Mouse ES cells maintained in different pluripotency-promoting conditions differ in their neural differentiation propensity

Prior to differentiation, embryonic stem (ES) cells in culture are maintained in a so-called “undifferentiated” state, allowing derivation of multiple downstream cell lineages when induced in a directed manner, which in turn grants these cells their “pluripotent” state. The current work is based on a simple observation that the initial culture condition for maintaining mouse ES cells in an “undifferentiated” state does impact on the differentiation propensity of these cells, in this case to a neuronal fate. We point out the importance in judging the “pluripotency” of a given stem cell culture, as this clearly demonstrated that the “undifferentiated” state of these cells is not necessarily a “pluripotent” state, even for a widely used mouse ES cell line. We partly attribute this difference in the initial value of ES cells to the naïve-to-primed status of pluripotency, which in turn may affect early events of the differentiation in vitro.     

Recombinant human thioredoxin suppresses lipopolysaccharide-induced bronchoalveolar neutrophil infiltration in rat

Human thioredoxin (TRX) is a multifunctional redox-active protein. We previously reported that the intraperitoneal administration of recombinant human thioredoxin (rhTRX) attenuates inflammatory cytokine- or bleomycin-induced lung injury in mice. In this study, the effect of rhTRX injected intravenously after lipopolysaccharide (LPS) injection was analyzed in rats. Rats were injected with LPS followed by treatment with rhTRX. Although the bolus injection exerted no protective effect, continuous intravenous administration of rhTRX significantly suppressed percentage number of neutrophils in bronchoalveolar lavage fluid. Histological examination also showed that rhTRX decreased neutrophil infiltration in the lung tissues. Administered rhTRX was mainly excreted into the urine and the tissue accumulation of rhTRX in the lung was marginal. LPS-induced oxidative stress in the lung was slight in this model. These results demonstrated that continuous intravenous administration of rhTRX suppresses LPS-induced bronchoalveolar neutrophil infiltration by an anti-chemotactic effect. Administration of rhTRX did not promote the tumor growth nor affect chemosensitivity in the xenotransplantation model, suggesting the safety of rhTRX therapy for cancer patients.     

Transmural and apicobasal gradients in repolarization contribute to T-wave genesis in human surface ECG

The cellular basis of the T-wave morphology of surface ECG remains controversial in clinical cardiology. We examined the effect of action potential duration (APD) distribution on T-wave morphology using a realistic model of the human ventricle and torso. We developed a finite-element model of the ventricle consisting of ～26 million elements, including the conduction system, each implemented with the ion current model of cardiomyocytes. This model was embedded in a torso model with distinct organ structures to obtain the standard ECG leads. The APD distribution was changed in the transmural direction by locating the M cells in either the endocardial or epicardial region. We also introduced apicobasal gradients by modifying the ion channel parameters. Both the transmural gradient (with M cells on the endocardial side) and the apicobasal gradient produced positive T waves, although a very large gradient was required for the apicobasal gradient. By contrast, T waves obtained with the transmural gradient were highly symmetric and, therefore, did not represent the true physiological state. Only combination of the transmural and the moderate apicobasal gradients produced physiological T waves in surface ECG. Positive T waves in surface ECG mainly originated from the transmural distribution of APD with M cells on the endocardial side, although the apicobasal gradient was also required to attain the physiological waveform.     

Two purified oligosaccharide elicitors,N-acetylchitohepatose and tetraglucosyl glucitol, derived fromMagnaporthe grisea cell walls, synergistically activate biosynthesis of phytoalexin in suspension-cultured rice cells

Two purified oligosaccharide elicitors generatable from fungal cell walls, N-acetylchitoheptaose and a tetraglucosyl glucitol from rice blast fungus (Magnaporthe grisea), synergistically activated phytoalexin biosynthesis in cultured rice cells. Inhibition experiments for the binding of radiolabeled N-acetylchitooligosaccharide elicitor to the plasma membrane from rice cells indicate that the two elicitors are recognized by different receptors. These results also indicate the presence of a positive interaction between the signal transduction cascade downstream of each elicitor/receptor, which enhances resistance against pathogens.     

Ethanol production from high cellulose concentration by the basidiomycete fungus Flammulina velutipes

Ethanol production by Flammulina velutipes from high substrate concentrations was evaluated. F. velutipes produces approximately 40–60 g l⁻¹ ethanol from 15 % (w/v) d-glucose, d-fructose, d-mannose, sucrose, maltose, and cellobiose, with the highest conversion rate of 83 % observed using cellobiose as a carbon source. We also attempted to assess direct ethanol fermentation from sugarcane bagasse cellulose (SCBC) by F. velutipes. The hydrolysis rate of 15 % (w/v) SCBC with commercial cellulase was approximately 20 %. In contrast, F. velutipes was able to produce a significant amount of ethanol from 15 % SCBC with the production of β-glucosidase, cellobohydrolase, and cellulase, although the addition of a small amount of commercial cellulase to the culture was required for the conversion. When 9 mg g⁻¹ biomass of commercial cellulase was added to cultures, 0.36 g of ethanol was produced from 1 g of cellulose, corresponding to an ethanol conversion rate of 69.6 %. These results indicate that F. velutipes would be useful for consolidated bioprocessing of lignocellulosic biomass to bioethanol.     

Intracellular delivery of serum-derived hepatitis C virus

A robust and reliable cell culture system for serum-derived HCV (HCVser) has not been established yet because of the presence of neutralizing antibody and tropism for infection. To overcome this obstacle, we employed a lipid-mediated protein intracellular delivery reagent (PIDR) that permits internalization of proteins into cells. Although entry of HCVcc was not enhanced by the treatment with PIDR, entry of HCVser into hepatoma cell lines (Huh7 and HepG2) and immortalized primary hepatocytes (Hc and HuS/E2) was significantly enhanced by the PIDR treatment. The entry of HCVser into Huh7 cells in the presence of PIDR was resistant to the neutralization by an anti-hCD81 antibody, suggesting that PIDR is capable of internalizing HCVser in a receptor-independent manner. Interestingly, the PIDR-mediated entry of HCVser and HCVcc was enhanced by the addition of sera from chronic hepatitis C patients but not from healthy donors. In addition, neutralization of HCVcc infection by anti-E2 antibody was canceled by the treatment with PIDR. In conclusion, the PIDR is a valuable tool to get over the obstacle of neutralizing antibodies to internalize HCV into cells and might be useful for the establishment of in vitro propagation HCVser.     

Identification of major histocompatibility complex genes in the guppy,Poecilia reticulata

The guppy, Poecilia reticulata, a teleostean fish of the order Cyprinodontiformes, has been used extensively in studies of host-parasite interactions, courtship behavior, and mating preference, as well as in ecological and evolutionary genetics. A related species was among the first poikilotherm vertebrates to be used in the study of histocompatibility genes. All these studies could benefit from the identification and characterization of the guppy major histocompatibility complex (Mhc) genes. Here, both class I and class II genes of the guppy are described. The number of expressed loci, as determined by representation of clones in a cDNA library, sequencing, and Southern blot analysis, may be low in both Mhc classes: combined evidence suggests that there may be one expressed class II locus only and one or two expressed class I loci. The variability of aquaristic guppy stocks is very low: only three and two genes have been detected at the class I and class II loci, respectively, in the stocks examined. This genetic paucity is most likely the consequence of breeding practices employed by aquarists and commercial establishments. Limited sampling of wild guppy populations revealed extensive Mhc polymorphism at loci of both classes in nature. Comparison of guppy Mhc sequences with those of other vertebrates has revealed the existence of a set of insertions/deletions which can be used as characters in cladistic analysis to infer phylogenetic relationships among vertebrate taxa and the Mhc genes themselves. These indels are particularly frequent in the regions coding for the loops of 1 and 2 domains of class I proteins.The nucleotide sequence data reported in this publication have been submitted to the EMBL nucleotide sequence database and have been assigned the accession numbers Z54076-Z54095     

Effects of hypoxic hypoxia on O2 uptake and heart rate kinetics during heavy exercise

Engelen, Marielle, Janos Porszasz, Marshall Riley, KarlmanWasserman, Kazuhira Maehara, and Thomas J. Barstow. Effects ofhypoxic hypoxia on O₂ uptake andheart rate kinetics during heavy exercise. J. Appl.Physiol. 81(6): 2500-2508, 1996.It is unclearwhether hypoxia alters the kinetics ofO₂ uptake(O₂) during heavy exercise[above the lactic acidosis threshold (LAT)] and how thesealterations might be linked to the rise in blood lactate. Eight healthyvolunteers performed transitions from unloaded cycling to the sameabsolute heavy work rate for 8 min while breathing one of threeinspired O₂ concentrations: 21%(room air), 15% (mild hypoxia), and 12% (moderate hypoxia). Breathing12% O₂ slowed the time constantbut did not affect the amplitude of the primary rise inO₂ (period of first2-3 min of exercise) and had no significant effect on either thetime constant or the amplitude of the slowO₂ component (beginning2-3 min into exercise). Baseline heart rate was elevated inproportion to the severity of the hypoxia, but the amplitude andkinetics of increase during exercise and in recovery were unaffected bylevel of inspired O₂.We conclude that the predominant effect of hypoxia during heavyexercise is on the early energetics as a slowed time constant forO₂ and an additionalanaerobic contribution. However, the sum total of the processesrepresenting the slow component of O₂ is unaffected.