Identification of Tannerella forsythia antigens specifically expressed in patients with periodontal disease

Molecular pathogenesis of Tannerella forsythia, a putative periodontal pathogen, has not yet been adequately elucidated due to limited information on its virulence factors. Here, identification of in vivo expressed antigens of T. forsythia is reported using in vivo-induced antigen technology (IVIAT). Among 13 000 recombinant clones screened, 16 positive clones were identified that reacted reproducibly with sera obtained from patients with periodontal disease. DNA sequences from 12 of these in vivo-induced genes were determined. IVIAT-identified protein antigens of T. forsythia include: BspA, a well-defined virulence factor of T. forsythia; enzymes involved in housekeeping functions (tRNA synthetases, glycine hydroxymethyltransferase, and glucoside glucohydrolase); enzymes implicated in tissue destruction (dipeptidyl peptidase IV); a DNA mismatch repair protein; and putative outer membrane proteins of unknown function. The in vivo gene expression of these IVIAT-identified antigens was confirmed by a quantitative real-time PCR analysis. This is, to the best of the authors' knowledge, the first report using IVIAT in T. forsythia. It is anticipated that detailed analysis of the in vivo-induced genes identified by IVIAT in this study will lead to a better understanding of the molecular mechanisms mediating periodontal infection by T. forsythia.     

Protein architecture chronology deduced from structures of amino acid synthases

Inferring the protein architecture chronology is one of central topics in origin of life study and has been given much attention. Based on an amino acid evolutionary model that late amino acids were bio-synthesized prior to early counterparts, we addressed the issue by examining the structures of amino acid synthases. Despite the limited structural information on amino acid synthases, our deduction revealed that alpha/beta was the oldest protein class, which is in good agreement with the prior fold-usage-based conclusion.     

Changes in components, glycyrrhizin and glycyrrhetinic acid, in raw Glycyrrhiza uralensis Fisch, modify insulin sensitizing and insulinotropic actions

We hypothesized that roasted Glycyrrhizae Radix (Glycyrrhizin Radix Praeparata, GRP) might modify anti-diabetic action due to compositional changes. Then we examined the anti-diabetic effect and mechanism of raw Glycyrrhizae Radix (GR) and GRP extracts and their major respective components, glycyrrhizin and glycyrrhetinic acid. In partial pancreatectomized (Px) diabetic mice, both GR and GRP improved glucose tolerance, but only GRP enhanced glucose-stimulated insulin secretion as much as exendin-4. Both GR and GRP extracts enhanced insulin-stimulated glucose uptake through peroxisome proliferation-activated receptor (PPAR)-gamma activation in 3T3-L1 adipocytes. Consistently with the results of the mice study, only GRP and glycyrrhetinic acid enhanced glucose-stimulated insulin secretion in isolated islets. In addition, they induced mRNA levels of insulin receptor substrate-2, pancreas duodenum homeobox-1, and glucokinase in the islets, which contributed to improving beta-cell viability. In conclusion, GRP extract containing glycyrrhetinic acid improved glucose tolerance better than GR extract by enhancing insulinotropic action. Thus, GRP had better anti-diabetic action than GR.     

Egg yolk soluble protein stimulates the proliferation and differentiation of osteoblastic MC3T3-E1 cells

We determined the effects of yolk water-soluble protein (YSP) on bone formation in pre-osteoblastic MC3T3-E1 cells. YSP (50-5,000 microg/ml) increased cell proliferation and collagen content. Alkaline phosphatase (ALP) activity was also increased by YSP treatment. After enhancement of ALP activity, significant augmentation of calcification was observed. These results suggest that YSP is a promising agent for the prevention and treatment of bone loss.     

Microbial response to salinity change in Lake Chaka, a hypersaline lake on Tibetan plateau

Previous investigations of the salinity effects on the microbial community composition have largely been limited to dynamic estuaries and coastal solar salterns. In this study, the effects of salinity and mineralogy on microbial community composition was studied by using a 900-cm sediment core collected from a stable, inland hypersaline lake, Lake Chaka, on the Tibetan Plateau, north-western China. This core, spanning a time of 17,000 years, was unique in that it possessed an entire range of salinity from freshwater clays and silty sands at the bottom to gypsum and glauberite in the middle, to halite at the top. Bacterial and archaeal communities were studied along the length of this core using an integrated approach combining mineralogy and geochemistry, molecular microbiology (16S rRNA gene analysis and quantitative polymerase chain reaction), cultivation and lipid biomarker analyses. Systematic changes in microbial community composition were correlated with the salinity gradient, but not with mineralogy. Bacterial community was dominated by the Firmicutes-related environmental sequences and known species (including sulfate-reducing bacteria) in the freshwater sediments at the bottom, but by halophilic and halotolerant Betaproteobacteria and Bacteroidetes in the hypersaline sediments at the top. Succession of proteobacterial groups along the salinity gradient, typically observed in free-living bacterial communities, was not observed in the sediment-associated community. Among Archaea, the Crenarchaeota were predominant in the bottom freshwater sediments, but the halophilic Halobacteriales of the Euryarchaeota was the most important group in the hypersaline sediments. Multiple isolates were obtained along the whole length of the core, and their salinity tolerance was consistent with the geochemical conditions. Iron-reducing bacteria were isolated in the freshwater sediments, which were capable of reducing structural Fe(III) in the Fe(III)-rich clay minerals predominant in the source sediment. These data have important implications for understanding how microorganisms respond to increased salinity in stable, inland water bodies.     

Life history response of <Emphasis Type="Italic">Daphnia galeata</Emphasis> to heterogeneous conditions within a reservoir as determined in a cross-designed laboratory experiment

Growth and reproduction were studied in the laboratory in a cross-designed experimental set-up in four Daphnia galeata subpopulations collected from different locations (with respect to water characteristics) in a reservoir (epilimnion, metalimnion and hypolimnion in the deepest part of the reservoir near the dam and epilimnion of the upstream part of the reservoir) and in a laboratory clone of the same species. The results of two-way ANOVA revealed significant effects of the two parameters manipulated – source of water used for cultures and Daphnia subpopulation – on the life history characteristics of growth and reproduction. The water from the upstream part of the reservoir was the most favourable culture medium for all characteristics of the Daphnia groups studied (the largest primiparae, clutches and eggs, the shortest postembryonic development time and filtering setae). The poorest performances were recorded in the downstream, epilimnetic and metalimnetic waters. The primiparae in the hypolimnetic water were smaller but had relatively larger clutches of smaller eggs and slightly longer postembryonic development times. The Daphnia subpopulation originating from the hypolimnion had the smallest primiparae, the largest clutches, the smallest eggs and the shortest postembryonic development, whereas the opposite was found in animals from the epilmnion. These differences in ecologically relevant traits were supported by analysis of the quasi-neutral genetic markers that indicated significant site-dependent differences in clonal structure between the subpopulations. There was no consistent trend to higher within-group variance in the life history traits in the genetically heterogeneous subpopulations from the reservoir compared to the laboratory clone.     

Filopodial actin bundles are not necessary for microtubule advance into the peripheral domain of Aplysia neuronal growth cones

Filopodial actin bundles guide microtubule assembly in the growth cone peripheral (P) domain and retrograde actin-network flow simultaneously transports microtubules rearward. Therefore, microtubule-end position is determined by the sum of microtubule assembly and retrograde transport rates. However, how filopodia actually affect microtubule assembly dynamics is unknown. To address this issue we quantitatively assessed microtubule and actin dynamics before and after selective removal of filopodia. Filopodium removal had surprisingly little effect on retrograde actin-flow rates or underlying network structures, but resulted in an approximate doubling of peripheral microtubule density and deeper penetration of microtubules into the P domain. The latter stemmed from less efficient coupling of microtubules to remaining actin networks and not from a change in microtubule polymer dynamics. Loss of filopodia also resulted in increased lateral microtubule movements and a more randomized microtubule distribution in the P domain. In summary, filopodia do not seem to be formally required for microtubule advance; however, their presence ensures radial distribution of microtubules in the P domain and facilitates microtubule transport by retrograde flow. The resulting dynamic steady state has interesting implications for rapid microtubule-positioning responses in the P domain.     

Phase 0 clinical trials in cancer drug development: from FDA guidance to clinical practice

The Food and Drug Administration (FDA) recently introduced the Exploratory Investigational New Drug Guidance to expedite the clinical evaluation of new therapeutic and imaging agents. Early clinical studies performed under the auspices of this guidance, so-called "Phase 0" trials, have been initiated at the National Cancer Institute to integrate qualified pharmacodynamic biomarker assays into first-in-human cancer clinical trials of molecularly targeted agents. The goal of this integration is to perform molecular proof-of-concept investigations at the earliest stage of cancer drug development. Phase 0 trials do not offer any possibility of patient benefit; instead, intensive, real-time pharmacodynamic and pharmacokinetic analyses of patient tumor samples and/or surrogate tissues are performed to inform subsequent trials. Phase 0 studies do not replace formal Phase I drug safety testing and require a substantial investment of resources in assay development early on; however, they offer the promise of more rational selection of agents for further, large-scale development as well as the molecular identification of potential therapeutic failures early in the development process.