Accuracy of an optical active-marker system to track the relative motion of rigid bodies

The measurement of relative motion between two moving bones is commonly accomplished for in vitro studies by attaching to each bone a series of either passive or active markers in a fixed orientation to create a rigid body (RB). This work determined the accuracy of motion between two RBs using an Optotrak optical motion capture system with active infrared LEDs. The stationary noise in the system was quantified by recording the apparent change in position with the RBs stationary and found to be 0.04 degrees and 0.03 mm. Incremental 10 degrees rotations and 10-mm translations were made using a more precise tool than the Optotrak. Increasing camera distance decreased the precision or increased the range of values observed for a set motion and increased the error in rotation or bias between the measured and actual rotation. The relative positions of the RBs with respect to the camera-viewing plane had a minimal effect on the kinematics and, therefore, for a given distance in the volume less than or close to the precalibrated camera distance, any motion was similarly reliable. For a typical operating set-up, a 10 degrees rotation showed a bias of 0.05 degrees and a 95% repeatability limit of 0.67 degrees. A 10-mm translation showed a bias of 0.03 mm and a 95% repeatability limit of 0.29 mm. To achieve a high level of accuracy it is important to keep the distance between the cameras and the markers near the distance the cameras are focused to during calibration.     

Identification of parasite DNA in common bile duct stones by PCR and DNA sequencing

We attempted to identify parasite DNA in the biliary stones of humans via PCR and DNA sequencing. Genomic DNA was isolated from each of 15 common bile duct (CBD) stones and 5 gallbladder (GB) stones. The patients who had the CBD stones suffered from cholangitis, and the patients with GB stones showed acute cholecystitis, respectively. The 28S and 18S rDNA genes were amplified successfully from 3 and/or 1 common bile duct stone samples, and then cloned and sequenced. The 28S and 18S rDNA sequences were highly conserved among isolates. Identity of the obtained 28S D1 rDNA with that of Clonorchis sinensis was higher than 97.6%, and identity of the 18S rDNA with that of other Ascarididae was 97.9%. Almost no intra-specific variations were detected in the 28S and 18S rDNA with the exception of a few nucleotide variations, i.e., substitution and deletion. These findings suggest that C. sinensis and Ascaris lumbricoides may be related with the biliary stone formation and development.     

Disruption of diacylglycerol kinase delta (DGKD) associated with seizures in humans and mice

We report a female patient with a de novo balanced translocation, 46,X,t(X;2)(p11.2;q37)dn, who exhibits seizures, capillary abnormality, developmental delay, infantile hypotonia, and obesity. The 2q37 breakpoint observed in association with the seizure phenotype is of particular interest, because it lies near loci implicated in epilepsy in humans and mice. Fluorescence in situ hybridization mapping of the translocation breakpoints showed that no known genes are disrupted at Xp11.2, whereas diacylglycerol kinase delta (DGKD) is disrupted at 2q37. Expression studies in Drosophila and mouse suggest that DGKD is involved in central nervous system development and function. Electroencephalographic assessment of Dgkd mutant mice revealed abnormal epileptic discharges and electrographic seizures in three of six homozygotes. These findings implicate DGKD disruption by the t(X;2)(p11.2;q37)dn in the observed phenotype and support a more general role for DGKD in the etiology of seizures.     

Sequence-based prioritization of nonsynonymous single-nucleotide polymorphisms for the study of disease mutations

The increasing demand for the identification of genetic variation responsible for common diseases has translated into a need for sophisticated methods for effectively prioritizing mutations occurring in disease-associated genetic regions. In this article, we prioritize candidate nonsynonymous single-nucleotide polymorphisms (nsSNPs) through a bioinformatics approach that takes advantages of a set of improved numeric features derived from protein-sequence information and a new statistical learning model called "multiple selection rule voting" (MSRV). The sequence-based features can maximize the scope of applications of our approach, and the MSRV model can capture subtle characteristics of individual mutations. Systematic validation of the approach demonstrates that this approach is capable of prioritizing causal mutations for both simple monogenic diseases and complex polygenic diseases. Further studies of familial Alzheimer diseases and diabetes show that the approach can enrich mutations underlying these polygenic diseases among the top of candidate mutations. Application of this approach to unclassified mutations suggests that there are 10 suspicious mutations likely to cause diseases, and there is strong support for this in the literature.     

Knockout of the gene (<Emphasis Type="Italic">ste15</Emphasis>) encoding a glycosyltransferase and its function in biosynthesis of exopolysaccharide in <Emphasis Type="Italic">Streptomyces</Emphasis> sp. 139

Streptomyces sp. 139 produces a novel exopolysaccharide (EPS) designated Ebosin which has antagonistic activity for IL-1R in vitro and remarkable anti-rheumatic arthritis activity in vivo. We previously identified a ste (Streptomyces eps) gene cluster consisting of 27 ORFs responsible for Ebosin biosynthesis. The gene product of ste15 shows high homology to known glycosyltransferases (GTFs). To elucidate its function in Ebosin biosynthesis, the ste15 gene was knocked out with a double crossover via homologous recombination. Our analysis of monosaccharide composition for EPS-m produced by the mutant strain Streptomyces sp. 139 (ste15 ⁻) showed that glucose was significantly diminished compared to its natural counterpart Ebosin. This derivative of Ebosin lost the antagonistic activity for IL-1R in vitro and its molecular mass was smaller than Ebosin. These results have demonstrated that the ste15 gene codes for a GTF for glucose, which is functionally involved in Ebosin biosynthesis.     

Identification Homologous Recombination Function from Haloarchaea Plasmid pHH205

Homologous recombination (HR) was found to be so frequent in haloarchaea that its significance in evolution and diversity of this clade of life might have been underestimated. However, so far there has been no report on recombination function carried on plasmid. Here we report that a 4.8-kb SnaBI-PvuII digested segment from pHH205 might carry such a function. Four constructed plasmids: pUN, pUN-205, pUM and pUM-205, with pUN and pUN205 containing Nov^R gene, pUM and pUM-205 carrying Mev^R gene, were used to transform Haloferax volcanii DS52 (radA⁻). The results showed that only pUN-205 and pUM-205 containing the 4.8-kb SnaBI-PvuII digested segment from pHH205 were able to shift Nov^R and Mev^R gene into the chromosome of Haloferax volcanii DS52 through HR, whereas those in pUN and pUM could not, which indicated that the segment from pHH205 does contain a recombination function.     

The pathways to tumor suppression via route p38

Besides its well-known functions in inflammation and other stresses, the p38 mitogen-activated protein kinase pathway also negatively regulates cell proliferation and tumorigenesis. Inactivation of the p38 pathway enhances cellular transformation and renders mice prone to tumor development with concurrent disruption of the induction of senescence. Conversely, persistent activation of p38 inhibits tumorigenesis. Mechanistic insights into this additional p38 function are starting to emerge. For example, p38 has been shown to have a crucial role in oncogene-induced senescence, replicative senescence, DNA-damage responses and contact-inhibition. In addition, the role of the p38 pathway in proliferative control and tumor suppression is mediated by its impact on several cell-cycle regulators. These findings reveal a tumor-suppressing function of the p38 pathway, and indicate that components of the p38 pathway are potential targets for novel cancer therapies.     

The class A chitin synthase gene of Spodoptera exigua: molecular cloning and expression patterns

Chitin synthase (CHS) is an important enzymatic component required for chitin formation in the cuticles and cuticular linings of other tissues. In the present study, a new CHS gene was characterized from the beet army worm Spodoptera exigua (Hübner) (Se). Homologous alignment and phylogenetic analysis of S. exigua CHS (SeCHS) with other related proteins suggest that SeCHS belongs to the class A CHS family (SeCHSA). Northern blot analysis revealed that SeCHSA is transcribed preferentially in the cuticle and tracheae. Further investigation indicated that SeCHSA mRNA is highly expressed in the early and late stages of each larval instar, and consistently expressed in high level during the pupal stage. Using antibody specific for CHS, SeCHS was further localized in the underlying epidermal cells of the integument and tracheal cells, but not in the fat body or Malpighian tubules. These data suggest that SeCHS plays an important role in cuticle formation and development of S. exigua.     

Synthesis and HMG CoA reductase inhibition of 4-thiophenyl quinolines as potential hypocholesterolemic agents

A series of novel 4-thiophenyl quinoline-based mevalonolactone derivatives were synthesized from ethyl 6,7,8-trisubstituted-4-chloro-quinoline-3-carboxylates by several reactions and evaluated for their ability to inhibit the rat HMG CoA reductase in vitro. It was found that substitution with a variety of thiophenyl groups at position 4 in quinoline resulted in retention or enhancement of the inhibition and the preferable groups were 4-isopropyl-thiophenyl and 3-methoxy-thiophenyl. (4R,6S)-6-[(E)-2-(6,7,8-trifluoro-4-isopropylthiophenyl-quinoline-3-yl)-ethenyl]-3,4,5,6-tetrahydro-4-hydroxy-2H-pyran-2-one (A16) and (4R, 6S)-6-[(E)-2-(6-fluoro-4,7-di-(3-methoxy-thiophenyl)-quinoline-3-yl)-ethenyl]-3,4,5,6-tetrahydro-4-hydroxy-2H-pyran-2-one (A23) were approximately three times more potent than rosuvastatin or pitavastatin in inhibiting HMG CoA reductase and selected as the hypocholesterolemic candidates for further evaluation.