Antibody conjugates mimic specific B cell presentation of antigen: relationship between T and B cell specificity

We developed antibody conjugates by covalently coupling antibodies against mouse mu-chain and monoclonal antibodies against nominal antigen, myoglobin, as a tool for antigen presentation and as a model of specific presentation of antigen by antigen-specific B cells and T-B interaction. In the presence of the antibody conjugates, myoglobin-specific Iad-restricted cloned T cells proliferated at 1000-fold lower concentration of myoglobin than the stimulatory concentration without the conjugates. This enhanced presentation was observed only when Iad spleen cells were 1000 R-irradiated but not 3300 R-irradiated, consistent with B cell presentation. The simple mixture of each component of the conjugates had no enhancement effects. The conjugates per se had no mitogenic effects on either splenic B cells or the cloned T cells at concentrations employed for antigen presentation. The conjugates reduced the number of antigen-presenting cells required for the maximal response but did not change the kinetics of response. The enhanced presentation by the conjugates required a genetically restricted interaction with B cells. Antigen specificity of the enhanced presentation was confirmed by using various T cell clones or lines with different antigen specificities and different conjugates constructed with monoclonal antibodies of known epitope specificity. The enhanced presentation was significantly inhibited by competition with exogenous mouse IgM or anti-mouse mu-chain but was not significantly inhibited by monoclonal antibodies against Fc receptor. Thus, conjugate-coated B cells serve as models for myoglobin-specific B cells in that they can take up specific antigens at extremely low concentration and can present the antigen to specific T cells. This model system can be applied to any antigen and any species without the need to develop antigen-specific B cell clones, which is not yet possible for most antigens and species of experimental animals. This system allowed us to investigate the relationship between T cell epitope and B cell epitope when these cells interact with each other in an antigen-specific and Ia-restricted manner. Experiments using antibody conjugates of different monoclonal antibodies against myoglobin and various myoglobin-specific cloned T cells of known antigen specificity revealed that there are some particular combinations in which much more limited enhancement of antigen presentation is observed.(ABSTRACT TRUNCATED AT 400 WORDS)     

Three-dimensional measurement of the occlusal surfaces of lower first molars of Australian aboriginals

The occlusal surfaces of lower first molars of Australian Aboriginals were measured in three dimensions with the aid of Moiré contourography. Molar cuspal heights in this population were higher than in Japanese (Mongoloid) but lower than in Dutch (Caucasoid) populations. Intercuspal distances were considerably larger than those in the two other populations. Populational differences in occlusal features may influence both craniofacial structures and jaw movements in the three populations. Low correlations between the cuspal heights and the intercuspal distances in the other two populations were also found in this population, indicating that human molar cuspal height is independent of the transverse size of the crown. Mean values for the height of the three principal cusps in lower first molars were less than those in upper ones. However, the height of the hypoconid, which was the highest in the lower molar cusps, showed almost the same mean value as the height of the upper three principal cusps, indicating that the height of the main functional cusp, in both upper and lower first molars, was almost the same.     

Intracellular Ca2+-calmodulin system involved in the palytoxin-induced K+ release from rabbit erythrocytes

Palytoxin (PTX) caused K+ release from rabbit erythrocytes which was dependent on the concentrations of extracellular Ca2+ and PTX. In a Ca2+-free solution, PTX still caused a slow K+ release. An intracellular Ca2+ antagonist, TMB-8, an intracellular Ca2+ chelator, quin 2, and calmodulin inhibitors, prenylamine, W-7 and W-5, inhibited the PTX-induced K+ release in a Ca2+-free solution. These results suggest that the PTX-induced K+ release is dependent on the process including intracellular Ca2+ and calmodulin.     

Purification and characterization of NADP+-linked isocitrate dehydrogenase from an alkalophilic Bacillus

We have succeeded in purifying to homogeneity a very labile NADP+-linked isocitrate dehydrogenase (isocitrate: NADP+ oxidoreductase (decarboxylating), EC 1.1.1.42) from a strain of alkalophilic Bacillus, by a simple method, with an overall yield over 76% of the original activity. The molecular weight on Sephadex G-200 was around 90,000; and that by electrophoresis on SDS-polyacrylamide gels was about 44,000. The sedimentation coefficient (s020,w) and isoelectric point of the enzyme were determined to be 3.22 S and pH 4.7, respectively. The enzyme required Mn2+ for the reaction and for stability. The optimum pH for the reaction was in the range 7.8-8.4 at 30 degrees C; the optimum temperature at pH 8.0 was 75 degrees C; the activation energy of the reaction was 6.2 kcal/mol. The Km values for threo-Ds-isocitrate, DL-isocitrate, and NADP+ were 5.4 microM, 9.9 microM, and 7.3 microM, respectively. This enzyme was inhibited by NADPH, glyceraldehyde 3-phosphate, 3-phosphoglycerate, phosphoenol pyruvate, cis-aconitate, alpha-ketoglutarate, and oxaloacetate. In addition, it was subject to a concerted inhibition by a combination of glyoxylate and oxaloacetate, and also to a cumulative inhibition by nucleoside triphosphates.     

Functional expression of the genes of Escherichia coli in gram-positive Corynebacterium glutamicum

Summary Hybrid plasmids were constructed by combining in vitro the Escherichia coli plasmid pGA22, which carries the genes determining resistance to kanamycin, tetracycline, chloramphenicol and ampicillin, with the cryptic plasmids, pCG1 and pCG2, of Corynebacterium glutamicum. The hybrid plasmids were introduced into C. glutamicum and E. coli and replicated in both hosts. They expressed all the E. coli resistance phenotypes except ampicillin resistance in C. glutamicum. The levels of antibiotic inactivating enzymes encoded on these plasmids were about four to ten times lower in C. glutamicum than in E. coli. Despite the lack of expression of ampicillin resistance, -lactamase activity was detected in C. glutamicum carrying hybrid plasmids.     

Simultaneous determination of uric and ascorbic acids in human serum by reversed-phase high-performance liquid chromatography with electrochemical detection

A rapid, easy, and accurate method for the determination of uric acid and ascorbic acid in human serum by reversed-phase high-performance liquid chromatography with electrochemical detection has been developed. Human serum (0.5 ml) was mixed with 1.5 ml of an aqueous solution containing 2.0% metaphosphoric acid and the mixture was centrifuged at 3000g for 30 min. The supernatant was passed through a membrane filter to remove the particulate matter. Ten microliters of the filtrate was injected into the chromatographic system employed in this study. Complete separation of uric acid and ascorbic acid was achieved in about 2 min. The assay limit for quantitation was about 10 pg for uric acid and ascorbic acid under the present chromatographic conditions. The analytical recoveries of uric acid and ascorbic acid in human serum samples were found to be almost 100%.     

Effects of gall volume on survival and fecundity of gall-making aphidsAdelges japonicus (Homoptera: Adelgidae)

The number of larvae reaching maturity within the gall of Adelges japonicus was positively related to gall volume, and the relationship between the number of mature larvae and gall volume did not change with different densities of colonized larvae. The population changes in the bud galls of A. japonicus were surveyed by collecting the galls, which did not suffer predation or parasitism within the galls, from young artificial plantations of Picea jezoensis over two years. In the year when the density of colonized larvae was high, they suffered a 42% mortality within the galls, whereas mortality was nearly zero in the low density year. The numbers of larvae per gall were positively correlated with gall volume. The regression lines of the number of colonized larvae on gall volume did not differ significantly in the regression coefficients between the two years, whereas the intercept was significantly higher in the year when the density of colonized larvae was higher. However, different within-gall mortality nullified this difference, and the regression lines of the number of mature larvae on gall volume had no significant difference both in the regression coefficients and the intercepts. This suggests that the number of mature larvae per gall was limited by available resources within the gall which were positively related to gall volume. In 25% of the galls in which mature larvae inhabited, the space within the galls were completely filled by the larvae, indicating that space was one of the limiting resources. Gall volume also affected the number of adults that emerged from the gall and the potential number of their progeny.     

Presumptive Primordial Germ Cells (pPGCs) and PGCs in Tadpoles from UV-irradiated embryos of Xenopus

In order to determine whether or not tadpoles that once lacked primordial germ cells (PGCs) in the genital ridges and dorsal mesentery as a result of ultraviolet (UV) irradiation subsequently contained germ cells at more advanced stages of larval development, the numbers of presumptive PGCs or PGCs were carefully examined in Xenopus tadpoles at Nieuwkoop and Faber's stage 35/36–52 that developed normally from UV-irradiated eggs.
No late-appearing germ cells were observed in almost all the UV-irradiated tadpoles examined at stages 49–52. This same population had completely lacked PGCs at about stage 46. Moreover, presumptive PGCs (pPGCs) or cells with granular cytoplasm that reacted with a monoclonal antibody specific for the germ plasm of cleaving Xenopus eggs stayed in the central part of the endoderm cell mass in the irradiated tadpoles at stage 35/36, when the majority of those cells were located in the dorsal part of the endoderm in unirradiated controls. Furthermore, in the irradiated embryos pPGCs were demonstrated to decrease in number with development and eventually to disappear in tadpoles at about stage 40. The results strongly suggest that UV irradiation under the conditions used here totally eliminated germline cells from the irradiated animals.     

Satellite tracking of the migration of the red-crowned crane Grus japonensis

Autumn migration routes of red-crowned cranes, Grus japonensis, from two continental east Asian sites were documented in detail by satellite tracking. Two routes were identified: a 2200km western route from Russias Khingansky Nature Reserve to coastal Jiangsu Province, China; and a 900km eastern route from Lake Khanka (Russia) to the Korean Peninsula and the Demilitarized Zone. The most important rest-sites were identified as Panjin Marsh (China), coastal mudflats south-east of Tangshan City (China), the Yellow River mouth (China), Tumen River mouth (North Korea/China/Russia), Kumya (North Korea) and Cholwon (Korean DMZ). Movements within the wintering range were also recorded, including complex commuting between sites by individual cranes and patterns of daily movements within sites. These data should prove useful for conservation of the flyway.     

Alkaline detergent enzymes from alkaliphiles: enzymatic properties, genetics, and structures

The cleaning power of detergents seems to have peaked; all detergents contain similar ingredients and are based on similar detergency mechanisms. To improve detergency, modern types of heavy-duty powder detegents and automatic dishwasher detergents usually contain one or more enzymes, such as protease, amylase, cellulase, and lipase. Alkaliphilic Bacillus strains are often good sources of alkaline extracellular enzymes, the properties of which fulfil the essential requirements for enzymes to be used in detergents. We have isolated numbers of alkaliphilic Bacillus that produce such alkaline detergent enzymes, including cellulase (CMCase), protease, α-amylase, and debranching enzymes, and have succeeded in large-scale industrial production of some of these enzymes. Here, we describe the enzymatic properties, genetics, and structures of the detergent enzymes that we have developed. Received: January 22, 1998 / Accepted: February 16, 1998