Induction of Manganese Superoxide Dismutase by Cytokines and Lipopolysaccharide in Cultured Mouse Astrocytes

Abstract: To determine whether cytokines or lipopolysaccharide (LPS) are involved in the induction of superoxide dismutase (SOD) in the nervous system, we examined the effects of these substances on the levels of SOD in cultured mouse astrocytes. Treatment of astrocytes with 10² to 10⁴ U/ml tumor necrosis factor-α for 3 days increased the levels of Mn SOD in a dose- and time-dependent manner to as much as six times the level under nontreated conditions. Treatment with 1.0 µg/ml LPS for 3 days elicited a fourfold increase in levels of Mn SOD, and the effect of LPS was also dose dependent. Furthermore, Mn SOD in astrocytes was induced by a 3-day exposure to interleukin-1α at concentrations of 10² or 10³ U/ml. However, these stimuli had no effect on levels of copper-zinc SOD (Cu/Zn SOD) in astrocytes. By contrast, interferon-γ did not change the levels of either Mn or Cu/Zn SOD in the cells. The results indicate that the selective induction of Mn SOD by cytokines and LPS, which has been observed in nonnervous tissues, may also occur in nervous tissues. The induction of Mn SOD may represent a mechanism for protection of cells from oxidative stress.     

Occurrence of thermolabile and regulatory NAD-linked glutamate dehydrogenase in Pseudomonas fluorescens.

NAD-linked glutamate dehydrogeanse [EC 1.4.1.2] was detected together with NADP-linked glutamate dehydrogenase [EC 1.4.1.4] and aspartase [EC 4.3.1.1] in Pseudomonas fluorescens cells. The three enzymes were distinctly separated by DEAE-Sephadex column chromatography. The NAD-linked enzyme was extremely thermolabile and was rapidly inactivated even at temperatures as low as 35--40 degrees C. The combined addition of NAD+ and glutamate, however, effectively stabilized the enzyme. The glutamate saturation profile of the NAD-linked enzyme exhibited cooperativity with a Hill coefficient (n) of 1.4. ATP inhibited the enzyme in an allosteric manner, increasing the n value to 2.2. These results suggest a novel type of metabolic regulation shared by the three enzymes in the biosynthesis and catabolism of amino acids.     

Purification and characterization of a new, ubiquitously distributed class of microtubule-associated protein with molecular mass 250 kDa.

A heat-stable microtubule-associated protein (MAP) with relative molecular mass 250 000, termed 250-kDa MAP, was purified from bovine adrenal cortex. It is classified as a MAP subspecies distinct from MAP1, MAP2, tau, and MAP4, as judged from its electrophoretic mobility, heat stability and immunoreactivity. Purified 250-kDa MAP was able to bind to taxol-stabilized microtubules, although it lacked the ability to polymerize purified tubulin into microtubules. Western-blot analysis showed that this MAP was expressed ubiquitously in mammalian tissues. Immunofluorescence microscopy revealed that polyclonal antibodies raised against 250-kDa MAP stained many punctate structures in the cytoplasm of cultured cells. Blurry cytosolic staining was also observed. Judging from the result of nocodazole treatment, the punctate structures were associated with the microtubule network throughout the cytoplasm, while cytosolic 250-kDa MAP colocalized with free tubulin. Under electron microscopy, 250-kDa MAP has the appearance of a hollow sphere of about 12 nm diameter.     

Oral and Anal Vaccination Confers Full Protection against Enteric Redmouth Disease (ERM) in Rainbow Trout

The effect of oral vaccines against bacterial fish diseases has been a topic for debate for decades. Recently both M-like cells and dendritic cells have been discovered in the intestine of rainbow trout. It is therefore likely that antigens reaching the intestine can be taken up and thereby induce immunity in orally vaccinated fish. The objective of this project was to investigate whether oral and anal vaccination of rainbow trout induces protection against an experimental waterborne infection with the pathogenic enterobacteria Yersinia ruckeri O1 biotype 1 the causative agent of enteric redmouth disease (ERM). Rainbow trout were orally vaccinated with AquaVac ERM Oral (MERCK Animal Health) or an experimental vaccine bacterin of Y. ruckeri O1. Both vaccines were tested with and without a booster vaccination four months post the primary vaccination. Furthermore, two groups of positive controls were included, one group receiving the experimental oral vaccine in a 50 times higher dose, and the other group receiving a single dose administered anally in order to bypass the stomach. Each group was bath challenged with 6.3×10⁸ CFU/ml Y. ruckeri, six months post the primary vaccination. The challenge induced significant mortality in all the infected groups except for the groups vaccinated anally with a single dose or orally with the high dose of bacterin. Both of these groups had 100% survival. These results show that a low dose of Y. ruckeri bacterin induces full protection when the bacterin is administered anally. Oral vaccination also induces full protection, however, at a dose 50 times higher than if the fish were to be vaccinated anally. This indicates that much of the orally fed antigen is digested in the stomach before it reaches the second segment of the intestine where it can be taken up as immunogenic antigens and presented to lymphocytes.     

Neurotrophic Activity of Organosulfur Compounds Having a Thioallyl Group on Cultured Rat Hippocampal Neurons

Several organosulfur compounds found in garlic extract promoted the survival of rat hippocampal neurons in vitro. From the analysis of structure-activity relationship, thioallyl group in these compounds is essential for the manifestation of neurotrophic activity. S-Allyl-L-cysteine (SAC), one of the organosulfur compounds having thioallyl group in garlic extract, also promoted the axonal branching of cultured neurons. These results suggest that thioallyl compounds make a unique group of neurotrophic factors.     

Genetic variation of pantropical Terminalia catappa plants with sea‐drifted seeds in the Bonin Islands: suggestions for transplantation guidelines

The Bonin Islands are endowed with endemic species. However, these species are at risk of extinction because of the exuberance of invasive alien plants. Therefore, native plant species should be revegetated after eradicating alien plants. We investigated the genetic variation of Terminalia catappa populations in the Bonin Islands by using nuclear (n) microsatellites (simple sequence repeats [SSRs]) and chloroplast (cp) DNA. No significant differences were observed in the genetic diversity of nSSRs among 22 populations. However, recent bottlenecks were detected in three populations on the Chichijima Island group. nSSR variation and cpDNA haplotypes suggested the presence of two genetically distinct groups in the Mukojima and Chichijima Island groups and the Hahajima Island group. A similar genetic structure was observed in plants and animals in the Bonin Islands. Populations on the three islands, which were separated from other islands in each island group when the water depth was 50‐m lower than the present level, were dominated by unique nSSRs clusters, suggesting that historical changes in island connections during the Pleistocene era affected genetic substructuring. These results suggested that different factors contributed to the genetic structure of T. catappa on different geographic scales. At the whole‐island level, the genetic structure was determined by long‐distance seed dispersal by ocean currents. At the island‐group level, the genetic structure was determined by historical changes in island connections caused by changes in the sea level due to glacial–interglacial transition. These findings would help in establishing transplantation zone borders for revegetating T. catappa on the Bonin Islands.     

Application of an enzyme-linked immunosorbent assay for screening of T-2 toxin-producing Fusarium spp.

Culture filtrates of Fusarium species were subjected without clean-up procedures to an indirect competitive enzyme-linked immunosorbent assay with anti-T-2 toxin monoclonal antibody. Fusarium sporotrichioides, F. poae, F. tricinctum, and F. culmorum strains were positive for T-2 toxin, with a minimum detection limit of 5 pg per assay (100 pg/ml of culture filtrate), and the assay data correlated well with the gas-liquid chromatographic data.     

Nuclear and chloroplast DNA phylogeography reveals Pleistocene divergence and subsequent secondary contact of two genetic lineages of the tropical rainforest tree species Shorea leprosula (Dipterocarpaceae) in South‐East Asia

Tropical rainforests in South‐East Asia have been affected by climatic fluctuations during past glacial eras. To examine how the accompanying changes in land areas and temperature have affected the genetic properties of rainforest trees in the region, we investigated the phylogeographic patterns of a widespread dipterocarp species, Shorea leprosula. Two types of DNA markers were used: expressed sequence tag‐based simple sequence repeats and chloroplast DNA (cpDNA) sequence variations. Both sets of markers revealed clear genetic differentiation between populations in Borneo and those in the Malay Peninsula and Sumatra (Malay/Sumatra). However, in the south‐western part of Borneo, genetic admixture of the lineages was observed in the two marker types. Coalescent simulation based on cpDNA sequence variation suggested that the two lineages arose 0.28–0.09 million years before present and that following their divergence migration from Malay/Sumatra to Borneo strongly exceeded migration in the opposite direction. We conclude that the genetic structure of S. leprosula was largely formed during the middle Pleistocene and was subsequently modified by eastward migration across the subaerially exposed Sunda Shelf.