N Akamatsu

Tokyo Metropolitan Institute of Medical Science, Edo, Tōkyō, Japan

Are you N Akamatsu?

Claim your profile

Publications (20)61.03 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: A defective platelet glycoprotein (GP) Ib/IX/V complex [von Willebrand factor (VWF) receptor] results in Bernard-Soulier syndrome (BSS), which is characterized by macrothrombocytopenia and impaired ristocetin- and thrombin-induced platelet aggregation. We found 2 independent BSS-variant families: Case I [compound heterozygous mutations, p.Glu331X and a frame shift by a deletion at c.1444delA of GPIbα (GP1BA) terminating at a premature stop codon (p.Thr452ProfsX58)], and case II [homozygous nonsense mutation at c.1723C>T, p.Gln545X]. Case I platelets expressed no GPIbα, resulting in absence of ristocetin-induced platelet aggregation (RIPA) and 50% reduction in thrombin-induced aggregation with no shape change. The mother's platelets had 50% the expression level of A-type GPIbα (4-repeated VNTR: variable number of tandem repeats, p.[Thr145Met; Ser399_Pro411[4]]); the father's platelets had the same expression level of C-type GPIbα (2-repeated VNTR, p.Ser399_Pro411dup) as the mother's platelets. The mother's RIPA was significantly higher than the father's. Thrombin-induced aggregation was normal in both parents. Case II platelets expressed a GPIbα with an abnormal cytoplasmic tail, p.Gln545X-truncated GPIbα, which complexed with GPIX and GPV on the cell surface; its expression level of the complex was normal. Case II platelets had reversible RIPA, with no ATP release, and weak thrombin-induced aggregation without shape change. These results suggest that a signaling process through the GPIbα cytoplasmic tail required for full platelet activation is defective in BSS variant case II and a length polymorphism of GPIbα is associated with a modified level of RIPA heterozygous BSS case I.
    Thrombosis Research 02/2013; · 3.13 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Flow cytometric studies demonstrated that the platelets from a patient with Bernard-Soulier syndrome expressed substantial amounts of both GPIX and GPV, although in reduced amounts.
    British Journal of Haematology 03/2008; 87(1):185 - 188. · 4.94 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Bone marrow transplantation (BMT) may be complicated by coagulation abnormalities. The present study evaluated whether platelets might be activated in patients who had undergone BMT without significant coagulopathy. The patients selected had received allogeneic BMTs a median of 39 months before the study (range, 11-124 months) and had not received cyclosporine, FK506 (tacrolimus), or other medication affecting cyclo-oxygenase for at least 3 months prior to the collection of blood samples. Furthermore, patients had platelet counts greater than 100 x 10(9) cells/L and normal serum creatinine levels. Twenty-five healthy volunteers acted as controls. Platelet aggregation studies and a mepacrine assay of platelets showed abnormal aggregation and decreased staining in some patients. The platelet storage-pool adenosine 5'-triphosphate (ATP) level in 15 patients after BMT was 0.45+/-0.24 micromol per 10(11) platelets, whereas the level in 18 controls was 1.03+/-0.36 micromol per 10(11) platelets (P = .00078). The total ATP levels of platelets in patients and controls were 4.33+/-1.14 and 5.63+/-1.51 micromol per 10(11) platelets, respectively (P = .016). With the exception of 1 patient, plasma levels of thrombomodulin and von Willebrand factor were all within the normal range. The average plasma level of 11-dehydrothromboxane B2 was significantly increased in 15 patients after BMT compared with controls, 20.6+/-8.2 and 10.3+/-1.2 pg/mL, respectively (P = .0004). These findings suggest a long-term process of platelet activation in patients after BMT and, following the cessation of cyclosporine, development of acquired storage-pool disorder of platelets.
    International Journal of Hematology 09/2001; 74(2):222-7. · 1.68 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: CD36, a multifunctional adhesive receptor on a variety of cells such as monocytes and platelets, has been implicated in clearance of modified LDL and in the removal of apoptotic or senescent cells. We recently developed a new anti-CD36 monoclonal antibody, GS95. We determined the binding site of phosphatidylserine (PS)-liposome on CD36 by flow cytometric analysis of competitive bindings between phospholipid-liposomes or synthetic CD36 peptides and FITC-labeled anti-CD36 antibodies (GS95, OKM5, and FA6-152). The epitope of GS95 was mapped to the amino acid sequence #162-183 of CD36 that was partially overlapped with, but distinct from, #155-183, which has been reported as the epitopes of two commercially available antibodies, OKM5 and FA6-152. Oxidized-LDL dose-dependently inhibited bindings of both GS95 and OKM5 antibodies to platelet CD36, while PS-liposome inhibited the binding of GS95 but not OKM5 or FA6-152. These results indicate that the binding site of PS-liposome on platelet CD36 is not identical to that of oxidized-LDL and may be located in the amino acid sequence #162-183.
    Thrombosis Research 04/2000; 97(5):317-26. · 3.13 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Using immunogold electron microscopy, we found that human neutrophilic sialyl Lewis x (sLe(x)), an adhesive ligand for selectins, detectable by a monoclonal antibody, KM-93, is present in the sacculi of the Golgi apparatus as well as on the membranes of large electron-lucent azurophilic granules and the plasma membrane, including surface projections and microvilli. Neutrophilic sLe(x), however, was not detected on the membranes of specific granules. In comparison with the distribution of sLe(x), CD18 was localized on the plasma membrane and specific granule membrane but not on the azurophilic granule membrane. We also found by immunogold electron microscopy and flow cytometry that treatment of neutrophils with sialidase resulted in a loss of sLex on the plasma membrane. In contrast, intracellular sLex on the azurophilic granule membrane was not destroyed by sialidase. When sialidase-treated neutrophils were stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP), an inflammatory mediator peptide, in the presence of cytochalasin B, we observed by immunogold electron microscopy and flow cytometry that sLe(x) again appeared on the plasma membrane. These results indicate that stimulation by fMLP induces the up-regulation of sLe(x) on the cell surface by promoting translocation of sLe(x) from the azurophilic granule membrane to the plasma membrane in human neutrophils.
    Journal of Electron Microscopy 02/2000; 49(2):359-70. · 1.44 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: CD36, a multifunctional adhesive receptor on a variety of cells such as monocytes and platelets, has been implicated in clearance of modified LDL and in the removal of apoptotic or senescent cells. We recently developed a new anti-CD36 monoclonal antibody, GS95. We determined the binding site of phosphatidylserine (PS)-1iposome on CD36 by flow cytometric analysis of competitive bindings between phospholipid-liposomes or synthetic CD36 peptides and FITC-1abeled anti-CD36 antibodies (GS95, OKM5, and FA6-152). The epitope of GS95 was mapped to the amino acid sequence #162–183 of CD36 that was partially overlapped with, but distinct from, #155–183, which has been reported as the epitopes of two commercially available antibodies, OKM5 and FA6-152. Oxidized-LDL dose-dependently inhibited bindings of both GS95 and OKM5 antibodies to platelet CD36, while PS-liposome inhibited the binding of GS95 but not OKM5 or FA6-152. These results indicate that the binding site of PS-liposome on platelet CD36 is not identical to that of oxidized-LDL and may be located in the amino acid sequence #162–183.
    Thrombosis Research - THROMB RES. 01/2000; 97(5):317-326.
  • [Show abstract] [Hide abstract]
    ABSTRACT: We studied the preventive effect against allergies in infants who and whose mothers consumed hypoallergenic formulas until 6 months after birth. Mother and infant pairs were divided into three groups, and the infants were monitored for the development of allergies for the first 2 years. In the MD group (n = 102; n = number of infants), the mothers were given a hypoallergenic formula for mothers (MOM HA), which contained hydrolyzed whey protein as the only protein source, as a substitution for cow's milk during late pregnancy and lactation. In the CD group (n = 127), the mothers were given cow's milk during the corresponding period. All infants in the MD and CD groups were exclusively breast-fed or mixed-fed with breast milk and hypoallergenic infant formula (NAN HA), which contains the same hydrolyzed protein as MOM HA. In the AF group (n = 54), the mothers consumed MOM HA and their infants were mixed-fed with breast milk and a cow's milk-based adopted infant formula during the corresponding period. In the MD group, no infants were positive to cow's milk-specific immunoglobulin E (RAST) at 4 months of age, in contrast to 6% and 3% of infants in the CD and AF groups, respectively. The infants in the MD group showed low incidence of various allergies, especially of eczema, as compared to the CD and AF groups. These results suggest that consumption of cow's milk by mothers and cow's milk-based formula feeding to infants elevate the risk of allergies in infants, and that consumption of hypoallergenic formula for pregnant and lactating women and for infants could be helpful in preventing allergy development in infants.
    Journal of Nutritional Science and Vitaminology 07/1997; 43(3):397-411. · 0.99 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Platelet integrin alpha IIb beta 3 (GPIIb-IIIa) plays important roles in platelet-mediated clot retraction. However, little is known about the mechanisms of clot retraction mediated by nucleated cells. In this report, we demonstrate that another member of the beta 3 integrin family, alpha v beta 3, is involved in clot retraction mediated by nucleated cells. Retraction of fibrin clots was observed using a human melanoma cell line, C32TG, which contains no alpha IIb beta 3 complex. This retraction was inhibited by RGD-containing peptide, monoclonal anti-beta 3, and anti-alpha v beta 3 antibodies. Immunoelectron microscopic studies revealed a direct interaction between beta 3 integrin and fibrin fibers at an early stage of clot retraction. We found that another human embryonal cell line, 293, which is known to express alpha v beta 1, but no alpha v beta 3, lacks fibrin gel retractile activity. Upon transfection of beta 3 DNA into 293 cells, the beta 3 subunit formed a complex with an endogenous alpha v subunit. The beta 3-bearing transfectants were found to retract fibrin gels, which was specifically inhibited by anti-beta 3 antibody. In addition, a point mutation at Asp119 in the beta 3 ligand binding domain abolished the clot retractile activity of 293 transfectants, indicating the requirement of alpha v beta 3 ligand-binding activity. Our findings suggest that alpha v beta 3 is involved in mediating the interaction between the three-dimensional fibrin network and nucleated cells and in promoting "post-receptor occupancy" events.
    Journal of Biological Chemistry 02/1995; 270(4):1785-90. · 4.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Platelet glycoprotein VI (GPVI), a 62kD membrane protein, has been identified as one of the platelet receptors for collagen, since GPVI-deficient platelets exhibit abnormal responses to collagen and an abnormal bleeding tendency. We report a female patient with a mild bleeding history whose platelets expressed 10% GPVI of normal platelets. Shape change, aggregation and ATP release of the patient's platelets were completely absent in response to 1-5 micrograms/ml collagen but present normally in response to ADP and Ca2+ ionophore A23187. Adhesion of the patient's platelets to coated collagen was mildly affected (40-60% of normal platelets) in spite of only 10% expression of GPVI. Flow cytometrical studies revealed that the patient's platelets expressed normal amounts of the GPIa/IIa complex. These results suggest that platelet GPVI is less involved in adhesion to collagen than shape change and aggregation induced by collagen.
    British Journal of Haematology 02/1995; 89(1):124-30. · 4.94 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Platelet integrin αβ3 (GPIIb-IIIa) plays important roles in platelet-mediated clot retraction. However, little is known about the mechanisms of clot retraction mediated by nucleated cells. In this report, we demonstrate that another member of the β3 integrin family, αvβ3, is involved in clot retraction mediated by nucleated cells. Retraction of fibrin clots was observed using a human melanoma cell line, C32TG, which contains no αβ3 complex. This retraction was inhibited by RGD-containing peptide, monoclonal anti-β3, and anti-αvβ3 antibodies. Immunoelectron microscopic studies revealed a direct interaction between β3 integrin and fibrin fibers at an early stage of clot retraction. We found that another human embryonal cell line, 293, which is known to express αvβ1, but no αvβ3, lacks fibrin gel retractile activity. Upon transfection of β3 DNA into 293 cells, the β3 subunit formed a complex with an endogenous αv subunit. The β3-bearing transfectants were found to retract fibrin gels, which was specifically inhibited by anti-β3 antibody. In addition, a point mutation at Asp in the β3 ligand binding domain abolished the clot retractile activity of 293 transfectants, indicating the requirement of αvβ3 ligand-binding activity. Our findings suggest that αvβ3 is involved in mediating the interaction between the three-dimensional fibrin network and nucleated cells and in promoting “post-receptor occupancy” events.
    Journal of Biological Chemistry 01/1995; 270(4):1785-1790. · 4.65 Impact Factor
  • British Journal of Haematology 06/1994; 87(1):185-8. · 4.94 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Platelet membrane glycoprotein (GP) IV (also called CD36 and GPIIb) deficiency is associated with N(aka)-negative platelets. Using flow-cytometric analysis of cells stained with the monoclonal anti-GPIV antibody OKM5, we have studied the expression of GPIV on the monocytes from 16 healthy Japanese individuals whose platelets were deficient in GPIV. GPIV was absent on the surface of monocytes from 2 platelet GPIV-negative donors (type I), whereas it was present on the monocytes from the remaining 14 platelet GPIV-negative donors (type II). The fluorescent intensity of OKM5-stained type II monocytes was significantly (P < .05) lower than that of normal monocytes derived from platelet GPIV-positive donors, suggesting that the expression of GPIV on the type II monocytes is also abnormally regulated as compared with that on normal monocytes. OKM5 induced an oxidative burst in the type II monocytes as well as in the normal monocytes, but it failed to induce it in the type I monocytes. Because the 2 individuals with the type I deficiency have been healthy and exhibited no immunologic problems, GPIV appears to be not essential for the normal physiologic functions of monocytes. An anti-GPIV antibody was detected in the serum from one of the type I GPIV-deficient women, who had never received any blood transfusions but had given birth to three apparently healthy children. These results suggest that type I GPIV-deficient individuals may be at risk of developing an anti-GPIV isoantibody upon blood transfusion or pregnancy.
    Blood 01/1994; 83(2):392-7. · 9.78 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Changes in platelet membrane glycoproteins (GPIb, GPIIb/IIIa, and GMP-140) were evaluated using flow cytometry after binding with monoclonal antibodies in 22 adult patients undergoing cardiopulmonary bypass (CPB) surgery. The amount of GPIb on platelets decreased significantly during CPB, reaching a minimum level of 64 +/- 26% of the pre CPB value at 120 min of CPB. There was no significant change in the amount of GPIIb/IIIa on platelets. In accordance with these changes, ristocetin induced agglutination decreased to 56.7 +/- 16.2% of the pre CPB value during CPB. However, there were no significant changes in ADP and collagen induced aggregation throughout the procedure. The number of the activated platelets expressing GMP-140 on their surfaces increased significantly during CPB. There was an upper limit to the amount of GMP-140 expression on each platelet in the circulating blood, suggesting that excessively activated platelets are removed from the circulation. The authors conclude that CPB reduces the amount of GPIb on platelets, which results in platelet dysfunction. In addition, removal of excessively activated platelets from the circulation may lead to thrombocytopenia after CPB.
    ASAIO Journal 01/1993; 39(3):M550-3. · 1.49 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: To detect in vivo activated platelets in humans as well as in animal models of thrombosis, we developed a new murine monoclonal antibody, 2T60, specific for activated human and rabbit platelets by immunizing with human thronibin-activated platelets. 2T60 (IgG(1) subclass) showed a great difference between binding to the thrombin-activated and resting human and rabbit platelets on ELISA and flow cytometer analysis. Immunoblotting analysis revealed that 2T60 reacted with a 130 or 106 kDa protein of human or rabbit platelets, respectively, only under non-reducing conditions. (125)I-labeled 2T60 inserted into the intermediate gel of CIE of solubilized human platelets was incorporated into a immunoprecipitation line. 2T60 immunoprecipitated a protein of 130 or 115 kDa from human or rabbit platelets, respectively, which had been activated and (125)I-surface-labeled. The N-terminal sequence of the affinity purified 2T60 antigen of human platelets was identical to that of GMP 140. There were differences in the carbohydrate chain content of GMP 140 between human and rabbit platelets. In experimental cerebral thrombosis of rabbits that had been injected with 2T60, the platelets adhering to the exposed subendothelium and contained in thrombi were found to bind 2T60 prominently. These results suggest that 2T60 may be a useful tool for clinical and experimental studies of thrombotic disease.
    Platelets 01/1993; 4(1):31-9. · 2.24 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Since glycoprotein IV (GPIV) has been shown to play an important role in the interaction of platelets with collagen and thrombospondin, the aggregation and secretion of GPIV-deficient platelets were examined. Using a binding assay with monoclonal 125I-OKM5 antibody against CD36 antigen and crossed immunoelectrophoresis of the solubilized platelets against anti-GPIV antibody, the platelets from seven (4.1%) out of 170 healthy Japanese donors were found to be deficient in GPIV. The GPIV-deficient platelets showed normal aggregations in response to collagen as well as ADP, epinephrine, arachidonic acid and thrombin in comparison with GPIV-positive platelets. Polyclonal anti-GPIV antibody aggregated GPIV-positive platelets but not the GPIV-negative ones. The F(ab')2 fragments of the anti-GPIV antibody competitively inhibited the anti-GPIV-induced aggregation, but did not affect the collagen-induced aggregation of GPIV-positive platelets. These results suggest that the deficiency of GPIV does not affect platelet aggregability.
    British Journal of Haematology 06/1992; 81(1):86-92. · 4.94 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Since glycoprotein IV (GPIV) has been shown to play an important role in the interaction of platelets with collagen and thrombospondin, the aggregation and secretion of GPIV-deficient platelets were examined. Using a binding assay with monoclonal 125I-OKM5 antibody against CD36 antigen and crossed immunoelectrophoresis of the solubilized platelets against anti-GPIV antibody, the platelets from seven (4.1%) out of 170 healthy Japanese donors were found to be deficient in GPIV. The GPIV-deficient platelets showed normal aggregations in response to collagen as well as ADP, epinephrine, arachidonic acid and thrombin in comparison with GPIV-positive platelets. Polyclonal anti-GPIV antibody aggregated GPIV-positive platelets but not the GPIV-negative ones. The F(ab')2 fragments of the anti-GPIV antibody competitively inhibited the anti-GPIV-induced aggregation, but did not affect the collagen-induced aggregation of GPIV-positive platelets. These results suggest that the deficiency of GPIV does not affect platelet aggregability.
    British Journal of Haematology 04/1992; 81(1):86 - 92. · 4.94 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: We have found that vascular injuries are induced by intravascular aggregation of platelets activated by arachidonic acid (AA) or ADP. The characteristic findings are the appearance of vacuoles in endothelial cells and eventual deendothelialization. In deendothelialized regions, formation of platelet thrombi was observed. The platelets in the thrombi were stained with 2T60, a monoclonal antibody that recognizes activated platelets. The change was more remarkable in the AA-injected animals because AA has a stronger platelet activating effect and a detergent effect on the endothelium. The ADP-injection experiments clarified the role of platelets in vascular injury. These findings suggest that activated platelets play a role in the genesis of vascular injuries, and that their role is related to thrombus formation.
    Japanese Circulation Journal 03/1992; 56(2):178-86.
  • [Show abstract] [Hide abstract]
    ABSTRACT: The clinical significance and detection methods for circulating activated platelets are reviewed. It has been recently demonstrated that three kinds of platelet granules, dense granule, alpha-granule and lysosome, have specific membrane proteins; granulophysin, GMP-140 (PADGEM) and CD63 antigen, respectively, and that these specific granule-membrane proteins become exposed on the surface of the activated platelets. It is believed that detection of circulating activated platelets, using monoclonal antibodies specific to these granule-membrane, may be reliable and diagnostic value in thrombotic and prethrombotic diseases. Preliminary clinical studies using 2T60, a newly developed anti-GMP-140 monoclonal antibody, was presented.
    Nippon rinsho. Japanese journal of clinical medicine 03/1992; 50(2):330-5.
  • Nippon Ketsueki Gakkai zasshi: journal of Japan Haematological Society 01/1988; 50(8):1714-22.
  • Thrombosis Research 65:S41. · 3.13 Impact Factor