Presence of residual beta cells and co-existing islet autoimmunity in the NOD mouse during longstanding diabetes: a combined histochemical and immunohistochemical study

During type 1 diabetes, most beta cells die by immune processes. However, the precise fate and characteristics of beta cells and islet autoimmunity after onset are unclear. Here, the extent of beta cell survival was determined in the non-obese diabetic (NOD) mouse during increasing duration of disease and correlated with insulitis. Pancreata from female NOD mice at diagnosis and at 1, 2, 3 and 4 weeks thereafter were analysed immunohistochemically for insulin, glucagon and somatostatin cells and glucose transporter-2 (glut2) and correlated with the degree of insulitis and islet immune cell phenotypes. Insulitis, although variable, persisted after diabetes and declined with increasing duration of disease. During this period, beta cells also declined sharply whereas glucagon and somatostatin cells increased, with occasional islet cells co-expressing insulin and glucagon. Glut2 was absent in insulin-containing cells from 1 week onwards. CD4 and CD8 T cells and macrophages persisted until 4 weeks, in islets with residual beta cells or extensive insulitis. We conclude that after diabetes onset, some beta cells survive for extended periods, with continuing autoimmunity and expansion of glucagon and somatostatin cells. The absence of glut2 in several insulin-positive cells suggests that some beta cells may be unresponsive to glucose.     

Étude des principaux facteurs influençant l‘émission d'attractifs sexuels synthétiques à partir de diffuseurs en caoutchouc et en plastique

Deux types de diffuseurs en caoutchouc (Michelin) et en plastique polystratifié (Hercon^®) sont testés en laboratoire pour étudier quelques aspects des phénomènes de diffusion. Trois composantes phéromonales, le Z9-12: Ac, le 12: Ac et le Z9-14: Ac imprégnées dans ces différents supports servent à détermineer le rôle joué par la longueur de la chaîne de la molécule et la présence de doubles liaisons sur la vitesse de diffusion. L'influence de la température, de la vitesse du vent et de l'humidité est déterminée en cellules climatisées.Parmi les facteurs relatifs à la structure chimique, l'influence de la longueur de la chaîne est prédominante par rapport à la présence d'une double liaison. Parmi les facteurs climatiques, le vent agit plus que la température sur la diffusion, alors que l'humidité paraît secondaire.     

New insights into the role of connexins in pancreatic islet function and diabetes

Multi-cellular systems require complex signaling mechanisms for proper tissue function, to mediate signaling between cells in close proximity and at distances. This holds true for the islets of Langerhans, which are multicellular micro-organs located in the pancreas responsible for glycemic control, through secretion of insulin and other hormones. Coupling of electrical and metabolic signaling between islet β-cells is required for proper insulin secretion and effective glycemic control. β-cell specific coupling is established through gap junctions composed of connexin36, which results in coordinated insulin release across the islet. Islet connexins have been implicated in both Type-1 and Type-2 diabetes; however a clear link remains to be determined. The goal of this review is to discuss recent discoveries regarding the role of connexins in regulating insulin secretion, the regulation of connexins within the islet, and recent studies which support a role for connexins in diabetes. Further studies which investigate the regulation of connexins in the islet and their role in diabetes may lead to novel diabetes therapies which regulate islet function and β-cell survival through modulation of gap junction coupling.     

氢火焰气相色谱仪内标法测定反式脂肪酸的条件的建立及验证

目的:研究氢火焰气相色谱仪(GC112A)内标法测定反式脂肪酸的实验室最佳条件。方法:采用气相色谱内标-标准曲线法,通过测定反式脂肪酸标准品工作液的标准曲线、重现性和分离度来确定和验证GC112A的最佳色谱条件。结果:实验室最佳色谱条件:毛细管柱,PC-88(100 m×0.25 mm×0.2μm);进样口温度:260℃;载气:高纯N2,压力220 k Pa,流速41.0 m L/min;检测器(FID)温度:290℃,氢气压力100 k Pa,流速21.0 m L/min;空气压力160 k Pa,流速215.0 m L/min;检测器(FID)灵敏度:1010;手动进样,进样量:1.0μL。结论:该方法避免了外标法(国标方法)的国产仪器局限性和操作者人工进样的不确定性,具有受仪器参数变化的影响较小的特点,适合在基层食品检验机构推广。     

Improving islet transplantation by gene delivery of hepatocyte growth factor (HGF) and its downstream target, protein kinase B (PKB)/Akt

Clinical studies have demonstrated that islet transplantation may be a useful procedure to replace beta cell function in patients with Type 1 diabetes. Islet transplantation faces many challenges, including complications associated with the procedure itself, the toxicity of immunosuppression regimens, and to the loss of islet function and insulin-independence with time. Despite the current successes, and residual challenges, these studies have pointed out an enormous scarcity of islet tissue that precludes the use of islet transplantation in a clinical setting on a wider scale. To address this problem, many research groups are trying to identify different islet growth factors and intracellular molecules capable of improving islet graft survival and function, therefore reducing the number of islets needed for successful transplantation. Among these growth factors, hepatocyte growth factor (HGF), a factor known to improve transplantation of a variety of organs/cells, has shown promising results in increasing islet graft survival and reducing the number of islets needed for successful transplantation in four different rodent models of islet transplantation. Protein kinase B (PKB)/Akt, a pro-survival intracellular signaling molecule is known to be activated in the beta cell by several different growth factors, including HGF. PKB/Akt has also shown promising results for improving human islet graft survival and function in a minimal islet mass model of islet transplantation in diabetic SCID mice. Increasing our knowledge on how HGF, PKB/Akt and other emerging molecules work for improving islet transplantation may provide substrate for future therapeutic approaches aimed at increasing the number of patients in which beta cell function can be successfully replaced.     

Communication disruption of maleAgrotis segetum moths with one or several sex pheromone components

Attraction of maleAgrotis segetum Dennis & Schiffermüller (Lepidoptera: Noctuidae) to sex pheromone traps in fields, which were treated with one or three pheromone components was investigated. Small plots of 1/4ha size were treated with synthetic pheromone, released by 25 evenly dispersed latex rubber tube dispensers. The dispensers were loaded with either 500 μg Z5-10:OAc (50 mg/ha), or 1000 μg Z7-12:OAc (100 mg/ha), or a 3-component mixture consisting of 500 μg Z5-10:OAc+1000 μg Z7-12:OAc+1000 μg Z9-14:OAc. Pheromone traps were placed both within and outside of the treated area in a cross design, with an intertrap spacing of 15 m. Release rates from disruption dispensers were measured in the laboratory after being exposed in the field. The release rates of the components were estimated to be 0.44, 0.11, and 0.06 μg/h/dispenser for Z5-10:OAc, Z7-12:OAc and Z9-14:OAc, respectively. The highest effect of disruption was achieved by the three-component blend, resulting in a significant suppression of trap catches extending 5 m outside of the treated area. The Z5-10:OAc treatment resulted in reduced trap catches inside the treated area, but the effect did not extend outside. Z7-12:OAc alone did not result in any significant reduction in trap catch. The results indicate that different mechanisms may explain the disruptive effect of the treatments and that the single pheromone components are not as effective as the three-component blend.     

Depletion of CUL4B in macrophages ameliorates diabetic kidney disease via miR-194-5p/ITGA9 axis

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Differential response of lymphocytes to free and sepharose-linked anti-immunoglobulin during different phases of the cell cycle

During proliferation induced by anti-immunoglobulins, B lymphocytes undergo cell volume increases prior to onset of DNA synthesis. Although both Sepharose-linked and free anti-immunoglobulin evoked essentially identical increases in cell volume, the Sepharose-linked antibody induced a significantly greater DNA synthesis than free antibody as judged from [3H]thymidine incorporation studies using mass cell culture technique, as well as by using autoradiographic analysis of individual cells. These findings are considered in terms of possible differences in triggering cell volume increases and DNA synthesis by free and linked anti-immunoglobulin and/or the possible existence of B cell subpopulations responding differentially to free and to linked antibody.     

Remodeling of phosphatidylglycerol in Synechocystis PCC6803

The phosphatidylglycerol deficient ΔpgsA mutant of Synechocystis PCC6803 provided a unique experimental system for investigating in vivo retailoring of exogenously added dioleoylphosphatidylglycerol in phosphatidylglycerol-depleted cells. Gas chromatographic analysis of fatty acid composition suggested that diacyl-phosphatidylglycerols were synthesized from the artificial synthetic precursor. The formation of new, retailored lipid species was confirmed by negative-ion electrospray ionization–Fourier-transform ion cyclotron resonance and ion trap tandem mass spectrometry. Various isomeric diacyl-phosphatidylglycerols were identified indicating transesterification of the exogenously added dioleoylphosphatidyl-glycerol at the sn-1 or sn-2 positions. Polyunsaturated fatty acids were incorporated selectively into the sn-1 position. Our experiments with Synechocystis PCC6803/ΔpgsA mutant cells demonstrated lipid remodeling in a prokaryotic photosynthetic bacterium. Our data suggest that the remodeling of diacylphosphatidylglycerol likely involves reactions catalyzed by phospholipase A₁ and A₂ or acyl-hydrolase, lysophosphatidylglycerol acyltransferase and acyl-lipid desaturases.     

An assay for angiotensin-converting enzyme using capillary zone electrophoresis

A sensitive and rapid method was developed for angiotensin-converting enzyme (ACE) activity determination by capillary zone electrophoresis. Hippuryl-l-histidyl-l-leucine, a synthetic tripeptide, was used as the ACE-specific substrate. Capillary zone electrophoresis was employed to separate the products of the enzymatic reaction and the ACE activity was determined by quantification of hippuric acid, a result of the enzymatic reaction on the tripeptide. The capillary electrophoresis was performed in a 27 cm x 75 micrometer i.d. fused-silica capillary using 200 mM boric acid-borate buffer (pH 9.0) as a run buffer with an applied voltage of 8.1 kV at a capillary temperature of 23 degrees C. The electrophoresis was monitored at 228 nm. Each electrophoretic run requires only a nanoliter of the enzymatic reactant solution, at only 6 min, rendering a powerful tool for the ACE assay.     

Tetranortriterpenoid derivatives from Turraea parvifolia (Meliaceae)

The methanol extract of the seeds of Turraea parvifolia has yielded seven novel triterpenoid derivatives: 12alpha-acetoxyazadironolide, turraparvin A (12alpha-acetoxy-7alpha,23-dihydroxy-24,25,26,27-tetranor-3-oxoapotirucalla-1,14,20(22)-trien-21,23-olide), turraparvin B (12alpha-acetoxy-7alpha,21-dihydroxy-24,25,26,27-tetranor-3-oxoapotirucalla-1,14,20(22)-trien-23,21-olide), turraparvin C (7alpha,12alpha-diacetoxy-21-hydroxy-24,25,26,27-tetranor-3-oxoapotirucalla-1,14,20(22)-trien-23,21-olide), 11-epi-21-hydroxytoonacilide, 11-epi-23-hydroxytoonacilide and turraparvin D (12alpha-acetoxy-7alpha-hydroxy-24,25,26,27-tetranor-3-oxoapotirucalla-1,14,20(22)-trien-21,23-lactam).     

Effects of the New Arginase Inhibitor N-Hydroxy-nor- -Arginine on NO Synthase Activity in Murine Macrophages

In stimulated murine macrophage, arginase and nitric oxide synthase (NOS) compete for their common substrate, -arginine. The objectives of this study were (i) to test the new α-amino acid N^ω-hydroxy-nor- -arginine (nor-NOHA) as a new selective arginase inhibitor and (ii) to elucidate the effects of arginase inhibition on -arginine utilization by an inducible NOS. Nor-NOHA is about 40-fold more potent than N^ω-hydroxy- -arginine (NOHA), an intermediate in the -arginine/NO pathway, to inhibit the hydrolysis of -arginine to -ornithine catalyzed by unstimulated murine macrophages (IC₅₀ values 12 ± 5 and 400 ± 50 μM, respectively). Stimulation of murine macrophages with interferon-γ and lipopolysaccharide (IFN-γ + LPS) results in clear expression of an inducible NOS (iNOS) and to an increase in arginase activity. Nor-NOHA is also a potent inhibitor of arginase in IFN-γ + LPS-stimulated macrophage (IC₅₀ value 10 ± 3 μM). In contrast to NOHA, nor-NOHA is neither a substrate nor an inhibitor for iNOS and it appears as a useful tool to study the interplays between arginase and NOS. Inhibition of arginase by nor-NOHA increases nitrite and -citrulline accumulation for incubation times higher than 12 h, under our conditions. Our results allow the determination of the kinetic parameters of the two competitive pathways and the proposal of a simple model which readily explains the differences observed between experiments. This model readily accounts for the observed effects and should be useful to predict the consequences of arginase inhibition in the presence of an active NOS on -arginine availability.     

Metabolic Engineering for Microbial Production of Aromatic Amino Acids and Derived Compounds

Metabolic engineering to design and construct microorganisms suitable for the production of aromatic amino acids and derivatives thereof requires control of a complicated network of metabolic reactions that partly act in parallel and frequently are in rapid equilibrium. Engineering the regulatory circuits, the uptake of carbon, the glycolytic pathway, the pentose phosphate pathway, and the common aromatic amino acid pathway as well as amino acid importers and exporters that have all been targeted to effect higher productivities of these compounds are discussed.     

Enantioselective synthesis and biological evaluation of 5-o-carboranyl pyrimidine nucleosides

Base-modified carborane-containing nucleosides such as 5-o-carboranyl-2'-deoxyuridine (CDU) when combined with neutrons have potential for the treatment of certain malignancies. Lack of toxicity in various cells, high accumulation in cancer cells and intracellular phosphorylation are desirable characteristics for modified nucleosides used in boron neutron capture therapy (BNCT) for brain tumors and other malignancies. The aim of this work was to synthesize the two beta-enantiomers of several 5-o-carboranyl-containing nucleosides. These derivatives may possess favorable properties such as high lipophilicity, high transportability, the ability to be phosphorylated, and resistance to catabolism. Beta-isomers of 2',3'-dihydroxynucleosides and analogues containing a heteroatom in the sugar moiety were also synthesized. Carboranyl pyrimidine nucleosides were prepared either from the parent beta-D-nucleoside, beta-L-nucleoside, or by a coupling reaction. The dioxolane derivative 7 was prepared by a coupling reaction between protected 5-o-carboranyluracil (8, CU) and the corresponding protected heterocycle. Specific catalysts were used during the N-glycosylation process to favor the formation of the beta-isomer. Biological evaluation of these new chiral 5-o-carboranyl pyrimidine derivatives indicated that most of these compounds have low toxicity in a variety of normal and malignant cells and achieved high cellular levels in a lymphoblastoid cell line. Increasing the number of hydroxyl groups on the sugar moiety decreased the cellular accumulation and serum binding to different extents. Five compounds were identified for further biological evaluation as potential agents for BNCT.     

Effect of 2-deoxy- -glucose on acetylcholine and histamine levels in gastric juice of pylorus-ligated rats anesthetized with urethane

Acetylcholine (ACh) in gastric juice was detected and measured by pretreatment of acetylcholinesterase inhibitor, 1 mM eserine (1 ml/rat, p.o.), in pylorus-ligated rats, by liquid chromatography with electrochemical detection. In order to elucidate whether or not the ACh level in gastric juice reflects the activity of cholinergic neurons, the effect of 2-deoxy- -glucose (2-DG), a vagus stimulant, on the levels of ACh, histamine and gastric acid in gastric juice was investigated in pylorus-ligated rats anesthetized with urethane (1.25 g/kg, i.p.). Under the non-anesthetic condition, ACh, histamine and gastric acid levels were 100±25 pmol/h, 120±10 ng/h, and 240±32 μequiv./h, respectively. These levels were completely inhibited by urethane anesthesia. Under the anesthetized condition, 2-DG (50–200 mg/kg, i.v.) significantly increased ACh and histamine levels in gastric juice, as well as acid secretion. The 2-DG (200 mg/kg, i.v.)-induced increases in these levels were completely inhibited by vagotomy. These results suggest that ACh level measured in gastric juice reflects the activity of cholinergic transmission. Furthermore, these results also support the conclusion that vagus stimulation facilitates not only cholinergic transmission but also histaminergic transmission related to gastric acid secretion.     

Evidence for the reactivity of fatty aldehydes released from oxidized plasmalogens with phosphatidylethanolamine to form Schiff base adducts in rat brain homogenates

The vinyl ether bond of plasmalogens could be among the first target of free radicals attack. Consequently, because of their location in the membranes of cells, plasmalogens represent a first shield against oxidative damages by protecting other macromolecules and are often considered as antioxidant molecules. However, under oxidative conditions their disruption leads to the release of fatty aldehydes. In this paper, we showed using gas chromatography-mass spectrometry (GC-MS) analyses that fatty aldehydes released from plasmalogens after oxidation (UV irradiation and Fe2+/ascorbate) of cerebral cortex homogenates can generate covalent modifications of endogenous macromolecules such as phosphatidylethanolamine (PE), like the very reactive and toxic malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE). These newly formed Schiff base adducts could be responsible for deleterious effects on cells thus making the protective role of plasmalogens potentially questionable.     

N-acetyl- -arginine ethyl ester synthesis catalysed by bovine trypsin in organic media

The bovine trypsin-catalysed synthesis of N-acetyl- -arginine ethyl ester from N-acetyl- -arginine and ethanol was studied in various organic solvents (dimethyl sulfoxide, dioxane, dimethylformamide, acetonitrile, acetone, tetrahydrofuran, chloroform, toluene, carbon tetrachloride, cyclohexane and n-hexane). The highest yield was achieved in acetonitrile after incubation for 6 or 24 h. The optimal conditions for ester synthesis in acetonitrile for 6 h were as follows: 5.0 mM N-acetyl- -arginine, 10.0 M ethanol, 7.2 mg trypsin, 2.87% water, total volume 10.3 ml, pH 7.0 and 30°C. The hydrolytic activity of trypsin was determined after incubation for 6 days, when 87.7% of the original activity remained, suggesting that acetonitrile caused little inactivation of the enzyme. The synthetic reaction resulted in a maximal 79.3% conversion under optimized conditions after incubation for 48 h.     

Biosynthetic intermediates and stereochemical aspects of aldehyde biosynthesis in the marine diatom Thalassiosira rotula

Intermediates of the aldehyde biosynthesis in Thalassiosira rotula are investigated. Use of labeled precursors and cell preparations proves production of 2E,4Z-octadienal from 6Z,9Z,12Z-hexadecatrienoic acid (C16:3 omega-4) through the lipoxygenase-dependent intermediate (9S)-9-hydroperoxyhexadeca-6,10,12-trienoic acid. On the contrary, synthesis of 2E,4Z,7Z-decatrienal involves mainly EPA (C20:5 omega-3) by a 11R-lipoxygenase, as suggested by identification of chiral 11R-HEPE (12% e.e.) in the diatom extracts. Consistently with the necessity to have a rapid transport and metabolization of the intermediate hydroperoxides, we show that lipoxygenase and lyase activities are both found in the same subcellular fraction of the microalga.