Characteristics of cytotoxic cells induced by Toxoplasma lysate antigen in mouse spleen.

Spleen cells from Toxoplasma lysate antigen (TLA)-sensitized BALB/c mice showed the strong cytotoxic activity against both natural killer (NK)-sensitive cells (YAC-1 and RL male-1) and NK-insensitive cells (P-815), when incubated with TLA or recombinant human IL-2 (rhIL-2). The increment of TLA concentration in culture medium increased the cytotoxic activity. Treatment of effector cells; spleen cells from TLA-sensitized mice incubated with TLA, with anti-asialo GM1 or anti-Thy-1 plus complement inhibited the cytotoxic activity of effector cells, whereas treatment with anti-mouse Lyt-2.2 serum plus complement had no effect on the cytotoxic activity. Treatment of spleen cells from TLA-sensitized mice with anti-asialo GM1 and/or anti-Thy-1 plus complement inhibited cytotoxic activities of effector cells. These results suggested that spleen cells sensitized with TLA both in vivo and in vitro were asialo GM1 positive and Thy-1 positive, and the majority of cytotoxic cells induced by TLA were similar to lymphokine-activated killer (LAK) cells induced by IL-2.     

Involvement of a host erythrocyte sialic acid content in Babesia bovis infection

Host sialic acid (SA) has recently been suggested to play an important role in erythrocyte (RBC) infection by Babesia spp. The present study attempted to further determine the specific type of SAs important in the RBC invasion. Bovine RBC was found to bear abundant alpha2-3-linked SA residues but not alpha2-6-linked SA in nature, confirmed by flow cytometric analysis of the neuraminidase (Nm)-treated RBCs. Lectin-blot analyses revealed the removal of alpha2-3-linked SAs from the 97-, 33-, and 31-kDa bands by the Nm treatment. Addition of the Nm-treated RBCs into an in vitro culture of B. bovis resulted in a decreased population of the parasitized RBCs. The thin smear samples from the cultures were then observed under a confocal laser scanning microscope after staining with the alpha2-3-linked SA-specific lectin: a selective invasion of B. bovis was found only in the intact RBCs bearing the SAs, but not in the desialylated RBCs. Furthermore, a significant reduction of the parasitized RBCs was also observed in the culture supplemented with exogenous 3'-sialyllactose containing the alpha2-3-linked SAs. However, the complete inhibition of parasite proliferation was not achieved in the culture. These findings indicate that while the alpha2-3-linked SA-dependent pathway is needed for highly efficient invasion of host RBCs by B. bovis, there might also be other potential alternative pathways.     

Antimicrobial activity of some N-(arylalkyl)maleimides

A series of N-(arylalkyl)maleimides was prepared for the reaction of maleic anhydride and N-(arylalkyl) amines, and their antimicrobial activities were examined. All compounds exhibited antibacterial activity against gram-positive bacteria such as Bacillus subtilis and Staphylococcus aureus. Almost all compounds exhibited antibacterial activity against the gram-negative bacterium, Escherichia coli, but were inactive against Pseudomonas aeruginosa. Activities against gram-positive bacteria were independent of the nature of the substituent on the benzene ring or the length of alkyl group, but that against gram-negative bacteria was influenced by these parameters. All N-(arylalkyl)maleimides showed activity against yeasts and mycelial fungi.     

In vitro characterization of the inhibitory effects of ketoconazole on metabolic activities of cytochrome P-450 in canine hepatic microsomes

OBJECTIVE: To evaluate the inhibitory potency of ketoconazole (KTZ) on the metabolic activities of isozymes of cytochrome P-450 (CYP) in dogs. ANIMALS: 4 healthy 1-year-old male Beagles. PROCEDURE: Hepatic microsomes were harvested from 4 dogs after euthanasia. To investigate the effects of KTZ on CYP metabolic activities, 7-ethoxyresorufin, tolbutamide, bufuralol, and midazolam hydrochloride were used as specific substrates for CYP1A1/2, CYP2C21, CYP2D15, and CYP3A12, respectively. The concentrations of metabolites formed by CYP were measured by high-performance liquid chromatography, except for the resorufin concentrations that were measured by a fluorometric method. The reaction velocity-substrate concentration data were analyzed to obtain kinetic variables, including maximum reaction velocity, Michaelis-Menten constant, and inhibitory constant (Ki). RESULTS: KTZ competitively inhibited 7-ethoxyresorufin O-deethylation and midazolam 4-hydroxylation; it noncompetitively inhibited tolbutamide methylhydroxylation. Bufuralol 1'-hydroxylation was inhibited slightly by KTZ. The mean Ki values of KTZ were 10.6+/-6.0, 170+/-2.5, and 0.180+/-0.131 microM for 7-ethoxyresorufin O-deethylation, tolbutamide methylhydroxylation, and midazolam 4-hydroxylation, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: In dogs, KTZ at a therapeutic dose may change the pharmacokinetics of CYP3A12 substrates as a result of inhibition of their biotransformation. Furthermore, no influence of KTZ on the pharmacokinetics of CYP1A1/2, CYP2C21, and CYP2D15 substrates are likely. In clinical practice, adverse drug effects may develop when KTZ is administered concomitantly with a drug that is primarily metabolized by CYP3A12.     

Comparison of macrophage scavenger receptor-A knockout mice with wild type ones in the immune response against repeated infestation with Haemaphysalis longicornis

Using macrophage scavenger receptor-A knockout (SRKO) mice, we examined the role of macrophage class A scavenger receptors (MRS-A) on the immune response and acquisition of host resistance against repeated infestation with Haemaphysalis longicornis. Except for one batch of nymphs that infested one of the SRKO (SR-/-) mice and showed no appreciable reduction in body weight, all the other groups of nymphs manifested significant decrease in body weight. Both SR-/- and wild type (SR+/+) mice showed a sustained increase in anti-tick antibody titers, but SR+/+ mice showed significantly higher titers. The IFN-gamma assayed in SR-/- mouse immune sera was substantially less compared with that in SR+/+ mice. Immune sera from SR-/- and SR+/+ mice recognized the 51 and 44 kDa, and 44 kDa proteins, respectively, of the salivary gland antigen. The difference in the level of anti-tick resistance manifested by both groups of mice may be influenced by less efficient trapping and processing of tick antigens by macrophages in mice lacking for the macrophage scavenger receptors, and consequently affected the cascade of Th1 and Th2 responses. We have thus obtained valuable data that strongly infer the role of MSR-A in enhancing host defense against repeated infestation with H. longicornis.     

Differentiation of Toxocara canis and T. cati eggs by light and scanning electron microscopy

In this study, we investigated the morphological identification of Toxocara canis and T. cati eggs on the basis of light and scanning electron microscopic (SEM) observations. T. canis and T. cati eggs used in this study were recovered from the uteri of respective gravid female worms. Measurement of egg size was not helpful in the differentiation of these species, because approximately 90% of eggs measured were of similar size. Using SEM, we were able to differentiate T. canis eggs from T. cati eggs based on their respective characteristic surface structures. Both species have subspherical eggs with markedly pitted surfaces like those of a golf ball, but the surface pitting in T. canis is more coarse than that in T. cati. In this study, however, these differences were not absolute, as 16% of T. canis and 29% of T. cati eggs showed surface pitting that was uncharacteristic of their species. Of the 16% of T. canis eggs that could not be differentiated by surface structure, 3% had pitting resembling T. cati, and the remaining 13% showed intermediate type surface pitting. Similarly, 5% of T. cati eggs resembled T. canis, and 25% of these were of intermediate type. Light microscopic observation yielded results similar to those of SEM, indicating that light microscopy is also a useful tool for the identification of Toxocara eggs.     

Sulfate--a candidate for the missing essential factor that is required for the formation of protein haze in white wine

Protein haze formation in white wine is dependent on the presence of both wine protein and other unknown wine components, termed factor(s) X. The ability to reconstitute protein haze upon heating artificial model wine solutions (500 mg/L thaumatin, 12% ethanol, 4 g/L tartaric acid) to which candidate components were added was employed to identify factor(s) X. No protein haze was formed in the absence of additives. The individual or combined addition of caffeic acid, caftaric acid, epicatechin, epigallocatechin-O-gallate, gallic acid, or ferulic acid at typical white wine concentrations did not generate protein haze. However, PVPP fining of commercial wines resulted in a reduction in protein haze, suggesting that phenolic compounds may play a modulating role in haze formation. To elucidate the nature of the unknown factor(s) wine was fractionated and fractions were back-added to model wine and tested for their essentiality. Wine fractions were generated by ultrafiltration, reverse-phase chromatography, and mixed-mode anion-exchange and reverse-phase chromatography. The only purified fraction containing the essential component(s) was free of phenolic compounds, and analysis by mass spectrometry identified sulfate anion as the dominant component. Reconstitution with KHSO4 using either commercially available thaumatin or wine proteins confirmed the role of sulfate in wine protein haze formation. The two main wine proteins, thaumatin-like protein and chitinase, differed in their haze response in model wines containing sulfate. Other common wine anions, acetate, chloride, citrate, phosphate, and tartrate, and wine cations, Fe(2+/3+) and Cu(+/2+), when added at typical white wine concentrations were not found to be essential for protein haze formation.     

Growth changes in bighand thornyhead <Emphasis Type="Italic">Sebastolobus macrochir</Emphasis> off the Pacific coast of northern Honshu,Japan

ABSTRACT:     The biomass of bighand thornyhead Sebastolobus macrochir was increased by the high recruitment success of the 1999–2002 year classes off the Pacific coast of northern Honshu, Japan. In this study, the growth of bighand thornyhead was examined over a 9-year period from 1996 to 2004 in this area. The growth of the 1999 year class and the 2000–2002 year classes was reduced at 3 and 2 years old, respectively, while the 1999–2002 year classes were smaller than the 1993–1998 year classes. In 2-, 3- and 4-year-old fish, the relationship between abundance and mean standard length was expressed by negative linear regressions, while fish became smaller when abundance of the year class was larger. Mean bottom temperatures were stable at depths of 350–900 m; variations in water temperature were small in the main distribution area of bighand thornyhead. We discuss the factors affecting the growth of bighand thornyhead via changes in the demersal fish community and feeding habits.     

Modeling and Evaluating the Performance of River Sediment on Immobilizing Arsenic from Hydrothermally Altered Rock in Laboratory Column Experiments with Hydrus-1D

Large volumes of excavated rock are produced as a result of road and railway tunnel construction in Hokkaido, Japan. Due to the geological condition of this region, these rocks have often undergone hydrothermal alterations, causing them to contain elevated amounts of hazardous elements including arsenic (As). Therefore, these excavated rocks are potentially hazardous waste, and proper disposal methods are required. In this article, performance of unsaturated river sediment on immobilizing As from hydrothermally altered rock is evaluated using laboratory column experiments and Hydrus-1D. The results reveal that the river sediment significantly reduces As migration. Arsenic retarded by river sediment was observed in three patterns. The first was an adsorption onto minerals originally contained in the river sediment. The next pattern was a combination of reduction of As generation by oxidation of As bearing-minerals, irreversible adsorption, and adsorption onto newly precipitated Fe oxy-hydroxide/oxide. The last pattern led to a further depletion of As leached from the rock layer due to a shift in the majority of the As generation mechanism from dissolution to oxidation in combination with a low concentration of oxygen in the rock layer. These patterns were satisfactorily evaluated by a Hydrus-1D model with reversible and irreversible adsorptions. The information from this work is effective in designing and establishing a reasonable technique for the disposal of hydrothermally altered rocks.     

Changes of lymphocyte subpopulations and natural killer cells in mice sensitized with Toxoplasma lysate antigen before and after Babesia infection

When 8-week-old BALB/c mice were sensitized with two intramuscular injections of Toxoplasma lysate antigen (TLA) at 2 week interval, the numbers of sIg(+), Thy-1,2(+), Lyt-1,2(+) Lyt-2,2(+), and Asialo GM1(ASGM1)(+) cells in the spleen, liver and peripheral blood increased by 2 to 4 times over those found in unsensitized mice of the same age. When TLA-sensitized and unsensitized mice were infected with Babesia, 4 of 10 (40%) of the TLA-sensitized mice survived infection, while none of the unsensitized control mice lived longer than 14 days after Babesia infection. By contrast, sensitization of nude mice with TLA had no effect on survival, and mice did not live more than 12 days. The number of thymic Thy-1,2(+) cells decreased in TLA-sensitized and unsensitized BALB/c mice by almost 80% within 10 days after infection (AI). During the same time, the numbers of B cells, T cells, and NK cells increased in the spleen, liver and peripheral blood of both sensitized and unsensitized mice. Especially notable were increases in numbers of Lyt-2,2(+) cells in the spleen and blood and increases in numbers of NK cells in the spleen, liver and blood in both TLA-sensitized and unsensitized mice. When spleen cells from TLA-sensitized and unsensitized mice were cultured in the presence or absence of TLA for 6 days, assays for cytotoxicity using NK-insensitive P-815 target cells and NK-sensitive YAC-1 target cells demonstrated higher rates of cytotoxicity in cultures of TLA-sensitized spleen cells.