A genome-wide screen for susceptibility loci in ankylosing spondylitis

Objective. To localize the regions containing genes that determine susceptibility to ankylosing spondylitis (AS). Methods. One hundred five white British families with 121 affected sibling pairs with AS were recruited, largely from the Royal National Hospital for Rheumatic Diseases AS database. A genome-wide linkage screen was undertaken using 254 highly polymorphic microsatellite markers from the Medical Research Council (UK) (MRC) set. The major histocompatibility complex (MHC) region was studied more intensively using 5 microsatellites lying within the HLA class III region and HLA-DRB1 typing. The Analyze package was used for 2-point analysis, and GeneHunter for multipoint analysis. Results. When only the MRC set was considered, 11 markers in 7 regions achieved a P value of ≤0.01. The maximum logarithm of odds score obtained was 3.8 (P = 1.4 x 10-5) using marker D6S273, which lies in the HLA class III region. A further marker used in mapping of the MHC class III region achieved a LOD score of 8.1 (P = 1 x 10-9). Nine of 118 affected sibling pairs (7.6%) did not share parental haplotypes identical by descent across the MHC, suggesting that only 31% of the susceptibility to AS is coded by genes linked to the MHC. The maximum non-MHC LOD score obtained was 2.6 (P = 0.0003) for marker D16S422. Conclusion. The results of this study confirm the strong linkage of the MHC with AS, and provide suggestive evidence regarding the presence and location of non-MHC genes influencing susceptibility to the disease.

Dissection of class III major histocompatibility complex haplotypes associated with rheumatoid arthritis

Objective. We have previously identified a single-nucleotide polymorphism (SNP) haplotype involving the lymphotoxin α (LTA) and tumor necrosis factor (TNF) loci (termed haplotype LTA-TNF2) on chromosome 6 that shows differential association with rheumatoid arthritis (RA) on HLA-DRB1*0404 and *0401 haplotypes, suggesting the presence of additional non-HLA-DRB1 RA susceptibility genes on these haplotypes. To refine this association, we performed a case-control association study using both SNPs and microsatellite markers in haplotypes matched either for HLA-DRB1*0404 or for HLA-DRB1*0401. Methods. Fourteen SNPs lying between HLA-DRB1 and LTA were genotyped in 87 DRB1*04-positive families. High-density microsatellite typing was performed using 24 markers spanning 2,500 kb centered around the TNF gene in 305 DRB1*0401 or *0404 cases and 400 DRB1*0401 or *0404 controls. Single-marker, 2-marker, and 3-marker minihaplotypes were constructed and their frequencies compared between the DRB1*0401 and DRB1*0404 matched case and control haplotypes. Results. Marked preservation of major histocompatibility complex haplotypes was seen, with chromosomes carrying LTA-TNF2 and either DRB1*0401 or DRB1*0404 both carrying an identical SNP haplotype across the 1-Mb region between TNF and HLA-DRB1. Using microsatellite markers, we observed two 3-marker minihaplotypes that were significantly overrepresented in the DRB1*0404 case haplotypes (P = 0.00024 and P = 0.00097). Conclusion. The presence of a single extended SNP haplotype between LTA-TNF2 and both DRB1*0401 and DRB1*0404 is evidence against this region harboring the genetic effects in linkage disequillbrium with LTA-TNF2. Two RA-associated haplotypes on the background of DRB1*0404 were identified in a 126-kb region surrounding and centromeric to the TNF locus.

Investigation of the role of ANKH in ankylosing spondylitis

Objective The ank/ank mouse develops a phenotype similar to ankylosing spondylitis (AS) in humans. ANKH, the human homolog of the mutated gene in the ank/ank mouse, has been implicated in familial autosomal-dominant chondrocalcinosis and autosomal-dominant craniometaphyseal dysplasia. This study was undertaken to investigate the role of ANKH in susceptibility to and clinical manifestations of AS. Methods Sequence variants were identified by genomic sequencing of the 12 ANKH exons and their flanking splice sites in 48 AS patients; variants were then screened in 233 patients and 478 controls. Linkage to the ANKH locus was assessed in 185 affected-sibling-pair families. Results Five single-nucleotide polymorphisms were identified within the coding region and flanking splice sites. No association between either susceptibility to AS or its clinical manifestations and these novel polymorphisms, or between disease susceptibility and 3 known promoter variants, was seen. No linkage between the ANKH locus and AS was observed. Multipoint exclusion mapping rejected the hypothesis of a locus of a magnitude λ≥1.4 (logarithm of odds score <-2) (equivalent to a genetic contribution of >10% to the AS sibling recurrence risk ratio) within this area contributing to AS. Conclusion These findings indicate that ANKH is not significantly involved in susceptibility to or clinical manifestations of AS.

Genome-wide association study for radiographic vertebral fractures: A potential role for the 16q24 BMD locus

Vertebral fracture risk is a heritable complex trait. The aim of this study was to identify genetic susceptibility factors for osteoporotic vertebral fractures applying a genome-wide association study (GWAS) approach. The GWAS discovery was based on the Rotterdam Study, a population-based study of elderly Dutch individuals aged >55years; and comprising 329 cases and 2666 controls with radiographic scoring (McCloskey-Kanis) and genetic data. Replication of one top-associated SNP was pursued by de-novo genotyping of 15 independent studies across Europe, the United States, and Australia and one Asian study. Radiographic vertebral fracture assessment was performed using McCloskey-Kanis or Genant semi-quantitative definitions. SNPs were analyzed in relation to vertebral fracture using logistic regression models corrected for age and sex. Fixed effects inverse variance and Han-Eskin alternative random effects meta-analyses were applied. Genome-wide significance was set at p<5×10-8. In the discovery, a SNP (rs11645938) on chromosome 16q24 was associated with the risk for vertebral fractures at p=4.6×10-8. However, the association was not significant across 5720 cases and 21,791 controls from 14 studies. Fixed-effects meta-analysis summary estimate was 1.06 (95% CI: 0.98-1.14; p=0.17), displaying high degree of heterogeneity (I2=57%; Qhet p=0.0006). Under Han-Eskin alternative random effects model the summary effect was significant (p=0.0005). The SNP maps to a region previously found associated with lumbar spine bone mineral density (LS-BMD) in two large meta-analyses from the GEFOS consortium. A false positive association in the GWAS discovery cannot be excluded, yet, the low-powered setting of the discovery and replication settings (appropriate to identify risk effect size >1.25) may still be consistent with an effect size <1.10, more of the type expected in complex traits. Larger effort in studies with standardized phenotype definitions is needed to confirm or reject the involvement of this locus on the risk for vertebral fractures.

Current trends in the etiology and diagnosis of HPV-related head and neck cancers

Human papilloma virus (HPV) infection is a major risk factor for a distinct subset of head and neck squamous cell carcinoma (HNSCC). The current review summarizes the epidemiology of HNSCC and the disease burden, the infectious cycle of HPV, the roles of viral oncoproteins, E6 and E7, and the downstream cellular events that lead to malignant transformation. Current techniques for the clinical diagnosis of HPV-associated HNSCC will also be discussed, that is, the detection of HPV DNA, RNA, and the HPV surrogate marker, p16 in tumor tissues, as well as HPV-specific antibodies in serum. Such methods do not allow for the early detection of HPV-associated HNSCC and most cases are at an advanced stage upon diagnosis. Novel noninvasive approaches using oral fluid, a clinically relevant biological fluid, allow for the detection of HPV and cellular alterations in infected cells, which may aid in the early detection and HPV-typing of HNSCC tumors. Noninvasive diagnostic methods will enable early detection and intervention, leading to a significant reduction in mortality and morbidity associated with HNSCC.

Suggestive linkage of 2p22-25 and 11q12-13 with low bone mineral density at the lumbar spine in the Irish population

Osteoporosis is a disease characterized by low bone mineral density (BMD) and poor bone quality. Peak bone density is achieved by the third decade of life, after which bone is maintained by a balanced cycle of bone resorption and synthesis. Age-related bone loss occurs as the bone resorption phase outweighs the bone synthesis phase of bone metabolism. Heritability accounts for up to 90% of the variability in BMD. Chromosomal loci including 1p36, 2p22-25, 11q12-13, parathyroid hormone receptor type 1 (PTHR1), interleukin-6 (IL-6), interleukin 1 alpha (IL-1α) and type II collagen A1/vitamin D receptor (COL11A1/VDR) have been linked or shown suggestive linkage with BMD in other populations. To determine whether these loci predispose to low BMD in the Irish population, we investigated 24 microsatellite markers at 7 chromosomal loci by linkage studies in 175 Irish families of probands with primary low BMD (T-score ≤ -1.5). Nonparametric analysis was performed using the maximum likelihood variance estimation and traditional Haseman-Elston tests on the Mapmaker/Sibs program. Suggestive evidence of linkage was observed with lumbar spine BMD at 2p22-25 (maximum LOD score 2.76) and 11q12-13 (MLS 2.55). One region, 1p36, approached suggestive linkage with femoral neck BMD (MLS 2.17). In addition, seven markers achieved LOD scores > 1.0, D2S149, D11S1313, D11S987, D11S1314 including those encompassing the PTHR1 (D3S3559, D3S1289) for lumbar spine BMD and D2S149 for femoral neck BMD. Our data suggest that genes within a these chromosomal regions are contributing to a predisposition to low BMD in the Irish population.

Genetics of ankylosing spondylitis

Purpose of Review Over the past 3 years, several new genes and gene deserts have been identified that are associated with ankylosing spondylitis (AS). The purpose of this review is to discuss the major findings of these studies, and the answers they provide and questions they raise about the pathogenesis of this common condition. Recent Findings: Five genes/genetic regions have now definitively been associated with AS [the major histocompatibility complex (MHC), IL23R, ERAP1, 2p15 and 21q22]. Strong evidence to support association with the disease has been demonstrated for the genes IL1R2, ANTXR2, TNFSF15, TNFR1 and a region on chromosome 16q including the gene TRADD. There is an overrepresentation of genes involved in Th17 lymphocyte differentiation/activation among genes associated with AS and the related diseases inflammatory bowel disease and psoriasis, pointing strongly to this pathway as playing a major causative role in the disease. Increasing information about differential association of HLA-B27 subtypes with disease suggests a hierarchy of strength of association of those alleles with AS, providing a useful test as to the validity of different potential mechanisms of association of HLA-B27 with AS. The mechanism underlying the association of the gene deserts, 2p15 and 21q22, suggests the involvement of noncoding RNA in AS etiopathogenesis. Summary: The increasing list of genes identified as being definitely involved in AS provides a useful platform for hypothesis-driven research in the field, providing a potential alternative route to determining the underlying mechanisms involved in the disease to research focusing on HLA-B27 alone.

After 10 years of clinical trials with liraglutide, do we know whether it is beneficial in diabetes?

Type 2 diabetes remains an escalating world-wide problem, despite a range of treatments. The revelation that insulin secretion is under the control of a gut hormone, glucagon-like peptide 1 (GLP-1), led to a new paradigm in the management of type 2 diabetes. Liraglutide is a long acting GLP-1 receptor agonist used in the treatment of type 2 diabetes. The review considers the clinical trials with liraglutide. There are many comparator trials between liraglutide and other medicines for the treatment of type 2 diabetes, and these trials have shown that liraglutide lowers HbA1c and body weight, and is well tolerated. A large cardiovascular safety trial with liraglutide is presently being undertaken. After 10 years of clinical trials with liraglutide, we do not know whether liraglutide has cardiovascular safety in subjects with type 2 diabetes and high cardiovascular risk. Although this is not a requirement for registration by the Food and Drug Administration (FDA), in my opinion, they should reconsider this. We also do not presently know whether liraglutide has any beneficial effects on clinical cardiovascular outcomes.

Dual acquisition of 18F-FMISO and 18F-FDOPA

Metabolic imaging using positron emission tomography (PET) has found increasing clinical use for the management of infiltrating tumours such as glioma. However, the heterogeneous biological nature of tumours and intrinsic treatment resistance in some regions means that knowledge of multiple biological factors is needed for effective treatment planning. For example, the use of 18F-FDOPA to identify infiltrative tumour and 18F-FMISO for localizing hypoxic regions. Performing multiple PET acquisitions is impractical in many clinical settings, but previous studies suggest multiplexed PET imaging could be viable. The fidelity of the two signals is affected by the injection interval, scan timing and injected dose. The contribution of this work is to propose a framework to explicitly trade-off signal fidelity with logistical constraints when designing the imaging protocol. The particular case of estimating 18F-FMISO from a single frame prior to injection of 18F-FDOPA is considered. Theoretical experiments using simulations for typical biological scenarios in humans demonstrate that results comparable to a pair of single-tracer acquisitions can be obtained provided protocol timings are carefully selected. These results were validated using a pre-clinical data set that was synthetically multiplexed. The results indicate that the dual acquisition of 18F-FMISO and 18F-FDOPA could be feasible in the clinical setting. The proposed framework could also be used to design protocols for other tracers.

New evidence of the reproductive organs of Glossopteris based on permineralized fossils from Queensland, Australia. II: pollen-bearing organ Ediea gen. nov

Ediea homevalensis H. Nishida, Kudo, Pigg & Rigby gen. et sp. nov. is proposed for permineralized pollen-bearing structures from the Late Permian Homevale Station locality of the Bowen Basin, Queensland, Australia. The taxon represents unisexual fertile shoots bearing helically arranged leaves on a central axis. The more apical leaves are fertile microsporophylls bearing a pair of multi-branched stalks on their adaxial surfaces that each supports a cluster of terminally borne pollen sacs. Proximal to the fertile leaves there are several rows of sterile scale-like leaves. The pollen sacs (microsporangia) have thickened and dark, striate walls that are typical of the Arberiella type found in most pollen organs presumed to be of glossopterid affinity. An examination of pollen organs at several developmental stages, including those containing in situ pollen of the Protohaploxypinus type, provides the basis for a detailed analysis of these types of structures, which bear similarities to both compression/impression Eretmonia-type glossopterid microsporangiate organs and permineralized Eretmonia macloughlinii from Antarctica. These fossils demonstrate that at least some Late Permian pollen organs were simple microsporophyll-bearing shoot systems and not borne directly on Glossopteris leaves.

Temporal and environmental influences on the vocal behaviour of a nocturnal bird

Temporal and environmental variation in vocal activity can provide information on avian behaviour and call function not available to short-term experimental studies. Intersexual differences in this variation can provide insight into selection effects. Yet factors influencing vocal behaviour have not been assessed in many birds, even those monitored by acoustic methods. This applies to the New Zealand kiwi (Apterygidae), for which call censuses are used extensively in conservation monitoring, yet which have poorly understood acoustic ecology. We investigated little spotted kiwi Apteryx owenii vocal behaviour over 3 yr, measuring influences on vocal activity in both sexes from time of night, season, weather conditions and lunar cycle. We tested hypotheses that call rate variation reflects call function, foraging efficiency, historic predation risk and variability in sound transmission, and that there are inter-sexual differences in call function. Significant seasonal variation showed that vocalisations were important in kiwi reproduction, and inter-sexual synchronisation of call rates indicated that contact, pair-bonding or resource defence were key functions. All weather variables significantly affected call rates, with elevated calling during increased humidity and ground moisture indicating a relation between vocal activity and foraging conditions. A significant decrease in calling activity on cloudy nights, combined with no moonlight effect, suggests an impact of light pollution in this species. These influences on vocal activity provide insight into kiwi call function, have direct consequences for conservation monitoring of kiwi, and have wider implications in understanding vocal behaviour in a range of nocturnal birds

A model to explain specific cellular communications and cellular harmony: - a hypothesis of coupled cells and interactive coupling molecules

Background The various cell types and their relative numbers in multicellular organisms are controlled by growth factors and related extracellular molecules which affect genetic expression pathways. However, these substances may have both/either inhibitory and/or stimulatory effects on cell division and cell differentiation depending on the cellular environment. It is not known how cells respond to these substances in such an ambiguous way. Many cellular effects have been investigated and reported using cell culture from cancer cell lines in an effort to define normal cellular behaviour using these abnormal cells. A model is offered to explain the harmony of cellular life in multicellular organisms involving interacting extracellular substances. Methods A basic model was proposed based on asymmetric cell division and evidence to support the hypothetical model was accumulated from the literature. In particular, relevant evidence was selected for the Insulin-Like Growth Factor system from the published data, especially from certain cell lines, to support the model. The evidence has been selective in an attempt to provide a picture of normal cellular responses, derived from the cell lines. Results The formation of a pair of coupled cells by asymmetric cell division is an integral part of the model as is the interaction of couplet molecules derived from these cells. Each couplet cell will have a receptor to measure the amount of the couplet molecule produced by the other cell; each cell will be receptor-positive or receptor-negative for the respective receptors. The couplet molecules will form a binary complex whose level is also measured by the cell. The hypothesis is heavily supported by selective collection of circumstantial evidence and by some direct evidence. The basic model can be expanded to other cellular interactions. Conclusions These couplet cells and interacting couplet molecules can be viewed as a mechanism that provides a controlled and balanced division-of-labour between the two progeny cells, and, in turn, their progeny. The presence or absence of a particular receptor for a couplet molecule will define a cell type and the presence or absence of many such receptors will define the cell types of the progeny within cell lineages.

Using quantitative wastewater analysis to measure daily usage of conventional and emerging illicit drugs at an annual music festival

Introduction and Aims Wastewater analysis provides a non-intrusive way of measuring drug use within a population. We used this approach to determine daily use of conventional illicit drugs [cannabis, cocaine, methamphetamine and 3,4-methylenedioxymethamphetamine (MDMA)] and emerging illicit psychostimulants (benzylpiperazine, mephedrone and methylone) in two consecutive years (2010 and 2011) at an annual music festival. Design and Methods Daily composite wastewater samples, representative of the festival, were collected from the on-site wastewater treatment plant and analysed for drug metabolites. Data over 2 years were compared using Wilcoxon matched-pair test. Data from 2010 festival were compared with data collected at the same time from a nearby urban community using equivalent methods. Results Conventional illicit drugs were detected in all samples whereas emerging illicit psychostimulants were found only on specific days. The estimated per capita consumption of MDMA, cocaine and cannabis was similar between the two festival years. Statistically significant (P < 0.05; Z = −2.0–2.2) decreases were observed in use of methamphetamine and one emerging illicit psychostimulant (benzyl piperazine). Only consumption of MDMA was elevated at the festival compared with the nearby urban community. Discussion and Conclusions Rates of substance use at this festival remained relatively consistent over two monitoring years. Compared with the urban community, drug use among festival goers was only elevated for MDMA, confirming its popularity in music settings. Our study demonstrated that wastewater analysis can objectively capture changes in substance use at a music setting without raising major ethical issues. It would potentially allow effective assessments of drug prevention strategies in such settings in the future.

A Model to Estimate the Population Contributing to the Wastewater Using Samples Collected on Census Day

An important uncertainty when estimating per capita consumption of, for example, illicit drugs by means of wastewater analysis (sometimes referred to as “sewage epidemiology”) relates to the size and variability of the de facto population in the catchment of interest. In the absence of a day-specific direct population count any indirect surrogate model to estimate population size lacks a standard to assess associated uncertainties. Therefore, the objective of this study was to collect wastewater samples at a unique opportunity, that is, on a census day, as a basis for a model to estimate the number of people contributing to a given wastewater sample. Mass loads for a wide range of pharmaceuticals and personal care products were quantified in influents of ten sewage treatment plants (STP) serving populations ranging from approximately 3500 to 500 000 people. Separate linear models for population size were estimated with the mass loads of the different chemical as the explanatory variable: 14 chemicals showed good, linear relationships, with highest correlations for acesulfame and gabapentin. De facto population was then estimated through Bayesian inference, by updating the population size provided by STP staff (prior knowledge) with measured chemical mass loads. Cross validation showed that large populations can be estimated fairly accurately with a few chemical mass loads quantified from 24-h composite samples. In contrast, the prior knowledge for small population sizes cannot be improved substantially despite the information of multiple chemical mass loads. In the future, observations other than chemical mass loads may improve this deficit, since Bayesian inference allows including any kind of information relating to population size.

Loading compliance of static load bearing exercises performed by transfemoral amputees fitted with an osseointegrated implant

Background To date bone-anchored prostheses are used to alleviate the concerns caused by socket suspended prostheses and to improve the quality of life of transfemoral amputees (TFA). Currently, two implants are commercially available (i.e., OPRA (Integrum AB, Sweden), ILP (Orthodynamics GmbH, Germany)). [1-17]The success of the OPRA technique is codetermined by the rehabilitation program. TFA fitted with an osseointegrated implant perform progressive mechanical loading (i.e. static load bearing exercises (LBE)) to facilitate bone remodelling around the implant.[18, 19] Aim This study investigated the trustworthiness of monitoring the load prescribed (LP) during experimental static LBEs using the vertical force provided by a mechanical bathroom scale that is considered a surrogate of the actual load applied. Method Eleven unilateral TFAs fitted with an OPRA implant performed five trials in four loading conditions. The forces and moments on the three axes of the implant were measured directly with an instrumented pylon including a six-channel transducer. The “axial” and “vectorial” comparisons corresponding to the difference between the force applied on the long axis of the fixation and LP as well as the resultant of the three components of the load applied and LP, respectively were analysed Results For each loading condition, Wilcoxon One-Sample Signed Rank Tests were used to investigate if significant differences (p<0.05) could be demonstrated between the force applied on the long axis and LP, and between the resultant of the force and LP. The results demonstrated that the raw axial and vectorial differences were significantly different from zero in all conditions (p<0.05), except for the vectorial difference for the 40 kg loading condition (p=0.182). The raw axial difference was negative for all the participants in every loading condition, except for TFA03 in the 10 kg condition (11.17 N). Discussion & Conclusion This study showed a significant lack of axial compliance. The load applied on the long axis was significantly smaller than LP in every loading condition. This led to a systematic underloading of the long axis of the implant during the proposed experimental LBE. Monitoring the vertical force might be only partially reflective of the actual load applied, particularly on the long axis of the implant.