A monoclonal antibody to the carbohydrate chain on human hepatocellular carcinoma-associated antigen which suppressed tumor growth in nude mice

Summary There have been few reports stating that monoclonal antibody alone inhibits human solid tumor growth in vivo. The present study demonstrated that monoclonal antibody S1 (IgG2a), which recognized the antigenic determinant of the carbohydrate moiety, showed antibody-dependent cell (or macrophage)-mediated cytotoxicity (ADCC or ADMC) in conjunction with murine splenocytes of both BALB/c and athymic mice. In vivo experiments demonstrated that the antibody S1 clearly prolonged the survival of athymic mice which had been inoculated with a human liver carcinoma cell line. In addition, the antibody S1 significantly suppressed the human hepatoma line transplanted s.c. into nude mice. ¹²⁵I-Labeled monoclonal antibody S1 revealed that the antibody accumulated significantly in the tumor mass. Many mononuclear cells were observed surrounding tumor cells when the antibody was given. This model system might be useful for analyzing the ADCC (or ADMC) mechanism in vivo.     

Degradation of methylmercury by selenium

When methylmercury was incubated in the presence of selenite and reduced glutathione (GSH), the mercury which was extracted into benzene under acidic condition decreased gradually with the elapse of time. This decrease was due to the cleavage of mercury-carbon bond of methylmercury. The reaction did not proceed when selenite or GSH was singly added to the reaction mixture. L-Cysteine, 2-mercaptoethanol and sodium sulfide in place of GSH also were effective for decomposition of methylmercury in combination with selenite, but oxidized glutathione (GSSG) and L-cystine were not. This suggests that reduction of selenite is needed for the degradation of methylmercury. Thus, the effect of reduced metabolites of selenite produced by GSH was investigated. Glutathione selenotrisulfide (GSSeSG) requierd GSH for the degradation of methylmercury, whereas H₂Se possessed a strong activity even in the absence of GSH. This may indicate that H₂Se is involved directly in the conversion of methylmercury to inorganic mercury. This phenomenon found in $\text{in vitro}$ experiments is discussed in relation to the biotransformation of methylmercury.     

Determination of green tea catechins in human plasma using liquid chromatography-electrospray ionization mass spectrometry

A method for the sensitive and specific determination of eight green tea catechins, consisting of catechin (C), epicatechin (EC), gallocatechin (GC), epigallocatechin (EGC), catechin-3-gallate (CG), epicatechin-3-gallate (ECG), gallocatechin-3-gallate (GCG) and epigallocatechin-3-gallate (EGCG), in human plasma was established. For optimization of conditions for LC-ESIMS, the separation of the eight catechins was achieved chromatographically using Inertsil ODS-2 column combined with a gradient elution system of 0.1M aqueous acetic acid and 0.1M acetic acid in acetonitrile. Detection using a mass spectrometer was performed with selected ion monitoring at m/z=289 for E and EC, 305 for GC and EGC, 441 for CG and ECG, and 457 for GCG and EGCG under negative ESI. A preparative procedure, consisting of the addition of perchloric acid and acetonitrile to the plasma for deproteinizing and the subsequent addition of potassium carbonate solution to remove excess acid, was developed. In six different plasma with the eight catechins spiked at two different concentrations, the average recoveries were in the range between 72.7 and 84.1%, which resulted from the matrix effect and preparative loss, with coefficients of variance being 8.2-19.8% among individuals. The levels of the catechins in prepared plasma solutions that were kept at 5 degrees C within 24h were stable, which allows us to simply analyze many prepared plasma solutions using an autosampler overnight. When using this method to analyze the eight catechins in human plasma after oral ingestion of a commercial green tea beverage, we detected all the catechins absorbed into human blood for the first time. This also suggested that extremely small amounts of the eight catechins orally ingested may be absorbed based on each absorptive property for the catechins. The method should enable pharmacokinetic studies of green tea catechins in humans.     

Distribution of single-channel conductances in cultured rat hippocampal neurons

1. The nonhomogeneous spatial distribution of ionic channels in neurons has been implied from intracellular recordings at somatic and dendritic locations. These reports indicate that Na- and Ca-dependent regenerative currents are distributed differently throughout the neuron. Although a variety of K conductances and a noninactivating Na conductance have been described in intracellular studies, little is known about the spatial distribution of inward and outward currents throughout different regions of the neuron. 2. We recorded from cell-attached patches from cultured hippocampal cells from 1-day-old rats. The cells were cultured for 3-21 days. The spatial distribution of a variety of ionic channels was determined by comparing the conductances from somatic and dendritic membranes. Single-channel currents obtained from cell-attached patches were identified by the time course of ensemble (averaged) responses, voltage dependence, and the effect of channel blocking agents. 3. We consistently observed that only the rapidly inactivating inward current was localized to the soma. The other channel types that we studied, including an inward noninactivating, delayed rectifier and transient A-type currents, were observed in both the somatic and dendritic regions. 4. We suggest that the distribution of ionic conductances that we have observed may be functional in limiting excitability during development of neurons.     

Liquid chromatography-electrospray ionization-tandem mass spectrometry for simultaneous analysis of chlorogenic acids and their metabolites in human plasma

A method using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) was developed for the simultaneous analysis of nine chlorogenic acids (CGAs), three isomers each of caffeoylquinic acids (CQAs), feruloylquinic acids (FQAs) and dicaffeoylquinic acids (dCQAs), and their two metabolites, caffeic acid (CA) and ferulic acid (FA), in human plasma. In simultaneous multiple reaction monitoring (MRM) measurements using ESI-MS/MS with a negative ion mode, a deprotonated molecular ion derived from each of the 11 molecules was used as a precursor ion while three diagnostic product ions characteristic for each were selected for the qualitative analysis. To obtain maximal intensities for all diagnostic product ions, the collision energy was optimized for each one. LC separation was achieved under conditions of a reversed-phase Inertsil ODS-2 column combined with a gradient elution system using 50mM acetic acid with 3% acetonitrile aqueous solution and 50 mM acetic acid with 100% acetonitrile. In the quantitative analysis, one of the three diagnostic product ions for each of the 11 molecules was selected. Application of simultaneous LC-ESI-MS/MS MRM measurements to analyze the 11 standards spiked into blank human plasma indicated that all diagnostic product ions were detected without any interference, and that the sensitivity, linearity and recovery of this method were acceptable. When using this method to analyze those 11 molecules in the plasma after oral ingestion of 250 ml of a drink containing a green coffee bean extract (300 mg CGAs), all 11 molecules were identified and CQAs, FQAs and FA were quantified. CQAs, FQAs and dCQAs in human plasma were detected for the first time. This method should be useful to understand the biological and pharmacological effects of CGAs, such as improvement of human hypertension.     

Promoting R &; D in Photobiological Hydrogen Production Utilizing Mariculture-Raised Cyanobacteria

This review article explores the potential of using mariculture-raised cyanobacteria as solar energy converters of hydrogen (H₂). The exploitation of the sea surface for large-scale renewable energy production and the reasons for selecting the economical, nitrogenase-based systems of cyanobacteria for H₂ production, are described in terms of societal benefits. Reports of cyanobacterial photobiological H₂ production are summarized with respect to specific activity, efficiency of solar energy conversion, and maximum H₂ concentration attainable. The need for further improvements in biological parameters such as low-light saturation properties, sustainability of H₂ production, and so forth, and the means to overcome these difficulties through the identification of promising wild-type strains followed by optimization of the selected strains using genetic engineering are also discussed. Finally, a possible mechanism for the development of economical large-scale mariculture operations in conjunction with international cooperation and social acceptance is outlined.     

High Photobiological Hydrogen Production Activity of a <Emphasis Type="Italic">Nostoc</Emphasis> sp. PCC 7422 Uptake Hydrogenase-Deficient Mutant with High Nitrogenase Activity

We describe a strategy to establish cyanobacterial strains with high levels of H₂ production that involves the identification of promising wild-type strains followed by optimization of the selected strains using genetic engineering. Nostoc sp. PCC 7422 was chosen from 12 other heterocystous strains, because it has the highest nitrogenase activity. We sequenced the uptake hydrogenase (Hup) gene cluster as well as the bidirectional hydrogenase gene cluster from the strain, and constructed a mutant (ΔhupL) by insertional disruption of the hupL gene. The ΔhupL mutant produced H₂ at 100 μmoles mg chlorophyll a ^-1 h^-1, a rate three times that of the wild-type. The ΔhupL cells could accumulate H₂ to about 29% (v/v) accompanied by O₂ evolution in 6 days, under a starting gas phase of Ar + 5% CO₂. The presence of 20% O₂ in the initial gas phase inhibited H₂ accumulation of the ΔhupL cells by less than 20% until day 7.     

Comprehensive quantification of ceramide species in human stratum corneum

One of the key challenges in lipidomics is to quantify lipidomes of interest, as it is practically impossible to collect all authentic materials covering the targeted lipidomes. For diverse ceramides (CER) in human stratum corneum (SC) that play important physicochemical roles in the skin, we developed a novel method for quantification of the overall CER species by improving our previously reported profiling technique using normal-phase liquid chromatog­raphy-electrospray ionization-mass spectrometry (NPLC-ESI-MS). The use of simultaneous selected ion monitoring measurement of as many as 182 kinds of molecular-related ions enables the highly sensitive detection of the overall CER species, as they can be analyzed in only one SC-stripped tape as small as 5 mm × 10 mm. To comprehensively quantify CERs, including those not available as authentic species, we designed a procedure to estimate their levels using relative responses of representative authentic species covering the species targeted, considering the systematic error based on intra-/inter-day analyses. The CER levels obtained by this method were comparable to those determined by conventional thin-layer chromatography (TLC), which guarantees the validity of this method. This method opens lipidomics approaches for CERs in the SC.