Fluorescent analysis of alpha-keto acids in serum and urine by high-performance liquid chromatography

A selective procedure using synthetic substrates for determination of exo-1,4,-beta-glucanases in a mixture of exoglucanases , endoglucanases , and beta-glucosidases is formulated. The heterobiosides , p- nithrophenyl -beta-D- cellobioside ( pNPC ) or p-nitrophenyl-beta-D-lactoside ( pNPL ), were used as selective substrates for the measurement of exoglucanase activity. The exoglucanases (especially cellobiohydrolases , which split off cellobiose units from the nonreducing end of the cellulose chain) specifically act on the agluconic bond (between p-nitrophenyl and the disaccharide moiety) and not on the holosidic bond (between the two glucose units of cellobiose). The interfering effect of beta-glucosidase, which acts on both agluconic and holosidic bonds, is overcome by the addition of D-glucono-1,5-delta-lactone, a specific inhibitor of beta-glucosidases. The interference of endoglucanases , which also act on both agluconic and holosidic bonds, can be compensated for by prior standardization of the assay procedure with a purified endoglucanase from the studied mixture of cellulases.     

Analysis of water in Autonomous Biological Systems (ABS) samples.

Several soluble components, peptidase and amino acids, and carbon isotopic ratio in the water retrieved from flight experiments of Autonomous Biological Systems (ABS) as well as ground control samples are analyzed to interpret the condition, dynamics, material balance of the ABS ecosystems. Organic carbons in flight samples were found to be more abundant compared with the control ones, which suggested the uniform ecosystems in low gravity might easily dissolve more soluble components. The Mir-1997 flight sample showed higher C/N ratio probably because of the dissolution of carbon-rich plant materials.     

Chloroplast DNA phylogeography of Fagus crenata (Fagaceae) in Japan

 CpDNA variation in Japanese beech, Fagus crenata (Fagaceae), was studied in 45 populations distributed throughout the species' range. Two cpDNA regions were sequenced: the non-coding region between the trnL (UAA) 5′exon and trnF (GAA), and the trnK region (including matK). Thirteen distinct cpDNA haplotypes were recognized and each haplotype was found to be geographically structured. Two major clades (I and II+III) were revealed in phylogenetic analyses among the haplotypes using F. sylvatica as an outgroup. The haplotypes of Clade I were distributed mainly along the Japan Sea side of the Japanese Archipelago, while those of Clade II+III occurred chiefly along the Pacific Ocean side. Consequently, the distribution of the two major cpDNA clades suggests that there were two migration routes in the history of F. crenata; one along the Japan Sea and the other along the Pacific Ocean side of the Japanese Islands. Received March 19, 2001 Accepted November 22, 2001     

Intrathoracic sources of caudal mediastinal lymph node efferent lymph in sheep

We investigated the intrathoracic contributions to the caudal mediastinal lymph node (CMN) efferent lymph in 12 anesthetized sheep after removing possible systemic contributions from below the diaphragm. We interrupted various pathways that may send lymph to the CMN (chest wall, esophagus, lung). Because the experiment is destructive, we did the resections in various combinations and waited 1 h between steps. Base-line CMN efferent lymph flow averaged 0.90 +/- 0.52 g/15 min (mean +/- SD). Cutting the pulmonary ligaments bilaterally caused lymph flow to decrease by an average of 58%. In five sheep, cauterizing around the lung hila reduced lymph flow by 16% of base line, cauterizing along the esophagus reduced lymph flow by 18% of base line, and cauterizing along the chest wall increased lymph flow by 6% of base line. After complete isolation of the node, except for the bronchoesophageal artery, dorsal mediastinal vein, and CMN efferent duct, 14% of base-line flow remained. The lymph-to-plasma total protein concentration ratios did not change significantly with any procedure. Under the conditions of our experiments, approximately 74% of base-line intrathoracic CMN efferent flow comes from the lung.     

Intracellular topography of glycine-extended pro-gastrin-processing intermediates in human antral mucosa: an electron-microscopic immunocytochemical study

To identify and characterize the subcellular topography of glycine-extended pro-gastrin-processing intermediates (G-Gly) in human antral mucosa, we performed an electron microscopic immunocytochemical study using region-specific antisera generated against the synthetic peptide, Tyr-Gly-Trp-Met-Asp-Phe-Gly (GL7), and C-terminal-specific anti-gastrin antisera. As has been previously reported, G-cells contained both electron-dense and electron-lucent granules, with a range of intermediate forms. Gastrin immunoreactivity was demonstrated in almost all granules of each type, whereas anti-GL7 antisera immunostained chiefly electron-dense granules. The relative ratio of GL7/gastrin granules varied among different cells but was approximately 1:10 on average. Other cytoplasmic organelles were devoid of specific labeling for GL7 or gastrin. As we have assumed that G-Gly serves as the immediate precursor for each molecular form of gastrin, electron-dense granules with high labeling for GL7 are regarded as the principal site for conversion of G-Gly to gastrin. This speculation supports many previous reports that electron-dense granules are immature and that the granules become less electron-dense with maturation.     

Mechanism of Electron Flow through the QB Site in Photosystem II. 4. Reaction Mechanism of Plastoquinone Derivatives at the QB Site in Spinach Photosystem II Membrane Fragments

Interactions of externally added plastoquinone (PQ) derivatives(PQ₀-PQ₃) with the photosystem II (PSII) acceptor side wereinvestigated in PSII membrane fragments prepared from spinachby measuring the photoreduction rates of PQ derivatives at variousPQ concentrations, and the following results were obtained. From the kinetic analysis, all the PQ derivatives (PQ₀-PQ₃)except PQ₃ were shown to accept electrons at two sites (theQ_B site and the PQ site) as in the case of Synechococcus vulcanusPSII particles with benzoquinone derivatives [Satoh et al. (1995)Plant Cell Physiol. 36: 597]. Affinities of PQ derivatives at the Q_B site increased as thelength of the isoprene side chain got longer, while those atthe PQ site were not very much different for all the PQ derivativestested in this study. The inhibitory effect of DCMU was noncompetitive, and, therefore,the affinity of PQ₃ for the PQ site was determined while thatfor the Q_B site could not be estimated presumably due to itsfairly high affinity to the site. Based on the results obtained using PQ derivatives, the mechanismof interaction of an authentic PQ, PQ₉, at the Q_B site is discussed. (Received May 2, 1996; Accepted July 24, 1996)     

Intravascular heavy chain-modification of hyaluronan during endotoxic shock

During inflammation, the covalent linking of the ubiquitous extracellular polysaccharide hyaluronan (HA) with the heavy chains (HC) of the serum protein inter alpha inhibitor (IαI) is exclusively mediated by the enzyme tumor necrosis factor α (TNFα)-stimulated-gene-6 (TSG-6). While significant advances have been made regarding how HC-modified HA (HC-HA) is an important regulator of inflammation, it remains unclear why HC-HA plays a critical role in promoting survival in intraperitoneal lipopolysaccharide (LPS)-induced endotoxemia while exerting only a modest role in the outcomes following intratracheal exposure to LPS. To address this gap, the two models of intraperitoneal LPS-induced endotoxic shock and intratracheal LPS-induced acute lung injury were directly compared in TSG-6 knockout mice and littermate controls. HC-HA formation, endogenous TSG-6 activity, and inflammatory markers were assessed in plasma and lung tissue. TSG-6 knockout mice exhibited accelerated mortality during endotoxic shock. While both intraperitoneal and intratracheal LPS induced HC-HA formation in lung parenchyma, only systemically-induced endotoxemia increased plasma TSG-6 levels and intravascular HC-HA formation. Cultured human lung microvascular endothelial cells secreted TSG-6 in response to both TNFα and IL1β stimulation, indicating that, in addition to inflammatory cells, the endothelium may secrete TSG-6 into circulation during systemic inflammation. These data show for the first time that LPS-induced systemic inflammation is uniquely characterized by significant vascular induction of TSG-6 and HC-HA, which may contribute to improved outcomes of endotoxemia.     

Synthesis and structure based optimization of 2-(4-phenoxybenzoyl)-5-hydroxyindole as a novel CaMKII inhibitor

Based on 2-(4-phenoxybenzoyl)-5-hydroxyindole (2), a novel structural class of CaMKII inhibitors were synthesized and further optimized. The strong acidity of the hydroxyl group and the lipophilic group at the 4 and 6-positions were found to be necessary for strong CaMKII inhibition. Compound 25 was identified as a promising compound with 50-fold more potent inhibitory activity for CaMKII than 2. Compound 25 also showed high selectivity for CaMKII over off-target kinases.     

Net release of dissolved organic matter by the scleractinian coral Acropora pulchra

The net production of dissolved organic matter (DOM) and dissolved combined and free amino acids (DCAA and DFAA, respectively) by the hermatypic coral Acropora pulchra was measured in the submerged condition, and the production rates were normalized to the coral surface area, tissue biomass, and net photosynthetic rates by zooxanthellae. When normalized to the unit surface area, the production rates of dissolved organic carbon and nitrogen (DOC and DON, respectively) were 37 and 4.4 nmol cm^− 2 h^− 1, respectively. Comparing with the photosynthetic rate by zooxanthellae, which was measured by ¹³C-tracer accumulation in the soft tissue of the coral colony, the release rate of DOC corresponded to 5.4% of the daily net photosynthetic production. The tissue biomass of the coral colony was 178 µmol C cm^− 2 and 23 µmol N cm^− 2, indicating that the release of DOC and DON accounted for 0.021% h^− 1 and 0.019% h^− 1 of the tissue C and N, respectively. The C:N ratios of the released DOM (average 8.4) were not significantly different from those of the soft tissue of the coral colonies (average 7.7). While DFAA did almost not accumulate in the incubated seawater, DCAA was considerably released by the coral colonies at the rate of 2.1 nmol cm^− 2 h^− 1 on average. Calculating C and N contents of the hydrolyzable DCAA, it was revealed that about 20% and 50%–60% of the released bulk DOC and DON, respectively, were composed of DCAA.