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Y Konno

Publications and source records attributed to Y Konno.

At least 37 records · Page 2Linked to original sources
PubMed1997

Electroresponsive pulsatile depot delivery of insulin from poly(dimethylaminopropylacrylamide) gel in rats.

We describe a model for pulsatile drug delivery with an electroresponsive polymer that is stimulated by an externally applied electrical field. Insulin loaded in an electroresponsive poly(dimethylaminopropylacrylamide) (PDMAPAA) gel was administered as a subcutaneous depot in rats. The gel induced a pulsatile plasma glucose decrease in correspondence to stimulation with a constant current of 1.0 mA (0.36 mA/cm2). The first drop occurred at 0.5 h after a 1-min application of current at 0 h and the second drop occurred at 3 h after a 10-min application of current at 2 h. Calculation of pharmacological bioavailability showed that the gel released 0.12% of the loaded insulin after these two stimuli. This in vivo study demonstrates the feasibility of this pulsatile delivery system. The mechanism of insulin release from the electroresponsive PDMAPAA gel is associated with electrokinetic flow of solvated insulin with water; that is, transportation process of counterions (electrophoresis) and water molecules (electroosmosis) in the crosslinked polyelectrolyte gel network.

Acrylic Resins↗
PubMed1997

Differential localization of s and e antigens in hepatitis B virus-associated glomerulonephritis.

We report here a case of membranous glomerulonephritis associated with chronic hepatitis B (HB) virus infection and describe differential localization of HB antigens in glomeruli. The patient showed mild proteinuria and was positive for hepatitis B surface (HBs) antigen, hepatitis B envelope (HBe) antigen, and antibody to hepatitis B core (HBc) antigen in the serum. The antibody against hepatitis C was negative. A renal biopsy revealed membranous glomerulonephritis with mesangial proliferation. The immunohistochemical studies using monoclonal antibodies localized the HBe antigen along the capillary wall and the HBs antigen in the mesangial area. The immunoelectron microscopic study confirmed the localization of HB antigens: HBe antigen was located in the subepithelial and intramembranous electron dense deposits and HBs antigen in the mesangial deposits. Our present results provide the first report of the differential localization of HB antigens in glomeruli at both the light and electron microscopic levels. The differential localization of HB antigens will provide insight into the pathogenesis of membranous glomerulonephritis.

Adult↗
PubMed1997

Closed-loop subcutaneous insulin infusion algorithm with a short-acting insulin analog for long-term clinical application of a wearable artificial endocrine pancreas.

Considering the management and safety of the insulin delivery route when a wearable artificial endocrine pancreas is applied to ambulatory diabetic patients on a long-term basis, we developed a s.c. insulin infusion algorithm by analyzing the dynamics of a s.c. injected short-acting insulin analog (Insulin Lispro) by a three-compartment model. Principally the insulin infusion algorithm was developed as a transfer function with the first-order delay in both proportional and derivative actions to blood glucose concentrations. The parameters for this algorithm were calculated to simulate a physiological plasma insulin profile as closely as possible. By applying this algorithm with regular insulin, diabetic patients showed a 2 h postprandial hyperglycemia and a delayed hyperinsulinemia, followed by hypoglycemic episodes 4-5 h after oral glucose load, just as observed in the computer simulation study. However, using Insulin Lispro, a near-physiological glycemic control (postprandial blood glucose of 153.1 +/- 8.3 mg/100 ml at 60 min and 90.3 +/- 7.1 mg/100 ml at 180 min, respectively) could be achieved without showing any delayed hyperinsulinemia or hypoglycemia. Daily glycemic excursions were also controlled near-physiologically and although the daily insulin requirement (731.7 +/- 160.5 mU/kg/day) was slightly higher, it was not significantly different from that with i.v. insulin infusion (622.3 +/- 142.6 mU/kg/day). These results indicate that the application of s.c. insulin infusion algorithm with Insulin Lispro is feasible for long-term glycemic control with a wearable artificial endocrine pancreas in ambulatory diabetic patients.

Adult↗
PubMed1996

cDNA cloning and genomic structure of human bone morphogenetic protein-3B (BMP-3b).

BMP-3b (also termed GDF-10) is a novel BMP-3 related protein recently discovered in rat femur tissue by molecular cloning. In this paper, we have isolated cDNA and the gene for human BMP-3b and determined their structure. Cloned human BMP-3b cDNA with a size of 2632 base pairs encodes a 478 amino acid precursor protein sharing 83% identity with rat BMP-3b. The human BMP-3b gene is composed of 3 exons and spans approximately 13 kilobases of DNA. The 5' flanking region carries no typical TATA box but G+C rich sequences. Southern blot analysis revealed that the BMP-3b gene is situated in a single locus of chromosome 10. By Northern analysis, human BMP-3b transcripts were detected primarily in femur, brain, lung, skeletal muscle, pancreas and testis.

Amino Acid Sequence↗
PubMed1996

Identification of rat bone morphogenetic protein-3b (BMP-3b), a new member of BMP-3.

Rat bone morphogenetic protein-3 (BMP-3) cDNA and BMP-3b cDNA were isolated from rat femur cDNA library by the RT-PCR method using degenerate oligonucleotide primers corresponding to human BMP-3. The deduced amino acid sequence of rat BMP-3 in the mature region reveals 2 amino acid changes compared to that of human BMP-3 (98% identity). The deduced BMP-3b amino acid sequence shows 81% similarity with the mature region of rat BMP-3 and only 37% similarity with the propeptide region. By Northern blot analysis, rat BMP-3b mRNA was detected in costa, costicartilage, femur, calvaria, trachea, aorta and brain. Among these tissues, BMP-3b transcripts were predominantly expressed in cerebellum. BMP-3 mRNA was found in femur, calvaria, trachea, lung and ovary. Although BMP-3b and BMP-3 are very closely related to each other, their transcripts are distributed in different tissues except that both are found in bone. The distribution pattern of BMP-3b mRNA suggests that BMP-3b plays an important role in the central nervous system as well as in bone formation and remodeling.

Aging↗
PubMed1996

A carboxy-terminal deletion mutant of protein kinase C beta II inhibits insulin-stimulated 2-deoxyglucose uptake in L6 rat skeletal muscle cells.

Alternative splicing of pre-mRNA encoding the carboxy-terminal (C-terminal) exons of protein kinase C beta (PKC beta) leads to the expression of two protein isoforms, PKC beta 1 and PKC beta II, with the potential for different functions. PKC beta II expression is regulated by insulin via alternative mRNA splicing. A physiological consequence of its activation was investigated in L6 rat skeletal muscle cells expressing GLUT4 transporters, a cell line in which PKC is involved in glucose transport. We examined the contribution of PKC beta II for insulin-stimulated [3H]2-deoxyglucose uptake by constructing three PKC beta II C-terminal deletion mutants designated M216, M217, and M218. When transiently expressed in COS1 cells, M217, with nine amino acids deleted, demonstrated autophosphorylation activity 10-fold less than full-length PKC beta II. The mutants M218, with 13 amino acids deleted, and M216, with 52 amino acids deleted, demonstrated no autophosphorylation activity and are kinase negative. When transiently expressed in L6 myotubes, M217 inhibited insulin-stimulated 2-deoxyglucose uptake by 45% (with a 45% transfection efficiency) whereas M216 and M218, kinase-negative mutants, had no effect compared with cells transfected with control plasmid. Cotransfection of full-length PKC beta II with M217 was able to rescue the inhibition of insulin-stimulated 2-deoxyglucose uptake as compared with cotransfection of M217 with the control plasmid, suggesting that M217 acts as a dominant-negative. In contrast, cotransfection of full-length PKC beta I, the other alternatively spliced form, did not rescue inhibition of insulin-stimulated 2-deoxyglucose uptake by M217. To further demonstrate the involvement of PKC, specifically PKC beta II, in insulin-stimulated 2-deoxyglucose uptake, we used two inhibitors, CG41251 (a specific PKC inhibitor) and CG53353 (a PKC beta II-specific inhibitor at 1 microM). Both inhibited insulin-stimulated 2-deoxyglucose uptake 50-60% in L6 myotubes. We conclude that M217 may act as a specific PKC beta II dominant-negative and that PKC beta II is more specific for insulin-stimulated 2-deoxyglucose uptake in these cells than PKC beta I.

Amino Acid Sequence↗
PubMed1995

Effects of early introduction of intensive insulin therapy on the clinical course in non-obese NIDDM patients.

In order to reconsider the extent of indication of insulin therapy in non-insulin dependent diabetes mellitus (NIDDM), we performed the following trial in a prospective fashion. At the beginning phase of treatment for diabetes, we introduced intensive insulin therapy in 22 non-obese (Body mass index approximately 24 kg/m2) NIDDM patients without proliferative retinopathy, who were selected in a standardized fashion, avoiding any arbitrary choice. None had received oral hypoglycemic agents (OHA) or insulin yet. By administering insulin 3 or 4 times a day, strict glycemic control was attained and maintained, and then the insulin dose was gradually lowered while keeping good glycemic control. In patients whose glycemic control was maintained at an excellent level for more than 7 days under an insulin dosage lower than 8 u/day, insulin therapy was discontinued. As a result, 15 patients (68%) attained good glycemic control both without insulin and OHA almost within a month and 6 patients (27%) shifted to OHA. It is recommended to introduce intensive insulin therapy in non-obese NIDDM patients without proliferative retinopathy and to aim at attaining good glycemic control both without insulin and OHA.

Blood Glucose↗
PubMed1994

Overproduction of a Ca(2+)-independent protein kinase C isozyme, nPKC epsilon, increases the secretion of prolactin from thyrotropin-releasing hormone-stimulated rat pituitary GH4C1 cells.

Rat pituitary GH4C1 cells express protein kinase C (PKC) transcripts for cPKC alpha, cPKC beta II, nPKC delta, nPKC epsilon, nPKC eta, and aPKC zeta, but not for cPKC gamma or nPKC theta. Of the transcripts produced, the nPKC epsilon isoform is the most abundant. Transfection of GH4C1 cells with an expression plasmid containing nPKC epsilon cDNA leads to the transient overexpression of cellular nPKC epsilon and confers enhanced phorbol ester binding activity. Transient expression of an inactive point mutant (nPKC epsilon K-->R) of nPKC epsilon, where Lys436 at the putative ATP-binding site is replaced with Arg, also confers elevated binding activity. However, only overproduction of the wild type in transfected cells increases the basal levels and stimulates the secretion of prolactin (PRL) by 12-O-tetradecanoylphorbol-13-acetate or thyrotropin-releasing hormone (TRH). In stable clones overexpressing nPKC epsilon, immunocytofluorescence and immunoblot experiments indicated that TRH causes the rapid translocation and down-regulation of an appreciable fraction of nPKC epsilon. Both the basal and TRH-stimulated levels of PRL secretion are clearly correlated with the expression level of nPKC epsilon but not with the TRH receptor densities in these clones. The dose dependence of TRH-stimulated secretion were similar in all cells overexpressing cPKC alpha, cPKC beta II, nPKC epsilon, and nPKC delta, but the enhancement of PRL secretion was specific for the overproduction of nPKC epsilon, no effect was found when other isozymes were overproduced. These findings clearly demonstrate that the expression level of nPKC epsilon in GH4C1 cells is rate-limiting for basal and TRH-stimulated PRL secretion, and they provide the first direct evidence that nPKC epsilon plays a key role in hormonal secretory processes.

Animals↗
PubMed1994

Artificial endocrine pancreas and optimal blood glucose regulation in diabetic patients--from bedside-type to wearable-type.

The artificial endocrine pancreas is a feedback control system regulating insulin delivery on a minute-by-minute basis according to the measured blood glucose levels. The bedside-type artificial endocrine pancreas has been proven to be useful not only as a therapeutic tool for diabetes mellitus but also as an elegant research tool for investigating the pathophysiology of the disease. With significant advances in the development of a subcutaneous tissue glucose monitoring system, the wearable-type artificial endocrine pancreas has been applied to diabetic patients. With this system, perfect glycemic control can be obtained for longer periods in ambulatory diabetic patients. The trend in the development of the artificial endocrine pancreas is now directed to implantable devices. Much efforts have been conducted to realize these devices.

Blood Glucose↗
PubMed1994

Antimetastatic effect of yeast mannan-bleomycin conjugate against mouse Lewis lung carcinoma.

A conjugate of bleomycin (BLM) and the mannan of bakers' yeast (Saccharomyces cerevisiae wild type strain) (WNM) was synthesized. The assay of its antimetastatic effect on Lewis lung carcinoma (3LL) implanted in C57BL/6 mice showed that this conjugate exhibited a higher antimetastatic effect and longer life-span elongation than those of free bleomycin and mannan with corresponding doses. This conjugate was also found to kill the 3LL cells in vitro. 14C-Labeled mannan-bleomycin conjugate was much more bound than 14C-labeled dextran-bleomycin conjugate to the 3LL. It was concluded that the anti-cancer mechanism of this conjugate, WNM-BLM possessed a specific binding effect to the tumor cells and exhibited a cytocidal effect on the 3LL target cells.

Animals↗
PubMed1993

Isolation and characterization of fruit vacuolar invertase genes from two tomato species and temporal differences in mRNA levels during fruit ripening.

To determine the relationship between invertase gene expression and glucose and fructose accumulation in ripening tomato fruit, fruit vacuolar invertase cDNA and genomic clones from the cultivated species, Lycopersicon esculentum cv. UC82B, and a wild species, Lycopersicon pimpinellifolium, were isolated and characterized. The coding sequences of all cDNA clones examined are identical. By comparison to the known amino acid sequence of mature L. esculentum fruit vacuolar invertase, a putative signal sequence and putative amino-terminal and carboxy-terminal propeptides were identified in the derived amino acid sequence. Of the residues 42% are identical with those of carrot cell wall invertase. A putative catalytic site and a five-residue motif found in carrot, yeast, and bacterial invertases are also present in the tomato sequence. Minor differences between the nucleotide sequences of the genomic clones from the two tomato species were found in one intron and in the putative regulatory region. The gene appears to be present in one copy per haploid genome. Northern analysis suggests a different temporal pattern of vacuolar invertase mRNA levels during fruit development in the two species, with the invertase mRNA appearing at an earlier stage of fruit development in the wild species. Nucleotide differences found in the putative regulatory regions may be involved in species differences in temporal regulation of this gene, which in turn may contribute to observed differences in hexose accumulation in ripening fruit.

Amino Acid Sequence↗
PubMed1993

Muscle relaxant action of dantrolene sodium in rats.

The relationship between the muscle relaxant action and intramuscular concentration of unchanged dantrolene sodium [1-[5-(p-natrophenyl) furfurylidene amino] hydantoin sodium hemiheptahydrate, dantrolene] by intravenous administration of dantrolene was examined in rats. The mechanism of the muscle relaxant action of dantrolene was also examined. Dantrolene, unlike tolperisone, a centrally acting muscle relaxant, dose-dependently inhibited twitch contraction evoked by electrical stimulation of both the muscle and the phrenic nerve in the rat isolated phrenic nerve-diaphragm preparation to a similar extent (IC50: 9.5 and 6.5 x 10(-6) M, respectively). Inhibition of both twitch contractions by dantrolene (50 microM) was antagonized by 4-aminopyridine (42.5 microM) and caffeine (4 mM). The twitch contraction of the gastrocnemius muscle elicited by sciatic nerve stimulation was inhibited by intravenous administration of dantrolene in a dose-dependent manner. The effective intramuscular concentration of dantrolene producing 50% depression (ID50) was 3.4 micrograms/g tissue, equal to about 1 x 10(-5) M. We thus found a close relationship between IC50 in vitro and ID50 in situ. Furthermore, this study raised the possibility that dantrolene produces its muscle relaxant action by affecting peripheral muscular Ca2+ mobilization.

4-Aminopyridine↗
PubMed1992

Purification and characterization of protein kinase C epsilon from rabbit brain.

Protein kinase C epsilon was chromatographically purified from rabbit brain to electrophoretic homogeneity. We identified the enzyme as the epsilon species of novel-type protein kinase C (nPKC epsilon), originally discovered and defined by cDNA cloning [Ohno, S., et al. (1988) Cell 53, 731-741], on the basis of the following observations: (i) the enzyme reacts specifically with an antipeptidic antiserum to nPKC epsilon but not with antisera to any of the other molecular species of PKC thus far known; (ii) it exhibits enzymatic behavior essentially identical to that of a recombinant nPKC epsilon purified from transfected COS cells [Konno, Y., et al. (1989) J. Biochem. 106, 673-678] and distinct from that of conventional PKC (alpha, beta I/II, and gamma) in its dependence on magnesium concentration and cofactors such as phospholipids, calcium, and phorbol ester; and (iii) it has an apparent molecular weight of 95.7K +/- 0.4K on SDS-PAGE, significantly greater than the other conventional and novel PKCs thus far identified. Notably, calcium exhibits a complex effect, both positive and negative, on the kinase activity of epsilon depending on the kind of substrate and the coexisting phospholipid, calling for a modification of the current notion that epsilon is a kinase unresponsive to calcium. The amount of epsilon species in the brain was estimated to be comparable to that of each conventional species, indicating that epsilon stands as one of the major PKC family members in brain. Furthermore, the enzyme shows a broader substrate spectrum than conventional PKC when examined with endogenous substrates, implying that it may cover a wider or different range of physiological functions.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1991

Selective up-regulation of type II inosine 5'-monophosphate dehydrogenase messenger RNA expression in human leukemias.

The discovery of isozymes (types I and II) of IMP dehydrogenase (IMPDH; EC 1.1.1.205), the rate-limiting enzyme of de novo GTP biosynthesis, has attracted attention as a possible novel approach to cancer diagnosis and selective tumor cell chemotherapy. To elucidate differences in expression and regulation of the two IMPDH isozymes, we examined the steady-state levels of these mRNAs in various types of leukemic cells from patients. Northern blot analysis revealed that type II IMPDH was more active transcriptionally (1.5- to 5.1-fold) in all the leukemic cells examined than in normal lymphocytes, whereas type I expression was similar. The increased expression of type II mRNA in leukemic cells was closely linked with the increase in total IMPDH activity (r = 0.92). When leukemic cells from a patient with chronic granulocytic leukemia in blast crisis were separated into blast-rich and mature leukocyte-rich fractions, the expression of type II mRNA correlated positively with the population of immature leukemic cells, whereas type I expression was unchanged. Treatment of leukemic blasts with 12-O-tetradecanoyl-phorbol-13-acetate for 5 days resulted in a 90% decrease in the expression of type II mRNA with macrophage-like differentiation, while the expression of type I mRNA was relatively stable. These observations suggest that expression of type II IMPDH is stringently linked with immature characteristics of leukemic cells; thus, it should be a selective target for antileukemic chemotherapy.

Cell Differentiation↗
PubMed1991

Expression of human IMP dehydrogenase types I and II in Escherichia coli and distribution in human normal lymphocytes and leukemic cell lines.

Two distinct cDNAs encoding proteins with 84% sequence identity have been isolated for human IMP dehydrogenase (EC 1.1.1.205) (Natsumeda, Y., Ohno, S., Kawasaki, H., Konno, Y., Weber, G., and Suzuki, K. (1990) J. Biol. Chem. 265, 5292-5295), an important target in antileukemic chemotherapy. We constructed expression plasmids containing these cDNAs in full length with pUC plasmids and produced lacZ'-IMP dehydrogenase fusion proteins in Escherichia coli. Both synthesized proteins exhibited IMP dehydrogenase activity and were partially separated from endogenous E. coli IMP dehydrogenase. By injecting the fusion proteins into mice we generated a polyclonal antibody specific to type I IMP dehydrogenase and an antibody which reacted with both types. Immunoblot analysis revealed that the total amounts of types I and II enzymes increased in human leukemic cell lines K562 and HL-60 in agreement with the increase in IMP dehydrogenase activity to 7.8- and 9.4-fold, respectively, above that of normal lymphocytes. The extent of expression of type I IMP dehydrogenase was similar in these cells, however, indicating that the increase in IMP dehydrogenase amount in leukemic cells was due to specific up-regulation of type II enzyme. Northern blot analysis also showed specific and predominant expression of type II in the leukemic cells. Thus, de novo GTP biosynthesis may be controlled differently in normal and neoplastic cells by different IMP dehydrogenase molecular species.

Cell Line↗
PubMed1991

Expression of cytochrome P-450lpr is developmentally regulated and limited to house fly.

Expression of house fly cytochrome P-450lpr was examined using immunoblotting in male and female adult LPR house flies, mixed sex adult house flies at 12 different ages, larvae, and pupae. P-450lpr was expressed in both male and female adult house flies. P-4501pr was clearly present in all adult stages examined, was barely detectable in pupae, and could not be detected in larvae. Thus, cytochrome P-450lpr is developmentally regulated and present in both sexes of house fly. Expression of cytochrome P-450, immunologically homologous to house fly cytochrome P-4501pr, was examined in other species using immunoblot analysis. Eleven animal species were tested in the orders Diptera, Hymenoptera, Lepidoptera, Orthoptera, Acari, and Rodentia, using microsomes in some species from both induced and noninduced animals or insecticide-resistant and susceptible strains. P-450lpr appears to be restricted to house flies, as none of these species contained cytochrome P-450 that reacted with antiserum to cytochrome P-450lpr.

Animals↗
PubMed1991

Structural and functional diversities of a family of signal transducing protein kinases, protein kinase C family; two distinct classes of PKC, conventional cPKC and novel nPKC.

Recent molecular cloning and biochemical experiments on the nature of protein kinase C (PKC) have revealed the existence of two distinct classes of phorbol ester (and diacylglycerol) receptor/protein kinase, conventional PKC (cPKC) and novel PKC (nPKC). Each of these classes contains multiple related molecules expressed in tissues and cells in a type-specific manner. Although nPKC does not show the typical PKC activity ascribable to conventional PKCs and thus was neglected in earlier studies, several lines of evidence suggest that nPKCs are involved in a variety of cell responses to physiological stimuli and phorbol esters. It is possible that in some cases nPKC is the major mediator of the so-called PKC-activators, such as phorbol esters, mezerein, and bryostatins. In addition to the clear difference between cPKC and nPKC, functional diversity among conventional PKCs has also been demonstrated; PKC gamma differs in its competence to mediate the signal toward transcriptional activation through TPA-responsive cis-acting elements from cPKC alpha and nPKC epsilon. The differences between cPKC and nPKC and among the individual members of each of these two classes, and their specific pattern of distribution in tissues and cells, provide a rationale by which to explain the specificity and diversity of cellular responses to external stimuli generating DAG and to phorbol esters. The results presented here also provide a means to dissect the complex signaling pathway in cells and to analyze the molecular basis underlying the signal transduction processes mediated by this family of protein kinases.

Animals↗