Identification and characterization of a novel murine beta-defensin-related gene

Beta-defensins comprise a family of cationic peptides, which are predominately expressed at epithelial surfaces and have a broad-range antimicrobial activity. We have assembled two BAC-based contigs from the chromosomal region 8A4 that contain the murine defensins, and we have mapped six reported beta-defensin genes. In addition, we have isolated and functionally characterized a novel beta-defensin gene that deviates from the canonical six cysteine motif present in the mature functional peptide of all other beta-defensins. This defensin-related gene (Defr1) is most highly expressed in testis and heart. The genomic organization is highly similar to Defb3, 4, 5, and 6, and the exon 1 sequence is very highly conserved. A synthetic Defr1 peptide displayed antimicrobial activity against Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, and Burkholderia cepacia. The antimicrobial activity of Defr1 against S. aureus, E.coli, and B. cepacia was found to be reduced in raised concentration of NaCl, but its action against P. aeruginosa was independent of NaCl concentration. This is the first report of a functional beta defensin that lacks one of the conserved cysteine residues in its predicted mature peptide. This study has major implications for the structure and functions of these important host defense molecules.     

Laser photoirradiation in digital flexor tendon repair.

This study evaluates tendon coaptation using Nd:YAG laser photoirradiation in an in vivo cockerel model. Using the intervinculum segments of the flexor profundus tendons, experimental transactions were performed. Tendon coaptation was then attempted using laser photoirradiation. Tendons were immediately examined for evidence of stable coaptation. After this assessment, specimens were excised and processed for electron microscopic examination and exposure to trypsin digestion. Despite varying multiple laser parameters, tissue welding was not observed. The subsequent functional and ultrastructural observations of irradiated tendon suggest that these changes are those of simple thermal denaturation. The results of this study suggest that when successful tissue welding has been observed in other tissue types, the mechanism is unlikely to be because of formation of intermolecular collagen bonds as hypothesized. An alternative hypothesis is that laser welding reflects photothermal coagulation of cytoplasmic peptides or nucleic acids liberated at the coaptation interface. This may explain the successful welding of cell-rich tissues such as bowel, vas deferens, and arteries and the observed failure of laser welding in collagen-rich but relatively hypocellular tendon.     

Plate 497. Lechenaultia tubiflora Goodeniaceae

Summary.  Lechenaultia tubiflora R.Br. ( Goodeniaceae ) heath leschenaultia has narrow, tubular flowers that show great variation in colour. It has proved difficult in cultivation.     

Plate 492. Lechenaultia acutiloba Goodeniaceae

Summary.  Lechenaultia acutiloba Benth. ( Goodeniaceae ) is a rare, greenish-yellow-flowered species of compact form, unknown in cultivation in Britain.     

Pseudomonas aeruginosa delays Kupffer cell death via stabilization of the X-chromosome-linked inhibitor of apoptosis protein

Kupffer cells are important for bacterial clearance and cytokine production during infection. We have previously shown that severe infection with Pseudomonas aeruginosa ultimately results in loss of Kupffer cells and hepatic bacterial clearance. This was associated with prolonged hepatic inflammation. However, there is a period of time during which there is both preserved hepatic bacterial clearance and increased circulating TNF-alpha. We hypothesized that early during infection, Kupffer cells are protected against TNF-alpha-induced cell death via activation of survival pathways. KC13-2 cells (a clonal Kupffer cell line) were treated with P. aeruginosa (strain PA103), TNF-alpha, or both. At early time points, TNF-alpha induced caspase-mediated cell death, but PA103 did not. When we combined the two exposures, PA103 protected KC13-2 cells from TNF-alpha-induced cell death. PA103, in the setting of TNF exposure, stabilized the X-chromosome-linked inhibitor of apoptosis protein (XIAP). Stabilization of XIAP can occur via PI3K and Akt. We found that PA103 activated Akt and that pretreatment with the PI3K inhibitor, LY294002, prevented PA103-induced protection against TNF-alpha-induced cell death. The effects of LY294002 included decreased levels of XIAP and increased amounts of cleaved caspase-3. Overexpression of Akt mimicked the effects of PA103 by protecting cells from TNF-alpha-induced cell death and XIAP cleavage. Transfection with a stable, nondegradable XIAP mutant also protected cells against TNF-alpha-induced cell death. These studies demonstrate that P. aeruginosa delays TNF-alpha-induced Kupffer cell death via stabilization of XIAP.     

Differential induction of type I interferon responses in myeloid dendritic cells by mosquito and mammalian-cell-derived alphaviruses

Dendritic cells (DCs) are an important early target cell for many mosquito-borne viruses, and in many cases mosquito-cell-derived arboviruses more efficiently infect DCs than viruses derived from mammalian cells. However, whether mosquito-cell-derived viruses differ from mammalian-cell-derived viruses in their ability to induce antiviral responses in the infected dendritic cell has not been evaluated. In this report, alphaviruses, which are mosquito-borne viruses that cause diseases ranging from encephalitis to arthritis, were used to determine whether viruses grown in mosquito cells differed from mammalian-cell-derived viruses in their ability to induce type I interferon (IFN) responses in infected primary dendritic cells. Consistent with previous results, mosquito-cell-derived Ross River virus (mos-RRV) and Venezuelan equine encephalitis virus (mos-VEE) exhibited enhanced infection of primary myeloid dendritic cells (mDCs) compared to mammalian-cell-derived virus preparations. However, unlike the mammalian-cell-derived viruses, which induced high levels of type I IFN in the infected mDC cultures, mos-RRV and mos-VEE were poor IFN inducers. Furthermore, the poor IFN induction by mos-RRV contributed to the enhanced infection of mDCs by mos-RRV. These results suggest that the viruses initially delivered by the mosquito vector differ from those generated in subsequent rounds of replication in the host, not just with respect to their ability to infect dendritic cells but also in their ability to induce or inhibit antiviral type I IFN responses. This difference may have an important impact on the mosquito-borne virus's ability to successfully make the transition from the arthropod vector to the vertebrate host.     

Demographic analysis of a declining pika Ochotona collaris population: linking survival to broad-scale climate patterns via spring snowmelt patterns

1. Demographic analysis is essential in order to determine which factors, such as survival, fertility and other life-history characteristics, have the greatest influence on a population's rate of growth (lambda). 2. We used life-table response experiments (LTREs) to assess the relative importance of survival and fertility rates for an alpine lagomorph, the collared pika Ochotona collaris, using 12 years (1995-2006) of census data. The LTRE analysis was repeated for each of three subpopulations within the main study site that were defined by aspect (east, west and south). 3. Across the entire study site, the survival and fertility of adults contributed 35.6 and 43.5%, respectively, to the variance observed in the projected population growth rate, V(lambda), whereas juvenile survival contributed 20.9%. Adult survival and fertility contributed approximately equal amounts for each subpopulation when considered separately, although their rank order varied spatially. 4. Adult survival across the entire site was positively correlated to the Pacific Decadal Oscillation (PDO) with a time lag of 1 year, and was uncorrelated to adult density. The PDO was negatively correlated to the timing of spring snowmelt at our site, implicating the importance of earlier spring conditions and plant phenology on the subsequent winter survival of adults and therefore, population growth. 5. When subpopulations were analysed separately, survivals and fertilities were variously correlated to lagged PDO and adult densities, but the patterns varied spatially. Therefore, the mechanisms underlying V(lambda) can vary substantially over relatively short distances.     

Identification of the major protein component of the pneumococcal eclipse complex

During genetic transformation of Streptococcus pneumoniae, single strands from native donor DNA enter competent cells, where they associate with an unidentified protein with a molecular mass of 15 to 20 kDa to form the eclipse complex. Using Western blotting, we identify the principal protein cofractionating with donor DNA in this complex as SsbB.     

Repeatability of 3D gait kinematics obtained from an electromagnetic tracking system during treadmill locomotion

The purpose of this paper was to describe a technique that enables three-dimensional (3D) gait kinematics to be obtained using an electromagnetic tracking system, and to report the intra-trial, intra-day/inter-tester and inter-day/intra-tester repeatability of kinematic gait data obtained using this technique. Ten able-bodied adults underwent four gait assessments; the same two testers tested each subject independently on two different days. Gait assessments were conducted on a custom-built long-bed treadmill with no metal components between the rollers. Each gait assessment involved familiarisation to treadmill walking, subject anatomical and functional calibration, and a period of steady-state treadmill walking at a self-selected speed. Following data collection, 3D joint kinematics were calculated using the joint coordinate system approach. 3D joint angle waveforms for 10 left and right strides were extracted and temporally normalised for each trial. Intra-trial, intra-day/inter-tester and inter-day/intra-tester repeatability of the temporally normalised kinematic waveforms were quantified using the coefficient of multiple determination (CMD). CMDs for joint kinematics averaged 0.942 intra-trial, 0.849 intra-day/inter-tester and 0.773 inter-day/intra-tester. In general, sagittal plane kinematics were more repeatable than frontal or transverse plane kinematics, and kinematics at the hip were more repeatable than at the knee or ankle. The level of repeatability of kinematic gait data obtained during treadmill walking using this protocol was equal or superior to that reported previously for overground walking using image-based protocols.