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Biomedical subjects

Y Kanoh

Publications and source records attributed to Y Kanoh.

At least 37 records · Page 2Linked to original sources
PubMed1999

Inhibitory effect of glyburide on thrombin-induced platelet aggregation and phosphoinositide metabolism in normal human platelets.

We previously reported the effects of diet, sulphonylureas or insulin on thrombin-induced platelet aggregation, phosphoinositide metabolism and protein phosphorylation in non-insulin-dependent diabetes mellitus (NIDDM) patients. To clarify the mechanism of glyburide and insulin on platelet function, here we studied the in vitro effects of glyburide and insulin on thrombin-induced metabolic changes using normal human platelets. Platelet aggregation stimulated with <0.5 U/ml thrombin, 0.75-3 microM adenosine diphosphate (ADP) or 1 microg/ml collagen was significantly lower in glyburide-treated platelets, but not in insulin-treated platelets, than in untreated ones (control). Thrombin-induced incorporation of 32P radioactivity into phosphatidic acid (PA) in glyburide-treated platelets was lower than that in control but not in insulin-treated platelets. Phosphorylated proteins of platelets induced by thrombin and 12- O -tetradecanoylphorbol 13-acetate (TPA) in glyburide-treated platelets were suppressed, but not in insulin-treated platelets, compared with control. These results suggest that glyburide induces suppression of thrombin-induced activation of phospholipase C, which mediates hydrolysis of PIP and PIP(2) and production of PA, and subsequently inhibits platelet aggregation.

Journal Article↗
PubMed1999

DHEA improves glucose uptake via activations of protein kinase C and phosphatidylinositol 3-kinase.

We have examined the effect of adrenal androgen, dehydroepiandrosterone (DHEA), on glucose uptake, phosphatidylinositol (PI) 3-kinase, and protein kinase C (PKC) activity in rat adipocytes. DHEA (1 microM) provoked a twofold increase in 2-[3H]deoxyglucose (DG) uptake for 30 min. Pretreatment with DHEA increased insulin-induced 2-[3H]DG uptake without alterations of insulin specific binding and autophosphorylation of insulin receptor. DHEA also stimulated PI 3-kinase activity. [3H]DHEA bound to purified PKC containing PKC-alpha, -beta, and -gamma. DHEA provoked the translocation of PKC-beta and -zeta from the cytosol to the membrane in rat adipocytes. These results suggest that DHEA stimulates both PI 3-kinase and PKCs and subsequently stimulates glucose uptake. Moreover, to clarify the in vivo effect of DHEA on Goto-Kakizaki (GK) and Otsuka Long-Evans fatty (OLETF) rats, animal models of non-insulin-dependent diabetes mellitus (NIDDM) were treated with 0.4% DHEA for 2 wk. Insulin- and 12-O-tetradecanoyl phorbol-13-acetate-induced 2-[3H]DG uptakes of adipocytes were significantly increased, but there was no significant increase in the soleus muscles in DHEA-treated GK/Wistar or OLETF/Long-Evans Tokushima (LETO) rats when compared with untreated GK/Wistar or OLETF/LETO rats. These results indicate that in vivo DHEA treatment can result in increased insulin-induced glucose uptake in two different NIDDM rat models.

Adipocytes↗
PubMed1999

[Quantitation and heterogeneity of serum alpha 2-macroglobulin].

alpha 2-Macroglobulin (alpha 2M) is a major protease inhibitor in human plasma. In this study, serum alpha 2M was determined by means of immunoelectrophoresis (IEP), rocket immunoelectrophoresis (R-IEP), laser nephelometry (LN) and single radial immunodiffusion (SRID) in 133 subjects. The values obtained by these methods coincided well in most specimens, but some specimens showed disagreement. The sera of alpha 2M deficiency found by IEP were analyzed by the other methods. We found that R-IEP gave the results of alpha 2M deficiency most frequently while the other methods gave the results of normal alpha 2M level for some specimens. We studied on the presumption that the phenomenon is due to heterogeneity in molecular weight of alpha 2M. After alpha 2M was purified by affinity chromatography and gel filtration chromatography, we further employed gel-filtration on a high-performance liquid chromatography (HPLC) to isolate alpha 2M with different molecular size. Two peaks of immunoreactive alpha 2M were found by the gel-filtration HPLC. When we performed IEP for both portions, the first portion with higher molecular size showed larger rate of migration than the second portion. Serum alpha 2M quantitation with SRID is not appropriate because molecular weight influences on the determination by SRID. We conclude that R-IEP is best suitable for the detection of decrease of serum alpha 2M, although this method requires expertness of manipulation.

Adolescent↗
PubMed1998

Glucose- and phorbol ester-induced insulin secretion in human insulinoma cells--association with protein kinase C activation.

This study examined the effect of glucose and 12-O-tetradecanoylphorbol-13-acetate (TPA) on insulin secretion in isolated human insulinoma cells. In addition, we analyzed conventional PKC alpha and beta activation in the membrane fractions, respectively. Treatment with 5 mM and 20 mM glucose for 5 min and 20 min resulted in 6-7-fold increases in insulin secretion, and treatment with 1 microM TPA for 5 min also resulted in 3-fold increases in insulin secretion from the basal level. Immunoblot analysis of membrane fractions showed increases in PKC alpha and beta immunoreactivities after treatment with 5 mM, 20 mM glucose and 1 microM TPA. Translocations of PKC alpha after treatment with glucose and TPA were greater than those of PKC beta in membrane fractions. These results suggest that TPA independently provokes insulin secretion via PKC activation and that PKC alpha and beta activation may be involved in insulin secretion in human insulinoma cells.

Adult↗
PubMed1998

Alterations in insulin-induced postreceptor signaling in adipocytes of the Otsuka Long-Evans Tokushima fatty rat strain.

The Otsuka Long-Evans Tokushima fatty (OLETF) rat is a new spontaneous non-insulin-dependent diabetes mellitus (NIDDM) model rat strain developed in Tokushima, Japan. After 18 weeks of age, decreases of 45% and 40% respectively in insulin- and phorbol ester-stimulated [3H]2-deoxyglucose (DOG) uptake were observed, compared with those in Long-Evans Tokushima (LETO) rats (control). Insulin-specific binding and 95 kDa autophosphorylation of insulin receptor in OLETF rats were not different from those in LETO rats. Insulin-induced diacylglycerol (DG) production and Mono Q column-purified protein kinase C (PKC) translocation in adipocytes of OLETF rats were decreased compared with those of LETO rats. Insulin-induced PKC beta translocation from cytosol to membrane was also decreased in adipocytes of OLETF rats. Increases of the PKC beta I, beta II, epsilon and zeta isoforms in membranes of OLETF rats were markedly smaller than those of LETO rats. Analysis of mRNA levels of PKC isoforms in adipocytes of OLETF rats showed decreases of basal level and insulin-induced delayed responses of PKC beta I, beta II, epsilon and zeta mRNA in OLETF rats. On the other hand, insulin- or phorbol ester-induced phosphatidylinositol 3-kinase (PI 3-kinase) activation was decreased in adipocytes of OLETF rats compared with those of LETO rats. These results suggest that insulin resistance in OLETF rats, a spontaneous NIDDM model rat, may be associated with deterioration of insulin-induced DG-PKC signaling and subsequent decrease in PI 3-kinase activation.

Adipocytes↗
PubMed1998

Differential effect of the antidiabetic thiazolidinediones troglitazone and pioglitazone on human platelet aggregation mechanism.

Troglitazone and pioglitazone, antidiabetic thiazolidinediones, are known to improve insulin resistance. However, the effect of these drugs on platelet aggregation remains unclear. The chemical structure of troglitazone contains vitamin E. Accordingly, we studied the effect of troglitazone, pioglitazone, and vitamin E on thrombin-induced platelet aggregation, metabolism of phosphoinositide, protein phosphorylation, protein kinase C (PKC)-alpha and -beta, and phosphatidylinositol (PI) 3-kinase activation in vitro in human platelets. Maximum platelet aggregation by ADP, collagen, and thrombin decreased in the presence of 0.1-1 micromol/l troglitazone and 500 nmol/l vitamin E for 60 min compared with controls. However, pioglitazone did not inhibit ADP-, collagen-, or thrombin-induced platelet aggregation. Pretreatment with troglitazone and vitamin E, but not with pioglitazone, resulted in decreases in thrombin-induced phosphatidic acid production, hydrolysis of phosphatidylinositol 4,5-bisphosphate by phospholipase C, and 47-kDa protein phosphorylation. Thrombin-induced PKC-alpha and -beta activation in membrane fraction was suppressed by pretreatment with troglitazone and vitamin E, but not with pioglitazone. Separately, troglitazone and pioglitazone stimulated PI 3-kinase activity, but thrombin-induced PI 3-kinase activation was suppressed by pretreatment with troglitazone and pioglitazone for 60 min. These results suggest that troglitazone and vitamin E, but not pioglitazone, have a potent inhibitory effect on platelet aggregation via suppression of the thrombin-induced activation of phosphoinositide signaling in human platelets. Finally, the chemical structure of vitamin E may contribute to the inhibitory effect of troglitazone on platelet aggregation in human platelets.

Adenosine Diphosphate↗
PubMed1998

[Concentrations of free form and complex form of prostate-specific antigen in serum of alpha 2 macroglobulin deficient patients with prostatic carcinoma].

Prostate-specific antigen (PSA) is present in blood in free form as well as in complex form with various protease inhibitors such as alpha 2 macroglobulin (alpha 2M) and alpha 1 antichymotrypsin (alpha 1 ACT). We had found that alpha 2M is deficient (below approximately 40 mg/dl) in some patients with prostatic carcinoma. Therefore, we investigated the levels of free and complex form of PSA in patients with prostatic disease and obtained the following results. The HPLC study showed that total (free plus complex) PSA level was much higher in the alpha 2M deficient patients with prostatic carcinoma stage D (n = 7, range 1,530-14,746 ng/ml, median value 6,800 ng/ml) than in the non-deficient patients with stage D (n = 16, range 121.6-4,210 ng/ml, median value 851 ng/ml). In the deficient patients, the complex of PSA with alpha 1 ACT increased extraordinarily while free PSA increased to only some extent. In the more severe cases of prostatic carcinoma, the ratio (complex/total PSA) was higher while the ratio (free/total PSA) was lower. The SDS-PAGE Western blotting showed that complex PSA increased extraordinarily and free PSA increased in sera of the deficient patients which was consistent with the HPLC results. Many bands appeared on the blotting at the positions smaller than alpha 2M molecule, which indicated that many fragments of alpha 2M were present in their sera. These bands were more intense than the bands with sera of controls or benign prostatic hypertrophy. The alpha 2M deficiency may be due to the rapid disappearance of the complex with PSA or any other prostate-originated proteases. The complex may undergo accelerated degradation or catabolism of alpha 2M.

Aged↗
PubMed1998

[Quantitative levels of serum amyloid A protein and other proteins in cerebrospinal fluid and serum of patients with meningitis].

Serum amyloid A protein (SAA), a putative precursor of the AA protein which constitutes amyloid fibrils in secondary amyloidosis, is evaluated as a sensitive acute-phase reactant in serum. At present, SAA concentration in serum is determined by latex nephelometry, but this assay cannot detect SAA in the low concentration range lower than 10 ng/ml. We have developed a sensitive enzyme immunoassay (EIA) for determining low concentrations of SAA. The assay is reproducible, reliable and requires no pretreatment of specimen prior to assay. We measured levels of SAA by this EIA in both cerebrospinal fluid (CSF) and serum of patients with suspected meningitis, measured also levels of albumin, alpha 2 macroglobulin and C-reactive protein (CRP) to investigate if these protein levels are useful for differential diagnoses of meningitis and for indicators damage of blood-CSF barrier. The SAA reference value in CSF is 3.99 +/- 1.74 ng/ml (mean +/- SD for nonmeningitis patients). The CRP concentration in CSF in bacterial meningitis was much higher than in viral meningitis, but CRP in CSF was also high in bacterial infection other than meningitis. On the other hand, SAA concentration in CSF in these patients with any meningitis are significantly higher than the reference values of SAA (p < 0.001). However, the differences in SAA concentrations among the three types (bacterial, viral and mycotic meningitis) of meningitis were not significant.

Acute-Phase Proteins↗
PubMed1997

Effect of glucocorticoid receptor antagonist RU 38486 on acute glucocorticoid-induced insulin resistance in rat adipocytes.

We examined the mechanism of acute glucocorticoid-induced insulin resistance in rat adipocytes using the glucocorticoid receptor antagonist RU 38486. Pretreatment with dexamethasone (DEX) and prednisolone for 60 minutes resulted in 50% inhibition of insulin-induced [3H]2-deoxyglucose (DOG) uptake at 10(-8) and 10(-7) mol/L, respectively, in rat adipocytes and 20% and 25% inhibition of insulin-induced [3H]2-DOG uptake, respectively, in soleus muscles. Our previous experiments indicated that DEX and prednisolone alone stimulate protein kinase C (PKC) in rat adipocytes. Accordingly, we examined [3H]DEX binding to PKC from MonoQ column-purified rat brain cytosol. Specific [3H]DEX binding to MonoQ column-purified PKC was observed (kd, 56.8 nmol/L; Bmax, 725 fmol/mg protein). Thus, insulin-induced PKC translocation from the cytosol to the membrane was suppressed by pretreatment with 10(-7) mol/L DEX and 10(-6) mol/L prednisolone for 80 minutes. During treatment with RU 38486 for 60 minutes, there was no change in the glucocorticoid-induced inhibitory effect on insulin-induced [3H]2-DOG uptake and PKC translocation from the cytosol to the membrane. Moreover, pretreatment with RU 38486 for 120 minutes slightly prevented the DEX-mediated inhibition of insulin-induced glucose uptake. These results suggest that acute glucocorticoid-induced insulin resistance may be mainly mediated through the other non-glucocorticoid receptor pathway.

Adipocytes↗
PubMed1997

Acute effects of phorbol ester and insulin on insulin-induced glucose uptake and protein kinase C activation in rat adipocytes.

We examined the acute effect of pretreatment with phorbol ester and insulin on insulin-induced glucose uptake and protein kinase C (PKC) translocation from cytosol to the membrane in rat adipocytes. Adipocytes were preincubated with 1 microM tetradecanoylphorbol 13-acetate (TPA) and 10 nM insulin for 60 min and then stimulated with 10 nM insulin for 10 and 30 min to measure PKC activity in cytosol and membrane fractions using a Mono Q column connected onto an HPLC system and [3H]2-deoxyglucose (DOG) uptake, respectively. Pretreatment with 1 microM TPA and 10 nM insulin for 60 min resulted in the marked decreases of insulin-induced [3H]2-DOG uptake. Translocation of Mono Q column-purified cytosolic PKC enzyme activity and PKC beta immunoreactivity from cytosol to the membrane was suppressed by pretreatment with TPA and insulin for 60 min. These results indicate that acute treatment with TPA and insulin which are PKC activators suppress translocation/activation of PKC, and accordingly inhibit insulin-induced glucose uptake. We suggest that a decrease of cytosolic PKC activity may mainly-contribute to the impaired responsiveness of the glucose transport system after acute TPA and insulin treatment.

Adipocytes↗
PubMed1997

Levels of interleukin-6, CRP and alpha 2 macroglobulin in cerebrospinal fluid (CSF) and serum as indicator of blood-CSF barrier damage.

We measured the levels of interleukin-6 (IL-6), albumin, C-reactive protein (CRP) and alpha 2 macroglobulin (alpha 2M), all of which have different spectrums of molecular weight, in the cerebrospinal fluid (CSF) and serum in 121 patients to evaluate damage to the blood-cerebrospinal fluid barrier (BCB) in meningitis. There was an extraordinary high level of IL-6 in the CSF when patients had bacterial or viral meningitis, but the level returned to a normal range within a week in almost all of these cases. There were no significant differences in CSF albumin levels among the different disease groups. The CRP level in CSF is considered to correlate with the serum level, and CSF CRP was higher in bacterial meningitis than in viral meningitis, however, CRP in CSF was increased in some of the infectious diseases without meningitis. The alpha 2M in CSF, which tends to be at extraordinarily high levels when there is damage to the BCB, correlated highly with CSF cell counts. CSF IL-6 seemed to be a useful indicator to identify the acute active phase of meningitis. CRP and alpha 2M in CSF are considered to be useful to differentiate bacterial meningitis, bacterial infection without meningitis and viral meningitis. Extraordinarily high levels of alpha 2M, which has a high molecular weight, in CSF is indicative of BCB damage.

Adult↗
PubMed1997

[Studies on alpha 2 macroglobulin deficiency in association with cancer metastasis].

We found 5 cases of prostatic carcinoma with metastasis with alpha 2 macroglobulin (alpha 2 M) concentration below approximately 40 mg/dl in serum. All these patients had bone metastasis, and none of them had DIC. We found no other cases with such a low concentration of alpha 2 M. Their alpha 2 M level increased to normal level after treatment with transurethral resection of prostate or hormone agents, and the level was correlated with the clinical symptom. During the clinical course, their alpha 2 M level was negatively correlated with prostate-specific antigen (PSA) and prostatic acid phosphate (PAP). All these results suggest that alpha 2 M concentration in serum reflects the severity of prostatic carcinoma with metastasis and that alpha 2 M deficiency is an indicator of metastasis. The acute phase proteins of CRP and serum amyloid A did not increase in spite of the presence of metastasis in these patients with extremely low alpha 2 M level (< 20 mg/dl), suggesting that alpha 2 M is involved in the metabolism of these acute phase proteins. On immunohistochemical studies, their specimens of prostatic carcinoma gave positive stain for PSA and urokinase-type plasminogen activator (u-PA). Both PSA and u-PA formed a complex with alpha 2 M in vitro. The alpha 2 M deficiency in these patients might be due to the complex formation between alpha 2 M and these prostate-originated proteases and to the rapid disappearance of the complex.

Aged↗
PubMed1996

Insulin regulated PKC isoform mRNA in rat adipocytes.

Insulin and 12-O-tetradecanoyl phorbol-13-acetate (TPA) induce both glucose uptake and translocation of protein kinase C (PKC) from cytosol to membrane in insulin-sensitive tissues as previously reported by several investigators. We examined insulin-mediated PKC beta I, beta II, and epsilon translocation from cytosol to cytoskeleton, and expression of PKC alpha, beta I, beta II, gamma, and epsilon isoforms using the reverse transcription polymerase chain reaction (RT-PCR) method during treatment with insulin for 240 min in rat adipocytes. Insulin-induced increases in PKC beta I, beta II, and epsilon were greater in the cytoskeleton fraction than those in the membrane fraction. Insulin induced time-dependent increases in PKC alpha, gamma, epsilon and zeta mRNA levels for up to 240 min (555%, 117%, 236% and 138% increase, respectively). TPA also induced time-dependent increases in PKC alpha and gamma (34% and 500% increase, respectively) but not in PKC zeta. However, PKC beta I mRNA was decreased for up to 60 min and then maintained at under the basal level during stimulation with insulin and TPA. On the other hand, PKC beta II mRNA was markedly increased for up to 240 min. These results suggest that insulin-regulated PKC alpha, gamma and epsilon mRNA levels and PKC beta mRNA alternative splicing may occur in rat adipocytes.

Adipocytes↗
PubMed1996

[Variations of serum amyloid A protein and interleukin-6 in patients with low concentration of serum C-reactive protein].

Both C-reactive protein (CRP) and Serum amyloid A protein(SAA) are determined as an indicator of inflammation and tissue damage. But serum CRP in the range less than 10 micrograms/dl was not correlated with SAA. We determined CRP, SAA and interleukin-6(IL-6) in sera of patients who had received corticosteroid therapy. CRP decreased extremely after administration of more than 50 mg/day corticosteroid. However, SAA and IL-6 levels changed independently from CRP levels. In the patients under long-term corticosteroid therapy, CRP decreased rapidly to the level below the reference range and remained low, while SAA decreased but to the level in the reference range, and IL-6 levels was unchanged. When they were complicated with infectious disease during the corticosteroid therapy, CRP increased to or above the reference level but not so markedly, however, SAA increased markedly and exceeded the upper reference range three or four days earlier than the day of CRP increase. It is suggested that serial SAA monitoring will be reliable for the early diagnosis of infection if the patients is taking corticosteroid therapy.

Adolescent↗
PubMed1994

[Phylogenetic relationships of mastomys to mouse and rat deduced from satellite DNA sequences].

Mastomys is a rodent with a intermediate size between mouse and rat, and classify as a subgenus of Praomys coucha. Our study determined the phylogenetic evolutional relationships of mastomys to mouse and rat deduced from satellite DNA sequences. Genomic DNA was extracted from each liver and digested by restriction enzymes (EcoRI and BglII). After separation and purification, we obtained these EcoRI/BglII-0.5 Kbp repeated elements and determined the nucleotide sequences. The similarities and speed of evolution were 86.06% and 0.1578 between mastomys and mouse, 85.56% and 0.1671 between mastomys and rat, and 83.91% and 0.1863 between rat and mouse, indicating that mastomys was evolutionary classified between mouse and rat, and closely related with mouse.

Animals↗
PubMed1992

Development of human monoclonal antibodies against human cytomegalovirus.

Human monoclonal antibodies (HMAbs) against human cytomegalovirus (HCMV) have been developed by fusion of human spleen cells and human lymphoblastoid cell lines (NP101 and NP197). The cell line NP101 had great advantages in its high fusion frequency and the stability of the resultant hybridomas. The specificity of HMAbs was confirmed by enzyme-linked immunosorbent assay (ELISA) and immunofluorescence staining. Two of the six HMAbs obtained, which were IgG3 subclass, neutralized viral infectivity in the absence of complement. The neutralizing activity of one of these two HMAbs was enhanced in the presence of human complement, whereas the other was not. Another IgG1 subclass HMAb neutralized viral infection only in the presence of complement. The remaining three HMAbs showed no neutralizing activity. Those HMAbs may provide an important approach to studying human immune responses to HCMV. HMAbs having neutralizing activity may prove to be useful for passive immunotherapy of HCMV diseases.

Antibodies, Monoclonal↗
PubMed1989

Comparison of efficacy, toxicity and pharmacokinetics of free adriamycin and adriamycin linked to oxidized dextran in rats.

Adriamycin linked to oxidized dextran (ADM-OXD) via Schiff's base formation was compared with free adriamycin with regard to antitumor activity, acute toxicity and plasma pharmacokinetics in rats following i.v. administration. ADM-OXD showed higher activity against Walker carcinosarcoma 256 than free adriamycin. On the other hand, the acute toxicity of ADM-OXD was about three times less than that of free adriamycin. In contrast to free adriamycin, a very high plasma level of adriamycin was found after i.v. administration of ADM-OXD. The area under the plasma concentration curve with ADM-OXD was about 160-fold higher than with free adriamycin. Thus, the improvement of the therapeutic index in the case of ADM-OXD might be due to the difference in the disposition of ADM-OXD and free adriamycin in rats.

Animals↗