Conditional activation of RhoA suppresses the epithelial to mesenchymal transition at the primitive streak during mouse gastrulation

Gastrulation is a pivotal event of mouse early embryogenesis. In telencephalin (TLCN)-Cre mice carrying the Cre recombinase gene inserted into the translational initiation site of the TLCN gene, Cre-mediated recombination took place at the postimplantation stage. To examine the role of RhoA signaling in early embryogenesis, we produced Rho36 mice carrying constitutively active RhoA(G14V) gene inducible by Cre recombinase and crossed with TLCN-Cre mice. In doubly transgenic embryos at the gastrulation stage, there appeared an abnormal bulge of cells protruded from the primitive streak region into the amniotic cavity. The bulged cell mass expressed the epiblast marker gene Oct3 and E-cadherin, but not the primitive streak marker gene T except for the basal portion. These results suggest that the conditional activation of RhoA signaling suppressed the epithelial to mesenchymal transition at the primitive streak during mouse gastrulation.     

Nonylphenol induces the death of neural stem cells due to activation of the caspase cascade and regulation of the cell cycle

Endocrine disruptors (EDs) are a great concern throughout the world, because they have adverse effects on human health and wildlife. In the present study, we investigated the effects of EDs on the proliferation and survival of murine neural stem cells (NSCs). In contrast to bisphenol A, phthalic acid benzyl n-butyl ester, phthalic acid di-n-butyl ester and phthalic acid di(2-ethylhexyl) ester, the treatment of NSCs with 4-nonylphenol for 24 h inhibited cell growth in a concentration-dependent manner. In addition, treatment with 4-nonylphenol resulted in nuclear condensation and DNA fragmentation (morphological changes due to apoptosis) in NSCs after 12 h of exposure, and activated caspase-3 after 6 h and 9 h of exposure. Furthermore, an exposure to 4-nonylphenol led to the accumulation of cells at the G2/M phase interface and down-regulated the protein levels of cyclin A and B1, which are the major regulatory proteins at the G2 to M transition of the cell cycle. Together, these results indicate that, in contrast to other EDs, 4-nonylphenol may exhibit a potent cytotoxicity through apoptosis via the caspase cascade and cell cycle arrest at the G2/M phase, and suggest that 4-nonylphenol may affect neurogenesis in the CNS.     

In vitro and in vivo effects of the overexpression of osteopontin on osteoblast differentiation using a recombinant adenoviral vector

Osteopontin (OPN) is a highly acidic secreted phosphoprotein that binds to cells via an RGD (arginine-glycine-aspartic acid) cell adhesion sequence that recognizes the alphaVbeta3 integrin. OPN may regulate the formation and remodeling of bone. To elucidate the function of OPN in bone tissue, we examined the overexpression of OPN in osteoblasts in vitro and in vivo using an adenoviral vector carrying an OPN cDNA (Adv-OPN). Rat bone marrow-derived osteoblasts infected with Adv-OPN were examined by Western blotting, immunofluorescence, nodule formation measurements, assay of alkaline phosphatase (ALP) activity, and Northern blotting. The results suggested that not only osteoblast differentiation markers such as osteocalcin and ALP, but nodule formation and ALP activity are markedly enhanced by OPN overexpression in the case of viral infection. On the contrary, when Adv-OPN and uninfected osteoblasts were implanted into subcutaneous sites with a porous ceramic scaffold, the ALP activity and calcium content of the OPN-infected composite were higher than in uninfected composites, however, the differences were smaller than expected from the in vitro experiments. We speculate that the difference in the result of in vitro and in vivo experiments originates from the inhibitory effect of secreted OPN on the crystal growth of apatite in vivo, which competes with the induced activity of osteoblasts.     

A neural model of predictive recognition in form pathway of visual cortex

We present a functional model of form pathway in visual cortex based on predictive coding scheme, in which the prediction is compared with feedforward signals filtered by two kinds of spatial resolution maps, broad and fine resolution map. We propose here the functional role of the prediction and of the two kinds of resolution maps in perception of object form in visual system. The prediction is represented based on memory of dynamical attractors in temporal cortex, categorized by an elemental figure in posterior temporal cortex. The prediction is generated by the feedforward signals of main neurons in broad resolution maps of V(1) and V(4), and then is compared with the feedforward signals of main neurons in fine resolution map of V(1) and V(4).     

Chemical constituents of Murraya siamensis: three coumarins and their anti-tumor promoting effect

Isolation and structure elucidation of three coumarins, murrayacoumarins A, B, and C, together with eight known coumarins, from the leaves of Murraya siamensis Craib collected in Thailand are described. Results of a primary screening of inhibitory effects of seven of these compounds on 12-O-tetradecanoylphorbol-13-acetate-induced Epstein-Barr virus early antigen activation in Raji cells are also presented.     

Gasdermin D Exerts Anti-inflammatory Effects by Promoting Neutrophil Death

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Inhibition of lens aldose reductase by flavonoids

The inhibitory activities of 73 flavonoids against rat aldose reductase were systematically investigated and cosmosiin, luteolin-7-glucuronide, lonicerin, 6-hydroxyluteolin, kaempferol-3-rhamnoside and avicularin were newly found to be highly active. The degree of inhibition appears to depend on the solvent system used. In general flavones are more active than flavonols and flavanones, glycosides are more active than aglycones, and the number of sugars present affects the activity.     

Activation of human plasma prekallikrein by Pseudomonas aeruginosa elastase. II. Kinetic analysis and identification of scissile bondof prekallikrein in the activation

Activation of human plasma prekallikrein by a bacterial metalloendopeptidase, Pseudomonas aeruginosa elastase, was reported (Shibuya et al. (1991) Biochim. Biophys. Acta 1097, 23–27). Details of the activation process were presently studied. The activation accompanied limited proteolysis of a peptide bond inside of a disulfide bridge of prekallikrein molecule. Amino acid sequencing analysis of the newly generated amino-terminal revealed that the cleavage site was Arg₃₇₁-Ile₃₇₂ bond which is the scissile bond in the activation of prekallikrein with trypsin-type proteinases. A pentapeptide substrate, 2-aminobenzoyl-Ser-Thr-Ile-Val-4-nitrobenzylamide, which contained the amino acid sequence identical to that around the scissile bond of prekallikrein was synthesized. Pseudomonal elastase, indeed, hydrolyzed the substrate at Arg-Ile bond with the kinetic parameters of K_m = 118 μM, k_cat = 1.56/s and k_cat/K_m = 1.33 · 10⁴/s M. These results indicated that the Arg₃₇₁-Ile₃₇₂ bond was sensitive not only to trypsin-type serine proteinases, but also a bacterial metalloproteinase. Kinetic analysis of the prekallikrein activation by psuedomonal elastase, however, revealed that the activation rate was show, though the K_m values was good enough to expect an occurence of this activation in vivo (K_m = 248 nM, k = 6.8 · 10^?4/s, and k_cat/K_m = 2.7 · 10³/s M. The activation rate of prekallikrein by pseudomonal elastase in Hageman factor deficient plasma was remarkably improved when the plasma was reconstituted with purified Hageman factor molecule. From the results, a biologuical significance of the proteinase cascade in the plasma kinin generation was also indicated. The present in vitro study might support the hypothesis that the Hageman factor/kallikrein-kinin system plays an important role in bacterial infection including the pseudomonal one.     

The Hageman factor-dependent system in the vascular permeability reaction

The mechanism by which the Hageman factor-dependent system induces vascular permeability has been analyzed. The Mr-28,000 active fragment of guinea pig Hageman factor (beta-HFa), injected intradermally, induces an increase in local vascular permeability. Inhibition of vascular permeability resulted from pretreatment of the beta-HFa with immunopurified anti-Hageman factor F(ab')2 antibody at concentrations of 10(-6)-10(-7) M as well as by incubation with corn and pumpkin seed inhibitors of beta-HFa. To determine whether prekallikrein and kallikrein participated in the permeability induced by beta-HFa, circulating prekallikrein was depleted by intra-arterial injections of anti-prekallikrein F(ab')2 antibody. This resulted in about 80% diminution of the vascular permeability response to beta-HFa, without affecting the permeability reaction to bradykinin. Soybean trypsin inhibitor (10(-6) M), injected at the same cutaneous site as the beta-HFa, inhibited the vascular permeability response to beta-HFa by more than 90%. This concentration of soybean inhibitor blocked more than 90% of the activity of guinea pig plasma kallikrein, but did not inhibit the amidolytic capacity of beta-HFa. The permeability activity of beta-HFa (but not its amidolytic activity) was augmented 10-fold by simultaneous injection of a synthetic kinin potentiator, SQ 20,881 (Glu-Tyr-Pro-Arg-Pro-Gln-Ile-Pro-Pro-OH), and was almost completely inhibited by the simultaneous injection of a kinin-destroying enzyme, carboxypeptidase B. These results support the hypothesis that the greatest proportion of vascular permeability induced by beta-HFa is produced by the activation of prekallikrein followed by the release of kinin in the cutaneous tissue. These data offer the first in vivo evidence that the Hageman factor-dependent system by itself can induce inflammatory changes.