Biomedical subjects

K Takata

Publications and source records attributed to K Takata.

At least 19 recordsLinked to original sources

PubMed1996

Modulatory role of drebrin on the cytoskeleton within dendritic spines in the rat cerebral cortex.

Morphological changes in the dendritic spines have been postulated to participate in the expression of synaptic plasticity. The cytoskeleton is likely to play a key role in regulating spine structure. Here we examine the molecular mechanisms responsible for the changes in spine morphology, focusing on drebrin, an actin-binding protein that is known to change the properties of actin filaments. We found that adult-type drebrin is localized in the dendritic spines of rat forebrain neurons, where it binds to the cytoskeleton. To identify the cytoskeletal proteins that associated with drebrin, we isolated drebrin-containing cytoskeletons using immunoprecipitation with a drebrin antibody. Drebrin, actin, myosin, and gelsolin were co-precipitated. We next examined the effect of drebrin on actomyosin interaction. In vitro, drebrin reduced the sliding velocity of actin filaments on immobilized myosin and inhibited the actin-activated ATPase activity of myosin. These results suggest that drebrin may modulate the actomyosin interaction within spines and may play a role in the structure-based plasticity of synapses.

Actins↗

PubMed1996

Vitamin D3 elicits calcium response and activates blood monocyte-derived macrophages from patients with vitamin D dependent rickets type II.

We studied the effects of vitamin D3 metabolites on intracellular free Ca2+ concentration ([Ca2+]i) and the respiratory burst of monocyte-derived macrophages (MDM) from patients with vitamin D dependent rickets type II. Treatment of MDM from the patients and healthy donors with 1 nM 1,25(OH)2D3 produced a rapid elevation of [Ca2+]i and similarly primed both types of cells for enhanced capacity for O2- release with phorbol diester. These results suggest that macrophages may have distinct non-genomic pathways of vitamin D3, which partly explain the absence of immunodeficiency and the disappearance of rickets after treatment with vitamin D3 in the patients.

24,25-Dihydroxyvitamin D 3↗

PubMed1996

Immuno-localization of H+/peptide cotransporter in rat digestive tract.

In the mammalian digestive tract, small peptides are absorbed by a H+-coupled peptide transport system. Using an antibody against the rat H+/peptide cotransporter (PepT1), we examined the localization of PepT1 immunohistochemically along the rat digestive tract. PepT1 was detected in the small intestine (duodenum, jejunum, and ileum), but not in the esophagus, stomach, colon, or rectum. PepT1 was especially enriched in the villi, where it was localized in the brush border of the absorptive epithelial cells. PepT1 was not detected in the mucus-secreting goblet cells or less-differentiated epithelial cells in the crypts. These observations show that PepT1 is specific to the brush border of the differentiated absorptive epithelial cells and suggest that H+-coupled uptake of small peptides and peptide-like drugs occurs at the apical membrane of these cells in the small intestine.

Animals↗

PubMed1996

Na(+)-dependent glucose transporter SGLT1 is localized in the apical plasma membrane upon completion of tight junction formation in MDCK cells.

SGLT1, an isoform of Na(+)-dependent glucose transporters, is localized at the apical plasma membrane in the epithelial cells of the small intestine and the kidney. In the present study we examined its location in SGLT1 cDNA-transfected MDCK cells, which form an epithelial sheet connected by tight junctions in culture. Formation of tight junctions was monitored by staining for occludin, an integral tight junction protein. In the cells demarcated by an uninterrupted occludin meshwork, SGLT1 was specifically localized at the apical plasma membrane, showing that SGLT1 has a signal to accomplish this restricted localization. In the cells with little or no occludin accumulation in the tight junction, however, SGLT1 was present along the entire aspect of the plasma membrane. Similar distribution of SGLT1 was observed in the cells as long as the occludin meshwork remained incomplete. These observations suggest that apical localization of SGLT1 occurs upon the completion of the uninterrupted meshwork of tight junctions.

Actins↗

PubMed1996

Connexin 43 and the glucose transporter, GLUT1, in the ciliary body of the rat.

To investigate the relationship between the gap junction protein connexin 43 and the glucose transporter GLUT1, their localization was visualized by double-immunofluorescence microscopy using frozen sections as well as immunogold staining of ultrathin frozen sections. In pigmented epithelial cells, most of the GLUT1 was localized along the plasma membrane facing the blood vessels, whereas in non-pigmented epithelial cells, it was present along the plasma membrane facing the aqueous humor. Connexin 43 was abundant in the ciliary body and localized mainly in the gap junctions connecting the pigmented and non-pigmented epithelial cells. Localization of GLUT1 and connexin 43 in the blood-aqueous barrier suggests that GLUT1, connexin 43, and GLUT1 disposed in this order could be a machinery responsible for the transport of glucose across the blood-aqueous barrier.

Animals↗

PubMed1996

Immunolocalization of GLUT1 and connexin 26 in the rat placenta.

Interhemal membrane in the rat placenta is composed of three trophoblastic layers and endothelial cells. GLUT1, an isoform of the facilitated-diffusion glucose transporter, is abundant in the cells of the placental barrier, i.e., syncytiotrophoblastic layers I and II. GLUT1 is localized at the plasma membranes of the maternal-blood side of syncytiotrophoblastic layer I, and of the fetal-blood side of syncytiotrophoblastic layer II. Double-immunofluorescence microscopy has shown that connexin 26 is present between these GLUT1-positive sites, i.e., between syncytiotrophoblastic layers I and II. Immunogold electron microscopy has revealed that connexin 26 is localized in the gap junctions connecting the two layers. Connexin 26 in these layers therefore makes them functionally a single syncytial layer for the transfer of small molecules such as glucose in the rat placental barrier. These results suggest that glucose transfer in the rat placental barrier is carried out as follows: GLUT1 is used for the entry of glucose into the cytoplasm of syncytiotrophoblastic layer I, connexin 26 for the transfer of glucose from syncytiotrophoblastic layer I to syncytiotrophoblastic layer II, and GLUT1 for the exit of glucose to the fetal circulation.

Animals↗

PubMed1996

Glucose transporters in the transepithelial transport of glucose.

Glucose transporters are integral membrane proteins that mediate the transport of glucose and structurally-related substances across the cellular membranes. Two families of glucose transporter have been identified: the facilitated-diffusion glucose transporter family (GLUT family), and the NA(+)-dependent glucose transporter one (SGLT family). These transporters play a pivotal role in the transfer of glucose across the epithelial cell layers that separate distinct compartments in the mammalian body. In the small intestine, a Na(+)-dependent glucose transporter, SGLT1, is localized at the apical plasma membrane of the absorptive epithelial cells, whereas a facilitated-diffusion glucose transporter, GLUT2, is at the basolateral membrane of the cells. Similar localization is seen in the kidney proximal tubules in the reabsorption of glucose. For the absorption of fructose in the small intestine, fructose transporter GLUT5 is localized at the apical membrane. The expressed GLUT5 in polarized cultured cells is targeted to the apical membrane, showing that the GLUT5 molecule itself has sufficient information to determine its cellular localization. In the blood-tissue barriers, such as the blood-brain barrier, blood-ocular barrier, and placental barrier, either endothelial or epithelial cell layers constitute the barrier. GLUT1 is abundant at the plasma membrane of these barrier cells, and plays a crucial role in the specific transfer of glucose across the barrier. When the barrier is composed of a two-cell layer, gap junctions connecting them could serve as intercellular channels for glucose transfer in addition to GLUT1. Proper localization of glucose transporters and gap junctions is a prerequisite for the successful transepithelial transport of sugars.

Animals↗

PubMed1996

Developmental expression of proprotein-processing endoprotease furin in rat pancreatic islets.

The expression of furin in pancreatic beta-cells induces faster cell growth and a decrease in differentiated beta-cell-specific characteristics. During the development of rat pancreatic islets, the prohormone convertases, PC2 and PC3, appear during late gestation and are expressed long after birth. We investigated the developmental change in another yeast Kex2 family endoprotease, furin, in rat islets in relation to islet cell growth. Furin had appeared by gestational day 18 and was distributed in islets as well as exocrine tissues. The expression of furin in islets increased toward the neonatal stage. Bromodeoxyuridine (BrdU) incorporation was also elevated in the perinatal period. On postnatal days 10 and 20, staining characteristics were attenuated. On day 25, immediately after weaning, furin staining began to localize in the beta-cell region, and staining in the alpha-cell region became fainter. On day 270, the staining in the alpha-cell region disappeared, and staining in the beta-cell region remained positive, but was weaker. We conclude that furin expression was greatest during the perinatal period, when BrdU incorporation into islets was maximal. Furin expression continued, however, even after the BrdU incorporation decreased. Thus, furin appears to control the proliferation as well as differentiation of islet cells.

Aging↗

PubMed1996

[A course of pirarubicin vesical irrigation at thirty mg effectively prevents post-TUR recurrence of superficial bladder cancer].

In order to study its safety and anti-cancer dose-effectiveness in the prevention of cancer recurrence, two groups of patients having superficial bladder cancer (TUR-Bt) had their bladder intravesically irrigated post-surgically with 20 or 30 mg aliquots of (2''R)-4'-0-tetrahydropyranyl-adriamycin (THP). A total of 74 patients with bladder cancer were entered in the study. Four cases proved ineligible, and 30 could not complete the planned treatment. THP vesical irrigation of 14-17 sessions was regarded as a condition for eligibility: Clinical evaluation was feasible with 19 patients of 20 mg and 21 of 30 mg THP. Non-recurrence rates at 1, 2, 3 and 4 years were 89.5, 53.3, and 26.6%, respectively, for the 20 mg group, and 95.2, 84.3, 75.9 and 75.9%, respectively, for the 30 mg THP group. The incidence of side effects appeared slightly greater in the 30 mg THP group than in the 20 mg group, and there were no systemic adverse reactions. Topical reaction of cystitis was noted to be as low as 12.5%. Thus, THP vesical irrigations at 30 mg were found tolerable and effective for the prevention of local recurrence following TUR-Bt.

Administration, Intravesical↗

PubMed1995

Structural and functional analysis of the canine histamine H2 receptor by site-directed mutagenesis: N-glycosylation is not vital for its action.

G-protein-coupled receptors generally share a similar structure containing seven membrane-spanning domains and extracellular site(s) for N-glycosylation. The histamine H2 receptor is a member of the family of G-protein-coupled receptors, and has three extracellular potential sites for N-glycosylation (Asn-4, Asn-162 and Asn-168). To date, however, no information has been presented regarding N-glycosylation of the H2 receptor. To investigate the presence, location and functional roles of N-glycosylation of the H2 receptor, site-directed mutagenesis was performed to eliminate the potential site(s) for N-glycosylation singly and collectively. The wild-type and mutated H2 receptors were expressed stably in Chinese hamster ovary (CHO) cells or transiently in COS7 cells. Immunoblotting of the wild-type and mutated H2 receptors with an antiserum directed against the C-terminus of the H2 receptor showed that mutation at Asn-162, but not at Asn-168, resulted in a substantial decrease in the molecular mass. A mutation at Asn-4 led to a further decrease in the molecular mass. Tunicamycin treatment of the transfected cells yielded a sharp band with a molecular mass identical to that of the mutant devoid of all three potential sites for N-glycosylation. These findings indicate that the H2 receptor is N-glycosylated, and that N-glycosylation takes place mainly at two sites, Asn-4 and Asn-162. Neither the affinity for tiotidine nor that for histamine was affected by the mutagenesis. Immunocytochemistry and tiotidine binding showed that the mutated receptors were exclusively distributed on the cell surface in a fashion similar to that of the wild-type. In addition, the glycosylation-defective receptor was capable of activating adenylate cyclase and elevating the intracellular Ca2+ concentration in response to histamine in stable CHO cell lines. Thus N-glycosylation of the H2 receptor is not required for cell surface localization, ligand binding or functional coupling to G-protein(s).

1-Methyl-3-isobutylxanthine↗

PubMed1995

Effects of fluvastatin on human biliary lipids.

The 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors have rapidly become widespread in the treatment of hypercholesterolemia and are known to be variable in efficacy. To investigate the effect on biliary lipids, a 3-month study using fluvastatin was devised. A total of 19 patients were enrolled in this study: all had hypercholesterolemia (7 men, 12 women; 13 with type IIa, 6 with type IIb). After an observation period of 4-6 weeks with placebo, fluvastatin at a daily dose of 30 mg was administered for 3 months. Fasting blood samples were taken early in the morning, before, and once a month during 3 months of fluvastatin treatment, for measurement of serum lipids. Cerulein-stimulated bile in the gallbladder was sampled using a duodenal tube, and the changes in biliary lipids were assessed. There was a marked decrease in serum total cholesterol after 12 weeks of treatment (21%; p < 0.001). However, there was no significant difference in the bile cholesterol saturation index (CSI): values before and after 3 months of drug administration were 0.93 and 0.99, respectively (Admirand-Small method). There were no significant changes in either the fatty acid composition of biliary lecithin or in the bile acid composition of bile. In conclusion, on the basis of these results, short-term (3 months) administration of fluvastatin does not appear to affect CSI.

Adult↗

PubMed1995

Pain drawing in the evaluation of low back pain.

Pain drawings were obtained from three groups of patients: 51 with lumbar disc herniation; 55 with lumbar stenosis, and 42 with benign low back pain. On grid assessment, patients with disc herniation and stenosis who complained of radiating pain or intermittent claudication had a significantly larger number of grids for the extremities than those with benign low back pain. In studies of the relation between the characteristics of the pain drawing and the outcome of treatment, most patients with 19 grids or less, or a score of 3 points or less, had a satisfactory outcome. Those with more grids or a higher score tended to be unsatisfactory. Pain drawing permits differentiation of the three patterns of pain and is useful for predicting the outcome of treatment.

Activities of Daily Living↗

PubMed1995

Peanut agglutinin binding to human prolactin-producing pituitary adenomas.

Peanut agglutinin (PNA)-binding sites in human prolactin (PRL)-producing pituitary adenomas were examined by light and electron microscopy together with immunoblot analysis. At the light microscopic level, the majority of the PRL-producing adenoma cells stained positively for PNA in 15 of 20 cases. PNA binding observed in the cytoplasm had a granular appearance. PRL-producing cells adjacent to the adenoma tissue showed negative PNA staining. In normal pituitary glands, the PRL-positive glandular cells were negative for PNA staining. By electron microscopy, reaction products showing PNA-binding sites were detected in some of the secretory granules. Immunoblotting analysis revealed that the PRL bands corresponded to PNA-stained ones with the exception of the main 23-kDa band. PNA-binding sites have some relation to the secretory granules containing glycosylated forms of PRL. These observations suggest that PNA staining can be used as a valuable method to analyze human PRL-producing pituitary adenomas.

Adenoma↗

PubMed1995

Origin of posterior pituitary high intensity on T1-weighted magnetic resonance imaging. Immunohistochemical, electron microscopic, and magnetic resonance studies of posterior pituitary lobe of dehydrated rabbits.

RATIONALE AND OBJECTIVES: To investigate the origin of posterior pituitary high intensity (PPHI) seen on T1-weighted magnetic resonance (MR) images. METHODS: Six rabbits, including four rabbits deprived of drinking water for 4 days and two control rabbits, were examined by MR imaging. Plasma vasopressin levels were sequentially measured by radioimmunoassay. Pituitary glands were immunostained with guinea pig anti-rabbit vasopressin antibody, and ultrathin sections of Epon/Araldite-embedded specimens were observed with a transmission electron microscope. RESULTS: In control rabbits, PPHI was noted on T1-weighted MR images, and the posterior pituitary lobe was positively immunostained with anti-vasopressin antibody. At the ultrastructural level, nerve terminals contained numerous neurosecretory granules bearing vasopressin. Conversely, plasma vasopressin levels gradually increased and PPHI was absent in 4-day dehydrated rabbits. The posterior lobe was scarcely stained with anti-vasopressin antibody, and neurosecretory granules were rarely observed. However, a number of small dispersed vesicles, possibly derived from the fragmentation of neurosecretory granule envelopes, were seen in the nerve terminal. CONCLUSIONS: Posterior pituitary high signal seen on T1-weighted MR images is attributed to neurosecretory granules bearing vasopressin.

Animals↗

PubMed1995

A new case of apoA-I deficiency showing codon 8 nonsense mutation of the apoA-I gene without evidence of coronary heart disease.

We report a 39-year-old Japanese man with HDL and apoA-I deficiency as well as data from members of his family. Corneal opacity and a stomatocyte were found but not tonsillar hypertrophy, xanthomas, or splenomegaly. His serum HDL cholesterol, apoA-I, apoA-II, and LDL cholesterol levels were t mg/dL, < 3 mg/dL, 6 mg/dL, and 175 mg/dL, respectively. Plasma triglyceride, phospholipid, apoB, apoC-III, and apoE levels were all within normal limits. Lecithin:cholesterol acyltransferase activity was half of normal, while lipoprotein lipase and hepatic triglyceride lipase activities were within normal limits. ApoA-I deficiency was confirmed by combined isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by an immunoblotting method. We surveyed the apoA-I gene of the patient and five of his family members by direct sequencing after amplification by polymerase chain reaction and found a codon 8 nonsense mutation (TGG --> TAG, Trp --> stop) in exon 3 of the apoA-I gene. The results of a pedigree analysis by DNA sequencing and restricted fragment length polymorphism (Sty I) were consistent with an autosomal codominant trait. Coronary angiography was performed to evaluate coronary atherosclerosis, but no significant luminal narrowing was detected. An intracoronary ultrasound study showed mild intimal hyperplasia in segment 6. In summary, this is a case of apoA-I deficiency without evidence of coronary heart disease.

Adolescent↗

PubMed1995

Different effects of subclasses of HDL containing apoA-I but not apoA-II (LpA-I) on cholesterol esterification in plasma and net cholesterol efflux from foam cells.

We investigated the effects of subclasses of plasma LpA-I (HDL containing apoA-I but not apoA-II) on cholesterol esterification in plasma and net cholesterol efflux from foam cells. LpA-I was composed of particles of three diameters: large (11.1 nm; Lg-LpA-I), medium (8.8 nm; Md-LpA-I), and small (7.7 nm; Sm-LpA-I). Plasma concentrations of LpA-I were positively correlated only with the level of Lg-LpA-I. Plasma concentrations of Lg-LpA-I were inversely correlated with the rate of cholesterol esterification in plasma and VLDL- and LDL-depleted plasma. Plasma concentrations of Md-LpA-I and Sm-LpA-I did not correlate with the rate of cholesterol esterification in plasma or VLDL- and LDL-depleted plasma. When macrophage foam cells were incubated with Md- and Sm-LpA-I, cellular cholesterol mass was reduced by approximately 70%. In contrast, the cellular cholesterol-reducing capacity of Lg-LpA-I was negligible. Lg-LpA-I inhibited net cholesterol removal from foam cells that was mediated by Md- and Sm-LpA-I and cholesteryl ester production with these particles. These results suggest that Md- and Sm-LpA-I may actively participate in cellular cholesterol removal and cholesterol esterification in plasma and HDL, while Lg-LpA-I may regulate these functions of Md- and Sm-LpA-I.

Adolescent↗

PubMed1995

Characterization of rat GLUT5 and functional analysis of chimeric proteins of GLUT1 glucose transporter and GLUT5 fructose transporter.

To investigate the biological and biochemical properties of GLUT5, rat GLUT5 complementary DNA was transfected into Chinese hamster ovary cells. Rat GLUT5 was exclusively targeted to the plasma membrane and exhibited a transport activity, not for glucose, but for fructose. The affinity for fructose (Km = 11.6mM) was much higher than that of GLUT2, the other glucose transporter with fructose transport activity. Interestingly, rat GLUT5 was not photolabeled with 0.5 microM cytochalasin B, whereas a similar amount of GLUT1 was adequately photolabeled under the same experimental conditions. Next, to investigate the domains required for transport of glucose/fructose in GLUT1 and/or GLUT5, several chimeric GLUT1/GLUT5 proteins were expressed, and their glucose and/or fructose transport activities were studied. The intracellular middle loop and the region encompassing the membrane spanning domains 7-12 were observed to have crucial roles in GLUT1 glucose transport, whereas replacement of the N-terminal half or the intracellular C-terminal region with the corresponding region of GLUT5 produced no marked effects on glucose transport activity. In contrast, both the N-terminal half encompassing the region from the N-terminus through the 6th membrane spanning domain and the intracellular C-terminal region were mandatory for GLUT5 fructose transport. In conclusion, GLUT5 is a transporter exclusively for fructose and the structural requirements for fructose transport are more stringent than those for glucose transport among hexose transporter proteins.

Affinity Labels↗

PubMed1995

The granin family--its role in sorting and secretory granule formation.

Two types of secretory pathways are present in mammalian cells: constitutive secretion and regulated secretion (1). In the constitutive secretory pathway, which is found in all types of cells, secretory products are packed in small vesicles. Most of the proteoglycans and glycoproteins of the extracellular matrix are secreted in this way. The regulated secretory pathway, found in the more differentiated secretory cells, is mediated by specialized secretory granules. Hormones and neuropeptides are secreted in this way. Both the constitutive and regulated secretory pathways emanate from the trans-Golgi network (TGN). Recent studies have indicated that the granin family (secretogranins/chromogranins) plays an important role in the sorting and aggregation of secretory products in the trans-Golgi network (TGN), and in the subsequent formation of secretory granules. The granin family is thought to be one of the mediators of the regulated secretory pathway (2-6). This family is widely expressed in endocrine (7-9) and neuronal cells (10-12), where they are stored in secretory granules together with various peptide hormones and neuropeptides. Three acidic sulfated proteins, chromogranin A (Cg A), secretogranin I (Sg I; also called chromogranin B; Cg B), and secretogranin II (Sg II) are well known as the major proteins in the family. Recently, three acidic secretory proteins, 1B1075 gene product (13), HISL-19 antigen (14) and 7B2 (15), have been thought to be members of the granin family, accordingly termed as Sg III, Sg IV and Sg V, respectively. However, it still remains open to discussion whether 1B1075 gene product, the HISL-19 antigen and 7B2 are true members of the granin family. In this review, the possible roles of granin family, especially Cg A, Sg I (Cg B) and Sg II in the regulated secretory pathway are discussed, focusing on the sorting, aggregation of proteins in the TGN, and subsequent secretory granule formation.

Animals↗