ProServer: a simple, extensible Perl DAS server

SUMMARY: The increasing size and complexity of biological databases has led to a growing trend to federate rather than duplicate them. In order to share data between federated databases, protocols for the exchange mechanism must be developed. One such data exchange protocol that is widely used is the Distributed Annotation System (DAS). For example, DAS has enabled small experimental groups to integrate their data into the Ensembl genome browser. We have developed ProServer, a simple, lightweight, Perl-based DAS server that does not depend on a separate HTTP server. The ProServer package is easily extensible, allowing data to be served from almost any underlying data model. Recent additions to the DAS protocol have enabled both structure and alignment (sequence and structural) data to be exchanged. ProServer allows both of these data types to be served. AVAILABILITY: ProServer can be downloaded from http://www.sanger.ac.uk/proserver/ or CPAN http://search.cpan.org/~rpettett/. Details on the system requirements and installation of ProServer can be found at http://www.sanger.ac.uk/proserver/.     

Coordination of cell proliferation and anterior-posterior axis establishment in the mouse embryo

During development, the growth of the embryo must be coupled to its patterning to ensure correct and timely morphogenesis. In the mouse embryo, migration of the anterior visceral endoderm (AVE) to the prospective anterior establishes the anterior-posterior (A-P) axis. By analysing the distribution of cells in S phase, M phase and G2 from the time just prior to the migration of the AVE until 18 hours after its movement, we show that there is no evidence for differential proliferation along the A-P axis of the mouse embryo. Rather, we have identified that as AVE movements are being initiated, the epiblast proliferates at a much higher rate than the visceral endoderm. We show that these high levels of proliferation in the epiblast are dependent on Nodal signalling and are required for A-P establishment, as blocking cell division in the epiblast inhibits AVE migration. Interestingly, inhibition of migration by blocking proliferation can be rescued by Dkk1. This suggests that the high levels of epiblast proliferation function to move the prospective AVE away from signals that are inhibitory to its migration. The finding that initiation of AVE movements requires a certain level of proliferation in the epiblast provides a mechanism whereby A-P axis development is coordinated with embryonic growth.     

Tiludronate treatment improves structural changes and symptoms of osteoarthritis in the canine anterior cruciate ligament model

Introduction

The aim of this prospective, randomized, controlled, double-blind study was to evaluate the effects of tiludronate (TLN), a bisphosphonate, on structural, biochemical and molecular changes and function in an experimental dog model of osteoarthritis (OA).

Methods

Baseline values were established the week preceding surgical transection of the right cranial/anterior cruciate ligament, with eight dogs serving as OA placebo controls and eight others receiving four TLN injections (2 mg/kg subcutaneously) at two-week intervals starting the day of surgery for eight weeks. At baseline, Week 4 and Week 8, the functional outcome was evaluated using kinetic gait analysis, telemetered locomotor actimetry and video-automated behaviour capture. Pain impairment was assessed using a composite numerical rating scale (NRS), a visual analog scale, and electrodermal activity (EDA). At necropsy (Week 8), macroscopic and histomorphological analyses of synovium, cartilage and subchondral bone of the femoral condyles and tibial plateaus were assessed. Immunohistochemistry of cartilage (matrix metalloproteinase (MMP)-1, MMP-13, and a disintegrin and metalloproteinase domain with thrombospondin motifs (ADAMTS5)) and subchondral bone (cathepsin K) was performed. Synovial fluid was analyzed for inflammatory (PGE₂ and nitrite/nitrate levels) biomarkers. Statistical analyses (mixed and generalized linear models) were performed with an α-threshold of 0.05.

Results

A better functional outcome was observed in TLN dogs than OA placebo controls. Hence, TLN dogs had lower gait disability (P = 0.04 at Week 8) and NRS score (P = 0.03, group effect), and demonstrated behaviours of painless condition with the video-capture (P < 0.04). Dogs treated with TLN demonstrated a trend toward improved actimetry and less pain according to EDA. Macroscopically, both groups had similar level of morphometric lesions, TLN-treated dogs having less joint effusion (P = 0.01), reduced synovial fluid levels of PGE₂ (P = 0.02), nitrites/nitrates (P = 0.01), lower synovitis score (P < 0.01) and a greater subchondral bone surface (P < 0.01). Immunohistochemical staining revealed lower levels in TLN-treated dogs of MMP-13 (P = 0.02), ADAMTS5 (P = 0.02) in cartilage and cathepsin K (P = 0.02) in subchondral bone.

Conclusion

Tiludronate treatment demonstrated a positive effect on gait disability and joint symptoms. This is likely related to the positive influence of the treatment at improving some OA structural changes and reducing the synthesis of catabolic and inflammatory mediators.     

Responses of benthic macroinvertebrate communities to natural geothermal discharges in Yellowstone National Park,USA

We examined responses of benthic macroinvertebrate communities to natural geothermal discharges in 32 streams in Yellowstone National Park (YNP), USA. Geothermal discharges played a major role in structuring benthic communities in YNP, as downstream communities were characterized by low species richness, reduced abundance of EPT taxa and increased abundance of tolerant caddisflies (Trichoptera), chironomids and non-insects. While some taxa were a subset of tolerant organisms that were also common at references sites, others (the damselfly Argia sp., the caddisfly Oxyethira sp. and the exotic New Zealand mudsnail Potamopyrgus antipodarum Gray 1843) were found almost exclusively in geothermal streams. Because geothermal waters are a common feature of YNP, monitoring programs designed to assess long-term status and trends of Yellowstone’s aquatic ecosystems must account for the influence of these discharges. To separate geothermal effects from other potential anthropogenic disturbances in YNP (e.g., atmospheric deposition, road construction, wastewater, global change), we developed a multimetric index based on responses of benthic communities to geothermal discharges. Streams were placed into one of four geothermal categories based on conductivity (reference = < 150 μS/cm; low, moderate and high = 151–300, 301–600 and >600 μS/cm, respectively). The index clearly distinguished among these categories and showed a well-defined threshold response to geothermal effects at very low levels of conductivity. Although the index was specific to geothermal effects, the approach used to develop the index has broad applicability for other systems where impacts of stressors must be assessed within the context of natural environmental gradients. Our findings may provide important insights into how benthic macroinvertebrate communities respond to global change. Reduced discharge and warmer temperatures predicted for Rocky Mountain streams may favor the establishment and expansion of exotic species such as New Zealand mudsnails (P. antipodarum), which are highly tolerant of geothermal influences.     

Correct patterning of the primitive streak requires the anterior visceral endoderm

Anterior-posterior axis specification in the mouse requires signalling from a specialised extra-embryonic tissue called the anterior visceral endoderm (AVE). AVE precursors are induced at the distal tip of the embryo and move to the prospective anterior. Embryological and genetic analysis has demonstrated that the AVE is required for anterior patterning and for correctly positioning the site of primitive streak formation by inhibiting Nodal activity. We have carried out a genetic ablation of the Hex-expressing cells of the AVE (Hex-AVE) by knocking the Diphtheria toxin subunit A into the Hex locus in an inducible manner. Using this model we have identified that, in addition to its requirement in the anterior of the embryo, the Hex-AVE sub-population has a novel role between 5.5 and 6.5dpc in patterning the primitive streak. Embryos lacking the Hex-AVE display delayed initiation of primitive streak formation and miss-patterning of the anterior primitive streak. We demonstrate that in the absence of the Hex-AVE the restriction of Bmp2 expression to the proximal visceral endoderm is also defective and expression of Wnt3 and Nodal is not correctly restricted to the posterior epiblast. These results, coupled with the observation that reducing Nodal signalling in Hex-AVE ablated embryos increases the frequency of phenotypes observed, suggests that these primitive streak patterning defects are due to defective Nodal signalling. Together, our experiments demonstrate that the AVE is not only required for anterior patterning, but also that specific sub-populations of this tissue are required to pattern the posterior of the embryo.     

Assessing the influence of water and substratum quality on benthic macroinvertebrate communities in a metal-polluted stream: an experimental approach

SUMMARY 1. Field and laboratory experiments were conducted to assess the relative influence of water quality and substratum quality on benthic macroinvertebrate communities in the Animas River, a metal-polluted stream in south-western Colorado (U.S.A.).
2. A community-level in situ toxicity test measured direct effects of Animas River water on benthic invertebrates collected from a reference stream (Elk Creek). The effects of metal-contaminated biofilm were examined by comparing macroinvertebrate colonisation of clean and contaminated substrata placed in Elk Creek. A feeding experiment with the mayfly Baetis tricaudatus Dodds (Ephemeroptera: Baetidae) examined metal bioaccumulation and effects of metal-contaminated biofilm on growth and survival.
3. Animas River water was acutely toxic to most taxa, with greatest effects observed on mayflies (Heptageniidae, Ephemerellidae) and stoneflies (Taeniopterygidae and Capniidae).
4. Although Animas River biofilm was characterised by high concentrations of metals and low algal biomass, most taxa colonised substratum from the reference stream and the Animas River equally. The exceptions were Ephemerellidae, Taeniopterygidae and Simuliidae, which were less abundant on Animas River substratum. Mayflies grazing Animas River biofilm accumulated significantly more metals and showed reduced growth compared with organisms feeding on Elk Creek biofilm.
5. Results of our experiments demonstrated that effects of heavy metals on benthic community structure in the Animas River were complex, and that responses to metals in water and contaminated substratum were species-specific. Predicting recovery of benthic communities following remediation requires an understanding of these species-specific responses.     

Rotavirus 2/6 Viruslike Particles Administered Intranasally with Cholera Toxin, Escherichia coli Heat-Labile Toxin (LT), and LT-R192G Induce Protection from Rotavirus Challenge

We have shown that rotavirus 2/6 viruslike particles composed of proteins VP2 and VP6 (2/6-VLPs) administered to mice intranasally with cholera toxin (CT) induced protection from rotavirus challenge, as measured by virus shedding. Since it is unclear if CT will be approved for human use, we evaluated the adjuvanticity of Escherichia coli heat-labile toxin (LT) and LT-R192G. Mice were inoculated intranasally with 10 μg of 2/6-VLPs combined with CT, LT, or LT-R192G. All three adjuvants induced equivalent geometric mean titers of rotavirus-specific serum antibody and intestinal immunoglobulin G (IgG). Mice inoculated with 2/6-VLPs with LT produced significantly higher titers of intestinal IgA than mice given CT as the adjuvant. All mice inoculated with 2/6-VLPs mixed with LT and LT-R192G were totally protected (100%) from rotavirus challenge, while mice inoculated with 2/6-VLPs mixed with CT showed a mean 91% protection from challenge. The availability of a safe, effective mucosal adjuvant such as LT-R192G will increase the practicality of administering recombinant vaccines mucosally.