Parametric Instability of Helical Oscillations with External Signals in a Positive Column

ArticleJournal of the Faculty of Textile Science and Technology, Shinshu University. Ser. F, Physics and mathematics 8: 1-10(1990)departmental bulletin pape

Tespa1 is a novel inositol 1,4,5-trisphosphate receptor binding protein in T and B lymphocytes

AbstractTespa1 has been recently reported to be a critical molecule in T-cell development, however, the precise molecular mechanisms of Tespa1 remain elusive. Here, we demonstrate that Tespa1 shows amino-acid sequence homology to KRAS-induced actin-interacting protein (KRAP), an inositol 1,4,5-trisphosphate receptor (IP3R) binding protein, and that Tespa1 physically associates with IP3R in T and B lymphocytes. Two-consecutive phenylalanine residues (Phe185/Phe186) in Tespa1, which are conserved between Tespa1 and KRAP, are indispensable for the association between Tespa1 and IP3R. These findings suggest that Tespa1 plays critical roles in the immune system through the regulation of the IP3R

Hepatitis B Virus e Antigen Physically Associates With Receptor-Interacting Serine/Threonine Protein Kinase 2 and Regulates IL-6 Gene Expression

We previously reported that hepatitis B virus (HBV) e antigen (HBeAg) inhibits production of interleukin 6 by suppressing NF-κB activation. NF-κB is known to be activated through receptor-interacting serine/threonine protein kinase 2 (RIPK2), and we examined the mechanisms of interleukin 6 regulation by HBeAg. HBeAg inhibits RIPK2 expression and interacts with RIPK2, which may represent 2 mechanisms through which HBeAg blocks nucleotide-binding oligomerization domain-containing protein 1 ligand-induced NF-κB activation in HepG2 cells. Our findings identified novel molecular mechanisms whereby HBeAg modulates intracellular signaling pathways by targeting RIPK2, supporting the concept that HBeAg could impair both innate and adaptive immune responses to promote chronic HBV infectio

Hepatitis C Virus Nonstructural 5A Protein Inhibits Lipopolysaccharide-Mediated Apoptosis of Hepatocytes by Decreasing Expression of Toll-Like Receptor 4

Background. Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) has been shown to modulate multiple cellular processes, including apoptosis. The aim of this study was to assess the effects of HCV NS5A on apoptosis induced by Toll-like receptor (TLR) 4 ligand, lipopolysaccharide (LPS). Methods. Apoptotic responses to TLR4 ligands and the expression of molecules involved in TLR signaling pathways in human hepatocytes were examined with or without expression of HCV NS5A. Results. HCV NS5A protected HepG2 hepatocytes against LPS-induced apoptosis, an effect linked to reduced TLR4 expression. A similar downregulation of TLR4 expression was observed in Huh-7-expressing genotype 1b and 2a. In agreement with these findings, NS5A inhibited the expression of numerous genes encoding for molecules involved in TLR4 signaling, such as CD14, MD-2, myeloid differentiation primary response gene 88, interferon regulatory factor 3, and nuclear factor-κB2. Consistent with a conferred prosurvival advantage, NS5A diminished the poly(adenosine diphosphate-ribose) polymerase cleavage and the activation of caspases 3, 7, 8, and 9 and increased the expression of anti-apoptotic molecules Bcl-2 and c-FLIP. Conclusions. HCV NS5A downregulates TLR4 signaling and LPS-induced apoptotic pathways in human hepatocytes, suggesting that disruption of TLR4-mediated apoptosis may play a role in the pathogenesis of HCV infectio

μ1B, a novel adaptor medium chain expressed in polarized epithelial cells11The nucleotide sequences reported in this paper have been submitted to GenBank with accession numbers AF020797 (human μ1B) and AF067146 (mouse μ1B).

AbstractThe apical and basolateral plasma membrane domains of polarized epithelial cells contain distinct sets of integral membrane proteins. Biosynthetic targeting of proteins to the basolateral plasma membrane is mediated by cytosolic tail determinants, many of which resemble signals involved in the rapid endocytosis or lysosomal targeting. Since these signals are recognized by adaptor proteins, we hypothesized that there could be epithelial-specific adaptors involved in polarized sorting. Here, we report the identification of a novel member of the adaptor medium chain family, named μ1B, which is closely related to the previously described μ1A (79% amino acid sequence identity). Northern blotting and in situ hybridization analyses reveal the specific expression of μ1B mRNA in a subset of polarized epithelial and exocrine cells. Yeast two-hybrid analyses show that μ1B is capable of interacting with generic tyrosine-based sorting signals. These observations suggest that μ1B may be involved in protein sorting events specific to polarized cells

Inhibition of Phosphodiesterase-4 (PDE4) activity triggers luminal apoptosis and AKT dephosphorylation in a 3-D colonic-crypt model

BACKGROUND: We previously established a three-dimensional (3-D) colonic crypt model using HKe3 cells which are human colorectal cancer (CRC) HCT116 cells with a disruption in oncogenic KRAS, and revealed the crucial roles of oncogenic KRAS both in inhibition of apoptosis and in disruption of cell polarity; however, the molecular mechanism of KRAS-induced these 3-D specific biological changes remains to be elucidated. RESULTS: Among the genes that were upregulated by oncogenic KRAS in this model, we focused on the phosphodiesterase 4B (PDE4B) of which expression levels were found to be higher in clinical tumor samples from CRC patients in comparison to those from healthy control in the public datasets of gene expression analysis. PDE4B2 was specifically overexpressed among other PDE4 isoforms, and re-expression of oncogenic KRAS in HKe3 cells resulted in PDE4B overexpression. Furthermore, the inhibition of PDE4 catalytic activity using rolipram reverted the disorganization of HCT116 cells into the normal physiologic state of the epithelial cell polarity by inducing the apical assembly of ZO-1 (a tight junction marker) and E-cadherin (an adherens junction marker) and by increasing the activity of caspase-3 (an apoptosis marker) in luminal cavities. Notably, rolipram reduced the AKT phosphorylation, which is known to be associated with the disruption of luminal cavity formation and CRC development. Similar results were also obtained using PDE4B2-shRNAs. In addition, increased expression of PDE4B mRNA was found to be correlated with relapsed CRC in a public datasets of gene expression analysis. CONCLUSIONS: These results collectively suggested that PDE4B is upregulated by oncogenic KRAS, and also that the inhibition of PDE4 catalytic activity can induce both epithelial cell polarity and luminal apoptosis in CRC, thus highlighting the utility of our 3-D culture (3 DC) model for the KRAS-induced development of CRC in 3-D microenvironment. Indeed, using this model, we found that PDE4B is a promising candidate for a therapeutic target as well as prognostic molecular marker in CRC. Further elucidation of the signaling network of PDE4B2 in 3 DC would provide a better understanding of CRC in vivo

Novel method for immunofluorescence staining of mammalian eggs using non-contact alternating-current electric-field mixing of microdroplets

Recently, a new technique was developed for non-catalytically mixing microdroplets. In this method, an alternating-current (AC) electric field is used to promote the antigen-antibody reaction within the microdroplet. Previously, this technique has only been applied to histological examinations of flat structures, such as surgical specimens. In this study, we applied this technique for the first time to immunofluorescence staining of three-dimensional structures, specifically, mammalian eggs. We diluted an antibody against microtubules from 1:1,000 to 1:16,000, and compared the chromatic degree and extent of fading across dilutions. In addition, we varied the frequency of AC electricfield mixing from 5 Hz to 46 Hz and evaluated the effect on microtubule staining. Microtubules were more strongly stained after AC electric-field mixing for only 5 minutes, even when the concentration of primary antibody was 10 times lower than in conventional methods. AC electric-field mixing also alleviated microtubule fading. At all frequencies tested, AC electric-field mixing resulted in stronger microtubule staining than in controls. There was no clear difference in a microtubule staining between frequencies. These results suggest that the novel method could reduce antibody consumption and shorten immunofluorescence staining time

Comparative Genetic Mapping and Discovery of Linkage Disequilibrium Across Linkage Groups in White Clover (Trifolium repens L.)

White clover (Trifolium repens L.) is an allotetraploid species (2n = 4X = 32) that is widely distributed in temperate regions and cultivated as a forage legume. In this study, we developed expressed sequence tag (EST)–derived simple sequence repeat (SSR) markers, constructed linkage maps, and performed comparative mapping with other legume species. A total of 7982 ESTs that could be assembled into 5400 contigs and 2582 singletons were generated. Using the EST sequences that were obtained, 1973 primer pairs to amplify EST-derived SSR markers were designed and used for linkage analysis of 188 F1 progenies, which were generated by a cross between two Japanese plants, ‘273-7’ and ‘T17-349,’ with previously published SSR markers. An integrated linkage map was constructed by combining parental-specific maps, which consisted of 1743 SSR loci on 16 homeologous linkage groups with a total length of 2511 cM. The primer sequences of the developed EST-SSR markers and their map positions are available on http://clovergarden.jp/. Linkage disequilibrium (LD) was observed on 9 of 16 linkage groups of a parental-specific map. The genome structures were compared among white clover, red clover (T. pratense L.), Medicago truncatula, and Lotus japonicus. Macrosynteny was observed across the four legume species. Surprisingly, the comparative genome structure between white clover and M. truncatula had a higher degree of conservation than that of the two clover species

The location of 8-shaped hatching influences inner cell mass formation in mouse blastocysts

The hatching of a blastocyst where the blastocyst portions on the inside and the outside of the zona pellucida feature a figure-of-eight shape is termed 8-shaped hatching; this type of hatching has been reported to affect the proper presentation of the inner cell mass (ICM) in both human and mouse embryos. Here, our aim was to investigate the factors that affect ICM presentation during 8-shaped hatching. We performed IVF by using B6D2F1 female mice and ICR male mice, and used the 104 captured blastocysts. Embryos were maintained in KSOM at 37 degrees C in a 5% CO2, 5% O-2, and 90% N-2 environment, and their growth behavior was monitored individually and continuously using time-lapse cinematography. At 120 h after insemination, embryos were immunostained and examined under a confocal microscope. We used the hatching form to identify 8-shaped hatching, and we classified the 8 shaped- hatching blastocysts into two groups, one in which the hatching site was near the ICM center, and the other in which the hatching site was far from the ICM center. We measured each group for ICM size and the number of Oct3/4-positive cells. Of the 95 hatching or hatched embryos, 74 were 8-shaped-hatching blastocysts, and in these embryos, the ICM was significantly wider when the hatching site was near the ICM than when the hatching site was far from the ICM (P = 0.0091). Moreover, in the 8-shaped-hatching blastocysts in which the ICM was included in the blastocyst portion outside the zona pellucida. the portion defined as the outside blastocyst. after the collapse of this outside blastocyst, the ICM adhered to the trophectoderm of the outside blastocyst, opposite the hatching site. Our results indicate that in 8-shaped-hatching blastocysts, the hatching site and the collapse of outside blastocyst affect ICM formation. Thus, the assessment of 8-shaped hatching behaviors could yield indices for accurately evaluating embryo quality