maximum for a further 12 h, and then gradually decreased. The effects of RA on cell surface TM activity and antigen levels were parallel in all experiments. TM expression was also increased by treatment with 10 microM-retinal or 10 microM-retinol for 24 h, though the increases were approx. 70% and 30% respectively of that produced by 10 microM-RA. Pretreatment of HUVECs with cycloheximide inhibited the effect of RA. When HUVECs were incubated with both 10 microM-RA and 5 mM-8-bromo cyclic AMP (or 1 mM-3-isobutyl-1-methylxanthine, a phosphodiesterase inhibitor), the increase in TM antigen was greater than that observed with either compound alone. Northern blot analysis showed that treatment of HUVECs with 8-bromo cyclic AMP, RA or RA plus 8-bromo cyclic AMP increased TM mRNA levels by 2.2-, 4.5- and 5.5-fold respectively compared with the untreated control. Furthermore, no significant difference in cellular cyclic AMP levels was observed between RA-treated and control cells. These results indicate that the expression of TM is not only controlled by the intracellular cyclic AMP level but is also affected by RA, and suggest that RA-induced up-regulation of TM on HUVECs is independent of cyclic AMP regulation.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1992

Clinical significance of the venous occlusion test on systemic lupus erythematosus patients with a focus on changes in blood levels of tissue plasminogen activator, von Willebrand factor antigen, and thrombomodulin.

We investigated the clinical significance of the venous occlusion (VO) test on patients with systemic lupus erythematosus (SLE) with or without circulating lupus anticoagulant (LA) concerning whether changes in the blood coagulation and fibrinolysis system in vivo subsequent to VO reflect mechanical stimulation of the endothelium or presence/development of endothelial damage. The tissue plasminogen activator antigen (tPA:Ag) before VO was much lower in the LA-positive patients than in the LA-negative ones (p < 0.01) and the von Willebrand factor antigen (VWF:Ag) pre-VO was significantly higher in the patient group, regardless of LA status, than in the control group (p < 0.01). But the mean increment in tPA:Ag and VWF:Ag post-VO, when expressed as the percentage of the baseline level, showed no appreciable difference between LA-positive and -negative groups. Thrombomodulin (TM) basically, on the other hand, was higher in the patients of either LA status than in the controls (p < 0.01) with a significant post-VO increase in the SLE group, which was more marked in the LA-positive patients, against no substantial change in the controls (p < 0.01). It is known that tPA and VWF:Ag are released simply as a result of endothelial stimulation and that the release of TM is preceded by endothelial damage. Based on the present results, we may well conclude that (1) the endothelium is functionally intact in SLE patients, (2) an injury of the endothelium, possibly as a consequence of vasculitis, preexists in LA-positive patients, and thus to measure the TM response to VO would offer a helpful tool in diagnosing the preexisting endothelial damage in these clinical settings.

Adult↗
PubMed1992

Effect of total-body cold exposure on plasma concentrations of von Willebrand factor, endothelin-1 and thrombomodulin in systemic lupus erythematosus patients with or without Raynaud's phenomenon.

The effect of total-body cold exposure on plasma concentrations of von Willebrand factor (vWF), endothelin-1 (ET) and thrombomodulin (TM), all of which are considered to be generated from the endothelium, was studied in systemic lupus erythematosus (SLE) patients with and without Raynaud's phenomenon. The plasma levels of vWF, ET and TM in SLE patients, irrespective of the presence of Raynaud's phenomenon, were significantly higher than in normal controls even before the cold provocation test. After the cold provocation test, plasma levels of vWF and ET were significantly higher in SLE patients with Raynaud's phenomenon than in those without and in normal controls. No significant increase in TM was observed in either the SLE patients or the controls. These results suggest that SLE patients, regardless of the presence of Raynaud's phenomenon, are in a hypercoagulable state and that this state may be further intensified by cold exposure. Hence, it is concluded that we should consider antithrombotic therapy for SLE patients, especially those with Raynaud's phenomenon, to prevent unwanted activation of the coagulation system and possible endothelial damage.

Adult↗
PubMed1992

[Improving effect of the synthetic protease inhibitor E-3123 on experimental DIC in dogs].

Disseminated intravascular coagulation (DIC) is a severe syndrome associated with generalized, intractable bleeding and multiple organ failure. Synthesized protease inhibitors such as gabexate mesilate and nafamostat mesilate show an improving effect on DIC, which develops by a chain reaction involving the coagulation, fibrinolysis, complement and kallikrein systems. Experimental DIC was developed in Beagle dogs by infusion of 150 U/kg tissue thromboplastin (Group I), and the improving effect of a new synthetic protease inhibitor, E-3123, was examined. The following groups of animals were treated with drugs: Group II (n = 4) was given with 5 mg/kg/hr of E-3123; group III (n = 4) was given 10 mg/kg/hr of E-3123; and group IV was given 6 mg/kg/hr of gabexate mesilate (GM). Although improvement of the hemodynamics or peripheral circulation was not apparent, a slight, but insignificant, improvement of lactate/pyruvate was noted in the treated groups. On the other hand, the hemostatic abnormalities such as prolongation of prothrombin time and activated thromboplastin time; decreases of platelet count, fibrinogen and alpha 2-antiplasmin; and increases of fibrin degradation products were significantly improved in the treated groups. These results indicate that E-3123 is effective for improving experimental DIC, and it is suggested that E-3123 is applicable for the treatment of clinical DIC.

Animals↗
PubMed1992

Plasma thrombomodulin and alpha 2-plasmin inhibitor-plasmin complex are elevated in active systemic lupus erythematosus.

Plasma levels of thrombomodulin and alpha 2-plasmin inhibitor-plasmin complex were measured by ELISA in patients with rheumatic diseases. Thrombomodulin levels in patients with active systemic lupus erythematosus (SLE) were significantly higher than those in patients with inactive SLE or in healthy controls. This suggests that thrombomodulin, normally a component of vascular endothelial cell membrane, is easily released to plasma in patients with active SLE. High titers of the thrombomodulin level and the correlated alpha 2-plasmin inhibitor-plasmin complex elevations imply vascular injury, and consequently, excessive fibrinolytic processes in active SLE.

Adolescent↗
PubMed1992

[Intracellular Ca2+ behavior and activation of calpain-I in activated platelets].

A relationship between intracellular Ca2+ concentration ([Ca2+]i) and calpain-I activation and change in subcellular localization of the enzyme in activated platelets were investigated. The [Ca2+]i exhibited a biphasic response after stimulation with thrombin. Activation of calpain-I was measured by determination of the appearance of active 76 and 78 kDa forms accompanying the disappearance of the 80 kDa form, the inactive form, on immunoblots. Calpain-I was activated dependent on the extent of the initial elevation of [Ca2+]i. For maximum activation (60%) 300-500 nM [Ca2+]i was required and half-maximal activation occurred at 160-220 nM [Ca2+]i. The active 76 kDa form was observed only in the fraction containing subcellular organelles and plasma membrane of activated platelets. It was demonstrated that the localization of calpain-I was changed from the cytosol to the membrane and calpain-I was activated on the membrane by Ca2+, elevated through the initial elevation after activation of platelets.

Calcium↗