Ao38, a new cell line from eggs of the black witch moth, Ascalapha odorata (Lepidoptera: Noctuidae), is permissive for AcMNPV infection and produces high levels of recombinant proteins

<p>Abstract</p> <p>Background</p> <p>The insect cell line is a critical component in the production of recombinant proteins in the baculovirus expression system and new cell lines hold the promise of increasing both quantity and quality of protein production.</p> <p>Results</p> <p>Seventy cell lines were established by single-cell cloning from a primary culture of cells derived from eggs of the black witch moth (<it>Ascalapha odorata</it>; Lepidoptera, Noctuidae). Among 8 rapidly growing lines, cell line 38 (Ao38) was selected for further analysis, based on susceptibility to AcMNPV infection and production of secreted alkaline phosphatase (SEAP) from a baculovirus expression vector. In comparisons with low-passage High Five (BTI-Tn-5B1-4) cells, infected Ao38 cells produced β-galactosidase and SEAP at levels higher (153% and 150%, respectively) than those measured from High Five cells. Analysis of N-glycans of SEAP produced in Ao38 cells revealed two N-glycosylation sites and glycosylation patterns similar to those reported for High Five and Sf9 cells. Glycopeptide isoforms consisted of pauci- or oligomannose, with and without fucose on N-acetylglucosamine(s) linked to asparagine residues. Estimates of Ao38 cell volume suggest that Ao38 cells are approximately 2.5× larger than Sf9 cells but only approximately 74% of the size of High Five cells. Ao38 cells were highly susceptible to AcMNPV infection, similar to infectivity of Sf9 cells. Production of infectious AcMNPV budded virions from Ao38 cells peaked at approximately 4.5 × 10⁷IU/ml, exceeding that from High Five cells while lower than that from Sf9 cells. Ao38 cells grew rapidly in stationary culture with a population doubling time of 20.2 hr, and Ao38 cells were readily adapted to serum-free medium (Sf-900III) and to a suspension culture system. Analysis of Ao38 and a parental <it>Ascalapha odorata </it>cell line indicated that these lines were free of the alphanodavirus that was recently identified as an adventitious agent in High Five cell lines.</p> <p>Conclusions</p> <p>Ao38 cells represent a highly productive new insect cell line that will be useful for heterologous protein expression and other applications in biotechnology.</p

Reduced-stress GaN epitaxial layers grown on Si(1 1 1) by using a porous GaN interlayer converted from GaAs

This paper reports the reduced-stress GaN epitaxial growth on Si (1 1 1) using a porous GaN interlayer which is formed from GaAs layer by a novel nitridation process. Initially a 2 μm thick GaAs layer is grown on a Si(1 1 1) substrate by MBE. Then, a GaN buffer layer of 20 nm thick is grown on the GaAs layer at 550°C in a MOVPE reactor. The GaAs layer capped with the GaN buffer layer is annealed in NH3 to 1000°C. Through this process, a porous GaN layer is formed beneath the GaN cap layer. An epitaxial GaN layer is grown on the GaN buffer layer at 1000°C in the MOVPE reactor. The epitaxial layer grown on the porous-GaN/Si(1 1 1) structure is found to have no cracks on the surface. In contrast, an epitaxial layer grown on the GaAs layer nitrated without a cap layer many cracks are found in the epilayer and the layer is sometimes peeled off from the substrate. It is found that the surface morphology of the GaN/porous-GaN/Si(1 1 1) sample is markedly improved by employing a 40 nm-thick interlayer grown at 800°C in addition to the above processes. A PL spectrum with a high intensity ratio between the excitonic emission and the deep yellow emission is obtained for the GaN/porous-GaN/Si(1 1 1) sample. E2 peak position in Raman scattering spectrum also shows a reduced stress for the GaN epilayers grown on the porous-GaN/Si(1 1 1)

Supertubes and Supercurves from M-Ribbons

We construct 1/4 BPS configurations, `M-ribbons', in M-theory on T^2, which give the supertubes and supercurves in type IIA theory upon dimensional reduction. These M-ribbons are generalized so as to be consistent with the SL(2,Z) modular transformation on T^2. In terms of the type IIB theory, the generalized M-ribbons are interpreted as an SL(2,Z) duality family of super D-helix. It is also shown that the BPS M-ribbons must be straight in one direction.Comment: 10 pages, 1 figure, references added, footnote added, BPS eq. (36) is examined without using the solution of field equations, some expressions are improve

Gravitational Dielectric Effect and Myers Effect

In this paper we study the gravitational dielectric phenomena of a D2-brane in the background of Kaluza-Klein monopoles and D6-branes. In both cases the spherical D2-brane with nonzero radius becomes classical solution of the D2-brane action. We also investigate the gravitational Myers effect in the background of D6-branes. This phenomenon occurs since the tension of the D2-brane balances with the repulsive force between D0-branes and D6-branes.Comment: 17 pages, 4 figures, The argument on the stability of the spehrical D2-D0 bound state is modified and clarified in the appendix, The effect of background R-R field are included, reference added, version appeared in PR

Effect of olmesartan on the levels of circulating endothelial progenitor cell after drug-eluting stent implantation in patients receiving statin therapy

AbstractBackgroundThe endothelial progenitor cell (EPC) plays an important role in repairing vascular injury. Statins and angiotensin II receptor blockers increase the level of circulating EPCs. However, it is unknown whether the angiotensin II receptor blocker olmesartan synergistically acts with statins to increase the levels of circulating EPCs. Moreover, the association between the levels of circulating EPCs and endothelial dysfunction after implantation of drug-eluting stents (DESs) has not been evaluated.MethodsNine patients with stable coronary artery disease underwent percutaneous coronary intervention (PCI) and received DES implantation. All patients received olmesartan in addition to statin therapy after PCI. The dose of olmesartan was based on the physician's discretion as per the patients’ blood pressure. The levels of circulating EPCs were analyzed at baseline, post-PCI, and 1, 2, 3, and 8 months after PCI. Coronary angiography and the acetylcholine provocation test were performed on all patients at 8 months.ResultsAlthough the angiotensin II level significantly changed, the levels of circulating EPCs did not change during 8 months of olmesartan treatment (3.1±0.6cells/ml, 2.5±0.8cells/ml, 2.0±0.6cells/ml, 2.9±0.9cells/ml, 3.0±0.4cells/ml, 3.4±0.8cells/ml, p=0.64). The patients were subsequently divided into two groups based on whether the level of circulating EPCs was less or greater than 4cells/ml at 8 months. There were no significant differences in the mean vessel diameter of each segment (proximal, proximal edge, distal edge, and distal) after the acetylcholine provocation test between the two groups.ConclusionsLow-to-moderate doses of olmesartan might not increase the level of circulating EPCs in patients receiving statin therapy. There might be no association between the levels of circulating EPCs and the degree of coronary vasospasm in the acetylcholine provocation test 8 months after DES implantation

Expanded Strings in the Background of NS5-branes via a M2-brane, a D2-brane and D0-branes

Classical configurations of a M2-brane, a D2-brane and D0-branes are investigated in the background of an infinite array of M5-branes or NS5-branes. On the M2-brane, we discuss three kinds of configurations, such as a sphere, a cylinder and a torus-like one. These are stabilized by virtue of the background fluxes of M5-branes. The torus-like M2-brane configuration has winding and momentum numbers of 11th direction, and in terms of the type IIA superstring theory, this corresponds to a torus-like D2-brane with electric and magnetic fluxes on it. We also reproduce the same configuration from a non-abelian Born-Infeld action for D0-branes. It will be a construction of closed strings from D0-branes. An electric flux quantization condition on the D2-brane is also discussed in terms of D0-branes.Comment: 33 pages, 6 figures, references and footnote added, confusing expressions and introduction are improved, version to appear in JHE

LuxS Coexpression Enhances Yields of Recombinant Proteins in Escherichia coli in Part through Posttranscriptional Control of GroEL

Cell-to-cell communication, or quorum sensing (QS), enables cell density-dependent regulation of bacterial gene expression which can be exploited for the autonomous-signal-guided expression of recombinant proteins (C. Y. Tsao, S. Hooshangi, H. C. Wu, J. J. Valdes, and W. E. Bentley, Metab. Eng. 12:291-297, 2010). Earlier observations that the metabolic potential of Escherichia coli is conveyed via the QS signaling molecule autoinducer-2 (AI-2) suggested that the capacity for protein synthesis could also be affected by AI-2 signaling (M. P. DeLisa, J. J. Valdes, and W. E. Bentley, J. Bacteriol. 183:2918-2928, 2001). In this work, we found that simply adding conditioned medium containing high levels of AI-2 at the same time as inducing the synthesis of recombinant proteins doubled the yield of active product. We have hypothesized that AI-2 signaling “conditions” cells as a natural consequence of cell-to-cell communication and that this could tweak the signal transduction cascade to alter the protein synthesis landscape. We inserted luxS (AI-2 synthase) into vectors which cosynthesized proteins of interest (organophosphorus hydrolase [OPH], chloramphenicol acetyltransferase [CAT], or UV-variant green fluorescent protein [GFPuv]) and evaluated the protein expression in luxS-deficient hosts. In this way, we altered the level of luxS in the cells in order to “tune” the synthesis of AI-2. We found conditions in which the protein yield was dramatically increased. Further studies demonstrated coincident upregulation of the chaperone GroEL, which may have facilitated higher yields and is shown for the first time to be positively regulated at the posttranscriptional level by AI-2. This report is the first to demonstrate that the protein synthesis capacity of E. coli can be altered by rewiring quorum sensing circuitry

Investigation of the effect of canagliflozin on the disposition index, a marker of pancreatic beta cell function, in patients with type 2 diabetes

This article is published in Diabetes, Metabolic Syndrome and Obesity: Targets and Therapy, 18 November 2020, Volume 2020:13, Pages 4457-4468

Improving Mathematics Lesson and Evaluation through Setting Inquiry Problem-Solving Activities

The purpose of this research is to make a theoretical framework for improving mathematics lesson and evaluation through setting inquiry problem-solving activities effectively in junior high and senior high mathematics lessons. To achieve this, we first carried out a questionnaire survey to capture how students perceive learning mathematics and, based on the results, made a framework incorporated these activities for lesson design and evaluation of student’s learning. Then, sample lesson conducted in a junior and senior high school based on the framework was analyzed to investigate its effectiveness and get some practical suggestions