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Biomedical subjects

H Ishihara

Publications and source records attributed to H Ishihara.

At least 217 records · Page 12Linked to original sources
PubMed1997

Evidence for functional roles of Crk-II in insulin and epidermal growth factor signaling in Rat-1 fibroblasts overexpressing insulin receptors.

We examined the potential role of Crk-II in insulin and epidermal growth factor (EGF) signaling in Rat-1 fibroblasts overexpressing insulin receptors. Crk is an SH2 and SH3 domain-containing adaptor protein that has been reported to associate with p130cas, paxillin, c-cbl, c-abl, Sos, and C3G in vitro. Insulin- and EGF-induced association of Crk-II with these molecules was assessed by immunoblotting of anti-Crk-II precipitates in Rat-1 fibroblasts overexpressing insulin receptors. Neither insulin nor EGF treatment induced Crk-II association with either Sos or C3G. Basal tyrosine phosphorylation of c-abl and its constitutive association with Crk-II were not further increased by insulin or EGF. p130cas and paxillin were heavily tyrosine phosphorylated in the basal state. Both insulin and EGF stimulated their dephosphorylation, followed by p130cas-Crk-II dissociation and paxillin-Crk-II association, although the magnitude of these effects was greater with insulin than with EGF. Interestingly, EGF, but not insulin, stimulated tyrosine phosphorylation of c-cbl and its association with Crk-II. To investigate the functional roles of Crk-II in mitogenesis and cytoskeletal rearrangement, we performed microinjection analysis. Cellular microinjection of anti-Crk-II antibody inhibited EGF-induced, but not insulin-induced, DNA synthesis. Insulin, but not EGF, stimulated cytoskeletal rearrangement in the cells, and microinjection of anti-Crk-II antibody effectively inhibited insulin-induced membrane ruffling, suggesting that Crk-II is involved in insulin-induced cytoskeletal rearrangement. These results indicate that Crk-II functions as a multifunctional adaptor molecule linking insulin and EGF receptors to their downstream signals. The presence of c-cbl-Crk-II association may partly determine the signal specificities initiated by insulin and EGF.

Animals↗
PubMed1997

Targeting of GLUT1-GLUT5 chimeric proteins in the polarized cell line Caco-2.

Caco-2, a human differentiated intestinal epithelial cell line, is a promising model for investigating the mechanism of polarized targeting of apical and basolateral membrane proteins. We stably transfected rat GLUT5 cDNA and rabbit GLUT1 cDNA into Caco-2 cells with an expression vector. Immunohistochemical study revealed that the GLUT5 protein expressed was localized at apical membranes and that the GLUT1 expressed was present primarily in the basolateral membranes of cells grown on permeable support. Next, to investigate the domain responsible for determining apical vs. basolateral sorting in glucose transporters, we prepared several GLUT1-GLUT5 chimeric cDNAs and transfected them into Caco-2 cells. A GLUT1 [N terminus approximately sixth transmembrane domain (TM6)]-GLUT5 [intracellular loop (IL) approximately C terminus] chimera was observed exclusively at the apical membrane, while GLUT1 (N terminus approximately IL)-GLUT5 (TM7 approximately C terminus) and GLUT1 (N terminus approximately TM12)-GLUT5 (C-terminal domain) chimeras were observed mainly at the basolateral membrane, a localization similar to that of GLUT1. Moreover, using a recombinant adenovirus expression system, we expressed a GLUT5 (N terminus approximately TM6)-GLUT1(IL)-GLUT5(TM7 approximately C-terminus) chimera, which was observed at the basolateral membrane. Based on these results, the C-terminal domain does not determine isoform-specific targeting of GLUT1 and GLUT5. Rather, it is the intracellular loop in glucose transporters that appears to play a pivotal role in apical-basolateral sorting signals in Caco-2 cells.

Animals↗
PubMed1997

Hepatic accumulation of glutamic acid branched neogalactosyllipid modified liposomes.

We synthesized branched type galactosyllipid derivatives for liposome modification for the targeting of asialoglycoprotein receptors on the surface of liver cells. Galactose was coupled to the alpha- and gamma-carboxyl groups of glutamic acid via a triethyleneglycol spacer, then this glutamic moiety was bound to the lipid anchor. Ricinus communis agglutinin (RCA120) induced the agglutination of liposomes modified with mono-, bi- and tri-antennary neogalactosyllipid. With the bi- or tri-antennary derivatives, agglutination was observed at fewer galactosyl residues on the liposomes. We examined the effect of the branching structure in vivo. The difference in accumulation of liposomes between non-branched type neogalactosyllipid and branched type neogalactosyllipid was not large. Liver accumulation of liposomes depended on the galactosyl residues. The number of galactosyl residues was more effective for accumulation in the liver than for branching. We studied the effect of asialofetuin preinjection on the hepatic accumulation of neogalactosyllipid modified liposomes. Hepatic accumulation of liposomes was inhibited by preinjection of asialofetuin. The effect of preinjection was almost equal among the ligands. These results show that the saccharide density on the liposome surface seemed to be a more important factor than the branching structure of the ligand for liver targeting.

Animals↗
PubMed1997

Synthesis and application of neoglycolipids for liposome modification.

We synthesized various glycolipid derivatives and examined the in vivo behaviors of liposomes modified with these novel glycolipid derivatives. Gal-t-psa (1,¿8-(2-hexadecyloctadecanoylamido)-3,6-dioxaoctyl¿-beta-D- galactoside), Lac-t-psa (3, 8-(2-hexadecyloctadecanoylamido)-3,6-dioxaoctyl beta-D-lactoside) and GalNAc-t-psa (4, 8-(2-hexadecyloctadecanoylamido)-3,6-dioxaoctyl 2-acetamido-beta-D-galactopyranoside) modified liposomes were recognized by the liver. Lac-t-psa (3) modified liposome was accumulated to the highest degree, followed by GalNAc-t-psa (4) modified liposome and then Gal-t-psa (1) modified liposome. The intrahepatic distributions of Gal-t-psa (1), GalNAc-t-psa (4), Glc-t-psa (2, 8-(2-hexadecyloctadecanoylamido)-3,6-dioxaoctyl beta-D-glucopyranoside) and Lac-t-psa (3) modified liposomes were investigated. GalNAc-t-psa (4) and Lac-t-psa (3) modified liposome were accumulated to greater extents than Gal-t-psa (1) modified liposome in hepatic parenchymal cells. The intrahepatic distribution of these liposomes showed that Lac-t-psa (3) and GalNAc-t-psa (4) were preferable to Gal-t-psa (1) for the selective delivery of liposomes to hepatic parenchymal cells.

Animals↗
PubMed1997

Acetylcholine-induced relaxation of rat aorta is greatest during estrus in the sexual cycle.

Acetylcholine (ACh)-induced relaxation in the aortae precontracted with norepinephrine was significantly enhanced in the aortae from estrus (E) rats, compared with that in those from metestrus (D-1), diestrus (D-2) and proestrus (PE) rats. NG-Nitro-L-arginine methyl ester (L-NAME) inhibited the endothelium-dependent relaxation in E rats. These results suggest that there is a difference in ACh-induced relaxation of the thoracic aorta during the sexual cycle of rats, and the relaxation is greatest in E of the sexual cycle; this may be due to a difference in nitric oxide synthesis in the endothelium in the sexual cycle.

Acetylcholine↗
PubMed1997

Generation of basidiomycetous hyphal cell-aggregates by addition of the Arg-Gly-Asp motif-containing fragment of high-molecular-weight cell-adhesion protein MFBA derived from the basidiomycete Lentinus edodes.

The Arg-Gly-Asp (RGD) motif-containing fragment of high-molecular-weight cell-adhesion protein MFBA derived from Lentinus edodes caused a significant aggregation of the fragmented hyphal cells of Schizophyllum commune. This fungal cell-aggregation was inhibited by a previous treatment of the cells with the Gly-Arg-Gly-Asp-Ser-Pro peptide, but not with the Gly-Arg-Gly-Glu-Ser-Pro peptide, showing that the RGD motif is essential for the cell-aggregation activity.

Basidiomycota↗
PubMed1997

Isolation and molecular characterization of four arginine/glutamate rich polypeptides from the seeds of sponge gourd (Luffa cylindrica).

Four arginine/glutamate rich polypeptides referred to as 5 k-, 6.5 k-, 12.5 k-, and 14 k-AGRPs were purified to homogeneity by gel filtration on Sephadex G-75 followed by CM-cellulose, butyl-Toyopearl 650 M, and reverse-phase HPLC from the seed of sponge gourd (Luffa cylindrica). Tricine SDS-PAGE indicated that 5 k- and 6.5 k-AGRPs are single polypeptides, but 12.5 k- and 14 k-AGRPs consist of two polypeptide chains, which are linked by disulfide bond(s). The N-terminal amino acid sequences of four AGRPs were analyzed by a gas-phase sequencer, and the result indicated that they are distinct molecules. Comparison of the sequences with those of proteins in the protein data base demonstrates that 5 k- and 6.5 k-AGRPs have a significant homology with a basic peptide from pumpkin seeds and with cocoa seed vicilin, respectively, and that 12.5 k- and 14 k-AGRPs are related to 2S seed storage proteins. Furthermore, it was assumed that the four AGRPs might occur in the protein bodies within cells of the seed.

Amino Acid Sequence↗
PubMed1997

[Preanesthetic meals in elective surgical patients].

This study investigated the effect of preanesthetic meals on the volume and pH of gastric contents in forty elective surgical patients ranging in ages from 20 to 60 years. Twenty patients who were given either isotonic beverage 250 ml or apple juice 250 ml on the morning of the operative day were subjected as control group and twenty patients of the breakfast group took two slices of bread with the above drink. About seven hours following drinking and feeding, the mean values of gastric volume were 20.9 +/- 18.3 ml in the control group and 19.2 +/- 16.3 ml in the breakfast group. The mean values of gastric pH were 4.3 +/- 2.3 in the drink group and 4.6 +/- 2.3 in the breakfast group. There were no significant differences in the gastric volume and pH between the two groups. However, very small amount of the bread was detected in the gastric fluid of three patients in the breakfast group. As preanesthetic drinking and feeding are advantageous for reducing the anxieties of preoperative patients and also for their nutrition during operation, it is encouraging that eating two slices of bread did not induce a significant effect of gastric volume or pH. The minute fragment of bread seems to have no clinically significant effect.

Adult↗
PubMed1997

[Simultaneous operation of emergency coronary artery bypass grafting and gastrotomy with suturing ulcer].

A 62-year-old man with complaints of severe chest pain came to our hospital. An emergency coronary angiography was performed and he was diagnosed as having acute myocardial infarction. Due to severe triple vessels disease he was referred to the department of Cardiovascular Surgery to undergo emergency coronary artery bypass grafting. In the coronary care unit, sudden hematoemesis due to hemorrhagic gastric ulcer occurred, however, just when he was going to be transferred to the operation room. Because the gastric bleeding was thought to be serious under extracorporeal circulation, which was indispensable for coronary artery bypass grafting, gastrotomy with suturing ulcer was performed prior to median sternotomy with use of intraaortic balloon pumping. Severe infection was not complicated. His postoperative course was uneventful.

Coronary Artery Bypass↗
PubMed1997

[A case of left atrial thrombi following mitral valvuloplasty for mitral regurgitation associated with idiopathic thrombocytopenic purpura].

A 58-year-old man, who had been submitted to an indication of mitral valvuloplasty (MVP) because of severe mitral regurgitation, was diagnosed as idiopathic thrombocytopenic purpura (ITP). In order to diminish the peri-operative blood loss, splenectomy and high-dose bolus administration of gamma-globulin (400 mg/kg/day) were performed at the time of two weeks and during last five days respectively, prior to MVP surgery. In early post-operative stage, anticoagulant therapy was held down considering ITP, and we were not troubled with bleeding throughout the peri-operative period. On the fourteenth post-operative day, however, mural thrombi in the left atrium were pointed out by transesophageal echocardiography (TEE). Then anticoagulant therapy was reinforced by increasing the warfarin dose. When TEE was restudied on 36th, post-operative day, mural thrombi had almost disappeared. Although it is commonly known that mitral valvuloplasty has low risk of thrombosis and less necessity of anticoagulation, hyper coagulability may be feasible in the early post-operative period of our case whose ITP is meticulously treated prior to surgery. Suitable anticoagulation should have been performed regardless of ITP immediately after MVP.

Heart Atria↗
PubMed1997

[Short-term results of leaving elephant trunk in type A aortic dissection].

Elephant trunk prosthesis was devised for treatment of multiple aortic aneurysm. But it has applied to type A aortic dissection. Because residual false lumen often dilates gradually and come to have the risk of rupture. Two patients who were diagnosed as type A aortic dissection were performed total aortic arch replacement with Elephant trunk prosthesis on distal anastmosis. Postoperative diagnostic images showed that residual false lumen of proximal descending aorta were thrombosed. Now it is not necessary to perform next extensive aortic replacement. Complication which is threatened in using Elephant trunk prosthesis, paraplegia, thromboembolism, kinking of prosthesis could have been avoided by setting suitable length and diameter of the prosthesis.

Aortic Dissection↗
PubMed1997

[Changes in epidural pressure during total intravenous anesthesia with propofol, fentanyl and ketamine].

Although ketamine elevates cerebrospinal fluid pressure (CSFP) with an increase in cerebral blood flow, sedatives such as benzodiazepines, barbiturates and opioids have been reported to inhibit it. In this study, we evaluated the changes in epidural pressure (EP) as a good index for CSFP during total intravenous anesthesia with propofol-fentanyl-ketamine (PFK) compared to isofluranenitrous oxide anesthesia (GOI). Twelve patients for gastrectomy were allocated to two groups: PFK (n = 6) and GOI group (n = 6). In PFK group, epidural pressure did not increase during the anesthesia, and was significantly lower than in GOI group (30 and 180 min after induction of anesthesia, and 0, 30 and 60 min after stopping anesthetic administration). The present data suggest that PFK may safely be used for patients with intracranial hypertension.

Aged↗
PubMed1997

Depressed mechanoenergetics and compensatory responses in idiopathic dilated cardiomyopathy.

The aim of this study was to clarify that the depressed mechanoenergetics in patients with idiopathic dilated cardiomyopathy (DCM) resulted from compensation for the decreased contractility. The study population consisted of eight control subjects, with normal left ventricular size and ejection fraction and 31 patients with DCM. Left ventricular end-systolic elastance (Ees), effective arterial elastance (Ea), external work (EW), and the pressure-volume area (PVA) were measured, using a dual-field volume conductance catheter equipped with a micromanometer-tipped catheter. Ea/Ees was evaluated as ventriculoarterial coupling. Normal hearts worked at almost optimal condition (defined as Ea/Ees = 1/2), while ventriculoarterial coupling was far from the optimum (Ea > Ees) in patients with DCM. Ees in patients with DCM was less than that in control subjects; however, Ea was similar in the two groups. The mismatch of Ea/Ees observed in DCM leads to depressed mechanoenergetics as a result of the compensatory response to maintain adequate blood pressure. Volume enlargement plays an important role in maintaining adequate blood pressure and cardiac output in the course of chronic deterioration of contractility.

Blood Pressure↗
PubMed1996

Mitochondrial DNA is required for regulation of glucose-stimulated insulin secretion in a mouse pancreatic beta cell line, MIN6.

To determine whether mtDNA and mitochondrial respiratory function in pancreatic beta cells are necessary for the phenotypic expression of glucose-stimulated insulin secretion, we used a cultured mouse pancreatic beta cell line, MIN6, and two derivative lines, mtDNA knockout MIN6 (rho0 MIN6) and mtDNA repopulated cybrid MIN6. The MIN6 cells retain the property of glucose-stimulated insulin secretion, but their mtDNA knockout induced the loss of mitochondrial transcription, translation, and respiration activity, without inhibition of transcription of the insulin gene or loss of succinate dehydrogenase activity, indicating that the observed mitochondrial dysfunction in rho0 MIN6 cells was not due to a cytotoxic side effect derived from the mtDNA knockout. Moreover, the mtDNA depletion also inhibited both the glucose-stimulated increase in the intracellular free Ca2+ content and the elevation of insulin secretion. The possibility of the involvement of nuclear genome-encoded factors in this process was excluded by the observation that the missing sensitivity to extracellular glucose stimulation in rho0 MIN6 cells was restored reversibly by repopulation with foreign mtDNA and isolating cybrid MIN6 clones. Therefore, these findings provide unambiguous evidence for the involvement of the mitochondrial dysfunction induced by mtDNA impairment in developing pathogeneses of some forms of diabetes mellitus.

Animals↗
PubMed1996

Function of Xenopus cystic fibrosis transmembrane conductance regulator (CFTR) Cl channels and use of human-Xenopus chimeras to investigate the pore properties of CFTR.

To explore the relationship between structure and function in the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel, we studied Xenopus CFTR. We found that the anion permeability sequence of cAMP-activated Cl- currents in the apical membrane of Xenopus A6 epithelia differed from that of cAMP-activated Cl- currents in human epithelia expressing CFTR. To understand the molecular basis for this difference and to learn whether CFTR from another species would have properties similar to human CFTR, we assembled a full-length Xenopus CFTR cDNA from A6 cells. Expression of Xenopus CFTR in HeLa cells generated cAMP-activated whole-cell currents and cAMP-dependent protein kinase-activated single channels that resembled those of human CFTR with the exception that the anion permeability sequence was different (Br- = I- > Cl- in Xenopus CFTR and Br- = Cl- > I- in human). In addition, the single-channel conductance of Xenopus CFTR was increased. To investigate protein regions that account for these differences, we constructed chimeric proteins by replacing either the first or second membrane-spanning domain of human CFTR with the equivalent region of Xenopus CFTR (hX1-6 and hX7-12, respectively) and examined their function in HeLa cells. We found that the anion permeability sequence (Br- = I- > Cl-) and single-channel conductance of hX1-6 resembled that of Xenopus CFTR expressed in HeLa cells, whereas hX7-12 had properties like those of human CFTR. However, the gating of hX1-6 showed a flickery behavior. The altered gating of hX1-6 was attributed to residues in the first extracellular loop of Xenopus CFTR because mutation of residues in that region to the corresponding residues of human CFTR produced gating behavior similar to that of human CFTR. These data suggest that sequence differences in the first membrane-spanning domains are responsible for the differences in the permeation properties of human and Xenopus CFTR and that the first extracellular loop influences channel gating.

Amino Acid Sequence↗