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991.
Hiroyuki Kayamuro Yasuo Yoshioka Yasuhiro Abe Kazufumi Katayama Kohei Yamashita Tomoaki Yoshikawa Takachika Hiroi Yuichi Kawai Haruhiko Kamada Shin-ichi Tsunoda Yasuo Tsutsumi 《Biochemical and biophysical research communications》2009,384(3):296-300
The identification of cytokine adjuvants capable of inducing an efficient mucosal immune response against viral pathogens has been long anticipated. Here, we attempted to identify the potential of tumor necrosis factor superfamily (TNFS) cytokines to function as mucosal vaccine adjuvants. Sixteen different TNFS cytokines were used to screen mucosal vaccine adjuvants, after which their immune responses were compared. Among the TNFS cytokines, intranasal immunization with OVA plus APRIL, TL1A, and TNF-α exhibited stronger immune response than those immunized with OVA alone. TL1A induced the strongest immune response and augmented OVA-specific IgG and IgA responses in serum and mucosal compartments, respectively. The OVA-specific immune response of TL1A was characterized by high levels of serum IgG1 and increased production of IL-4 and IL-5 from splenocytes of immunized mice, suggesting that TL1A might induce Th2-type responses. These findings indicate that TL1A has the most potential as a mucosal adjuvant among the TNFS cytokines. 相似文献
992.
Taiji Adachi Yuki Aonuma Shin-ichi Ito Mototsugu Tanaka Masaki Hojo Teruko Takano-Yamamoto Hiroshi Kamioka 《Journal of biomechanics》2009,42(15):2507-2512
Osteocytes embedded in calcified bone matrix have been widely believed to play important roles in mechanosensing to achieve adaptive bone remodeling in a changing mechanical environment. In vitro studies have clarified several types of mechanical stimuli such as hydrostatic pressure, fluid shear stress, and direct deformation influence osteocyte functions. However, osteocyte response to mechanical stimuli in the bone matrix has not been clearly understood. In this study, we observed the osteocyte calcium signaling response to the quantitatively applied deformation in the bone matrix. A novel experimental system was developed to apply deformation to cultured bone tissue with osteocytes on a microscope stage. As a mechanical stimulus to the osteocytes in bone matrix, in-plane shear deformation was applied using a pair of glass microneedles to bone fragments, obtained from 13-day-old embryonic chick calvariae. Deformation of bone matrix and cells was quantitatively evaluated using an image correlation method by applying for differential interference contrast images of the matrix and fluorescent images of immunolabeled osteocytes, together with imaging of the cellular calcium transient using a ratiometric method. As a result, it was confirmed that the newly developed system enables us to apply deformation to bone matrix and osteocytes successfully under the microscope without significant focal plane shift or deviation from the observation view field. The system could be a basis for further development to investigate the mechanosensing mechanism of osteocytes in bone matrix through examination of various types of rapid biochemical signaling responses and intercellular communication induced by matrix deformation. 相似文献
993.
994.
Takema Fukatsu Takahiro Hosokawa Ryuichi Koga Naruo Nikoh Takuya Kato Shin-ichi Hayama Haruo Takefushi Ichirou Tanaka 《Applied and environmental microbiology》2009,75(11):3796-3799
A symbiotic bacterium of the macaque louse, Pedicinus obtusus, was characterized. The symbiont constituted a gammaproteobacterial lineage distinct from the symbionts of anthropoid primate lice, localized in the midgut epithelium and the ovaries and exhibiting AT-biased genes and accelerated molecular evolution. The designation “Candidatus Puchtella pedicinophila” was proposed for it.Sucking lice (Insecta: Phthiraptera: Anoplura), ectoparasitic insects that feed exclusively on the blood of their specific mammalian hosts, are generally associated with endosymbiotic bacteria (2, 13). Recent molecular studies have demonstrated that symbiotic bacteria of sucking lice are of multiple evolutionary origins (1, 8, 10, 16).From primates, three louse genera, Pediculus, Pthirus, and Pedicinus, have been recorded. Three Pediculus and two Pthirus species are known from anthropoid primates, harboring gammaproteobacterial “Candidatus Riesia spp.” in the stomach disc (1, 16). Meanwhile, 14 Pedicinus species, whose symbiotic bacteria have not been characterized, have been recorded from Old World monkeys (3).Samples of Pedicinus obtusus were collected in 2008 at the Jigokudani Monkey Park, Nagano, Japan, where a wild population of the Japanese macaque, Macaca fuscata, is exhibited to the public under careful control. When animals were caught and inspected, samples of P. obtusus were collected from the anesthetized animals under the supervision of a veterinary doctor (permission number 19-26-24, Nagano Prefecture, Japan). Samples of P. obtusus were collected in November 2008 by courtesy of Kiyoyasu Kowaki from a subspecies of the Japanese macaque, M. fuscata yakui, that is endemic in Yakushima Island, Kagoshima, Japan. Each of the insect samples was subjected to DNA extraction, and a 1.5-kb segment of the 16S rRNA gene (5) and a 1.6-kb segment of the groEL gene (7) were amplified by PCR, cloned, and sequenced. Molecular phylogenetic analyses were performed using the programs PAUP 4.0b10 (Sinauer Associates, Sunderland, MA), RAxML version 7.0.0 (17), and MrBayes 3.1.2 (15).The 1,490-bp 16S rRNA gene sequences formed a distinct lineage in the Gammaproteobacteria, exhibiting no phylogenetic affinity to the symbionts of other louse species, including those of human lice and the chimpanzee louse (see Fig. S1 in the supplemental material). The 1,601-bp groEL gene sequences also constituted a gammaproteobacterial lineage, exhibiting no phylogenetic affinity to the symbiont of human lice (Fig. (Fig.1)1) These results indicated that the endosymbiotic bacterium of the Old World monkey louse evolved independently of the endosymbiotic bacteria of the anthropoid primate lice. Considering that the date of divergence of Old World monkeys and anthropoid primates has been inferred as 23 to 31 million years ago, the endosymbiotic evolution in the primate lice must have occurred within this time scale (12).Open in a separate windowFIG. 1.groEL gene sequence-based molecular phylogenetic analysis of the symbiont of P. obtusus in the Gammaproteobacteria. A maximum-likelihood tree inferred from 1,040 unambiguously aligned nucleotide sites is shown. Bayesian and neighbor-joining analyses gave essentially the same results (data not shown). Statistical support values higher than 70% are indicated at the nodes in the order of maximum-likelihood/Bayesian/neighbor-joining values. Asterisks indicate statistical support values lower than 70%. Sequence accession numbers are shown in brackets. AT contents of the sequences are also shown. The sequences from the symbionts of human and monkey lice are highlighted in boldface. P-symbiont, primary symbiont; S-symbiont, secondary symbiont.The molecular evolutionary rates of the symbiont gene sequences were analyzed with a relative rate test using the program RRTree (14). The evolutionary rates of the 16S rRNA and groEL gene sequences in the lineage of the P. obtusus symbiont were significantly higher than those in the lineages of related free-living gammaproteobacteria (see Table S1 in the supplemental material).The AT contents of the 16S rRNA and groEL gene sequences of the P. obtusus symbiont were 53.5% and 64.8%, respectively. These values were equivalent to those of obligate endosymbionts of other insects (>50% for the 16S rRNA gene and >60% for the groEL gene) and were remarkably higher than those of allied free-living gammaproteobacteria (∼45% for the 16S rRNA gene and 45 to 50% for the groEL gene) (Fig. (Fig.1;1; see Fig. S1 in the supplemental material).Obligate endosymbiotic bacteria that cospeciate with their host insects commonly exhibit peculiar genetic traits, including AT-biased nucleotide composition, an accelerated rate of molecular evolution, and significant genome reduction (18). The AT-biased nucleotide composition and the accelerated evolution observed with the P. obtusus symbiont (Fig. (Fig.1;1; see Fig. S1 and Table S1 in the supplemental material) are suggestive of a stable and intimate host-symbiont association over evolutionary time.Fluorescent in situ hybridization targeting 16S rRNA of the symbiont was performed using the Alexa Fluor 555-labeled oligonucleotide probes Al555-ML439 (Al555-5′-ATAATATCTTCTTTCCTACCGA-3′) and Al555-ML1256 (Al555-5′-GCTAATTCTTGCGAATTTGCTT-3′) as described previously (9). In first-, second-, and third-instar nymphs, the symbiont signals were detected in the posterior half of the stomach in the abdomen (Fig. (Fig.2A).2A). In the posterior stomach, the signals exhibited a periodical and striated pattern (Fig. (Fig.2B).2B). Magnified images located the symbiont signals endocellularly in the intestinal wall tissue (Fig. (Fig.2C).2C). In some of the third-instar nymphs, the symbiont signals were found not only in the posterior stomach but also in the ovaries (Fig. (Fig.2A).2A). In adult females, the symbiont signals were restricted to the lateral oviducts (Fig. (Fig.2A),2A), where many bacteriocytes formed a pair of symbiotic organs called ovarial ampullae (Fig. (Fig.2D).2D). The ovarial ampullae were located adjacent to developing oocytes in the ovarioles, where the symbiont was passed to the developing eggs (Fig. (Fig.2E2E).Open in a separate windowFIG. 2.Localization of the symbiont in nymphs and adults of P. obtusus. (A) General localization of the symbiont in nymphal and adult insects. The symbiont signals are seen in the posterior stomach (green arrows) and the ovarial ampullae (yellow arrows). (B) An image of the posterior stomach of a second-instar nymph. Periodical and striped regions are densely populated by the symbiont. (C) An image of the posterior stomach of a third-instar nymph. The symbiont signals are restricted to the gut epithelial cells and not detected in the stomach lumen. Bacteriocytes are intercalated with uninfected cells, forming a striated pattern. (D) An enlarged image of the ovarial ampulla, consisting of a number of uninucleated bacteriocytes. (E) An image of the ovary in an adult female, wherein developing oocytes are found in the ovarioles. Ovarial ampullae (yellow arrows) are located at the anterior tip of the lateral oviducts, where symbiont transmission to oocytes occurs (white arrow). Panel A shows epifluorescent images, while panels B to E are confocal optical sectioning images. Red and blue signals reflect the symbiont 16S rRNA and the host nuclear DNA, respectively. Abbreviations: lu, stomach lumen; oc, oocyte.These results suggested that in third-instar female nymphs of P. obtusus, the symbiont localization drastically changes, from the posterior stomach to the ovarial ampullae (Fig. (Fig.2A).2A). Presumably, the endocellular symbiont escapes the host cells and somehow moves to the female reproductive organ, establishing a new endocellular association and finding a way to the next host generation. Interestingly, such symbiont migrations from a symbiotic organ to the ovaries in third-instar female nymphs have been reported for the human body louse Pediculus humanus (4, 13) and the slender pigeon louse Columbicola columbae (6, 13). Here it is notable that the symbiotic bacteria of P. obtusus, P. humanus, and C. columbae are phylogenetically not related to each other (see Fig. S1 in the supplemental material). The mechanisms underpinning the symbiont migration are currently unknown. Eberle and McLean (4) suggested by a series of experiments that the ovary of the human body louse might emanate a humoral factor that attracts the symbiotic bacteria to induce the migration. Whether or not this hypothesis is true deserves future experimental studies of these louse species and their symbiotic bacteria.On the basis of these results, we propose the designation “Candidatus Puchtella pedicinophila” for the novel endosymbiont lineage. The generic name honors Otto Puchta, who identified the biological role of the human louse symbiont as the provision of B vitamins (11). The specific name indicates association with a Pedicinus monkey louse. Whether the other monkey lice harbor symbiotic bacteria allied to the symbiont of P. obtusus deserves future studies. 相似文献
995.
996.
Karseno Kazuo Harada Takeshi Bamba Susilaningsih Dwi Aparat Mahakhant Tomoaki Yoshikawa Kazumasa Hirata 《Biotechnology letters》2009,31(7):999-1003
During growth of the freshwater cyanobacteria, Oscillatoria sp. BTCC/A0004, and Scytonema sp. TISTR 8208, a pink pigment is released into the growth medium. The pigment from each source had a molecular weight of
approximately 250 kDa and had adsorption maxima at 560 and 620 nm. These results suggest that pink pigment is a phycoerythrin-like
protein. It inhibited the growth of green algae, Chlorella fusca and Chlamydomonas reinhardtii, but not other cyanobacteria or true bacteria. The concentration at which growth inhibition 50% occurred was 0.5, 6 and more
than 10 mg ml−1, respectively. 相似文献
997.
998.
Yui Harada Yasuji Ueda Hiroaki Kinoh Atsushi Komaru Terumi Fuji-Ogawa Aki Furuya Akihiro Iida Mamoru Hasegawa Tomohiko Ichikawa Yoshikazu Yonemitsu 《PloS one》2009,4(8)
Background
Limitations of the clinical efficacy of dendritic cell (DC)-based immunotherapy, as well as difficulties in their industrial production, are largely related to the limited number of autologous DCs from each patient. We here established a possible breakthrough, a simple and cytokine-based culture method to realize a log-scale order of functional murine DCs (>1,000-fold), which cells were used as a model before moving to human studies.Methodology/Principal Findings
Floating cultivation of lineage-negative hematopoietic progenitors from bone marrow in an optimized cytokine cocktail (FLT3-L, IL-3, IL-6, and SCF) led to a stable log-scale proliferation of these cells, and a subsequent differentiation study using IL-4/GM-CSF revealed that 3-weeks of expansion was optimal to produce CD11b+/CD11c+ DC-like cells. The expanded DCs had typical features of conventional myeloid DCs in vitro and in vivo, including identical efficacy as tumor vaccines.Conclusions/Significance
The concept of DC expansion should make a significant contribution to the progress of DC-based immunotherapy. 相似文献999.
John L. Markley David J. Aceti Craig A. Bingman Brian G. Fox Ronnie O. Frederick Shin-ichi Makino Karl W. Nichols George N. Phillips Jr. John G. Primm Sarata C. Sahu Frank C. Vojtik Brian F. Volkman Russell L. Wrobel Zsolt Zolnai 《Journal of structural and functional genomics》2009,10(2):165-179
The Center for Eukaryotic Structural Genomics (CESG) is a “specialized” or “technology development” center supported by the
Protein Structure Initiative (PSI). CESG’s mission is to develop improved methods for the high-throughput solution of structures
from eukaryotic proteins, with a very strong weighting toward human proteins of biomedical relevance. During the first three
years of PSI-2, CESG selected targets representing 601 proteins from Homo sapiens, 33 from mouse, 10 from rat, 139 from Galdieria sulphuraria, 35 from Arabidopsis thaliana, 96 from Cyanidioschyzon merolae, 80 from Plasmodium falciparum, 24 from yeast, and about 25 from other eukaryotes. Notably, 30% of all structures of human proteins solved by the PSI Centers
were determined at CESG. Whereas eukaryotic proteins generally are considered to be much more challenging targets than prokaryotic
proteins, the technology now in place at CESG yields success rates that are comparable to those of the large production centers
that work primarily on prokaryotic proteins. We describe here the technological innovations that underlie CESG’s platforms
for bioinformatics and laboratory information management, target selection, protein production, and structure determination
by X-ray crystallography or NMR spectroscopy.
相似文献
John L. MarkleyEmail: |
1000.
Takeshi Kobayakawa Shin-ichi Yamada Akio Mizuno Yuko Ohara-Nemoto Tomomi T. Baba Takayuki K. Nemoto 《The protein journal》2009,28(1):24-28
A single nucleotide polymorphism (SNP) that causes a missense mutation of highly conserved Gln488 to His of the α isoform
of the 90-kDa heat shock protein (Hsp90α) molecular chaperone is observed in Caucasians. The mutated Hsp90α severely reduced
the growth of yeast cells. To investigate this molecular mechanism, we examined the domain–domain interactions of human Hsp90α
by using bacterial 2-hybrid system. Hsp90α was expressed as a full-length form, N-terminal domain (residues 1–400), or middle
(residues 401–617) plus C-terminal (residues 618–732) domains (MC domain/amino acids 401–732). The Gln488His substitution
in MC domain did not affect the intra-molecular interaction with N-terminal domain, whereas the dimeric interaction-mediated
by the inter-molecular interaction between MC domains was decreased to 32%. Gln488Ala caused a similar change, whereas Gln488Thr,
which exceptionally occurs in mitochondrial Hsp90 paralog, fully maintained the dimeric interaction. Therefore, the SNP causing
Gln488His mutation could abrogate the Hsp90 function due to reduced dimerization. 相似文献