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M Sakon

Publications and source records attributed to M Sakon.

At least 91 records · Page 5Linked to original sources
PubMed1998

Possible involvement of m-calpain in vascular smooth muscle cell proliferation.

Vascular smooth muscle cell (VSMC) proliferation still remains a poorly understood process, although it is believed to play a critical role in pathological states, including atherosclerosis and hypertension. Several reports have suggested that proteases may be directly involved in this process; however, it was still unclear which protease is responsible for VSMC proliferation. In this study, by use of a cell-permeable calpain inhibitor (calpeptin; benzyloxycarbonyl-Leu-nLeu-H), its analogue (benzyloxycarbonyl-Leu-Met-H), the cell-impermeable serine protease inhibitor leupeptin, and antisense oligonucleotide against m-calpain to inhibit proliferation of primarily cultured human VSMCs, we investigated whether calcium-activated neutral protease (calpain) is involved in VSMC proliferation. Calpeptin and its analogue, more specific for m-calpain, equally inhibited the proliferation of VSMCs in a dose-related manner, whereas a more limited antiproliferative effect was observed in leupeptin-treated VSMCs. Antisense oligonucleotide against m-calpain, but not scrambled antisense, dose-dependently inhibited m-calpain expression and proliferation of VSMCs. Maximal inhibition was an approximately 50% reduction of cell number and m-calpain antigen observed at 50 micromol/L of antisense oligonucleotide. Calpeptin or antisense oligonucleotide against m-calpain increased the expression of the endogenous calpain substrate pp125FAK (focal adhesion kinase), whereas the expression of the endogenous calpain inhibitor calpastatin was not affected. These results suggest that the proliferation of VSMCs requires protease activity, some of which is due to m-calpain.

Calcium-Binding Proteins↗
PubMed1998

Detection of K-ras gene mutations in plasma DNA of patients with pancreatic adenocarcinoma: correlation with clinicopathological features.

We investigated the presence of K-ras gene mutation in plasma DNA and assessed its clinical value in patients with pancreatic adenocarcinoma. Mutations in codon 12 of the K-ras gene were examined by mutant allele-specific amplification method using DNA extracted from surgical specimens and plasma samples of 21 patients with pancreatic adenocarcinoma. K-ras gene mutation was detected in 15 of 21 (71%) primary tumors. In 9 of 15 (60%) patients with K-ras gene mutation-positive tumors, an identical mutation was detected in the plasma DNA. None of four patients with chronic pancreatitis or five healthy subjects had such mutations in plasma DNA. Tumors positive for K-ras gene mutation in plasma DNA were significantly larger (P = 0.04) and less likely to result in a curative cure after surgical resection (P = 0.09) than those negative for the mutation. Other clinicopathological features, including age, sex, histological type, mode of invasion, and metastasis, did not correlate with K-ras gene mutations in plasma DNA. Treatment resulted in disappearance of K-ras gene mutations in plasma DNA in six of nine (67%) patients. Three patients with a persistently positive K-ras gene mutation in pre- and post-treatment plasma samples were likely to show early recurrence or have a progressive disease. Our findings suggest that K-ras gene mutation can be detected in plasma DNA of patients with pancreatic adenocarcinoma. Detection of K-ras mutations in plasma may be clinically useful for evaluating tumor burden and efficacy of treatment.

Adenocarcinoma↗
PubMed1998

[Results of clinical study with epirubicin hydrochloride injectable solution in hepatoma].

A 10-center cooperative clinical study with a new formulation of epirubicin hydrochloride injectable solution (Epirubicin-RTU) was conducted in patients with hepatocellular carcinoma. Epirubicin-RTU 60 mg/m2 was injected into the hepatic artery and a three-week drug-free interval followed. Of 15 patients with hepatocellular carcinoma registered in this study, 14 patients were eligible, and they all completed the entire course. The objective was to investigate the safety of treatment with Epirubicin-RTU in 14 eligible patients. The adverse drug reactions frequently observed in these 14 eligible cases were leukopenia, neutropenia, thrombocytopenia, alopecia, and fever. They were all reversible and tolerable. With these results. Epirubicin-RTU was considered to be a safe pharmaceutical product to inject into the hepatic artery.

Adult↗
PubMed1997

Expanded polytetrafluoroethylene (ePTFE) vascular graft loses its thrombogenicity six months after implantation.

Although expanded polytetrafluoroethylene (ePTFE) has been believed to be an inert material for vascular prosthesis, it shows less tendency of graft maturation by means of endothelialization. The aim of this study is to evaluate long-term alteration in thrombogenicity of ePTFE grafts after implantation. Serial levels of thrombin-antithrombin III complex (TAT), D-dimer, C-reactive protein (CRP), and prothrombin time (PT) were examined in 77 patients following ePTFE Y-graft implantation for up to five years. TAT showed biphasic elevation after implantation; TAT increased from 16.4+/-8.6 ng/ml to 27.4+/-10.5 ng/ml at one day, decreased to 18.5+/-4.5 ng/ml at one week, and increased again to 25.3+/-8.5 ng/ml at two weeks. Elevated TAT gradually decreased after the second peak to reach a lower level than that before surgery after six months. There was no significant difference in TAT level after six months due to the difference in diagnosis or anti-thrombotic therapy. We suggest that ePTFE grafts lose their thrombogenicity six months after implantation, after which anti-thrombotic therapy might be unnecessary.

Adult↗
PubMed1997

Awareness of donor alloantigens in antiadhesion therapy induces antigen-specific unresponsiveness to islet allografts.

BACKGROUND: Antiadhesion therapy using monoclonal antibodies (mAbs) to adhesion molecules in vivo has been shown to produce significant prolongation of graft survival in various transplantation models. However, it remains unclear whether antiadhesion therapy operates by merely blocking adhesion between antigen-presenting cells and T cells physically and/or by blocking costimulatory signals while preserving signals mediated through T-cell receptors in vivo. We examined antigen-specific T-cell responses during and after antiadhesion therapy. METHODS: BALB/c islets were transplanted into the renal subcapsular space of streptozotocin-induced diabetic C57BL/6 mice given anti-lymphocyte function-associated antigen (LFA)-1 and/or anti-intercellular adhesion molecule-1 mAb treatment. The animals bearing surviving islet allografts were challenged with BALB/c or third-party islets on day 7 or more than 100 days after transplantation. RESULTS: Islet allografts were acutely rejected in untreated animals, with a mean survival time (MST) of 19+/-8 days. Administration of anti-LFA-1 mAb induced significant prolongation of graft survival with a mean survival time of 72+/-33 days, and half of the allografts showed indefinite survival. The animals given anti-LFA-1 mAb alone 7 days before transplantation showed acute rejection of BALB/c islets, whereas a significant number of animals given anti-LFA-1 mAb and the BALB/c islet allograft simultaneously accepted secondary BALB/c islets, but rejected third-party islets. Likewise, most of the animals bearing long-term functioning BALB/c allografts for more than 100 days accepted secondary BALB/c islets, but rejected C3H islets acutely. Interestingly, the spleen cells from these animals transferred unresponsiveness to BALB/c islets into the 2.5-Gy x-irradiated recipients, whereas those from naive animals induced acute rejection. CONCLUSIONS: These results indicate that anti-LFA-1 mAb treatment prevents T-cell activation leading to rejection, but results in a T-cell receptor engagement leading to antigen-specific unresponsiveness maintained by transferrable suppressor cells.

Animals↗
PubMed1997

The release mechanism of platelet-activating factor during shear-stress induced platelet aggregation.

We previously reported that 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (platelet-activating factor, PAF), released from activated platelets stimulated with thrombin plus collagen, is associated with platelet microparticles. In the present study, we found that PAF is concentrated and associated with microparticles released from high shear stress-induced activated platelets. The total amount of PAF released from 3 x 10(8) platelets under high shear stress (108 dyne/cm2) was 3.2 +/- 0.6 x 10(-15)mol (n = 5, mean +/- S.D.). Eighty percent of the PAF released from the platelets was recovered in the microparticle fraction after ultracentrifugation in the presence of albumin. Under high shear stress, PAF was not released from platelets within 3 minutes, although microparticles were released. In conclusion, microparticles released from activated platelets in the rheological condition of high shear stress are major carriers of PAF.

Animals↗
PubMed1997

Hypercholesterolemia as a risk factor for deep-vein thrombosis.

Our retrospective study has shown that hyperlipidemia is a novel etiologic factor in deep-vein thrombosis (DVT) and that most of the idiopathic DVT patients were hyperlipidemic (Thrombosis Research 79, 147-151, 1995). The aim of our current study is to analyze the interrelationship between hyperlipidemia and DVT by means of a case-control study. A series of lipid parameters were analyzed using serum from 109 patients with deep vein thrombosis (DVT). One hundred nine age- and sex-matched subjects served as controls. Diagnosis of hyperlipidemia was made if the serum cholesterol level was above 220 mg/dL or if the triglyceride level was above 150 mg/dL. Among several types of hyperlipidemia examined, the risk factor associated with the highest estimated odds ratio was carriage of hypercholesterolemia associated with hypertriglyceridemia (odds ratio 5.1) followed in order by hypercholesterolemia without hypertriglyceridemia (odds ratio 2.6) and hypertriglyceridemia without hypercholesterolemia (odds ratio 0.9). These findings support the hypothesis that hypercholesterolemia plays an important role in the pathogenesis of DVT.

Adult↗
PubMed1997

Identification of mu-, m-calpains and calpastatin and capture of mu-calpain activation in endothelial cells.

The presence of the calpain-calpastatin system in human umbilical vein endothelial cells (HUVEC) was investigated by means of ion exchange chromatography, Western blot analysis, and Northern blot analysis. On DEAE anion exchange chromatography, calpain and calpastatin activities were eluted at approximately 0.30 M and 0.15-0.25 M NaCl, respectively. For half-maximal activity, the protease required 800 microM Ca2+, comparable to the Ca2+ requirement of m-calpain. By Western blot analysis, the large subunit of mu-calpain (80 kDa) was found to be eluted with calpastatin (110 kDa). Both the large subunit of m-calpain (80 kDa) and calpastatin were detected in the respective active fractions. By Northern blot analysis, mRNAs for large subunits of mu- and m-calpains were detected in single bands, each corresponding to approximately 3.5 Kb. Calpastatin mRNA was observed in two bands corresponding to approximately 3.8 and 2.6 Kb. Furthermore, the activation of mu-calpain in HUVEC by a calcium ionophore was examined, using an antibody specifically recognizing an autolytic intermediate form of mu-calpain large subunit (78 kDa). Both talin and filamin of HUVEC were proteolyzed in a calcium-dependent manner, and the reactions were inhibited by calpeptin, a cell-permeable calpain specific inhibitor. Proteolysis of the cytoskeleton was preceded by the appearance of the autolytic intermediate form of mu-calpain, while the fully autolyzed postautolysis form of mu-calpain (76 kDa) remained below detectable levels at all time points examined. These results indicate that the calpain-calpastatin system is present in human endothelial cells and that mu-calpain may be involved in endothelial cell function mediated by Ca2+ via the limited proteolysis of various proteins.

Antibody Specificity↗
PubMed1997

Calpain activation in shear-induced platelet aggregation.

Fluid shear stress has been known to activate platelet reaction such as aggregation, but the exact mechanism of shear-induced platelet aggregation (SIPA) has not been fully understood. Calpain, an intracellular calcium-activated cysteine protease, is abundant in platelets and is considered to be activated and involved in the proteolytic processes during platelet activation. A possible activation of calpain in SIPA was investigated, employing a newly developed aggregometer and specific monoclonal antibodies to detect activation of calpain. When a shear stress gradient varying between 6 and 108 dyn/cm2 was applied to platelets, activation of mu-calpain was observed only in high-shear-stressed platelets, resulting in the proteolysis of talin. At 1 min after the onset of constant high shear stress of 108 dyn/cm2, mu-calpain activation and proteolysis of talin were detected and increased in a time-dependent manner. Constant shear stress more than 50 dyn/cm2, applied for 5 min, caused mu-calpain activation and proteolysis of talin, which were increased in a shear-force-dependent manner. Calpeptin, a calpain-specific peptide antagonist, caused the complete inhibition of both mu-calpain activation and proteolysis of talin, while SIPA profiles with calpeptin showed almost no change compared to those without calpeptin. These results suggest the possibility of calpain involvement in late phases of shear-induced platelet activation such as cytoskeletal reorganization.

Calpain↗
PubMed1997

Prostacyclin analog-suppressed ischemia-reperfusion injury of the rat liver: evaluation by calpain mu activation.

Prostaglandin I2 has a protective effect on hepatic ischemia-reperfusion injury. However, the exact intracellular mechanisms of this effect have not been elucidated. Calpain micro, a Ca2+-dependent protease, has been found to play a role in the ischemia-reperfusion injury of various organs. The hilar area of the left lateral and median lobes of rat livers was clamped for 60 min. A prostaglandin I2 analog (OP2507, C35H41NO4) was intravenously administered at 0.1, 0.32, or 1.0 microg/kg/min from 20 min before the ischemia. In addition to biochemical and microscopic analyses, the activation of calpain mu was investigated using specific antibodies against the intermediate (activated) and preactivated forms of calpain mu. The degradation of talin was also studied by Western blotting. When OP2507 was infused at 0.32 and 1.0 microg/kg/min, bile flow significantly increased after reperfusion compared with the control group, consistent with the decrease in serum transaminase levels. Membrane bleb formation and the appearance of the intermediate form of calpain mu were observed at 60 min of ischemia in the control and OP2507 (0.1 microg/kg/min) groups and remained present until 120 min after reperfusion. OP2507 (1.0 microg/kg/min) markedly suppressed not only membrane bleb formation but also calpain mu activation and the degradation of talin. In conclusion, OP2507 suppresses ischemia-reperfusion injury of the rat liver, and its cytoprotective effect is closely associated with the inhibition of calpain mu activation.

Alanine Transaminase↗
PubMed1997

Effect of low concentration of epinephrine on human platelet aggregation analyzed by particle counting method and confocal microscopy.

The effect of a physiologic concentration of epinephrine (Ep) on platelet activation was studied by using a novel method that simultaneously measures platelet aggregation by changes in light transmission and counts particles of various sizes by using light scattering. Detailed morphologic changes associated with activation process were studied by using confocal laser scanning microscopy (CLSM). A low concentration of Ep (20 nmol/L) corresponding to a high physiologic concentration triggered the formation of small platelet aggregates (diameter 7 to 30 microm) without any change in light transmission. The redistribution of filamentous actin (F-actin) and the expression of activated glycoprotein IIb-IIIa complex (GPIIb-IIIa), detected by PAC-1 binding, were also observed in the platelets comprising the small aggregates. Attempts were then made to detect changes in cytoplasmic ionized Ca2+ ((Ca2+)i) in individual platelets involved in the aggregate formation by CLSM with fluo-3, a Ca2+-indicating dye. Ep caused a weak (Ca2+)i increase in some individual platelets involved in the formation of small aggregates. This (Ca2+)i increase was associated with platelet aggregation, because no (Ca2+)i rise was detected in single platelets. Furthermore, platelets stimulated by Ep in the presence of RGDS or Ro 44-9883, a GPIIb-IIIa antagonist, did not form small aggregates or trigger a (Ca2+)i rise. Prior incubation with low concentrations of Ep (20, 100 nmol/L) enhanced the initial formation of small (diameter 7 to 30 microm), medium (diameter 30 to 50 microm), or large (diameter 50 to 70 microm) aggregates induced by a subthreshold concentration of adenosine diphosphate (0.25 micromol/L) as determined by the particle counting method. However, no apparent synergistic (Ca2+)i increase was observed in platelets involved in aggregate formation. From these observations the following conclusions have been reached. (1) A high physiologic concentration of Ep (20 nmol/L) is capable of triggering the formation of small aggregates, resulting in the redistribution of F-actin and the expression of activated GPIIb-IIIa complex. (2) An increase in (Ca2+)i is observed in platelets comprising the small aggregates. This increase is not related to the binding of Ep to its receptor but most likely is triggered by platelet-platelet association. (3) The characteristic potentiating effect of Ep is not due to the synergistic increase in (Ca2+)i.

Actins↗
PubMed1997

Shear-stress causes polarized change in cytoplasmic calcium concentration in human umbilical vein endothelial cells (HUVECs).

Using a newly developed, parallel-plate flow-chamber for confocal laser scanning microscopy (CLSM), we studied the distribution and temporal changes in intracellular Ca2+ concentration ([Ca2+]i) in individual HUVECs stimulated by shear-stress. In the presence of ATP, shear-stress (1-10 dyne/cm2) caused a rise in [Ca2+]i, whereas no such response was observed in the absence of ATP or in the presence of Ni2+, a nonspecific, plasma membrane Ca2+ channel blocker. These results suggest that both ATP and Ca2+ influx are essential for the increase in [Ca2+]i in response to shear stress at less than 10 dyne/cm2. Analysis of [Ca2+]i distribution revealed a repetitive intracellular 'Ca2+ wave' originating from the upstream edge of the cell in some populations of HUVECS, which was transmitted to the downstream of the cell. The polarized [Ca2+]i response induced by shear-stress might be integral to polarized cellular reactions such as remodeling of endothelial lining.

Adenosine Triphosphate↗
PubMed1997

Renal pelvic cancer with tumor thrombus in the vena cava inferior. A case report and review of the literature.

Renal pelvic cancer with a large vein tumor thrombus is very rare. We report a case of transitional cell carcinoma of the right renal pelvis with tumor thrombus in the inferior vena cava. Based on the radiological findings, the patient was diagnosed having renal cell carcinoma with tumor thrombus in the vena cava preoperatively. Radical nephrectomy with thrombectomy was performed. A literature review of 17 reported cases revealed that 13 cases (76.5%) were diagnosed as renal cancer preoperatively. Prognosis of transitional cell carcinoma with vena cava tumor thrombus is relatively poor compared to that of renal cell carcinoma with a large vein tumor thrombus.

Carcinoma, Renal Cell↗
PubMed1997

[The change of hepatic energy status after transcatheter arterial embolization (TAE) for hepatocellular carcinoma--a study using 31P-MRS].

In six patients with hepatocellular carcinoma, adenosine triphosphate (ATP) phosphomonoester (PME), phosphoric acid (Pi) and pH were measured before and after transcatheter arterial embolization (TAE), using 31P-magnetic resonance spectroscopy (MRS). The effectiveness of TAE was judged by the extent of lipiodol accumulation within the tumor and change in serum alphafetoprotein level. The pH within the tumor decreased 2-9 days after TAE in all the cases, and beta ATP/Pi ratio decreased when TAE was effective, whereas the ratio did not decrease when TAE was ineffective. The beta ATP/Pi ratio measured by 31P-MRS seems to be a useful, non-invasive marker of the anti-tumor effect of TAE, although further investigation is needed.

Adenosine Triphosphate↗
PubMed1997

[Transarterial immuno-embolization therapy in patients with hepatocellular carcinoma].

Surgical resection, transcatheter arterial embolization (TAE) and percutaneous ethanol injection therapy (PEIT) are effective for hepatocellular carcinoma (HCC), but the recurrence rate is high. We have devised a new therapy of transarterial immuno-embolization (TIE) with OK-432, fibrinogen and thrombin, and 2 cases are reported. Case 1: A 78-year-old Japanese male with HCC (diameter, 4 cm in subsegment 5) received TIE. The tumor size was markedly decreased, and the patient survived for more than 3 years without recurrence. Case 2: A 61-year-old Japanese male with HCC (diameter, 4.5 cm in segment 5) received hepatic subsegmentectomy following TIE. Histological examination of resected specimens following TIE showed massive infiltration of mononuclear cells in the main tumor. Tumor recurrence had developed three times thereafter, but was effectively treated by TIE. TIE may be an effective therapy for HCC.

Aged↗
PubMed1996

Ca&sup2+;-Dependent Neutral Protease (Calpain) Activity in Breast Cancer Tissue and Estrogen Receptor Status.

Cancer invasion and metastasis require action of tumor-associated proteases, which degrade the extracellular matrix. It has been reported that calpain, a calcium-activated neutral protease and a thiol protease regulated by Ca&sup2+;, proteolyzes estrogen recepor (ER) and that calpain may play an important role in the regulation of ER function. In the present study, the activities of calpain were measured in human normal breast tissues and breast cancer tissues stratified by estrogen receptor levels. There were no correlations between calpain activity and tumor size or lymph node involvement. Activities of calpain were significantly higher in breast cancer tissues compared with those of normal breast tissues, and were higher in the ER-positive tumors than in ER-negative ones. These results indicate that calpain is related to mammary malignant transformation and is involved in the regulation of the ER function in breast cancer tissues.

Journal Article↗
PubMed1996

LFA-1/ICAM-3 mediates neutrophil homotypic aggregation under fluid shear stress.

We found that human neutrophils undergo homotypic aggregation by loading the physiological range of fluid shear stress (12-30 dynes/cm2). Under the fluid shear stress, an increase of intracellular Ca2+ concentration of neutrophils was observed. This increase of intracellular Ca2+ concentration was caused by Ca2+ influx, and the blockage of the flux by NiCl2 suppressed the neutrophil homotypic aggregation. Furthermore, this neutrophil aggregation under fluid shear stress was completely inhibited by pretreatment with antibody against LFA-1 or ICAM-3. These results suggested that NiCl2-sensitive Ca2+ channel played an important role in LFA-1/ICAM-3-mediated neutrophil homotypic aggregation under fluid shear stress.

Antigens, CD↗