Autophagy inhibition in early but not in later stages prevents 3T3-L1 differentiation: Effect on mitochondrial remodeling

Autophagy is essential for successful white adipocyte differentiation but the data regarding the timing and relevance of autophagy action during different phases of adipogenesis are limited.     

Effect of239Pu on mouse hemopoietic stem cells in different types of bone marrow cavities

Summary Repopulative activity of hemopoietic stem cells of mice given i.v. 5 kBq²³⁹Pu/mouse (166.5 kBq/kg) was followed. The activity retained was measured in the whole mouse, the skeleton and the liver. Simultaneously average cumulative skeletal dose was calculated. Quantitative parameters of the stem cell compartment and the marrow cellularity were studied in variously arranged bones (femur, pelvis, lumbar vertebra) using the exocolonizing test and cytological techniques. The effects of radiation were most marked in lumbar vertebra, less serious changes were found in pelvis and only a moderate response was present in femur. The bone marrow hemopoiesis is damaged in various bone sites to different degrees and the percentage of cells at risk appears higher in trabecular than in cortical bone.     

RNAi in mouse oocytes and preimplantation embryos: effectiveness of hairpin dsRNA

Wolbachia are intracellular symbionts mainly found in arthropods, causing various sexual alterations on their hosts by unknown mechanisms. Here we report the results that strongly suggest that Wolbachia have virus-like particles of phage WO, which was previously identified as a prophage-like element in the Wolbachia genome. Wolbachia (strain wTai) infection in an insect was detected with the antibody against Wsp, an outer surface protein of Wolbachia, by fluorescence microscopy and immunoelectron-microscopy for the first time. Virus-like particles in Wolbachia were observed by electron-microscopy. The 0.22-microm filtrate of insect ovary contained DAPI-positive particles, and PCR analysis demonstrated that a phage WO DNA passed through the filter while Wolbachia DNA were eliminated, suggesting that the DAPI-positive particles were phage WO.     

Activity-dependent synaptogenesis in the adult Mammalian cortex

Recent electron microscopic studies provide evidence that the adult cortex generates new synapses in response to sensory activity and that these structural changes can occur rapidly, within 24 hr of sensory stimulation. Together with progress imaging synapses in vivo, the stage appears set for advances in understanding the dynamics and mechanisms of experience-dependent synaptogenesis.     

The Accumulation of Risk and Essential Elements in Edible Mushrooms Chlorophyllum rhacodes,Suillus grevillei,Imleria badia,and Xerocomellus chrysenteron Growing in the Czech Republic

Risk and essential elements were determined in fruiting bodies of wild growing edible mushrooms Chlorophyllum rhacodes, Suillus grevillei, Imleria badia, and Xerocomellus chrysenteron collected in an unpolluted site in South Bohemia, the Czech Republic. The elements were also determined in underlying soils and the bioconcentration factors were calculated. The analyses revealed that C. rhacodes accumulated Ag, Cu, Rb, Se, Zn, As, Cd, and Tl. On the other hand, S. grevillei accumulated Cd, Rb, Ag, Se, and Cs. I. badia and X. chrysenteron strongly accumulated Rb, Cs, and Ag; these species showed the ability to accumulate Cu and Zn as well. Contents of detrimental Cr^VI were in all cases below the quantification limit (0.003 mg kg^?1 dry matter). Studied mushroom species (mainly C. rhacodes) accumulated some toxic elements. However, no considerable effect on human health is expected since they are usually consumed as a delicacy and do not represent a major component of diet.     

Characterization of dicyclohexylcarbodiimide binding sites in beef-heart mitochondria.

Binding studies with [¹⁴C]-dicyclohexylcarbodiimide showed the presence of binding sites in the beef-heart mitochondrial membrane at a concentration of 1.8 nmol/mg protein (1.4 sites per cytochrome a+a₃). Saturation of these sites correlated with the inhibition of the ATPase activity. The maximum binding capacity could be related with the amount of F₁-ATPase in mitochondria from various tissues.     

Effect of Bordetella pertussis toxin on ADP-ribosylation of membrane proteins, adenylate cyclase activity and insulin release in rat pancreatic islets

Exposure of rat pancreatic islet membranes to [alpha-32P]-NAD+ in the presence of Bordetella Pertussis toxin (islet-activating protein) reveals the ADP-ribosylation of a peptide with a Mr close to 41 kDa, which corresponds to the alpha-subunit of the guanine nucleotide regulatory protein Ni. Islets removed from rats pretreated with the Bordetella Pertussis toxin display a specific increase in adenylate cyclase responsiveness to GTP and are characterized by a resistance to the inhibitory action of alpha2-adrenergic agonists upon either adenylate cyclase activity or glucose-induced insulin release.     

Suppression of Mexican bean beetle development by foliar applications of azasterols and reversal by cholesterol

Suppression of growth and molting occurred in Mexican bean beetle (Epilachna varivestis) larvae confined with bean plants treated with 25-azacholesterol (25-A) or 3-methoxy-25-azacholesterol (3-M-25-A). No larvae confined with bean plants dipped into 10 ppm of 3-M-25-A completed development and over half failed to complete the first ecdysis. Three larvae treated with 25-A developed everted wing pads on the last instar. Addition of an equal weight of cholesterol to the dipping solution completely nullified the inhibitive effects of a dose of 25-A 100 x greater than that required for inhibitive effects. Treatment of the bean plants with a combination of 25-A and a JH mimic resulted in additive and possibly synergistic pathological developmental effects in confined larvae.

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Résumé Des larves d'Epilachna varivestis élevées sur des plants de haricots trempés dans des solutions d'acétone et eau contenant soit du 25-azacholesterol (25-A) ou du 3-methoxy-25-azacholesterol (3-M-25A), cessent de muer et leur croissance est stoppée. Avec des plants de haricots trempés dans une solution à 10 p.p.m. de. 3 M-25A, aucune larve n'acheva son développement et plus de la moitié des 100 larves expérimentées ne purent achever leur lère mue. Les pupes obtenues ne pesaient fréquemment que la moitié du poids des témoins. Trois larves traitées avec 25-A développèrent des ébauches alaires inversées au cours du dernier stade.Il a été reconnu que ces azasterols inhibent les D²² et D^22,24 sterolreductases dans le tissu larvaire de l'intestin moyen des larves de Manduca sexta (Tobacco Hornworm). Ces enzymes interviennent dans la transformation des principaux phytostérols des feuilles de haricots (stigmasterol, -sitosterol et campesterol) en cholesterol. Comme il y a moins de 1% de cholesterol dans les feuilles de haricots, nous avons essayé de compenser l'effet inhibiteur du 25-A en ajoutant du cholesterol aux solutions du bain. L'addition de 1000 ppm de cholesterol annule complètement les effets inhibiteurs de 1000 ppm de 25-A (100 fois plus que la dose nécessaire pour provoquer des effets inhibiteurs).Des larves de Manduca sexta élevées sur plants de tabac à l'azasterol ne sont pas affectées, bien que leur développement soit grandement retardé par de petites quantités de ces azasterols ajoutés à un aliment synthétique dépourvu de cholestérol. Sans doute, la haute teneur en cholesterol des feuilles de tabac (9% des sterols totaux) est suffisante pour satisfaire les besoins en cholesterol des larves. Des traitements de plants de haricots avec une combinaison de 25-A plus la substance mimétique de l'hormone juvénile (E)-4-[(6,7-epoxy-3-ethyl-7-methyl-2-nonenyl) oxy]-1,2-(methylenedioxy) benzène, ont provoqué des effets pathologiques surjoutés et peut être synergiques sur le développement des larves d'Epilachna.

     

The molecular mechanism of induction of unfolded protein response by Chlamydia

The unfolded protein response (UPR) contributes to chlamydial pathogenesis, as a source of lipids and ATP during replication, and for establishing the initial anti-apoptotic state of host cell that ensures successful inclusion development. The molecular mechanism(s) of UPR induction by Chlamydia is unknown. Chlamydia use type III secretion system (T3SS) effector proteins (e.g, the Translocated Actin-Recruiting Phosphoprotein (Tarp) to stimulate host cell's cytoskeletal reorganization that facilitates invasion and inclusion development. We investigated the hypothesis that T3SS effector-mediated assembly of myosin-II complex produces activated non-muscle myosin heavy chain II (NMMHC-II), which then binds the UPR master regulator (BiP) and/or transducers to induce UPR. Our results revealed the interaction of the chlamydial effector proteins (CT228 and Tarp) with components of the myosin II complex and UPR regulator and transducer during infection. These interactions caused the activation and binding of NMMHC-II to BiP and IRE1α leading to UPR induction. In addition, specific inhibitors of myosin light chain kinase, Tarp oligomerization and myosin ATPase significantly reduced UPR activation and Chlamydia replication. Thus, Chlamydia induce UPR through T3SS effector-mediated activation of NMMHC-II components of the myosin complex to facilitate infectivity. The finding provides greater insights into chlamydial pathogenesis with the potential to identify therapeutic targets and formulations.     

Characterization and subcellular targeting of GCaMP-type genetically-encoded calcium indicators

Genetically-encoded calcium indicators (GECIs) hold the promise of monitoring [Ca(2+)] in selected populations of neurons and in specific cellular compartments. Relating GECI fluorescence to neuronal activity requires quantitative characterization. We have characterized a promising new genetically-encoded calcium indicator-GCaMP2-in mammalian pyramidal neurons. Fluorescence changes in response to single action potentials (17+/-10% DeltaF/F [mean+/-SD]) could be detected in some, but not all, neurons. Trains of high-frequency action potentials yielded robust responses (302+/-50% for trains of 40 action potentials at 83 Hz). Responses were similar in acute brain slices from in utero electroporated mice, indicating that long-term expression did not interfere with GCaMP2 function. Membrane-targeted versions of GCaMP2 did not yield larger signals than their non-targeted counterparts. We further targeted GCaMP2 to dendritic spines to monitor Ca(2+) accumulations evoked by activation of synaptic NMDA receptors. We observed robust DeltaF/F responses (range: 37%-264%) to single spine uncaging stimuli that were correlated with NMDA receptor currents measured through a somatic patch pipette. One major drawback of GCaMP2 was its low baseline fluorescence. Our results show that GCaMP2 is improved from the previous versions of GCaMP and may be suited to detect bursts of high-frequency action potentials and synaptic currents in vivo.