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Biomedical subjects

S Kume

Publications and source records attributed to S Kume.

At least 19 recordsLinked to original sources
PubMed1998

A cross-sectional evaluation of spontaneous platelet aggregation in relation to complications in patients with type II diabetes mellitus.

To clarify the relationship between platelet function and diabetic complications, we investigated spontaneous platelet aggregation (SPA) and agonist-induced platelet aggregation by a particle counting method using light scattering (LS) and by a conventional light transmission method (LT) in 23 age- and sex-matched control subjects and 74 patients with type II diabetes mellitus. We also observed platelets using the FIC-2 (TOA Medical Electronics, Kobe, Japan) flow cytometer and imaging device. Observation by the FIC-2 device showed microaggregates of platelets in samples with increased SPA-LS. SPA-LS was significantly elevated in patients with type II diabetes mellitus as a whole compared with control subjects. SPA-LS also showed significant differences between control subjects and three diabetic patient subgroups with a varying severity of retinopathy, nephropathy, or neuropathy, and the mean values increased along with the increasing severity of complications. On the other hand, although SPA-LT also showed significant differences between these groups, the absolute values were all less than 10%, which we believe does not warrant quantitative analysis. Adenosine-5'-diphosphate (ADP)-induced platelet aggregation failed to show significant differences between controls and subjects with a varying severity of retinopathy by either LS or LT, which indicates that SPA is more sensitive than agonist-induced platelet aggregation in relation to diabetic complications. We observed significant correlations between SPA-LS and the patients' age, hemoglobin A1c (HbA1c) level, plasma fibrinogen level, or 6-keto-PGF1alpha (6KF) to 11-dehydro-thromboxane B2 (TXB2) ratio. Our study demonstrated a close relationship between platelet hyperaggregability and diabetic complications, and a longitudinal prospective study of SPA-LS in diabetic patients is warranted to clarify cause-and-effect relationships.

Cross-Sectional Studies↗
PubMed1998

Quantification of sphingosine derivatives in human platelets: inducible formation of free sphingosine.

To elucidate the physiologic role of sphingolipid-derived products as signaling molecules, we analyzed the levels of endogenous sphingosine (Sph) derivatives in human platelets. When the platelets were stimulated with thrombin or 12-O-tetradecanoylphorbol 13-acetate, neither ceramide formation nor sphingomyelin hydrolysis was observed, which suggests that the sphingomyelin cycle may not be an essential part of the signaling pathway under these conditions. In contrast, Sph was found to increase in platelets upon stimulation. The level of Sph 1-phosphate, which is formed from Sph by the action of Sph kinase, was not affected under our conditions. Although it has been established that Sph inhibits protein kinase C, which regulates the functional responses of the platelets, Sph levels which exert an inhibitory effect on protein kinase C cannot be attained under physiological conditions (without exogenous Sph). Considering the stimulation of the synthesis of Sph by the physiological agonist thrombin, we speculate that Sph is a signaling molecule of physiological importance in platelets, but protein kinase C may not be its target.

Adult↗
PubMed1998

Effect of restricted feed intake of dams and heat stress on mineral status of newborn calves.

To clarify the effects of restricted feed intake, heat stress, and parity on the mineral status of cows and heifers around parturition and on the mineral status of their calves during 1 wk of age, data were collected from 66 Holstein cows and heifers and their calves. In Experiment 1, 36 heifers and mature cows that calved during hot or cool weather were fed to meet requirements for total digestible nutrients (TDN), protein, and minerals. In Experiment 2, 15 mature cows that calved during hot or cool weather were fed to meet maintenance requirements for TDN plus requirements for TDN for the last 2 mo of gestation, and 15 heifers were fed to meet TDN requirements. Heat stress increased rectal temperatures of newborn calves. Blood hematocrit and hemoglobin of heifers around parturition were higher than those of mature cows, but blood hematocrit and hemoglobin of calves born from heifers were lower. The restricted feed intake of dams decreased blood hematocrit and hemoglobin as well as plasma Fe of calves in hot and cool weather. Plasma Ca, inorganic P, and alkaline phosphatase as well as colostral Ca, P, Mg, and Zn of heifers were higher than those of mature cows, but plasma Mg of heifers was lower. Plasma Mg of calves and their dams was lower in hot weather than in cool weather, and restricted feed intake accelerated the reduction in plasma Mg of calves and their dams during hot weather. Plasma Na of calves and their dams was higher in hot weather than in cool weather. Heat stress increased plasma K of heifers and their calves. Heat stress increased Ca concentration in meconium of calves born from cows, and the restricted feed intake increased P concentrations in meconium. These results suggest that the maintenance of optimum erythropoiesis and mineral status in heatstressed periparturient cows and heifers and their calves must be met by dietary energy and minerals that are fed at maintenance concentrations plus excess requirements necessary during the gestation period.

Animals↗
PubMed1998

[Immunohistochemical study on keratin no. 8, 18 and 19 expression of carcinoid tumors].

The purpose of this study is to evaluate keratin expression in neuroendocrine cells and carcinoid tumor of the gastrointestinal tract and lung. Neuroendocrine cells in the lung and the gastrointestinal tract reacted with low-molecular keratin strongly, but not with high-molecular keratin. The low-molecular keratin expression of carcinoid tumor in the lung and the gastrointestinal tract, which varied in degree, were identical to that of neuroendocrine cells. Keratin 8 and 18 were found abundantly in neuroendocrine tumor and carcinoid tumor of the gastrointestinal tract, and in a lesser amount in carcinoid tumor of the lung, whereas keratin 19 was found in all cases with variable amounts. Based on these results, the combination of antibodies for several types of keratin is useful for examining keratin expression as a marker of epithelium in carcinoid tumor.

Adult↗
PubMed1998

[Appearance of extrahepatic lesions in the superior lip and orbit of a patient with liver metastases of gastric cancer following partial response by hepatic arterial infusion chemotherapy].

A 76-year-old man was totally gastrectomized in June, 1996 for advanced gastric cancer. In February, 1997, multiple liver metastases were found by abdominal CT scan, and hepatic arterial infusion chemotherapy was started with 5-FU, EPI and MMC by an implantable reservoir indwelled via the left subclavian artery. This treatment was judged to have led to a partial response because reduced focus size of the liver metastasis was revealed by CT scan and levels of tumor markers decreased significantly. However, metastatic foci were found in the skin of the superior lip and the orbit in December 1997. He was treated with a variety of therapies, but died in January, 1998. Many cases with metastatic hepatic cancer have a poor prognosis because of the appearance of extrahepatic lesions in spite of the fact that a partial response can be obtained by hepatic arterial infusion chemotherapy. The present case was unique since extrahepatic lesions appeared at very rare sites such as the superior lip and the orbit.

Aged↗
PubMed1997

Glycoprotein Ib-von Willebrand factor interactions activate tyrosine kinases in human platelets.

von Willebrand factor (vWF ) in the presence of botrocetin induces p72(syk) activation, assessed as its autophosphorylated level and in vitro kinase assays, the transient association of p72(syk) with p60(c-src), and the translocation of p60(c-src) and p54/58(lyn) to cytoskeletal fractions. Jararaca glycoprotein Ib-binding protein (GPIb-BP), which specifically binds to GPIb, abolished these phenomena, suggesting that they are mediated by the vWF-GPIb interaction. These tyrosine kinase-related events were not inhibited by GRGDS peptide (plus EGTA), indicating that GPIIb/IIIa is not involved in the observed responses. Shc, an adaptor protein, was also tyrosine phosphorylated by the botrocetin-vWF activation. When GPIb was immunoprecipitated with nonfunctional monoclonal antibodies (MoAbs) directed against GPIb, a kinase activity was found to associate with GPIb upon botrocetin-vWF activation. On the other hand, anti-GPIb MoAbs that inhibit the vWF-GPIb interaction did not coprecipitate a kinase activity. Because the recovery of GPIb did not differ significantly, it is suggested that the excessive presence of inhibitory anti-GPIb MoAb dissociated a kinase activity from GPIb. Phosphoamino acid analysis showed that the kinase activity was that of a tyrosine kinase. The identity of the tyrosine kinase and the mode of interaction with the cytoplasmic region of GPIb await to be determined. Our findings suggest that the tyrosine kinase associated with GPIb serves at a most proximal step in the signal transduction pathway involved in the vWF-GPIb-induced platelet activation, which leads to other tyrosine kinase-related intracellular signals.

Blood Platelets↗
PubMed1997

Role of inositol 1,4,5-trisphosphate receptor in ventral signaling in Xenopus embryos.

The inositol 1,4,5-trisphosphate (IP3) receptor is a calcium ion channel involved in the release of free Ca2+ from intracellular stores. For analysis of the role of IP3-induced Ca2+ release (IICR) on patterning of the embryonic body, monoclonal antibodies that inhibit IICR were produced. Injection of these blocking antibodies into the ventral part of early Xenopus embryos induced modest dorsal differentiation. A close correlation between IICR blocking potencies and ectopic dorsal axis induction frequency suggests that an active IP3-Ca2+ signal may participate in the modulation of ventral differentiation.

Activins↗
PubMed1997

N,N-dimethylsphingosine 1-phosphate activates human platelets.

We recently reported that N,N-dimethylsphingosine 1-phosphate (DMS-1-P) can be formed from N,N-dimethylsphingosine (DMS) in activated platelets [Y. Yatomi et al., Biochem. Biophys. Res. Commun. 231 (1997) 848-851]. In this study, we synthesized, for the first time, DMS-1-P and examined the functional effects of DMS-1-P and its related sphingolipids on platelets. Although exogenous DMS was inactive, its phosphorylated derivative, DMS-1-P, induced platelet intracellular Ca2+ mobilization and shape change, but not aggregation or release reactions. Since sphingosine 1-phosphate (Sph-1-P) is structurally related to DMS-1-P and activates platelets more strongly than DMS-1-P, a competitive binding experiment for [3H]Sph-1-P was performed using DMS-1-P. DMS-1-P reduced the binding of [3H]Sph-1-P to platelets almost as much as unlabeled Sph-1-P did. These results suggest that DMS-1-P activates platelets via an interaction with a platelet surface receptor for Sph-1-P.

Blood Platelets↗
PubMed1997

Activation of the G protein Gq/11 through tyrosine phosphorylation of the alpha subunit.

Various receptors coupled to the heterotrimeric guanine nucleotide-binding protein Gq/11 stimulate formation of inositol-1,4,5-trisphosphate (IP3). Activation of these receptors also induces protein tyrosine phosphorylation. Formation of IP3 in response to stimulated receptors that couple to Gq/11 was blocked by protein tyrosine kinase inhibitors. These inhibitors appeared to act before activation of Gq/11. Moreover, stimulation of receptors coupled to Gq/11 induced phosphorylation on a tyrosine residue (Tyr356) of the Galphaq/11 subunit, and this tyrosine phosphorylation event was essential for Gq/11 activation. Tyrosine phosphorylation of Galphaq/11 induced changes in its interaction with receptors. Therefore, tyrosine phosphorylation of Galphaq/11 appears to regulate the activation of Gq/11 protein.

Animals↗
PubMed1997

Tyrosine phosphorylation and p72syk activation by an anti-glycoprotein Ib monoclonal antibody.

NNKY5-5, an IgG monoclonal antibody directed against the von Willebrand factor-binding domain of glycoprotein (GP) Ib alpha, induced weak but irreversible aggregation (or association) of platelets in citrate-anticoagulated platelet-rich plasma. This phenomenon was defined as small aggregate formation (SAF). Platelets in hirudin-anticoagulated plasma or washed platelets showed little response to NNKY5-5 alone, but the antibody potentiated aggregation induced by low concentrations of adenosine diphosphate or platelet-activating factor. NNKY5-5 did not induce granule release or intracellular Ca2+ mobilization. However, NNKY5-5 caused tyrosine phosphorylation of a 64-kD protein and activation of a tyrosine kinase, p72syk. An anti-Fc gamma II receptor antibody had no effect on SAF, suggesting that NNKY5-5 activated platelets by interacting with glycoprotein Ib. Fab' fragments of NNKY5-5 did not induce SAF, but potentiated aggregation induced by other agonists. The Fab' fragment of NNKY5-5 induced the activation of p72syk, suggesting that such activation was independent of the Fc gamma II receptor. Cross-linking of the receptor-bound Fab' fragment of NNKY5-5 with a secondary antibody induced SAF. GRGDS peptide, chelation of extracellular Ca2+, and an anti-GPIIb/IIIa antibody inhibited NNKY5-5-induced SAF, but had no effect on 64-kD protein tyrosine phosphorylation or p72syk activations. Various inhibitors, including aspirin and protein kinase C, had no effect on SAF, protein tyrosine phosphorylation, or p72syk activation. In contrast, tyrphostin 47, a potent tyrosine kinase inhibitor, inhibited NNKY5-5-induced SAF as well as tyrosine phosphorylation and p72syk activation. Our findings suggest that binding of NNKY5-5 to GPIb potentiates platelet aggregation by facilitating the interaction between fibrinogen and GPIIb/IIIa through a mechanism associated with p72syk activation and tyrosine phosphorylation of a 64-kD protein.

Antibodies, Monoclonal↗
PubMed1997

N,N-dimethylsphingosine phosphorylation in human platelets.

Metabolism of sphingosine (Sph) derivatives in human platelets was examined. [3H]Sph was rapidly and heavily phosphorylated into sphingosine 1-phosphate, similarly in resting and stimulated platelets. [14C]N,N-dimethylsphingosine was stable in resting platelets, while it was converted into N,N-dimethylsphingosine 1-phosphate (DMS-1-P), although weakly, in platelets stimulated with thrombin or 12-O-tetradecanoylphorbol 13-acetate. This DMS-1-P formation was inhibited by staurosporine, a potent protein kinase inhibitor. [3H]C2-ceramide was unchanged both in resting and stimulated platelets. Our report is the first to describe production of DMS-1-P in a biological system.

Blood Platelets↗
PubMed1997

Developmental expression of the inositol 1,4,5-trisphosphate receptor and structural changes in the endoplasmic reticulum during oogenesis and meiotic maturation of Xenopus laevis.

To study the development of the calcium release mechanism, we examined the temporal and spatial expression of inositol 1,4,5-trisphosphate receptor (IP3R) during the oogenesis and meiotic maturation of Xenopus laevis. Observation of a series of fixed samples by immunofluoresence microscopy revealed a relocalization of Xenopus IP3R (XIP3R)-positive structures during meiotic maturation. We visualized the endoplasmic reticulum (ER) using ER-sensitive dye, DiI, by observation under confocal laser scanning microscopy. Time-lapse visualization of the living oocytes revealed that while the ER of immature fully grown oocytes underwent relatively little movement, the ER of maturing oocytes and mature eggs moved rapidly. A possible role for the increase of ER mobility in the dynamic redistribution of XIP3R during oocyte maturation is also discussed.

Animals↗
PubMed1997

Developmental expression of the inositol 1,4,5-trisphosphate receptor and localization of inositol 1,4,5-trisphosphate during early embryogenesis in Xenopus laevis.

To study the role of inositol 1,4,5-trisphosphate (IP3) receptors during early embryogenesis in Xenopus, we examined the temporal-spatial localization of Xenopus IP3 receptor (XIP3R). XIP3R protein is enriched in the animal hemisphere of early cleavage stage embryos and becomes localized in the ectoderm and involuted mesoderm in gastrula stage embryos. Up to tailbud stages, expression of XIP3R is observed in the mesodermal tissues and in most subregions of the central nervous system. A quantitative analysis of endogenous IP3 mass during normal early embryogenesis revealed an increase in IP3 mass first observed at early gastrula stage of 10.5 with an enrichment in the ectoderm throughout the gastrula stages, implying a potential role during gastrulation.

Animals↗
PubMed1997

Outside-in signaling from integrin alpha IIb beta 3 into platelets in the absence of agonist-induced signaling.

Platelet agonists generate intracellular signals which lead to activation of the platelet membrane glycoprotein IIb-IIIa (integrin alpha IIb beta 3). The resulting occupancy of alpha IIb beta 3 by ligands also generates signals into the cell (outside-in signaling). We reported previously that unlike platelet agonists, the F(ab')2 fragments of an anti-alpha IIb beta 3 monoclonal antibody, PMA4, induced fibrinogen binding to alpha IIb beta 3 without causing intracellular activation. In this study, in order to determine whether outside-in signaling occurs in the absence of agonist-induced intracellular signals, we used PMA4 F(ab')2 as an inducer of fibrinogen binding to alpha IIb beta 3. PMA4 F(ab')2-induced fibrinogen binding and subsequent platelet aggregation triggered tyrosine phosphorylation of several proteins including pp72syk but not pp125FAK. No Ca2+ influx or mobilization, thromboxane B2 synthesis, phosphorylation of pleckstrin or the myosin light chain, cytoplasmic alkalinization, or platelet shape changes, were detected. These findings suggest that, in the absence of agonist-induced signaling, alpha IIb beta 3 occupied by soluble fibrinogen generates only a limited outside-in signal.

Antibodies, Monoclonal↗
PubMed1997

Sphingosine 1-phosphate, a bioactive sphingolipid abundantly stored in platelets, is a normal constituent of human plasma and serum.

Although sphingosine 1-phosphate (Sph-1-P) is reportedly involved in diverse cellular processes and the physiological roles of this bioactive sphingolipid have been strongly suggested, few studies have revealed the presence of Sph-1-P in human samples, including body fluids and cells, under physiological conditions. In this study, we identified Sph-1-P as a normal constituent of human plasma and serum. The Sph-1-P levels in plasma and serum were 191+/-79 and 484+/-82 pmol/ml (mean+/-SD, n=8), respectively. Furthermore, when Sph-1-P was measured in paired plasma and serum samples obtained from 6 healthy adults, the serum Sph-1-P/plasma Sph-1-P ratio was found to be 2.65+/-1.26 (mean+/-SD). It is most likely that the source of discharged Sph-1-P during blood clotting is platelets, because platelets abundantly store Sph-1-P compared with other blood cells, and release part of their stored Sph-1-P extracellularly upon stimulation. We also studied Sph-1-P-related metabolism in plasma. [3H]Sph was stable and not metabolized at all in plasma, but was rapidly incorporated into platelets and metabolized mainly to Sph-1-P in platelet-rich plasma. [3H]Sph-1-P was found to be unchanged in plasma, revealing that plasma does not contain the enzymes needed for Sph-1-P degradation. In summary, platelets can convert Sph into Sph-1-P, and are storage sites for the latter in the blood. In view of the diverse biological effects of Sph-1-P, the release of Sph-1-P from activated platelets may be involved in a variety of physiological and pathophysiological processes, including thrombosis, hemostasis, atherosclerosis and wound healing.

Blood Cells↗
PubMed1997

A case of hepatocellular carcinoma with marked hyperfibrinogenemia.

The present report concerns a case of hepatocellular carcinoma (HCC) with marked hyperfibrinogenemia. The plasma fibrinogen level reached as high as 1,704 mg/dl. Since treatment against HCC resulted in reduction of plasma fibrinogen and PIVKA-II, an HCC marker, the hyperfibrinogenemia appears to be related to HCC. Immunohistochemically, the HCC specimen from this patient reacted strongly with antiserum to human fibrinogen, suggesting that the elevated fibrinogen was due to synthesis of this protein by the carcinoma cells, not to decreased fibrinolytic activity.

Adult↗
PubMed1997

Phosphorylation of myosin light chain in resting platelets from NIDDM patients is enhanced: correlation with spontaneous aggregation.

Platelet function in patients with NIDDM is enhanced. We have found that spontaneous aggregation (i.e., the formation of small-sized aggregates in the absence of agonist stimulation) occurs at a high rate in platelets from NIDDM patients. We then investigated basal myosin light chain 20 (MLC) phosphorylation, which plays a key role in platelet shape change and aggregation, using a monoclonal antibody against a phosphorylation site (serine 19 residue) in the MLC molecule in platelets from these patients. Standard calibration curves obtained from purified MLC or the phosphorylated form of myosin light chain 20 (MLC-P) were linear within the range of 0-150 ng for MLC and 0-3 ng for MLC-P. The amount of MLC or MLC-P in platelets was estimated, and basal MLC phosphorylation was calculated. Platelets were obtained from 9 young healthy control subjects, 13 age- and sex-matched nondiabetic control subjects, and 13 patients with NIDDM. The basal MLC phosphorylation in platelets was significantly higher in the NIDDM patients than in the control subjects, irrespective of age. These findings suggest that platelets from NIDDM patients are activated in vivo. Platelets obtained from NIDDM patients generated spontaneous aggregation, the degree of which was significantly higher than that in control subjects. Platelet spontaneous aggregation correlated well with basal MLC phosphorylation. These findings suggest that increases in basal MLC in platelets may be one factor leading to hyperaggregability of platelets in these patients.

Adult↗