Intratracheal immunization with pili protein protects against mortality associated with Pseudomonas aeruginosa pneumonia in mice

We examined the protective effect of intratracheal immunization with Pseudomonas aeruginosa pili protein against respiratory infection caused by P. aeruginosa. Mice were immunized intratracheally or subcutaneously with purified pili protein or bovine serum albumin as a control. Intratracheally but not subcutaneously pili protein-immunized mice showed significant improvement of survival after intratracheal challenge with the PAO1 strain. Furthermore, bacterial cell counts in pili protein-immunized murine lungs were significantly decreased compared to controls at 18 h after the challenge. Antipili protein antibody titers in bronchoalveolar lavage fluid of intratracheally pili protein-immunized mice were higher than in bovine serum albumin immunized mice. However, antipili antibody titers were not increased in bronchoalveolar lavage fluid of subcutaneously pili protein-immunized mice, despite the high serum antipili antibody titers. Inoculation of P. aeruginosa induced immediate increases in interleukin-12 and interferon-gamma in bronchoalveolar lavage fluid of pili protein-immunized mice, reflecting an adequate and rapid immune response against P. aeruginosa respiratory tract infection. Our findings suggest that intratracheal pili protein immunization is effective against respiratory tract infection caused by P. aeruginosa in mice.     

A knowledge base for predicting protein localization sites in eukaryotic cells

To automate examination of massive amounts of sequence data for biological function, it is important to computerize interpretation based on empirical knowledge of sequence-function relationships. For this purpose, we have been constructing a knowledge base by organizing various experimental and computational observations as a collection of if-then rules. Here we report an expert system, which utilizes this knowledge base, for predicting localization sites of proteins only from the information on the amino acid sequence and the source origin. We collected data for 401 eukaryotic proteins with known localization sites (subcellular and extracellular) and divided them into training data and testing data. Fourteen localization sites were distinguished for animal cells and 17 for plant cells. When sorting signals were not well characterized experimentally, various sequence features were computationally derived from the training data. It was found that 66% of the training data and 59% of the testing data were correctly predicted by our expert system. This artificial intelligence approach is powerful and flexible enough to be used in genome analyses.     

Availability of Energy in Ester and Ether Derivatives of Glycols by Growing Chicks

Biological availability of 33 esters, 17 ethers and 2 acetals of ethanediol, 1,2-propanediol, 1,3-butanediol and 1,4-butanediol was compared by mini-test with chicks. Chicks can utilize esters of ethanediol, 1,2-propanediol and 1,3-butanediol with acetic acid and fatty acids of carbon chain length from 5 to 12 with more improved palatability than that of free acids, while availability of esters of these glycols with propionic and butyric acids was low. Esters of 1,4-butanediol and ether derivatives of these glycols was not available, except ethyl ether of di-ethanediol which was partially available. Acetacetal of ethanediol was partially available but n-butyracetal was not.     

Molecular pathogenesis of malignant melanoma: a different perspective from the studies of melanocytic nevus and acral melanoma

The Clark model for melanoma progression emphasizes a series of histopathological changes beginning from benign melanocytic nevus to melanoma via dysplastic nevus. Several models of the genetic basis of melanoma development and progression are based on this Clark’s multi-step model, and predict that the acquisition of a BRAF mutation can be a founder event in melanocytic neoplasia. However, our recent investigations have challenged this view, showing the polyclonality of BRAF mutations in melanocytic nevi. Furthermore, it is suggested that many melanomas, including acral and mucosal melanomas, arise de novo, not from melanocytic nevus. While mutations of the BRAF gene are frequent in melanomas on non-chronic sun damaged skin which are prevalent in Caucasians, acral and mucosal melanomas harbor mutations of the KIT gene as well as the amplifications of cyclin D1 or cyclin-dependent kinase 4 gene. Amplifications of the cyclin D1 gene are detected in normal-looking ‘field melanocytes’, which represent a latent progression phase of acral melanoma that precedes the stage of atypical melanocyte proliferation in the epidermis. Based on these observations, we propose an alternative genetic progression model for melanoma.     

Oxygen concentration-dependent induction of a 140-kDa protein in magnetic bacterium Magnetospirillum magnetotacticum MS-1

Abstract The magnetic bacterium Magnetospirillum magnetotacticum prefers a microaerobic habitat and should be able to sense oxygen. Therefore, the bacterium was cultured under atmospheres containing 0–5% O₂ and analyzed for oxygen-dependent changes in the levels of its protein components by sodium dodecyl sulfate-polyccrylamide gel electrophoresis (SDS-PAGE). The analysis revealed a marked anaerobic induction of a 140-kDa protein, which was suppressed when M. magnetotacticum was switched from microaerobic (<1% O₂) to aerobic (>1% O₂) growth conditions. Although its function remains to be determined, the 140-kDa protein may serve as a useful tool to gain insight into the physiology of the organism.     

Response of isolated rat iris dilator to adrenergic and cholinergic agents and electrical stimulation

Responses of isolated rat iris dilator to some agents and to electrical stimulation were examined. Norepinephrine and epinephrine produced contraction, which was antagonized by 0.03 μM phentolamine. Acetylcholine produced relaxation at low concentrations (1 nM ? 1 μM) as great as 80 % of the resting tone while contraction at high concentrations (≥1 μM). Both responses were suppressed by 0.02 μM atropine and enhanced by 0.03 μM physostigmine. Electrical stimulation at low voltage or low frequency (up to 10 Hz) elicited relaxation while stimulation at high voltage or high frequency (30 Hz) produced contraction. Stimulation with intermediate strength elicited biphasic response. The contraction and relaxation induced by electrical stimulation were abolished by 3 μM phentolamine or by 0.05 μM atropine, respectively. Both phases were abolished by tetrodotoxin (0.3 μM). It is suggested that in the rat the cholinergic relaxation of the dilator may assist the cholinergic contraction of the sphincter (1). The pronounced cholinergic relaxation of nonvascular tissue is to be noted.     

2-Ketogluconate Reductase Inhibition by Oxalate

The active site of α-glucosidase from Mucor javanicus IFO 4570 was investigated by kinetic studies. Competition between maltose and soluble starch, and linearity of Lineweaver-Burk plots for the mixed substrates were observed. The dependence of the apparent maximum velocities agreed with those predicted for a single active site mechanism. These results suggest that the enzyme hydrolyzes maltose and soluble starch at a single active site.     

Studies on the Protection Test of Pertussis Vaccine. Immunization Period and Protectivity

In order to confirm the data reported in the previous papers, variously prepared pertussis vaccines were employed in the present investigation. Pertussis organisms grown either on a solid or in a liquid semisynthetic medium were treated by: (1) heating at 56 C for 30 min, (2) storage in 0.1% formalin at 37 C for 5 days, (3) storage in 0.1% formalin at 25 C for 5 days, and (4) simple addition of sodium ethyl-mercuri thiosalicylate (merthiolate) as a preservative. Freeze-dried vaccines were made from these preparations four and a half months later. Mice were immunized intraperitoneally and challenged intracerebrally with a virulent strain of Bordetella pertussis 10 or 17 days later. The data were analyzed statistically assuming the probit corresponding to the percentage of survivors at any dose be a linear function of the logarithms of the dose. The 50% effective doses (ED₅₀) and slopes of each vaccine were found to be uniform. More accurate estimates of ED₅₀ were obtained by employing a pooled slope in each experiment. From these ED₅₀ values, the relative potency was estimated by comparing the value of a vaccine to that of a dried merthiolate-vaccine. For vaccines derived from solid cultures, with an immunization period of 17 days, the relative potency of the vaccine heated at 56 C was 0.63 (95% fiducial limits=0.52 to 0.76); the value for the formalinized vaccine at 37 C was 0.40 (0.30 to 0.53) and one at 25 C was 0.51 (0.34 to 0.77). Vaccines derived from liquid cultures showed a relative potency of 20 to 50% less than that of corresponding vaccine derived from a solid culture. The potency obtained for the 17 day immunization period was usually higher than that for the 10 day period. Using the overall-pooled slope, an experimental design which will be appropriate for statistical analysis is discussed.     

Human platelet aggregation induced by prostaglandin endodisulfide

The effect of human platelet functions of 9,11-dithio analogues of prostaglandin endoperoxide was investigated. Methyl (5z,9α,11α,13e,15S)-9,11-epidithio-15-hydroxyprosta- 5,13-dienoate induced platelet aggregation, while the 9β,11β-epimer was inactive. The platelet aggregation caused by the 9α,11α-dithio analogue was associated with serotonin release from platelets, and was inhibited by methyl ester of prostaglandin I₂ (prostacyclin) but not by indomethacin.