Nonlinear Response of Underground Duct Structures with Due Attention to Seismic Input Ground Motions

Under destructive earthquake motions, structures behave heavily in nonlinear manner that is quite different from the linear response computation. The underground structures such as tunnels, ducts are susceptible to the ground motions so that the nonlinear interaction analysis should be performed for the reliable design. In case of irregular soil profile, due to the surface wave generation, the vertical motions come out to a less negligible extent compared to the horizontal motions. The present paper has concerned with such nonlinear response evaluation of a duct structure under strong motions. The effect of the transient characteristics in the inland type and ocean type earthquake motions is investigated. The effect of the vertical component in the ground motions is also evaluated

Enhancement effect of poly(amino acid)s on insulin uptake in alveolar epithelial cells.

In this study, we elucidated the effect of poly(amino acid)s such as poly-L-ornithine (PLO) on FITC-insulin uptake in cultured alveolar type II epithelial cells, RLE-6TN. FITC-insulin uptake by RLE-6TN cells as well as its cell surface binding was markedly increased by PLO without cytotoxicity. The uptake of FITC-insulin in the presence of PLO was shown to be mediated by endocytosis, but in contrast to the uptake in the absence of PLO, the contribution of macropinocytosis emerged. Colocalization of FITC-insulin and LysoTracker Red was observed by confocal laser scanning microscopy both in the absence and presence of PLO, indicating that FITC-insulin was partly targeted to lysosomes in the cells and degraded. The half-life of the intracellular degradation of FITC-insulin was, however, prolonged by the presence of PLO. PLO also stimulated the uptake of other FITC-labeled compounds. Among them, the enhancement effects of PLO on FITC-albumin and FITC-insulin uptake were prominent. The effect of PLO on insulin absorption was also examined in in-vivo pulmonary administration in rats, and co-administration of PLO enhanced the hypoglycemic action of insulin. These findings suggest that co-administration of poly(amino acid)s such as PLO is a useful strategy for enhancing insulin uptake by alveolar epithelial cells and subsequent absorption from the lung.In this study, we elucidated the effect of poly(amino acid)s such as poly-L-ornithine (PLO) on FITC-insulin uptake in cultured alveolar type II epithelial cells, RLE-6TN. FITC-insulin uptake by RLE-6TN cells as well as its cell surface binding was markedly increased by PLO without cytotoxicity. The uptake of FITC-insulin in the presence of PLO was shown to be mediated by endocytosis, but in contrast to the uptake in the absence of PLO, the contribution of macropinocytosis emerged. Colocalization of FITC-insulin and LysoTracker Red was observed by confocal laser scanning microscopy both in the absence and presence of PLO, indicating that FITC-insulin was partly targeted to lysosomes in the cells and degraded. The half-life of the intracellular degradation of FITC-insulin was, however, prolonged by the presence of PLO. PLO also stimulated the uptake of other FITC-labeled compounds. Among them, the enhancement effects of PLO on FITC-albumin and FITC-insulin uptake were prominent. The effect of PLO on insulin absorption was also examined in in-vivo pulmonary administration in rats, and co-administration of PLO enhanced the hypoglycemic action of insulin. These findings suggest that co-administration of poly(amino acid)s such as PLO is a useful strategy for enhancing insulin uptake by alveolar epithelial cells and subsequent absorption from the lung

Mechanism underlying insulin uptake in alveolar epithelial cell line RLE-6TN

For the development of efficient pulmonary delivery systems for protein and peptide drugs, it is important to understand their transport mechanisms in alveolar epithelial cells. In this study, the uptake mechanism for FITC-insulin in cultured alveolar epithelial cell line RLE-6TN was elucidated. FITC-insulin uptake by RLE-6TN cells was time-dependent, temperature-sensitive, and concentration-dependent. The uptake was inhibited by metabolic inhibitors, cytochalasin D, clathrin-mediated endocytosis inhibitors, and dynasore, an inhibitor of dynamin GTPase. On the other hand, no inhibitory effect was observed with caveolae-mediated endocytosis inhibitors and a macropinocytosis inhibitor. Intracellular FITC-insulin was found to be partly transported to the basal side of the epithelial cell monolayers. In addition, colocalization of FITC-insulin and LysoTracker Red was observed on confocal laser scanning microscopy, indicating that FITC-insulin was partly targeted to lysosomes. In accordance with these findings, SDS-PAGE/fluoroimage analysis showed that intact FITC-insulin in the cells was eliminated with time. The possible receptor involved in FITC-insulin uptake by RLE-6TN cells was examined by using siRNA. Transfection of the cells with megalin or insulin receptor siRNA successfully reduced the corresponding mRNA expression. FITC-insulin uptake decreased on the transfection with insulin receptor siRNA, but not that with megalin siRNA. These results suggest that insulin is taken up through endocytosis in RLE-6TN cells, and after the endocytosis, the intracellular insulin is partly degraded in lysosomes and partly transported to the basal side. Insulin receptor, but not megalin, may be involved at least partly in insulin endocytosis in RLE-6TN cells

Mechanism underlying insulin uptake in alveolar epithelial cell line RLE-6TN.

For the development of efficient pulmonary delivery systems for protein and peptide drugs, it is important to understand their transport mechanisms in alveolar epithelial cells. In this study, the uptake mechanism for FITC-insulin in cultured alveolar epithelial cell line RLE-6TN was elucidated. FITC-insulin uptake by RLE-6TN cells was time-dependent, temperature-sensitive, and concentration-dependent. The uptake was inhibited by metabolic inhibitors, cytochalasin D, clathrin-mediated endocytosis inhibitors, and dynasore, an inhibitor of dynamin GTPase. On the other hand, no inhibitory effect was observed with caveolae-mediated endocytosis inhibitors and a macropinocytosis inhibitor. Intracellular FITC-insulin was found to be partly transported to the basal side of the epithelial cell monolayers. In addition, colocalization of FITC-insulin and LysoTracker Red was observed on confocal laser scanning microscopy, indicating that FITC-insulin was partly targeted to lysosomes. In accordance with these findings, SDS-PAGE/fluoroimage analysis showed that intact FITC-insulin in the cells was eliminated with time. The possible receptor involved in FITC-insulin uptake by RLE-6TN cells was examined by using siRNA. Transfection of the cells with megalin or insulin receptor siRNA successfully reduced the corresponding mRNA expression. FITC-insulin uptake decreased on the transfection with insulin receptor siRNA, but not that with megalin siRNA. These results suggest that insulin is taken up through endocytosis in RLE-6TN cells, and after the endocytosis, the intracellular insulin is partly degraded in lysosomes and partly transported to the basal side. Insulin receptor, but not megalin, may be involved at least partly in insulin endocytosis in RLE-6TN cells.For the development of efficient pulmonary delivery systems for protein and peptide drugs, it is important to understand their transport mechanisms in alveolar epithelial cells. In this study, the uptake mechanism for FITC-insulin in cultured alveolar epithelial cell line RLE-6TN was elucidated. FITC-insulin uptake by RLE-6TN cells was time-dependent, temperature-sensitive, and concentration-dependent. The uptake was inhibited by metabolic inhibitors, cytochalasin D, clathrin-mediated endocytosis inhibitors, and dynasore, an inhibitor of dynamin GTPase. On the other hand, no inhibitory effect was observed with caveolae-mediated endocytosis inhibitors and a macropinocytosis inhibitor. Intracellular FITC-insulin was found to be partly transported to the basal side of the epithelial cell monolayers. In addition, colocalization of FITC-insulin and LysoTracker Red was observed on confocal laser scanning microscopy, indicating that FITC-insulin was partly targeted to lysosomes. In accordance with these findings, SDS-PAGE/fluoroimage analysis showed that intact FITC-insulin in the cells was eliminated with time. The possible receptor involved in FITC-insulin uptake by RLE-6TN cells was examined by using siRNA. Transfection of the cells with megalin or insulin receptor siRNA successfully reduced the corresponding mRNA expression. FITC-insulin uptake decreased on the transfection with insulin receptor siRNA, but not that with megalin siRNA. These results suggest that insulin is taken up through endocytosis in RLE-6TN cells, and after the endocytosis, the intracellular insulin is partly degraded in lysosomes and partly transported to the basal side. Insulin receptor, but not megalin, may be involved at least partly in insulin endocytosis in RLE-6TN cells

Vibrational spectroscopy analysis of ligand efficacy in human M₂ muscarinic acetylcholine receptor (M₂R)

振動分光法を駆使した薬剤効能測定法の開発 --アセチルコリン受容体を標的とした神経疾患の治療薬開発への期待--. 京都大学プレスリリース. 2021-12-01.The intrinsic efficacy of ligand binding to G protein-coupled receptors (GPCRs) reflects the ability of the ligand to differentially activate its receptor to cause a physiological effect. Here we use attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy to examine the ligand-dependent conformational changes in the human M₂ muscarinic acetylcholine receptor (M₂R)). We show that different ligands affect conformational alteration appearing at the C=O stretch of amide-I band in M2R. Notably, ATR-FTIR signals strongly correlated with G-protein activation levels in cells. Together, we propose that amide-I band serves as an infrared probe to distinguish the ligand efficacy in M₂R) and paves the path to rationally design ligands with varied efficacy towards the target GPCR

Development of the analog ASIC for multi-channel readout X-ray CCD camera

We report on the performance of an analog application-specific integrated circuit (ASIC) developed aiming for the front-end electronics of the X-ray CCDcamera system onboard the next X-ray astronomical satellite, ASTRO-H. It has four identical channels that simultaneously process the CCD signals. Distinctive capability of analog-to-digital conversion enables us to construct a CCD camera body that outputs only digital signals. As the result of the front-end electronics test, it works properly with low input noise of =<30 uV at the pixel rate below 100 kHz. The power consumption is sufficiently low of about 150 mW/chip. The input signal range of 720 mV covers the effective energy range of the typical X-ray photon counting CCD (up to 20 keV). The integrated non-linearity is 0.2% that is similar as those of the conventional CCDs in orbit. We also performed a radiation tolerance test against the total ionizing dose (TID) effect and the single event effect. The irradiation test using 60Co and proton beam showed that the ASIC has the sufficient tolerance against TID up to 200 krad, which absolutely exceeds the expected amount of dose during the period of operating in a low-inclination low-earth orbit. The irradiation of Fe ions with the fluence of 5.2x10^8 Ion/cm2 resulted in no single event latchup (SEL), although there were some possible single event upsets. The threshold against SEL is higher than 1.68 MeV cm^2/mg, which is sufficiently high enough that the SEL event should not be one of major causes of instrument downtime in orbit.Comment: 16 pages, 6 figure

Molecular Evolutionary Analyses of the RNA-Dependent RNA Polymerase Region in Norovirus Genogroup II

Noroviruses are the leading cause of viral gastroenteritis in humans across the world. RNA-dependent RNA polymerase (RdRp) plays a critical role in the replication of the viral genome. Although there have been some reports on a limited number of genotypes with respect to the norovirus evolution of the RdRp region, no comprehensive molecular evolution examination of the norovirus GII genotype has yet been undertaken. Therefore, we conducted an evolutionary analysis of the 25 genotypes of the norovirus GII RdRp region (full-length), collected globally using different bioinformatics technologies. The time-scaled phylogenetic tree, generated using the Bayesian Markov Chain Monte Carlo (MCMC) method, indicated that the common ancestor of GII diverged from GIV around 1443 CE [95% highest posterior density (HPD), 1336–1542]. The GII RdRp region emerged around 1731 CE (95% HPD, 1703–1757), forming three lineages. The evolutionary rate of the RdRp region of the norovirus GII strains was estimated at over 10−3 substitutions/site/year. The evolutionary rates were significantly distinct in each genotype. The composition of the phylogenetic distances differed among the strains for each genotype. Furthermore, we mapped the negative selection sites on the RdRp protein and many of these were predicted in the GII.P4 RdRp proteins. The phylodynamics of GII.P4, GII.P12, GII.P16, and GII.Pe showed that their effective population sizes increased during the period from 2003 to 2014. Our results cumulatively suggest that the RdRp region of the norovirus GII rapidly and uniquely evolved with a high divergence similar to that of the norovirus VP1 gene