Insulin inhibits glucose production by a direct effect in diabetic depancreatized dogs during euglycemia

In our previous studies in nondiabetic dogs and humans, insulin suppressed glucose production (GP) by both an indirect extrahepatic and a direct hepatic effect. However, insulin had no direct effect on GP in diabetic depancreatized dogs under conditions of moderate hyperglycemia. The present study was designed to investigate whether insulin can inhibit GP by a direct effect in this model under conditions of euglycemia. Depancreatized dogs were made euglycemic (approximately 6 mmol/l), rather than moderately hyperglycemic (approximately 10 mmol/l) as in our previous studies, by basal portal insulin infusion. After approximately 100 min of euglycemia, a hyperinsulinemic euglycemic clamp was performed by giving an additional infusion of insulin either portally (POR) or peripherally at about one-half the rate (1/2 PER) to match the peripheral venous insulin concentrations. The greater hepatic insulin load in POR resulted in greater suppression of GP (from 16.5 +/- 1.8 to 12.2 +/- 1.6 micromol. kg(-1). min(-1)) than 1/2 PER (from 17.8 +/- 1.9 to 15.6 +/- 2.0 micromol. kg(-1). min(-1), P < 0.001 vs. POR), consistent with insulin having a direct hepatic effect in suppressing GP. We conclude that the direct effect of insulin to inhibit GP is present in diabetic depancreatized dogs under conditions of acutely induced euglycemia. These results suggest that, in diabetes, the prevailing glycemic level is a determinant of the balance between insulin's direct and indirect effects on GP.     

Wound healing ability of Xenopus laevis embryos. I. Rapid wound closure achieved by bisectional half embryos

We examined wound closure in 'half embryos' produced by the transverse bisection of Xenopus laevis embryos at the primary eye vesicle stage. Both the anterior- and posterior-half embryos survived for more than 6 days, and grew into 'half tadpoles'. Histology and videomicroscopy revealed that the open wound in the half embryo was rapidly closed by an epithelial sheet movement in the wound marginal zone. The time-course of wound closure showed a downward convex curve: the wound area decreased to one-fifth of the original area within 30 min, and the wound continued to contract slowly thereafter. The rapidity of closure of the epidermis as well as the absence of inflammatory cells are typical features of an embryonic type of wound healing. There was a dorso-ventral polarity in the motility of the epidermis: the wound was predominantly closed by the ventral and lateral epidermis. The change in the contour of the wound edge with time suggested a complex mechanism involved in the wound closure that could not be explained only by the purse-string theory. The present experimental system would be a unique and useful model for analyses of cellular movements in the embryonic epithelia.     

Crystal transformation from anhydrous alpha-maltose to hydrous beta-maltose and from anhydrous trehalose to hydrous trehalose

Anhydrous sugars such as maltose and trehalose are useful for making dry powder of foods and liquids. The crystal-transformation rate of maltose and trehalose were investigated under humid conditions and by kneading. The enthalpy for solubilization was 7.0 kJ/mol for the anhydrous maltose. The crystal-transformation rate of anhydrous alpha-maltose to hydrous beta-maltose depended on the temperature at 75% humidity. However, that of anhydrous trehalose did not depend on the temperature, and transformation was very rapid. An anomeric change to maltose and no such change to trehalose might have caused this. The activation energy of crystal transformation was 79 kJ/mol for maltose and zero for trehalose. The rate of crystal transformation of anhydrous maltose while kneading depended on the purity of the anhydrous alpha-maltose and the amount of water present. This crystal transformation rate fitted the Avrami equation.     

Refolding of denatured/reduced lysozyme at high concentration with diafiltration

Refolding of reduced and denatured protein in vitro has been an important issue for both basic research and applied biotechnology. Refolding at low protein concentration requires large volumes of refolding buffer. Among various refolding methods, diafiltration is very useful to control the denaturant and red/ox reagents in a refolding solution. We constructed a refolding procedure of high lysozyme concentration (0.5-10 mg/ml) based on the linear reduction of the urea concentration during diafiltration under oxygen pressure. When the urea concentration in the refolding vessel was decreased from 4 M with a rate of 0.167 M/h, the refolding yields were 85% and 63% at protein concentrations, 5 mg/ml and 10 mg/ml, respectively, after 11 h. This method gave a high productivity of 40.1,microM/h of the refolding lysozyme. The change in refolding yields during the diafiltration could be simulated using the model of Hevehan and Clark.     

Bile acid sulfonate and 7-alkylated bile acid analogs: effect on intestinal absorption of taurocholate and cholesterol 7alpha-hydroxylase activity in cultured rat hepatocytes

The effects of sulfonate analogs of cholic (C), chenodeoxycholic (CDC), and ursodeoxycholic acid (UDC) and three 7-alkylated CDCs--7-methyl-, 7-ethyl-, and 7-propyl-CDCs--on taurocholate absorption from rat terminal ileum in situ and on cholesterol 7alpha-hydroxylase activity in primary culture of the rat liver were investigated. The sulfonate analogs of two dihydroxy bile acids CDC and UDC, but not C, significantly decreased the absorption of taurocholate. Taurine conjugates of 7-alkylated CDC slightly decreased the taurocholate absorption, and tauro-7-propyl-CDC significantly suppressed the absorption. Although the sulfonate analogs of C and CDC reduced cholesterol 7alpha-hydroxylase activity by 40% and 60% compared to control, UDC-sulfonate analog did not affect enzymatic activity. These results were consistent with those of the lead compounds, C, CDC, and UDC. The introduction of methyl group at C-7 position of CDC attenuated the reduction in cholesterol 7alpha-hydroxylase activity by CDC. However, elongation of the alkyl group resulted in an inhibitory effect. The present study revealed the following: 1) bile acid sulfonates act on cholesterol and bile acid metabolism in a similar manner as taurine conjugated bile acids; and 2) the biologic properties of CDC could be altered by the introduction of alkyl group at C-7 position.     

The creation of a database of odorous compounds focused on molecular rigidity and analysis of the molecular features of the compounds in the database

It is important to select odorous molecules for experiments on olfaction and for the development of an electronic nose. Odorous molecules having a small number of conformers, namely structurally rigid molecules, are assumed to interact with a small number of types of olfactory receptor proteins or to interact with the proteins in a simpler manner than that of fairly flexible molecules. Focusing on the rigidity of molecular structures, we collected 287 odorous molecules from data sources, which included 1205 chemicals in total and a database of the 287 odorous molecules (DB_odMOL) was created using CS ChemFinder Pro (version 5.0). The logarithmic value of the octanol/water partition coefficient (log P) and melting point, boiling point and vapour pressure of the molecules were estimated using CS Chem3D Pro. The database DB_odMOL accumulates these estimated data in addition to literature values for odour quality, odour detection thresholds and the safety of molecules. The rigidity of the 287 molecules was further analysed by conformational analysis performed by molecular mechanics using Conformer in CS Chem3D Pro (version 5) and 72 rigid odorous molecules were selected. The 287 molecules were also analysed based on atomic composition, substructure and molecular size. Sixty-two odorous molecules among the 72 rigid odorous molecules were further selected based on their atomic composition. The 62 rigid molecules with simple atomic composition that were finally selected should be useful for researchers in choosing odorous molecules for the study of olfaction, including the fields of molecular biology, physiology, structure-odour relationships and other fields of the study concerning odour.     

Multiple Administrations of 64Cu-ATSM as a Novel Therapeutic Option for Glioblastoma: a Translational Study Using Mice with Xenografts

Glioblastoma is the most aggressive malignant brain tumor in humans and is difficult to cure using current treatment options. Hypoxic regions are frequently found in glioblastoma, and increased levels of hypoxia are associated with poor clinical outcomes of glioblastoma patients. Hypoxia plays important roles in the progression and recurrence of glioblastoma because of drug delivery deficiencies and induction of hypoxia-inducible factor-1α in tumor cells, which lead to poor prognosis. We focused on a promising hypoxia-targeted internal radiotherapy agent, ⁶⁴Cu-diacetyl-bis (N⁴-methylthiosemicarbazone) (⁶⁴Cu-ATSM), to address the need for additional treatment for glioblastoma. This compound can target the overreduced state under hypoxic conditions within tumors. Clinical positron emission tomography studies using radiolabeled Cu-ATSM have shown that Cu-ATSM accumulates in glioblastoma and its uptake is associated with high hypoxia-inducible factor-1α expression. To evaluate the therapeutic potential of this agent for glioblastoma, we examined the efficacy of ⁶⁴Cu-ATSM in mice bearing U87MG glioblastoma tumors. Administration of single dosage (18.5, 37, 74, 111, and 148 MBq) and multiple dosages (37 MBq × 4) of ⁶⁴Cu-ATSM was investigated. Single administration of ⁶⁴Cu-ATSM in high-dose groups dose-dependently inhibited tumor growth and prolonged survival, with slight and reverse signs of adverse events. Multiple dosages of ⁶⁴Cu-ATSM remarkably inhibited tumor growth and prolonged survival. By splitting the dose of ⁶⁴Cu-ATSM, no adverse effects were observed. Our findings indicate that multiple administrations of ⁶⁴Cu-ATSM have effective antitumor effects in glioblastoma without side effects, indicating its potential for treating this fatal disease.     

Biosynthesis of auxin-induced ethylene in mung bean hypocotyls

The pathway of ethylene biosynthesis in auxin-treated mung beanhypocotyls was investigated by comparing the specific radioactivitiesof ethylene produced and S-adenosylmethionine (SAM) in the tissuefollowing the administration of 3,4-¹⁴C-methionine, and by analyzingthe methionine metabolites. When the rate of auxin-induced ethyleneproduction was low due to a low concentration of auxin, thespecific radioactivity of ethylene released was always higherthan that of SAM in the tissue. When the tissue was treatedwith auxin, the tissue produced and accumulated a methioninemetabolite which was converted into ethylene more efficientlythan methionine. The metabolite was identified as 1-aminocyclopropane-l-carboxylicacid (ACC) by means of paper and thin-layer chromatography,high voltage paper electrophoresis and co-crystallization. ACCformation was neither inhibited by low oxygen nor by the inhibitoryprotein of ethylene synthesis, but inhibited by aminoethoxyvinylglycine(AVG). ACC application to the tissue greatly reduced incorporationof 3,4-¹⁴C-methionine into ethylene. The control tissue thatwas not treated with auxin also converted ACC into ethyleneindicating that the enzyme which converts ACC into ethyleneis already present in the tissue and that auxin induced productionof the enzymatic system responsible for the conversion of methionineinto ACC. Ethylene synthesis from ACC was not inhibited by AVG,abscisic acid, cycloheximide or actinomycin D, but inhibitedby low oxygen and the inhibitory protein. (Received November 21, 1979; )     

Stabilization of Photosystem II (O(2) Evolution) of Spinach Chloroplasts by Radiation-induced Immobilization

Spinach chloroplasts were immobilized with vinyl monomers by radiation-induced polymerization at low temperature and stored in buffer containing bovine serum albumin. The lifetime of O₂ evolution activity in photosystem II was prolonged remarkably in immobilized chloroplasts. Thermostability of immobilized chloroplasts stored in buffer containing bovine serum albumin was far better than that of immobilized chloroplasts in pure buffer and that of intact chloroplasts. When immobilized chloroplasts were stored in buffer including polyethylene glycol, the lifetime of O₂ evolution activity was longer than for those stored in buffer containing bovine serum albumin.     

Cattle cDNA clone encoding a new allele of the MHC class II DQA1 gene

The nucleotide sequence data reported in this paper have been submitted to the DDBJ, EMBL, and GenBank nucleotide sequence databases and have been assigned the accession number D50454