Life history, enamel formation, and linear enamel hypoplasia in the Ceboidea

Linear enamel hypoplasia (LEH), a developmental defect of enamel, increases in frequency from prosimian to monkey to lesser ape to great ape grades (Guatelli-Steinberg 2000 Am. J. Phys. Anthropol. 112:395-410, [2001] Evol. Anthropol. 10:138-151; Newell 1998 Ph.D. dissertation, Temple University). This taxonomic pattern in the distribution of LEH is closely related to maturation length across the primate order (Newell 1998 Ph.D. dissertation, Temple University, 2000 Am. J. Phys. Anthropol. [Suppl.] 30:236). Longer maturation periods are associated with higher LEH frequencies; they appear to provide greater opportunity for defects to form. The present study explores the relationship between maturation length and LEH frequency within the Ceboidea. Because of its prolonged period of growth, Cebus is predicted to manifest LEH at a higher frequency than the more rapidly maturing ceboid genera. To test this hypothesis, two separate researchers (E.A.N. and D.G.-S.) examined LEH in nonoverlapping museum series of ceboids. The results support the hypothesis: in 13 genera (n = 1,276), E.A.N. found that LEH frequencies ranged from 0% in Callicebus, Cebuella, and Saimiri to 20% in Cebus. D.G.-S. found similar frequencies among five genera (n = 107), from 0% in Saimiri to 32% in Cebus. Thus, the broad pattern of LEH distribution evident across major taxonomic groups of primates is repeated within the Ceboidea. We also examined a related hypothesis linking the spacing of perikymata, which is influenced by enamel extension rates (Shellis 1998 J. Hum. Evol. 35:387-400), to LEH. The most likely areas of tooth crowns to exhibit LEH in human teeth are those in which perikymata are most closely spaced (Hillson and Bond 1997 Am. J. Phys. Anthropol. 104:89-103). We hypothesized that the longer-maturing Cebus, with its elevated LEH frequency, will also exhibit more closely spaced perikymata than other ceboids. Analysis of a small microscopic subsample (n = 8) lends limited support to this second hypothesis.     

The stimulus-induced tyrosine phosphorylation of Munc18c facilitates vesicle exocytosis

Stimulus-induced tyrosine phosphorylation of Munc18c was investigated as a potential regulatory mechanism by which the Munc18c-Syntaxin 4 complex can be dissociated in response to divergent stimuli in multiple cell types. Use of [(32)P]orthophosphate incorporation, pervanadate treatment, and phosphotyrosine-specific antibodies demonstrated that Munc18c underwent tyrosine phosphorylation. Phosphorylation was apparent under basal conditions, but levels were significantly increased within 5 min of glucose stimulation in MIN6 beta cells. Tyrosine phosphorylation of Munc18c was also detected in 3T3L1 adipocytes and increased with insulin stimulation, suggesting that this may be a conserved mechanism. Syntaxin 4 binding to Munc18c decreased as Munc18c phosphorylation levels increased in pervanadate-treated cells, suggesting that phosphorylation dissociates the Munc18c-Syntaxin 4 complex. Munc18c phosphorylation was localized to the N-terminal 255 residues. Mutagenesis of one residue in this region, Y219F, significantly increased the affinity of Munc18c for Syntaxin 4, whereas mutation of three other candidate sites was without effect. Moreover, Munc18c-Y219F expression in MIN6 cells functionally inhibited glucose-stimulated SNARE complex formation and insulin granule exocytosis. These data support a novel and conserved mechanism for the dissociation of Munc18c-Syntaxin 4 complexes in a stimulus-dependent manner to facilitate the increase in Syntaxin 4-VAMP2 association and to promote vesicle/granule fusion.     

Cloning, sequence and expression of the lactate dehydrogenase gene from the human malaria parasite, Plasmodium vivax

Increased drug resistance to anti-malarials highlights the need for the development of new therapeutics for the treatment of malaria. To this end, the lactate dehydrogenase (LDH) gene was cloned and sequenced from genomic DNA of Plasmodium vivax ( PvLDH) Belem strain. The 316 amino acid protein-coding region of the PvLDH gene was inserted into the prokaryotic expression vector pKK223-3 and a 34 kDa protein with LDH activity was expressed in E. coli. Structural differences between human LDHs and PfLDH make the latter an attractive target for inhibitors leading to novel anti-malarial drugs. The sequence similarity between PvLDH and PfLDH (90% residue identity and no insertions or deletions) indicate that the same approach could be applied to Plasmodium vivax, the most common human malaria parasite in the world.     

Analysis of the Relationship between Peak Stress and Proteoglycan Loss following Injurious Compression of Human Post-mortem Knee and Ankle Cartilage

While traumatic joint injuries are known to increase the risk of osteoarthritis (OA), the mechanism is not known. Models for injurious compression of cartilage may identify predictors of injury that suggest a clinical mechanism. We investigated the relationship between peak stress during compression and glycosaminoglycan (GAG) loss after injury for knee and ankle cartilages. Human cartilage explant disks were harvested post-mortem from the knee and ankle of three organ donors with no history of OA and subjected to injurious compression to 65% strain in uniaxial unconfined compression at 2 mm/s (400%/s). The GAG content of the conditioned medium was measured 3 days after injury. After injury of knee cartilage disks, damage was visible in 18 of 39 disks (36%). Three days after injury, the increase in GAG loss to the medium (GAG loss from injured disks minus GAG loss from location-matched uncompressed controls) was 1.5±0.3 μg/disk (mean ± SEM). With final strain and compression velocity held constant, we observed that increasing peak stress during injury was associated with less GAG loss after injury (P<0.001). In contrast, ankle cartilage appeared damaged after injury in only 1 of 16 disks (6%), there was no increase in GAG loss (0.0±0.3 μg/disk), and no relationship between peak stress and increase in GAG loss was detected (P=0.51). By itself, increasing peak stress did not appear to be an important cause of GAG loss from human cartilage in our injurious compression model. However, we observed further evidence for differences in the response of knee and ankle cartilages to injury.     

The OmpL porin does not modulate redox potential in the periplasmic space of Escherichia coli

The Escherichia coli DsbA protein is the major oxidative catalyst in the periplasm. Dartigalongue et al. (EMBO J., 19, 5980-5988, 2000) reported that null mutations in the ompL gene of E.coli fully suppress all phenotypes associated with dsbA mutants, i.e. sensitivity to the reducing agent dithiothreitol (DTT) and the antibiotic benzylpenicillin, lack of motility, reduced alkaline phosphatase activity and mucoidy. They showed that OmpL is a porin and hypothesized that ompL null mutations exert their suppressive effect by preventing efflux of a putative oxidizing-reducing compound into the medium. We have repeated these experiments using two different ompL null alleles in at least three different E.coli K-12 genetic backgrounds and have failed to reproduce any of the ompL suppressive effects noted above. Also, we show that, contrary to earlier results, ompL null mutations alone do not result in partial DTT sensitivity or partial motility, nor do they appreciably affect bacterial growth rates or block propagation of the male-specific bacteriophage M13. Thus, our findings clearly demonstrate that ompL plays no perceptible role in modulating redox potential in the periplasm of E.coli.     

Heterogeneity and randomness of DNA methylation patterns in human embryonic stem cells

DNA methylation has been proposed to be important in many biological processes and is the subject of intense study. Traditional bisulfite genomic sequencing allows detailed high-resolution methylation pattern analysis of each molecule with haplotype information across a few hundred bases at each locus, but lacks the capacity to gather voluminous data. Although recent technological developments are aimed at assessing DNA methylation patterns in a high-throughput manner across the genome, the haplotype information cannot be accurately assembled when the sequencing reads are short or when each hybridization target only includes one or two cytosine-phosphate-guanine (CpG) sites. Whether a distinct and nonrandom DNA methylation pattern is present at a given locus is difficult to discern without the haplotype information, and the DNA methylation patterns are much less apparent because the data are often obtained only as methylation frequencies at each CpG site with some of these methods. It would facilitate the interpretation of data obtained from high-throughput bisulfite sequencing if the loci with nonrandom DNA methylation patterns could be distinguished from those that are randomly methylated. In this study, we carried out traditional genomic bisulfite sequencing using the normal diploid human embryonic stem (hES) cell lines, and utilized Hamming distance analysis to evaluate the existence of a distinct and nonrandom DNA methylation pattern at each locus studied. Our findings suggest that Hamming distance is a simple, quick, and useful tool to identify loci with nonrandom DNA methylation patterns and may be utilized to discern links between biological changes and DNA methylation patterns in the high-throughput bisulfite sequencing data sets.     

Impact of NHS walk-in centres on the workload of other local healthcare providers: time series analysis

ObjectivesTo assess the impact of NHS walk-in centres on the workload of local accident and emergency departments, general practices, and out of hours services.DesignTime series analysis in walk-in centre sites with no-treatment control series in matched sites.SettingWalk-in centres and matched control towns without walk-in centres in England.Participants20 accident and emergency departments, 40 general practices, and 14 out of hours services within 3 km of a walk-in centre or the centre of a control town.ResultsA reduction in consultations at emergency departments (–175 (95% confidence interval –387 to 36) consultations per department per month) and general practices (–19.8 (−53.3 to 13.8) consultations per 1000 patients per month) close to walk-in centres became apparent, although these reductions were not statistically significant. Walk-in centres did not have any impact on consultations on out of hours services.ConclusionIt will be necessary to assess the impact of walk-in centres in a larger number of sites and over a prolonged period, to determine whether they reduce the demand on other local NHS providers.

What is already known on this topic

One of the objectives for NHS walk-in centres was to reduce demand on other NHS services, particularly general practitioners'' services and accident and emergency departmentsStudies of walk-in centres in North America have indicated that such centres do not reduce demand on other healthcare servicesStudies of minor injuries units in the United Kingdom (which have some similarities with walk-in centres) indicate that these units substitute mainly for consultations in accident and emergency departments

What this study adds

The data imply that walk-in centres may moderate the increasing demand on general practice and reduce the number of consultations in accident and emergency departmentsThe high level of background variability in consultation rates means that any impact of a walk-in centre is not statistically significantTo draw robust conclusions about the impact of walk-in centres on other health providers will require study of a large number of sites over an extended period of time     

The Prevalence of Angiostrongylus cantonensis/mackerrasae Complex in Molluscs from the Sydney Region

Angiostrongylus cantonensis and Angiostrongylus mackerrasae are metastrongyloid nematodes that infect various rat species. Terrestrial and aquatic molluscs are intermediate hosts of these worms while humans and dogs are accidental hosts. Angiostrongylus cantonensis is the major cause of angiostrongyliasis, a disease characterised by eosinophilic meningitis. Although both A. cantonensis and A. mackerrasae are found in Australia, A. cantonensis appears to account for most infections in humans and animals. Due to the occurrence of several severe clinical cases in Sydney and Brisbane, the need for epidemiological studies on angiostrongyliasis in this region has become apparent. In the present study, a conventional PCR and a TaqMan assay were compared for their ability to amplify Angiostrongylus DNA from DNA extracted from molluscs. The TaqMan assay was more sensitive, capable of detecting the DNA equivalent to one hundredth of a nematode larva. Therefore, the TaqMan assay was used to screen molluscs (n=500) of 14 species collected from the Sydney region. Angiostrongylus DNA was detected in 2 of the 14 mollusc species; Cornu aspersum [14/312 (4.5%)], and Bradybaenia similaris [1/10 (10%)], which are non-native terrestrial snails commonly found in urban habitats. The prevalence of Angiostrongylus spp. was 3.0% ± 0.8% (CI 95%). Additionally, experimentally infected Austropeplea lessoni snails shed A. cantonensis larvae in their mucus, implicating mucus as a source of infection. This is the first Australian study to survey molluscs using real-time PCR and confirms that the garden snail, C. aspersum, is a common intermediate host for Angiostrongylus spp. in Sydney.     

Superior abstract-concept learning by Clark's nutcrackers (Nucifraga columbiana)

The ability to learn abstract relational concepts is fundamental to higher level cognition. In contrast to item-specific concepts (e.g. pictures containing trees versus pictures containing cars), abstract relational concepts are not bound to particular stimulus features, but instead involve the relationship between stimuli and therefore may be extrapolated to novel stimuli. Previous research investigating the same/different abstract concept has suggested that primates might be specially adapted to extract relations among items and would require fewer exemplars of a rule to learn an abstract concept than non-primate species. We assessed abstract-concept learning in an avian species, Clark''s nutcracker (Nucifraga columbiana), using a small number of exemplars (eight pairs of the same rule, and 56 pairs of the different rule) identical to that previously used to compare rhesus monkeys, capuchin monkeys and pigeons. Nutcrackers as a group (N = 9) showed more novel stimulus transfer than any previous species tested with this small number of exemplars. Two nutcrackers showed full concept learning and four more showed transfer considerably above chance performance, indicating partial concept learning. These results show that the Clark''s nutcracker, a corvid species well known for its amazing feats of spatial memory, learns the same/different abstract concept better than any non-human species (including non-human primates) yet tested on this same task.     

Loss of the Mono-ADP-ribosyltransferase,Tiparp, Increases Sensitivity to Dioxin-induced Steatohepatitis and Lethality

The aryl hydrocarbon receptor (AHR) mediates the toxic effects of the environmental contaminant dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin; TCDD). Dioxin causes a range of toxic responses, including hepatic damage, steatohepatitis, and a lethal wasting syndrome; however, the mechanisms are still unknown. Here, we show that the loss of TCDD-inducible poly(ADP-ribose) polymerase (Tiparp), an ADP-ribosyltransferase and AHR repressor, increases sensitivity to dioxin-induced toxicity, steatohepatitis, and lethality. Tiparp^−/− mice given a single injection of 100 μg/kg dioxin did not survive beyond day 5; all Tiparp^+/+ mice survived the 30-day treatment. Dioxin-treated Tiparp^−/− mice exhibited increased liver steatosis and hepatotoxicity. Tiparp ADP-ribosylated AHR but not its dimerization partner, the AHR nuclear translocator, and the repressive effects of TIPARP on AHR were reversed by the macrodomain containing mono-ADP-ribosylase MACROD1 but not MACROD2. These results reveal previously unidentified roles for Tiparp, MacroD1, and ADP-ribosylation in AHR-mediated steatohepatitis and lethality in response to dioxin.