Biomedical subjects

J Takagi

Publications and source records attributed to J Takagi.

At least 55 records · Page 3Linked to original sources

PubMed1997

Structural interlock between ligand-binding site and stalk-like region of beta1 integrin revealed by a monoclonal antibody recognizing conformation-dependent epitope.

Integrin activation and sebsequent ligand binding to it are regulated by intracellular mechanisms called inside-out signaling, which are not fully understood and are accompanied by dynamic structural changes of the integrin molecule itself. A monoclonal antibody recognizing a conformation-dependent epitope on human beta1 integrin was produced and characterized in detail. This antibody, AG89, reacted with human integrin beta1 chain regardless of the alpha subunit. AG89 can recognize resting state beta1 integrin on the cells, but the reactivity is increased approximately 2-fold upon integrin activation by activating anti-beta1 antibodies and approximately 3-fold by Mn2+. Furthermore, occupation of the ligand-binding pocket by a soluble ligand (RGD peptide for alpha(v)beta1 and CS-1 peptide for alpha4beta1) resulted in maximum binding of AG89, indicating that the epitope for AG89 is exposed during the conformational changes of beta1 integrin upon activation/ligation. Epitope mapping by using interspecies chimeric beta1 revealed that the epitope for AG89 lies within residues 426-587, which corresponds to the cysteine-rich repeat structure located in the middle of the beta1 chain. The fact that binding of AG89 itself could activate the resting beta1 integrin indicates that exposure of the AG89 epitope in the membrane-proximal stalk-like domain and "opening" of the ligand-binding pocket at the outermost domain are physically linked. We propose that the integrin "signaling" is mediated by this direct physical transduction of conformational information along the integrin molecule.

Antibodies, Monoclonal↗

PubMed1997

Central facial weakness due to medial medullary infarction: the course of facial corticobulbar fibres.

Two patients are reported with contralateral hemiparesis including a face of supranuclear type, caused by an infarct of the unilateral ventromedial part of the upper medulla. Data from these patients support the hypothesis that part of the corticobulbar fibres supplying the lower facial muscles descend ipsilaterally in the ventromedial part of the upper medulla and then, after decussation, ascend rostrally to the contralateral facial nucleus.

Adult↗

PubMed1997

DISTRIBUTION OF SOMATOSTATIN RECEPTOR TYPE 3 IN THE RAT: IMMUNOHISTOCHEMICAL STUDY.

Somatostatin receptor type 3 (SSTR-3) was identified immunohistochemically in the rat tissues using specific anti-SSTR-3 serum which was raised in New Zealand white rabbits immunized with a conjugate of synthetic SSTR-3 peptide (28-41) with bovine serum albumin. Immunohistochemical analysis was performed by avidin-biotin complex method. SSTR-3 immunoreactivity was visualized in the central nervous system, anterior pituitary, gastric and duodenal mucosa, Auerbach's and Meissner's nervous branch of gastrointestinal tract, adrenal medulla, testis and pancreas. Significant staining was detected in neural perikarya, axons and dendrites. When using antiserum preincubated with synthetic SSTR-3 peptide (28-41) or rat anterior pituitary homogenate which contains SSTR-3 peptide, no significant stain of the anterior pituitary or neurons in the hypothalamus was detected. These findings suggest that SSTR-3 is widely distributed and that this method is valuable in studying the distribution of SSTR-3 in rats.

Journal Article↗

PubMed1997

Organ distribution of iodide transporter (symporter) in the rat: immunohistochemical study.

Iodide transporter/symporter (NIS) was identified immunohistochemically in rat tissues using specific antipeptide serum. Anti-NIS serum was raised in New Zealand white rabbits immunized with a conjugate of synthetic NIS peptide (39-53) with bovine serum albumin. Immunohistochemical analysis was performed by avidine-biotin complex method. NIS immunoreactivity was visualized in the thyroid gland, gastric and small intestine mucosa, anterior pituitary, adrenal medulla, pancreatic islets, kidney, chorioid plexus and several brain and spinal cord nuclei. When using antiserum preincubated with synthetic NIS peptide (39-53) or rat thyroid homogenate containing NIS, no significant stain of the thyroid gland was detected. These findings suggest that NIS is widely distributed and that the method used is suitable for studying the distribution of NIS in rats.

Journal Article↗

PubMed1996

Inhibition of adenylyl cyclases by 12(S)-hydroxyeicosatetraenoic acid.

The inhibition of adenylyl cyclase (AC) by a 12-lipoxygenase metabolite of arachidonic acid, 12(S)-hydroxy-5Z,8Z,10E,14Z-eicosatetraenoic acid (12-HETE), was investigated using three different kinds of cells: NRK-49F (normal rat kidney fibroblasts), AtT-20 (mouse pituitary tumor cell line) and HL-60 (human leukemia cells) cells. The inhibition was very obvious in NRK-49F and AtT-20 cells, but it was almost negligible in HL-60 cells. There was no difference in terms of the binding of 12-HETE to NRK-49F and HL-60 cells. Pretreatment of NRK-49F cells with pertussis toxin almost completely ADP-ribosylated Gi proteins, but it did not affect the inhibition of 12-HETE on AC in this cell. This result excludes the involvement of Gi proteins in 12-HETE-mediated inhibition of AC. It was revealed that the characteristics of ACs in these cells were quite different in response to agonists and forskolin, suggesting that these cells do have different isoform of AC. We conclude that 12-HETE inhibits the activity of AC depending upon the isoform.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗

PubMed1996

Gas transfer and blood compatibility of fluorinated polyimide membranes.

Fluorinated polyimide derived from 2,2'-bis(3,4-dicarboxyphenyl) hexafluoropropane dianhydride (6FDA) and bis[4-(4-aminophenoxy) phenyl]sulfone (APPS) was synthesized to develop a novel membrane oxygenator combining excellent gas transfer and blood compatibility. The asymmetric gas exchange membranes of 6FDA-APPS made by a dry/wet process consisted of an ultrathin and defect-free skin layer supported by a porous substructure. O2 transfer through the 6FDA-APPS membrane was extremely augmented as compared with that of the presently available membrane, poly(dimethylsiloxane), and the previously reported 6FDA-DDS membrane. Since CO2 transfer through the 6FDA-APPS membrane increased with a decrease in CO2 pressure according to dual-mode transport theory, CO2 from the membrane was selectively removed at low CO2 pressure. For the evaluation of in vitro blood compatibility, the platelet adhesion and the plasma protein adsorption on the surface of the 6FDA-APPS membrane were observed by using scanning electron microscopy and the amounts of platelet and plasma protein were determined by an amino acid analyzer. The results indicated that the fluorinated polyimide membranes showed excellent blood compatibility.

Blood Gas Analysis↗

PubMed1996

Dual functions of transglutaminase in novel cell adhesion.

Transglutaminases (TGases) are enzymes which catalyze cross-link formation between glutamine residues and lysine residues in substrate proteins. In the present study, we report for the first time that a representative enzyme, blood coagulation factor XIIIa (FXIIIa), is capable of mediating adhesion of various cells. When coated on plastic surfaces FXIIIa promoted adhesion and spreading of various cells of both normal and tumor origin, in a concentration-dependent manner. The adhesion was not inhibited by antibodies against possible contaminants in the enzyme preparation such as fibronectin and vitronectin, but was completely inhibited by a polyclonal antibody against the enzyme. Therefore, if there were any contaminating cell adhesive substrates in the enzyme preparation, they cannot account for the observed cell adhesion to the enzyme; FXIIIa itself mediates the cell adhesion. Furthermore, phosphorylation of tyrosine residues in 120 kDa and 70 kDa proteins was clearly shown in human fibroblasts adhering to the enzyme. Formation of actin stress fibers was also unambiguously observed in the adhering cells. These biochemical reactions, which are also observed when cells adhere to a typical cell adhesion protein, fibronectin, are believed to be of importance in the process of cell adhesion. This adhesion activity of FXIIIa was dependent on its TGase activity, because both a modification of the active center cysteine with iodoacetamide and the addition of ammonium ion abolished the cell adhesion activity along with the enzyme activity. The cell adhesion to fibronectin, however, was not affected by these treatments. The effects of various anti-integrin antibodies suggested that both alpha v beta 3 and beta 1 family integrins participated in the cell adhesion to FXIIIa. Taken together, these data demonstrate for the first time that there is a unique TGase activity-mediated cell adhesion. This novel function of the enzyme may be of physiological importance.

Alkylating Agents↗

PubMed1996

A possible involvement of monoaminergic and opioidergic systems in the analgesia induced by electro-acupuncture in rabbits.

To elucidate mechanisms involved in analgesia induced by effects of electro-acupuncture (EAP), effects of EAP on evoked potentials and release of substance P (SP) following tooth pulp stimulation (ST) in the superficial layers of the trigeminal nucleus caudalis (Vc-I-II) were studied in the rabbit. The potentials evoked by ST were composed of two main components with conduction velocity of ca. 30 m/sec (fast component) and ca. 12 m/sec (late component). The late component was significantly inhibited by morphine (10 mg/kg, i.v.) or CP-96,345 (5 mg/kg, i.v.), an SP antagonist. This inhibitory effect of morphine was antagonized by naloxone (1 mg/kg, i.v.) or methysergide (5 mg/kg, i.v.). In addition, the late component was significantly inhibited by EAP, which was observed in ca. 70% of the rabbits examined. This EAP-induced inhibitory effect was antagonized by naloxone (1 mg/kg, i.v.) or methysergide (5 mg/kg, i.v.), but not by prazosin (5 mg/kg, i.v.) and yohimbine (1 mg/kg, i.v.). The stimulus-evoked SP release was inhibited by EAP, which was significantly antagonized by pretreatment with naloxone (1 mg/kg, i.v.) or methysergide (5 mg/kg, i.v.). These results suggest that one of the mechanisms of analgesia induced by EAP is due to inhibition of the stimulus-evoked SP release in the Vc-I-II through activation of the descending serotonergic systems linking up with opioidergic systems.

Acupuncture Analgesia↗

PubMed1996

[Favorable prognosis in a case of recurrent cerebral infarctions caused by chronic disseminated intravascular coagulation due to ovarian carcinoma].

We report a 53-year-old woman who developed a cerebral infarction in the left middle cerebral artery area. Although she did not have a hemorrhagic tendency, the results of her examination showed a chronic disseminated intravascular coagulation (DIC) and a left ovarian carcinoma. Gabexate mesilate (FOY) and nafamostat mesilate (FUT) were not effective for her DIC. One month after admission, she had another cerebral infarction in the right posterior cerebral artery area. She was treated by heparin sodium and her chronic DIC improved. Then, she was operated on for ovarian carcinoma; the histologic finding was clear cell adenocarcinoma. No recurrence of DIC nor cerebral infarction was observed postoperatively. Patients with cerebral infarctions caused by chronic DIC due to ovarian carcinoma have been reported in the literature, but few patients had a favorable prognosis such as the present case. Heparin therapy appears to be the treatment of choice for chronic DIC rather than FOY or FUT, if there is no hemorrhagic tendency. When a patient with DIC develops cerebral infarction, one should always investigate for possible malignant tumors.

Adenocarcinoma, Clear Cell↗

PubMed1995

Identification of factor-XIIIa-reactive glutaminyl residues in the propolypeptide of bovine von Willebrand factor.

von Willebrand factor is a large multimeric plasma protein which plays important roles in platelet aggregation, blood coagulation and probably also in the adhesion of endothelial cells. A 100-kDa propeptide, called the propolypeptide of von Willebrand factor (pp-vWF), is generated during biosynthesis. We found that pp-vWF served as a substrate for transglutaminases including human factor XIIIa and guinea pig liver transglutaminase [Usui, T., Takagi, J. & Saito, Y. (1993) J. Biol. Chem. 268, 12311-12316]. As such, it could form cross-linked copolymers with the extracellular matrix protein, laminin, making it all the more likely that pp-vWF plays a role in cell adhesion phenomena [Takagi, J., Sudo, Y., Saito, T. & Saito, Y. (1994) Eur. J. Biochem. 222, 861-867]. In this work, we identified the Gln residues in pp-vWF specifically reacting with blood coagulation factor XIIIa as amine acceptors. The fluorescent amine, dansylcadaverine, was employed for labeling the enzyme-reactive sites of the protein. Following partial proteolysis, fragments containing the labeled Gln residues were isolated by passage through an anti-dansyl affinity chromatographic column. Amino acid sequence analyses of the fragments revealed that, out of about 40 Gln residues in pp-vWF, only four could be modified in the factor-XIIIa-catalyzed reaction.

Amino Acid Sequence↗

PubMed1995

A fruiting body-specific cDNA, mfbAc, from the mushroom Lentinus edodes encodes a high-molecular-weight cell-adhesion protein containing an Arg-Gly-Asp motif.

A cDNA clone (designated mfbAc), encoding 2157 amino acids (aa), was isolated from a mature fruiting-body cDNA library of the edible mushroom Lentinus edodes. The mfbA transcript was abundant in mature fruiting bodies, detectable in immature fruiting bodies but absent in earlier developmental stages and in the vegetative mycelium. Although more abundant in the pileus than the stipe, only low levels were found in the gill tissue. The deduced MFBA protein (234.5 kDa) contained a cell-surface attachment-promoting Arg-Gly-Asp (RGD) motif. MFBA was produced in Escherichia coli using a maltose-binding protein (MBP) fusion vector, but it was cleaved into four fragments even in a protease-deficient host. A 425-aa MFBA peptide containing the RGD motif (named MFBA(582-1006) peptide) was successfully produced using the phage T7 expression system. This MFBA(582-1006) peptide exhibited a cell adhesion and spreading activity toward mammalian cells. This activity of the MFBA fragment was competitively inhibited by the Gly-Arg-Gly-Asp-Ser-Pro peptide but not by the Gly-Arg-Gly-Glu-Ser-Pro peptide, showing that the RGD motif of MFBA is essential for the cell-binding activity.

Amino Acid Sequence↗

PubMed1995

Differential role of protein tyrosine phosphorylation/dephosphorylation in affinity regulation of beta 1 and beta 3 integrin in human fibroblasts.

We investigated the effects of protein tyrosine phosphatase inhibitors, pervanadate and phenylarsine oxide (PAO), on the beta 1 and beta 3 integrin-mediated cell-substrate interaction using normal diploid human fibroblast. Pervanadate treatment of the cells in suspension state resulted in highly elevated levels of cellular protein tyrosine phosphorylation accompanied by loss of beta 1 integrin-mediated adhesion to substrata (i.e., collagen and laminin). In contrast, beta 3 integrin-mediated adhesion to substrata (i.e., fibronectin and vitronectin) of these cells was less affected. Moreover, pervanadate could reverse beta 1 integrin-dependent adhesion, and cells already adhered on collagen or laminin, but not on fibronectin or vitronectin, came off within 30 min upon pervanadate treatment. These effects are likely to be directly mediated by increased cellular protein tyrosine phosphorylation, because another chemical compound, PAO, which also inhibits protein tyrosine phosphatase through a quite different mechanism, also exhibited the specific deterioration of beta 1 integrin-mediated cell-substrate interaction. Upon treatment with these protein tyrosine phosphatase inhibitors, the well developed actin stress fibers were disrupted resulting in the rounding up of cells on fibronectin and vitronectin substrate though they were still attached to the beta 3 integrin-dependent substrates. Using immunoprecipitation and anti-phosphotyrosine immunoblotting, beta 1 integrin itself was shown not to be tyrosine phosphorylated. These results indicate that affinity regulation of beta 1 and beta 3 integrin is differentially controlled and that the specific regulation of beta 1 integrin is due to certain cellular component(s) whose activity is modulated by tyrosine phosphorylation/dephosphorylation.

Antibodies, Monoclonal↗

PubMed1994

Beta 1-integrin-mediated adhesion of melanoma cells to the propolypeptide of von Willebrand factor.

Cell-adhesion activity of the bovine propolypeptide of von Willebrand factor (pp-vWF) was assessed by means of an in vitro assay with several cell lines of both normal and tumor-cell origin. pp-vWF promoted adhesion and spreading of B16 mouse melanoma cells and G-361 human melanoma cells. However, it could not induce adhesion of any other cell lines tested including endothelial cells, normal fibroblasts, and tumor cells of sarcoma, carcinoma, neuroblastoma and leukemia origin. A monospecific polyclonal antibody against pp-vWF, but not against fibronectin, laminin, and von Willebrand factor (vWF), completely blocked the pp-vWF-mediated adhesion, indicating that the cell adhesion was due to the pp-vWF molecule and not due to possible contamination of these three well-known adhesive proteins. The cell-adhesion activity was also observed with human pp-vWF and, furthermore, the adhesion to both bovine and human pp-vWF was not affected by a peptide containing the Arg-Gly-Asp sequence while the peptide abolished the cell adhesion to vWF. The adhesion was completely dependent on Mg2+ and inhibited by Ca2+. Inhibition by an anti-(beta 1 integrin) mAb (4B4) indicates that the receptor for this protein belongs to the beta 1-integrin family. A monoclonal antibody (TC4) among several antibodies directed against bovine pp-vWF inhibited the B16 adhesion to immobilized pp-vWF. The epitope for this monoclonal antibody lies in a central 8-kDa portion of pp-vWF, suggesting that this region is important for the cell-adhesion activity. This idea was supported by the finding that purified 8-kDa fragment promoted adhesion of B16 cells in a concentration-dependent manner. As pp-vWF shows unique cell-type specificity in its adhesion activity, which is completely different from that of fibronectin, laminin, vWF and collagen, it may be a novel type of adhesive glycoprotein that utilizes a beta 1-integrin receptor.

Amino Acid Sequence↗

PubMed1994

Intravascular ultrasound of coronary arteries in children. Assessment of the wall morphology and the lumen after Kawasaki disease.

BACKGROUND: The long-term clinical issue in Kawasaki disease (KD) concerns the coronary artery lesion. Two-dimensional echocardiography and coronary angiography are routine examinations to evaluate the coronary lesions; however, these are not adequate to assess the wall morphology of the coronary artery (CA). Intravascular ultrasound imaging (IVUS), a new technology for the evaluation of the coronary artery lumen and wall morphology in vivo, was performed for patients after KD in their long-term follow-up, and we examined the new insights it gave. METHODS AND RESULTS: IVUS was performed during cardiac catheterization in 20 subjects (10 patients after KD who still had coronary aneurysms or regressed coronary aneurysms, 2 after KD who had no coronary abnormal lesion, and 8 control patients with congenital heart disease and normal CA). We evaluated the wall structure at 10 to 15 sites of the CA in each patient. IVUS was performed with a commercially available ultrasound imaging catheter. Four sites of a CA aneurysm in KD demonstrated a markedly dilated lumen without thickened intima. One site of a CA aneurysm with calcification demonstrated an asymmetrical lumen by a dense echo with acoustic shadows. Twenty-two sites of a regressed CA aneurysm demonstrated a marked symmetrical or asymmetrical thickening of the intima with a dense echo, in which the size of the lumen was similar to that at a site near a regressed aneurysm. The sites of angiographically normal CA revealed normal structures and a thin intima in many instances. Nine of 28 sites in KD with a CA abnormal lesion, particularly near a coronary aneurysm or regressed aneurysm, demonstrated a mild thickening of the intima. All the 10 sites in KD without a CA abnormal lesion and all the 25 sites in patients with congenital heart disease with normal CA demonstrated a smooth intima. CONCLUSIONS: This study demonstrated that the site of a regressed coronary aneurysm has a markedly thickened but smooth intima. The sites of angiographically normal CA after KD with or without a coronary lesion demonstrated normal IVUS findings in most instances but in some cases revealed a mild intimal thickening. IVUS is useful to evaluate the CA wall morphology and may contribute to the assessment of long-term CA sequelae and the possible development of arteriosclerotic changes in KD.

Adolescent↗

PubMed1994

A new "complex exercise test" for children with cardiac diseases.

In view of the characteristics of children's play or exercise activity, we designed a new method of exercise testing for children, called the "Complex Exercise Test" (CET), which includes three kinds of exercise test: (1) a graded exercise test on the treadmill, (2) a simple bending and stretching test to evaluate momentary exercise tolerance and (3) a swimming test with the ECG in the water. The CET was applied to a total of 25 cases which included 10 cases of postoperative tetralogy of Fallot and 15 cases with ventricular premature contraction (VPC). A two-way exercise test, which included the treadmill test and the simple bending and stretching test, was also applied to 36 postoperative cases (21 cyanotic and 15 non-cyanotic), 20 VPC cases and 16 control children without any organic cardiac disorders, for a total of 72 cases. The treadmill test was useful to evaluate the maximal exercise tolerance. The simple bending and stretching test, on the other hand, seemed useful for observation of VPC changes by exercise (including postoperative VPC), since the heart rate immediately after bending and stretching exceeded that observed at the time VPC disappeared during the treadmill test, thus proving VPC disappearance by this testing. With ECG during diving, it is possible to detect VPC associated with possible risks which could not be identified by the above two methods of testing. Since each one of the three different types of exercise tests has different characteristics, it seems desirable to control children's exercise after performing the CET.

Adolescent↗

PubMed1994

[A histopathological study on intraepithelial extensivity of carcinoma in situ of the uterine cervix in the period of sexual maturity].

To investigate intraepithelial extension of CIS, thirty-two cases of carcinoma in situ (CIS) of the uterine cervix in the period of sexual maturity were examined histopathologically and compared with ten cases of microinvasive carcinoma of the uterine cervix (restricted to Japanese criteria). The results were summarized as follows. 1) A positive correlation was found between the circular spreads and longitudinal ones in the lesion in CIS cases. 2) A positive correlation was found between circular spread and the depth of gland involvement in the lesion in CIS cases. 3) Circular spread of the lesion in microinvasive carcinoma cases was significantly more extended than that in CIS cases. These results suggest that CIS in the period of sexual maturity is intraepithelially extensive and that there is a high risk possibility of invading stroma when circular spread of the CIS lesion is greater than one fourth of the whole circle of the uterine cervix.

Adult↗

PubMed1993

A single chain 19-kDa fragment from bovine thrombospondin binds to type V collagen and heparin.

Location of the type V collagen-binding domain within bovine thrombospondin (TSP) was investigated by using fragments of reduced and alkylated TSP. A fragment of relative molecular mass (19 kDa) was isolated, which inhibited binding of 125I-TSP to type V collagen in a solid-phase binding assay. A direct binding assay using the 125I-labeled fragment confirmed that the fragment actually bound to collagen. The fragment retained specificity for the native structure of type V collagen like the intact TSP molecule. Its binding to the collagen, however, was not inhibited by Ca2+ in contrast to intact TSP. Amino acid sequence analysis of the 19-kDa fragment suggested that this fragment corresponded to Val333-Lys412, a part of the stalklike region in the human TSP primary structure. It was found that the fragment also bound well to heparin in a specific and saturable manner and, furthermore, binding to type V collagen was inhibited by soluble heparin. Removal of the N-linked sugar chain from this fragment resulted in a 14-kDa fragment. The deglycosylated fragment retained the ability to bind to type V collagen as well as heparin. These results suggest that a type V collagen-binding site is present in the 80-residue portion of bovine TSP (Val333-Lys412) and is likely to be identical or lie very close to a heparin-binding site, which exists in the type I repeat structure.

Amino Acid Sequence↗

PubMed1993

Propolypeptide of von Willebrand factor serves as a substrate for factor XIIIa and is cross-linked to laminin.

The propolypeptide of von Willebrand factor (pp-vWF) was found to serve as a substrate for Factor XIIIa (FXIIIa). FXIIIa catalyzed polymerization of pp-vWF as well as the incorporation of monodansylcadaverine and [14C]putrescine into pp-vWF in a Ca(2+)- and time-dependent manner. The amount of putrescine incorporated into pp-vWF was increased as a function of putrescine concentration and reached a maximum of 3 mol/mol of pp-vWF, indicating that there are at least 3 glutamine residues in the pp-vWF molecule responsible for the reaction. 125I-pp-vWF was incubated with FXIIIa in the presence of various adhesive glycoproteins. Laminin, among the proteins tested, specifically formed a large molecular weight complex with 125I-pp-vWF in a Ca(2+)- and time-dependent manner. It was also found that FXIIIa catalyzed polymerization of laminin as well as the incorporation of monodansylca-daverine into laminin. The high molecular weight complexes of 125I-pp-vWF (or 125I-laminin) formed in the presence of laminin (or pp-vWF) were immunoprecipitated with anti-laminin (or anti-pp-vWF) antibodies. Taking all the data together it can be concluded that both pp-vWF and laminin have glutamine and lysine residues responsive to FXIIIa and make copolymers by virtue of FXIIIa.

Animals↗