Reliability of computerized image analysis for the evaluation of serial synovial biopsies in randomized controlled trials in rheumatoid arthritis

Analysis of biomarkers in synovial tissue is increasingly used in the evaluation of new targeted therapies for patients with rheumatoid arthritis (RA). This study determined the intrarater and inter-rater reliability of digital image analysis (DIA) of synovial biopsies from RA patients participating in clinical trials. Arthroscopic synovial biopsies were obtained before and after treatment from 19 RA patients participating in a randomized controlled trial with prednisolone. Immunohistochemistry was used to detect CD3⁺ T cells, CD38⁺ plasma cells and CD68⁺ macrophages. The mean change in positive cells per square millimetre for each marker was determined by different operators and at different times using DIA. Nonparametric tests were used to determine differences between observers and assessments, and to determine changes after treatment. The intraclass correlations (ICCs) were calculated to determine the intrarater and inter-rater reliability. Intrarater ICCs showed good reliability for measuring changes in T lymphocytes (R = 0.87), plasma cells (R = 0.62) and macrophages (R = 0.73). Analysis by Bland–Altman plots showed no systemic differences between measurements. The smallest detectable changes were calculated and their discriminatory power revealed good response in the prednisolone group compared with the placebo group. Similarly, inter-rater ICCs also revealed good reliability for measuring T lymphocytes (R = 0.68), plasma cells (R = 0.69) and macrophages (R = 0.72). All measurements identified the same cell types as changing significantly in the treated patients compared with the placebo group. The measurement of change in total positive cell numbers in synovial tissue can be determined reproducibly for various cell types by DIA in RA clinical trials.     

The Role of Tryptophan Catabolism along the Kynurenine Pathway in Acute Ischemic Stroke

Post-stroke inflammation may induce upregulation of the kynurenine (KYN) pathway for tryptophan (TRP) oxidation, resulting in neuroprotective (kynurenic acid, KA) and neurotoxic metabolites (3-hydroxyanthranillic acid, 3-HAA). We investigated whether activity of the kynurenine pathway in acute ischemic stroke is related to initial stroke severity, long-term stroke outcome and the ischemia-induced inflammatory response. Plasma concentrations of TRP and its metabolites were measured in 149 stroke patients at admission, at 24 h, at 72 h and at day 7 after stroke onset. We evaluated the relation between the KYN/TRP ratio, the KA/3-HAA ratio and stroke severity, outcome and inflammatory parameters (C-reactive protein (CRP), erythrocyte sedimentation rate (ESR) and neutrophil/lymphocyte ratio (NLR)). KYN/TRP but not KA/3-HAA correlated with the NIHSS score and with the infarct volume. Patients with poor outcome had higher mean KYN/TRP ratios than patients with more favourable outcome. The KYN/TRP ratio at admission correlated with CRP levels, ESR and NLR. The activity of the kynurenine pathway for tryptophan degradation in acute ischemic stroke correlates with stroke severity and long-term stroke outcome. Tryptophan oxidation is related to the stroke-induced inflammatory response.     

Nanomolar affinity,iminosugar-based chemical probes for specific labeling of lysosomal glucocerebrosidase

Three different photoprobes were synthesized to label β-glucosidases; one probe was based on glucose, two probes on the iminosugar deoxynojirimycin. The affinity of the probes for three different β-glucosidases was determined. Furthermore, their labeling efficiencies, binding specificities through competition with deoxynojirimycin, and binding specificities in the presence of cell lysate, were evaluated. Especially one showed very high affinity towards non-lysosomal glucoceramidase (IC₅₀ = 20 nM).     

Moss Responses to Elevated CO2 and Variation in Hydrology in a Temperate Lowland Peatland

We studied the effects of elevated CO₂ (180–200 ppmv above ambient) on growth and chemistry of three moss species (Sphagnum palustre, S. recurvum and Polytrichum commune) in a lowland peatland in the Netherlands. Thereto, we conducted both a greenhouse experiment with both Sphagnum species and a field experiment with all three species using MiniFACE (Free Air CO₂ Enrichment) technology during 3 years. The greenhouse experiment showed that Sphagnum growth was stimulated by elevated CO₂ in the short term, but that in the longer term (≥1 year) growth was probably inhibited by low water tables and/or down-regulation of photosynthesis. In the field experiment, we did not find significant changes in moss abundance in response to elevated CO₂, although CO₂ enrichment appeared to reduce S. recurvum abundance. Both Sphagnum species showed stronger responses to spatial variation in hydrology than to increased atmospheric CO₂ concentrations. Polytrichum was insensitive to changes in hydrology. Apart from the confounding effects of hydrology, the relative lack of growth response of the moss species may also have been due to the relatively small increase in assimilated CO₂ as achieved by the experimentally added CO₂. We calculated that the added CO₂ contributed at most 32% to the carbon assimilation of the mosses, while our estimates based on stable C isotope data even suggest lower contributions for Sphagnum (24–27%). Chemical analyses of the mosses showed only small elevated CO₂ effects on living tissue N concentration and C/N ratio of the mosses, but the C/N ratio of Polytrichum was substantially lower than those of the Sphagnum species. Continuing expansion of Polytrichum at the expense of Sphagnum could reduce the C sink function of this lowland Sphagnum peatland, and similar ones elsewhere, as litter decomposition rates would probably be enhanced. Such a reduction in sink function would be driven mostly by increased atmospheric N deposition, water table regulation for agricultural purposes and land management to preserve the early successional stage (mowing, tree and shrub removal), since these anthropogenic factors will probably exert a greater control on competition between Polytrichum and Sphagnum than increased atmospheric CO₂ concentrations.     

Plant Performance in a Warmer World: General Responses of Plants from Cold, Northern Biomes and the Importance of Winter and Spring Events

During the past three decades the Earth has warmed with a rate unprecedented during the past 1000 years. There is already ample evidence that this fast climate warming has affected a broad range of organisms, including plants. Plants from high-latitude and high-altitude sites (‘cold biomes’) are especially sensitive to climate warming. In this paper we (1) review the response in the phenology of plants, changes in their range and distribution, soil nutrient availability, and the effects on the structure and dynamics of plant communities for cold, northern biomes; and (2) we show, by using data from an ongoing snow and temperature manipulation experiment in northern Sweden, that also winter and spring events have a profound influence on plant performance. Both long-term phenological data sets, experimental warming studies (performed in summer or year-round), natural gradient studies and satellite images show that key phenological events are responsive to temperature increases and that recent climate warming does indeed lead to changes in plant phenology. However, data from a warming and snow manipulation study that we are conducting in northern Sweden show that plants respond differently to the various climatic scenarios that we had imposed on these species and that especially winter and spring events have a profound impact. This indicates that it is necessary to include several scenarios of both summer and winter climate change in experimental climate change studies, and that we need detailed projections of future climate at a regional scale to be able to assess their impacts on natural ecosystems. There is also ample evidence that the range shift of herbs and shrubs to more northern regions is for the vast majority of species mainly caused by changes in the climate. This is in line with the observed ‘up-greening’ of northern tundra sites. These rapid northern shifts in distribution of plants as a result of climate warming may have substantial consequences for the structure and dynamics of high-latitude ecosystems. An analysis of warming studies at 9 tundra sites shows that heating during at least 3 years increased net N-mineralization from 0.32±0.31 (SE) g N m⁻² yr⁻¹ in the controls to 0.53±0.31 (SE) g N m⁻² yr⁻¹ in the heated plots (p<0.05), an increase of about 70%. Thus, warming leads to higher N availability in high-latitude northern tundra sites, but the variability is substantial. Higher nutrient availability affects in turn the species composition of high-latitude sites, which has important consequences for the carbon and water balance of these systems.     

Neuropsychological and behavioural correlates of CSF biomarkers in dementia

To improve clinical, neuropsychological and behavioural characterisation of the cerebrospinal fluid (CSF) biomarkers beta-amyloid((1-42)) protein (Abeta42), protein tau (tau) and tau phosphorylated at threonine 181 (P-tau181) across diagnostic dementia categories, a prospective study was set up. Patients with probable Alzheimer's disease (AD) (n=201), AD with cerebrovascular disease (CVD) (AD+CVD) (n=33), frontotemporal dementia (FTD) (n=27), dementia with Lewy bodies (DLB) (n=22) and healthy controls (n=148) were included. All patients underwent neuropsychological examination and behavioural assessment by means of a battery of behavioural assessment scales. CSF was obtained by lumbar puncture and levels of Abeta42, tau and P-tau181 were determined with commercially available ELISA kits. Negative correlations between CSF Abeta42 levels and aggressiveness (Spearman: r=-0.223; p=0.002) and positive correlations with age at inclusion (r=0.195; p=0.006), age at onset (r=0.205; p=0.003) and MMSE scores (r=0.198; p=0.005) were found in AD. In AD+CVD, CSF Abeta42 levels were correlated with MMSE (r=0.482; p=0.006), Hierarchic Dementia Scale (r=0.503; p=0.017) and Boston Naming Test (r=0.516; p=0.012) scores. In controls, age was positively correlated with CSF tau (r=0.465; p<0.001) and P-tau181 levels (r=0.312; p<0.001). CSF tau and P-tau181 levels correlated significantly in all groups, whereas CSF Abeta42 correlated with tau and P-tau181 levels in healthy controls only. Negative correlations between CSF Abeta42 levels and aggressiveness were found in AD patients. CSF Abeta42 seems to be a stage marker for AD (+/-CVD) given the positive correlations with neuropsychological test results suggesting that CSF Abeta42 might be of help for monitoring disease progression. Different correlations between age and CSF biomarker levels were obtained in healthy controls compared to AD patients, indicating that AD-induced pathophysiological processes change age-dependent regulation of CSF biomarker levels.     

Nitrogen supply differentially affects litter decomposition rates and nitrogen dynamics of sub-arctic bog species

High-latitude peatlands are important soil carbon sinks. In these ecosystems, the mineralization of carbon and nitrogen are constrained by low temperatures and low nutrient concentrations in plant litter and soil organic matter. Global warming is predicted to increase soil N availability for plants at high-latitude sites. We applied N fertilizer as an experimental analogue for this increase. In a three-year field experiment we studied N fertilization effects on leaf litter decomposition and N dynamics of the four dominant plant species (comprising >75% of total aboveground biomass) in a sub-arctic bog in northern Sweden. The species were Empetrum nigrum (evergreen shrub), Eriophorum vaginatum (graminoid), Betula nana (deciduous shrub) and Rubus chamaemorus (perennial forb). In the controls, litter mass loss rates increased in the order: Empetrum < Eriophorum < Betula < Rubus. Increased N availability had variable, species-specific effects: litter mass loss rates (expressed per unit litter mass) increased in Empetrum, did not change in Eriophorum and Betula and decreased in Rubus. In the leaf litter from the controls, we measured no or only slight net N mineralization even after three years. In the N-fertilized treatments we found strong net N immobilization, especially in Eriophorum and Betula. This suggests that, probably owing to substantial chemical and/or microbial immobilization, additional N supply does not increase the rate of N cycling for at least the first three years.     

Foliar pH as a new plant trait: can it explain variation in foliar chemistry and carbon cycling processes among subarctic plant species and types?

Plant traits have become popular as predictors of interspecific variation in important ecosystem properties and processes. Here we introduce foliar pH as a possible new plant trait, and tested whether (1) green leaf pH or leaf litter pH correlates with biochemical and structural foliar traits that are linked to biogeochemical cycling; (2) there is consistent variation in green leaf pH or leaf litter pH among plant types as defined by nutrient uptake mode and higher taxonomy; (3) green leaf pH can predict a significant proportion of variation in leaf digestibility among plant species and types; (4) leaf litter pH can predict a significant proportion of variation in leaf litter decomposability among plant species and types. We found some evidence in support of all four hypotheses for a wide range of species in a subarctic flora, although cryptogams (fern allies and a moss) tended to weaken the patterns by showing relatively poor leaf digestibility or litter decomposability at a given pH. Among seed plant species, green leaf pH itself explained only up to a third of the interspecific variation in leaf digestibility and leaf litter up to a quarter of the interspecific variation in leaf litter decomposability. However, foliar pH substantially improved the power of foliar lignin and/or cellulose concentrations as predictors of these processes when added to regression models as a second variable. When species were aggregated into plant types as defined by higher taxonomy and nutrient uptake mode, green-specific leaf area was a more powerful predictor of digestibility or decomposability than any of the biochemical traits including pH. The usefulness of foliar pH as a new predictive trait, whether or not in combination with other traits, remains to be tested across more plant species, types and biomes, and also in relation to other plant or ecosystem traits and processes.     

A dynamic force analysis system for climbing of large primates

Registering substrate reaction forces from primates during climbing requires the design and construction of customized recording devices. The technical difficulties in constructing a reliable apparatus hinder research on the kinetics of primate locomotion. This is unfortunate since arboreal locomotion, especially vertical climbing, is an important component of the hominoid locomotor repertoire. In this technical paper, we describe a custom-built climbing pole that allows recordings of dynamic 3-dimensional forces during locomotion on horizontal and sloping substrates and during vertical climbing. The pole contains an instrumented section that can readily be modified and enables us to register forces of a single limb or multiple limbs in a broad range of primates. For verification, we constructed a similar set-up (which would not be usable for primates) using a conventional force plate. Data for a human subject walking on both set-ups were compared. The experimental set-up records accurate and reliable substrate reaction forces in three orthogonal directions. Because of its adjustability, this type of modular set-up can be used for a great variety of primate studies. When combining such kinetic measurements together with kinematic information, data of great biomechanical value can be generated. These data will hopefully allow biological anthropologists to answer current questions about primate behaviours on vertical substrates.