Solid-gas carbonylation of aryloxide rhodium(I) complexes: stepwise reaction forming Vaska-type complexes

Aryloxide rhodium(I) complexes Rh(OAr)(PPh₃)₃ (1a: Ar=C₆Cl₅, 1b: Ar=C₆F₅, 1c: Ar=C₆H₄-NO₂-4) react with CO in toluene solutions to produce Vaska-type complexes trans-Rh(OAr)(CO)(PPh₃)₂ (2a: Ar=C₆Cl₅, 2b: Ar=C₆F₅, 2c: Ar=C₆H₄-NO₂-4). Carbonylation of a similar complex with PMe₃ ligands, Rh(OC₆H₄-NO₂-4)(PMe₃)₃ (3c), also forms trans-Rh(OC₆H₄-NO₂-4)(CO)(PMe₃)₂ (4c). Molecular structures of the complexes are determined by X-ray crystallography and NMR spectroscopy. Complex 1a reacts with CO in the absence of solvent to produce a mixture of 2a and complex A, the latter of which shows the IR and ¹³C{¹H} signals due to the carbonyl ligand at different positions from those of 2a. Addition of Et₂O to the above mixture turns it into analytically pure 2a. Carbonylation of 1b and 1c under the solvent-free conditions produces complexes B and C as the respective products of the solid-gas reaction. Recrystallization of B and C turns them into 2b and 2c, respectively. Complex 3c also reacts with CO in the solid state to form a mixture of 4c and complex D, although the latter complex is converted slowly into 4c even in the solid state.     

In vivo MR detection of vascular endothelial injury using a new class of MRI contrast agent

A new class of dye-based MRI contrast agents, EB-DTPA-Gd, was designed and synthesized. The contrast agent was found to accumulate at the site of endothelial injury when the reagent was applied to isolated porcine blood vessels or in an ex vivo experiment using rat. In vivo MR detection of vascular endothelial injury was also successful in rat with its common carotid artery injured by balloon treatment. These results indicate that EB-DTPA-Gd is potentially useful for the diagnosis of vascular diseases.     

Prion Replication Elicits Cytopathic Changes in Differentiated Neurosphere Cultures

The molecular mechanisms of prion-induced cytotoxicity remain largely obscure. Currently, only a few cell culture models have exhibited the cytopathic changes associated with prion infection. In this study, we introduced a cell culture model based on differentiated neurosphere cultures isolated from the brains of neonatal prion protein (PrP)-null mice and transgenic mice expressing murine PrP (dNP0 and dNP20 cultures). Upon exposure to mouse Chandler prions, dNP20 cultures supported the de novo formation of abnormal PrP and the resulting infectivity, as assessed by bioassays. Furthermore, this culture was susceptible to various prion strains, including mouse-adapted scrapie, bovine spongiform encephalopathy, and Gerstmann-Sträussler-Scheinker syndrome prions. Importantly, a subset of the cells in the infected culture that was mainly composed of astrocyte lineage cells consistently displayed late-occurring, progressive signs of cytotoxicity as evidenced by morphological alterations, decreased cell viability, and increased lactate dehydrogenase release. These signs of cytotoxicity were not observed in infected dNP0 cultures, suggesting the requirement of endogenous PrP expression for prion-induced cytotoxicity. Degenerated cells positive for glial fibrillary acidic protein accumulated abnormal PrP and exhibited features of apoptotic death as assessed by active caspase-3 and terminal deoxynucleotidyltransferase nick-end staining. Furthermore, caspase inhibition provided partial protection from prion-mediated cell death. These results suggest that differentiated neurosphere cultures can provide an in vitro bioassay for mouse prions and permit the study of the molecular basis for prion-induced cytotoxicity at the cellular level.     

Nasal cholera toxin elicits IL-5 and IL-5 receptor alpha-chain expressing B-1a B cells for innate mucosal IgA antibody responses

In this study, we examine whether native cholera toxin (nCT) as a mucosal adjuvant can support trinitrophenyl (TNP)-LPS-specific mucosal immune responses. C57BL/6 mice were given nasal TNP-LPS in the presence or absence of nCT. Five days later, significantly higher levels of TNP-specific mucosal IgA Ab responses were induced in the nasal washes, saliva, and plasma of mice given nCT plus TNP-LPS than in those given TNP-LPS alone. High numbers of TNP-specific IgA Ab-forming cells were also detected in mucosal tissues such as the nasal passages (NPs), the submandibular glands (SMGs), and nasopharyngeal-associated lymphoreticular tissue of mice given nCT. Flow cytometric analysis showed that higher numbers of surface IgA+, CD5+ B cells (B-1a B cells) in SMGs and NPs of mice given nasal TNP-LPS plus nCT than in those given TNP-LPS alone. Furthermore, increased levels of IL-5R alpha-chain were expressed by B-1a B cells in SMGs and NPs of mice given nasal TNP-LPS plus nCT. Thus, CD4+ T cells from these mucosal effector lymphoid tissues produce high levels of IL-5 at both protein and mRNA levels. When mice were treated with anti-IL-5 mAb, significant reductions in TNP-specific mucosal IgA Ab responses were noted in external secretions. These findings show that nasal nCT as an adjuvant enhances mucosal immune responses to a T cell-independent Ag due to the cross-talk between IL-5Ralpha+ B-1a B cells and IL-5-producing CD4+ T cells in the mucosal effector lymphoid tissues.     

Calcium-induced synergistic inhibition of a translational factor eEF2 in nerve growth cones

Local protein synthesis in nerve growth cones has been suggested, but how it is controlled remains largely unknown. We found eukaryotic elongation factor-2 (eEF2), a key component of mRNA translation, in growth cones by immunocytochemistry. While phosphorylated eEF2 was weakly distributed in advancing growth cones, eEF2 phosphorylation was increased by high potassium-evoked calcium influx. In the growth cone, calcium elevation increased eEF2 kinase (EF2K), a calcim-calmodulin-dependent enzyme. Calcium also decreased the level of phosphorylated p70-S6 kinase (S6K), a kinase known to inhibit EF2K. Moreover, calcium elevation decreased total eEF2 in growth cones. Since phosphorylated eEF2 inhibits mRNA translation, calcium elevation appears to inhibit mRNA translation in growth cones by a synergistic mechanism involving regulation of EF2K, S6K, and eEF2 itself. Time-lapse imaging showed that calcium elevation induced growth arrest of neurites. The inhibitory effect on mRNA translation may thus be involved in the regulation of neurite outgrowth.     

Regioselective and enzymatic production of γ-resorcylic acid from resorcinol using recombinant <Emphasis Type="Italic">Escherichia coli</Emphasis> cells expressing a novel decarboxylase gene

A recombinant Escherichia coli, expressing the rdc gene, which encodes a γ-resorcylic acid decarboxylase (Rdc) reversibly catalyzing regioselective carboxylation of resorcinol derived from Rhizobium radiobacter WU-0108, converted 20 mM resorcinol to γ-resorcylic acid with a 44% (mol/mol) yield at 30°C for 7 h. The recombinant E. coli cells were recyclable at least five times for use as biocatalysts.     

A major locus involved in the formation of the radial oxygen loss barrier in adventitious roots of teosinte Zea nicaraguensis is located on the short‐arm of chromosome 3

A radial oxygen loss (ROL) barrier in roots of waterlogging‐tolerant plants promotes oxygen movement via aerenchyma to the root tip, and impedes soil phytotoxin entry. The molecular mechanism and genetic regulation of ROL barrier formation are largely unknown. Zea nicaraguensis, a waterlogging‐tolerant wild relative of maize (Zea mays ssp. mays), forms a tight ROL barrier in its roots when waterlogged. We used Z. nicaraguensis chromosome segment introgression lines (ILs) in maize (inbred line Mi29) to elucidate the chromosomal region involved in regulating root ROL barrier formation. A segment of the short‐arm of chromosome 3 of Z. nicaraguensis conferred ROL barrier formation in the genetic background of maize. This chromosome segment also decreased apoplastic solute permeability across the hypodermis/exodermis. However, the IL and maize were similar for suberin staining in the hypodermis/exodermis at 40 mm and further behind the root tip. Z. nicaraguensis contained suberin in the hypodermis/exodermis at 20 mm and lignin at the epidermis. The IL with ROL barrier, however, did not contain lignin in the epidermis. Discovery of the Z. nicaraguensis chromosomal region responsible for root ROL barrier formation has improved knowledge of this trait and is an important step towards improvement of waterlogging tolerance in maize.     

Susceptibility of GT1–7 cells to mouse-passaged field scrapie isolates with a long incubation

A typical feature of scrapie in sheep and goats is the accumulation of disease-associated prion protein. Scrapie consists of many strains with different biological properties. Nine natural sheep scrapie cases were transmitted to wild-type mice and mouse-passaged isolates were classified into 2 types based on incubation time: short and long. These 2 types displayed a distinct difference in their pathology. We attempted to transmit these mouse-passaged isolates to 2 murine cell lines (GT1–7 and L929) to compare their properties. All of the isolates were transmitted to L929 cells. However, only mouse-passaged field isolates with a long incubation time were transmitted to GT1–7 cells. This specific susceptibility of GT1–7 cells was also confirmed with a primary-passaged isolate that was not completely adapted to the new host species. Characterization of the mechanisms of the specific susceptibility of GT1–7 cells to isolates with a long incubation time may lead to a greater understanding of the differences among prion strains.