Effects of six diterpenes on macrophage eicosanoid biosynthesis

Six diterpenes (three clerodanes, two abietanes and one rosane) were tested for interactions with the cyclooxygenase and 5-lipoxygenase pathways of arachidonate metabolism and for effects of nitric oxide production. Two abietane diterpenes, aethiopinone and 11,12-dihydroxy-6-oxo-8,11,13-abietatriene and the rosane lagascatriol showed a remarkable effect on COX-1 pathway of PGE2 release in calcium ionophore A23187-stimulated peritoneal macrophages. Only the two latter diterpenes showed inhibition on COX-2 pathway of PGE2 release in E. coli LPS-stimulated peritoneal macrophages. In addition, all compounds assayed were inhibitors of LTC4 release with IC50 < or = 10 microM. Clerodane diterpenes were inactive in COX assay. None of the diterpenes assayed, except 11,12-dihydroxy-6-oxo-8,11,13-abietatriene, affected NO production. The results obtained suggest that the cellular mechanisms of action of some of these substances may involve inhibition of cyclooxygenase/lipoxygenase pathways and nitric oxide production.     

Chromosome length polymorphism in Cryptococcus neoformans clinical and environmental isolates

A protocol for intact DNA preparation from the basidiomycetous yeast Cryptococcus neoformans has been developed and applied to karyotyping C. neoformans isolates displaying different degrees of capsule formation. A total of 46 strains have been analyzed: 23 (50%) isolated from environmental samples (pigeon droppings), all of them belonging to C. neoformans var. neoformans; and 23 (50%) from clinical samples (human and veterinarian) including 10 isolates of C. neoformans var. neoformans and 13 isolates of C. neoformans var. gattii. Our results showed a global genome size ranging from 14.2 to 20.9 Mb for variety neoformans and from 7.9 to 16.8 Mb for variety gattii. The karyotype diversity was very high for variety neoformans (29 different patterns for the 33 analyzed strains) and lower for variety gattii (six different patterns for 13 strains). No grouping among variety neoformans strains from the same origin was found indicating very high genome diversity for this variety, irrespectively of the origin of the strains.     

The C terminus of the nuclear protein NuMA: phylogenetic distribution and structure

The C terminus of the nuclear protein NuMA, NuMA-CT, has a well-known function in mitosis via its proximal segment, but it seems also involved in the control of differentiation. To further investigate the structure and function of NuMA, we exploited established computational techniques and tools to collate and characterize proteins with regions similar to the distal portion of NuMA-CT (NuMA-CTDP). The phylogenetic distribution of NuMA-CTDP was examined by PSI-BLAST- and TBLASTN-based analysis of genome and protein sequence databases. Proteins and open reading frames with a NuMA-CTDP-like region were found in a diverse set of vertebrate species including mammals, birds, amphibia, and early teleost fish. The potential structure of NuMA-CTDP was investigated by searching a database of protein sequences of known three-dimensional structure with a hidden Markov model (HMM) estimated using representative (human, frog, chicken, and pufferfish) sequences. The two highest scoring sequences that aligned to the HMM were the extracellular domains of beta3-integrin and Her2, suggesting that NuMA-CTDP may have a primarily beta fold structure. These data indicate that NuMA-CTDP may represent an important functional sequence conserved in vertebrates, where it may act as a receptor to coordinate cellular events.     

Simulated bioprosthetic heart valve deformation under quasi-static loading

For more than 40 years, the replacement of diseased natural heart valves with prosthetic devices has dramatically extended the quality and length of the lives of millions of patients worldwide. However, bioprosthetic heart valves (BHV) continue to fail due to structural failure resulting from poor tissue durability and faulty design. Clearly, an in-depth understanding of the biomechanical behavior of BHV at both the tissue and functional prosthesis levels is essential to improving BHV design and to reduce rates of failure. In this study, we simulated quasi-static BHV leaflet deformation under 40, 80, and 120 mm Hg quasi-static transvalvular pressures. A Fung-elastic material model was used that incorporated material parameters and axes derived from actual leaflet biaxial tests and measured leaflet collagen fiber structure. Rigorous experimental validation of predicted leaflet strain field was used to validate the model results. An overall maximum discrepancy of 2.36% strain between the finite element (FE) results and experiment measurements was obtained, indicating good agreement between computed and measured major principal strains. Parametric studies utilizing the material parameter set from one leaflet for all three leaflets resulted in substantial variations in leaflet stress and strain distributions. This result suggests that utilization of actual leaflet material properties is essential for accurate BHV FE simulations. The present study also underscores the need for rigorous experimentation and accurate constitutive models in simulating BHV function and design.     

Design and synthesis of an optimized positional scanning library of peptoids: identification of novel multidrug resistance reversal agents

Herein is reported the optimized solid-phase synthesis of a library of 5,120 trimeric N-alkylglycines (peptoids) using the positional scanning format and the submonomer strategy. Diversity at the N-terminal position was generated from 20 commercially available primary amines, whereas 16 primary amines were employed for the middle and C-terminal positions of the trimers. Formation of undesirable side-products observed in a previous library synthesis (Humet, M. et al. J. Comb. Chem. 2003, 5, 597-605) was averted by restricting the use of primary amines functionalized with tertiary amino groups to the third amination step. Screening of the new library for the identification of chemosensitizers yielded two peptoids, compounds 1 and 2, with potent in vitro activity as multidrug resistance (MDR) reversal agents. The structures of the lead peptoids are consistent with a pharmacophore model generated from the interaction of various known inhibitors with the MDR-implicated transmembrane glycoprotein P-gp.     

Genomic analysis of Drosophila melanogaster telomeres: full-length copies of HeT-A and TART elements at telomeres

The repetitive nature of heterochromatin hampers its analysis in general genome-sequencing projects. Specific studies are needed to extend the sequence into telomeric and centromeric heterochromatin. Drosophila telomeres lack the telomerase-generated repeats that are characteristic of other eukaryotic chromosomes. Instead, they consist of tandem arrays of HeT-A and TART elements. Herein, we present the genomic organization of the telomeres in the isogenic strain (y; cn bw sp) that was used for the Drosophila melanogaster sequencing project. The data indicate that the canonical features of telomere organization are widely conserved in evolution. In addition, we have identified full-length elements, likely competent elements, for HeT-A and TART.     

Phylogenetic analysis of the functional domains of mariner-like element (MLE) transposases

We have analyzed the sequences of mariner-like element (MLE) transposases, in order to obtain a clearer picture of their phylogenetic relationships. In particular, we have considered their two known structural domains, as well as the nucleic acid sequences of the MLE inverted terminal repeats (ITR). The most consistent tree was obtained using sequences of the catalytic domain of the transposase. The trees obtained with the amino acid sequences of the ITR-binding domain and the ITR sequences themselves were similar to that obtained with the catalytic domain. However, a major difference indicated that the cecropia sub-family is divided into two sub-groups. These new trees were used to examine the evolutionary divergence of mariner-like transposable elements, with particular reference to the possibility that recombination events or gene conversions created mosaic elements during the evolution of transposons.     

Pericentromeric regions containing 1.688 satellite DNA sequences show anti-kinetochore antibody staining in prometaphase chromosomes of Drosophila melanogaster

A striking characteristic of the centromeric heterochromatin of Drosophila melanogaster is that each chromosome carries different satellite DNA sequences. Here we show that while the major component of the 1.688 satellite DNA family expands across the centromere of the X chromosome the rest of the minor variants are located at pericentromeric positions in the large autosomes. Immunostaining of prometaphase chromosomes with the kinetocore-specific anti-BUB1 antibody reveals the transient presence of this centromeric protein in all the regions containing the 1.688 satellite.