Plant communities of the montane mesophilous grasslands (Polygono bistortae–Trisetion flavescentis alliance) in central Europe: Formalized classification and syntaxonomical revision

Abstract

The research conducted here presents a syntaxonomical revision of the montane mesophilous meadows of the Polygono bistortae-Trisetion flavescentis alliance in central Europe – the Slovak part of the western and eastern Carpathians. These typical semi-natural grasslands occur mainly as small islands over the calcareous bedrocks. Associations of this alliance have tight relationships to the Arrhenatherion, Mesobromion and Nardo-Agrostion alliances. Formal definitions based on combination of the species groups were used regarding the diversity and geographical range of the Polygono bistortae-Trisetion flavescentis alliance. Following the formal definitions from the eight associations reported previously for Slovakia, only four of them can be placed within the Polygono bistortae-Trisetion flavescentis: Campanulo glomeratae-Geranietum sylvatici, Geranio sylvatici-Trisetetum, Crepido mollis-Agrostietum capillaries and Geranio-Alchemilletum crinitae. A comparison of traditional and formalized classification, and the advantages/disadvantages of the formalized classification are discussed.     

C/EBP homologous protein drives pro-catabolic responses in chondrocytes

Introduction

Excess C/EBP homologous protein (CHOP) expression is one feature of the unfolded protein response (UPR) to endoplasmic reticulum (ER) stress. Here, we focused on CHOP expression and function in chondrocytes.

Methods

We studied human knee osteoarthritis (OA) cartilage, bovine chondrocytes cultured in alginate and subjected to sub-lethal biomechanical injury, and knee chondrocytes of human autopsy donors. We performed siRNA knockdown and transfection.

Results

UPR activation was increased in human knee OA cartilage in situ, and in biomechanically injured cultured chondrocytes in vitro. In normal human chondrocytes, CHOP “gain of function” sensitized chondrocytes to IL-1β induced nitric oxide (NO) and matrix metalloproteinase (MMP)-3 release without inducing these responses by itself. Excess CHOP expression, by itself, induced superoxide production and apoptosis. Conversely, siRNA knockdown of CHOP and the UPR-specific mediator X-box binding protein (XBP1) inhibited NO release by >80% (P <0.0005) in response to IL-1β, and blunted MMP-3 release, whereas there were only minimal effects of the UPR mediator GRP78 on these responses. The anti-inflammatory metabolic “super-regulator” AMP kinase (AMPK) is known to limit UPR activation in vascular muscle cells. Here, CHOP supported the capacity of IL-1β to suppress AMPK activity in chondrocytes. We also observed that inhibition of AMPK activity promoted an increase in chondrocyte CHOP expression. Conversely, pharmacologic activation of AMPK by 5-Aminoimidazole-4-carboxamide ribonucleotide (AICAR) blunted chondrocyte CHOP expression in response to biomechanical injury.

Conclusions

Biomechanical injury and IL-1 signaling stimulate UPR activation in chondrocytes. CHOP mediates chondrocyte catabolic and apoptotic responses to IL-1β, and does so partly by inhibiting AMPK activity. Conversely, development of excess CHOP activity is limited by AMPK activity in chondrocytes. Our findings suggest a mechanism for potential chondroprotection by AICAR and other AMPK activators. The work is of translational relevance for OA, since several drugs that activate AMPK are already in the clinic for arthritis (for example, allosteric AMPK activators sodium salicylate and high dose aspirin, and methotrexate, which activates AMPK by generating AICAR).     

Microwell bioreactor system for cell-based high throughput proliferation and cytotoxicity assays

Cell-based high throughput proliferation and cytotoxicity assays are increasingly used in drug screening and bioprocess development. However, online monitoring of cell proliferation, pH, and dissolved oxygen (DO) has been a challenge in 3D cell-based assays. In this work, a 40-microwell bioreactor (40-MBR) system was developed from a 384-well plate for real-time, noninvasive monitoring of pH, DO, and cell proliferation in 3D microenvironments. Chinese hamster ovary (CHO) and MCF-07 breast cancer cells cultured in 40-MBR confirmed that the 40-MBR was capable of simultaneously monitoring DO and cell proliferation based on culture fluorescence and pH by measuring the absorbance of phenol red. Cytotoxicity studies of sodium butyrate on CHO cells demonstrated that 40-MBR with dynamic background fluorescence correction gave more reliable and highly reproducible growth kinetic data compared to conventional multiwells with static background correction. Furthermore, the dosage effects of two new anticancer drug candidates, 5,7-dihydroxy-2-(4-hydroxyphenyl)-8-[(E)-2-phenylethenyl]-3,4-dihydro-2H-1-benzopyran-4-one (DH-8P-DB) and 5,7-dihydroxy-2-(4-hydroxyphenyl)-6-[(E)-2-phenylethenyl]-3,4-dihydro-2H-1-benzopyran-4-one (DH-6P-DB), on HT-29 colon cancer cells were assessed using the 40-MBR, and the results indicated that DH-6P-DB would be a more potent drug in treating colon cancer than DH-8P-DB. These studies demonstrated that 40-MBR could serve as a high throughput platform for screening potential cancer drugs in early-stage drug discovery.     

Cell-based high-throughput proliferation and cytotoxicity assays for screening traditional Chinese herbal medicines

A simple, reliable, high-throughput screening method was developed and used to assess the pharmaceutical effects of extracts of traditional Chinese herbal medicines (TCHMs). This method is based on 3-dimensional (3-D) cultures of mouse embryonic stem (mES) and human colon cancer and breast cancer cells expressing enhanced green fluorescent protein (EGFP) in polyethylene terephthalate (PET) fibrous scaffolds on modified 384-well plates with online monitoring of culture fluorescence for dynamic responses of cells to drugs present in culture media. Cell responses to deoxycholic acid and the extracts of 3 TCHMs (Ganoderma lucidum spores, Ginkgo biloba, and Epimedium brevicornum) at various concentrations were investigated for their effects on proliferation and cytotoxicity. The screening results, i.e., the growth responses of cancer cells to those drugs, were consistent with what have been reported in the literature, confirming the reliability of the new screening approach. Different from previous screening methods for both TCHMs and western medicines that used animal models or 2-D cell-based assays with single cell lines, this 3-D cell-based screening method employs both cancer and normal cells and thereby provides a way for quick, direct evaluation of the anticancer effects of TCHMs. This method also offers assessment on the side effects of TCHMs.     

Rice Performance and Water Use Efficiency under Plastic Mulching with Drip Irrigation

Plastic mulching with drip irrigation is a new water-saving rice cultivation technology, but little is known on its productivity and water-saving capacity. This study aimed to assess the production potential, performance, and water use efficiency (WUE) of rice under plastic mulching with drip irrigation. Field experiments were conducted over 2 years with two rice cultivars under different cultivation systems: conventional flooding (CF), non-flooded irrigation incorporating plastic mulching with furrow irrigation (FIM), non-mulching with furrow irrigation (FIN), and plastic mulching with drip irrigation (DI). Compared with the CF treatment, grain yields were reduced by 31.76–52.19% under the DI treatment, by 57.16–61.02% under the FIM treatment, by 74.40–75.73% under the FIN treatment, which were mainly from source limitation, especially a low dry matter accumulation during post-anthesis, in non-flooded irrigation. WUE was the highest in the DI treatment, being 1.52–2.12 times higher than with the CF treatment, 1.35–1.89 times higher than with the FIM treatment, and 2.37–3.78 times higher than with the FIN treatment. The yield contribution from tillers (YCFTs) was 50.65–62.47% for the CF treatment and 12.07–20.62% for the non-flooded irrigation treatments. These low YCFTs values were attributed to the poor performance in tiller panicles rather than the total tiller number. Under non-flooded irrigation, root length was significantly reduced with more roots distributed in deep soil layers compared with the CF treatment; the DI treatment had more roots in the topsoil layer than the FIM and FIN treatments. The experiment demonstrates that the DI treatment has greater water saving capacity and lower yield and economic benefit gaps than the FIM and FIN treatments compared with the CF treatment, and would therefore be a better water-saving technology in areas of water scarcity.     

Does Serum 25-Hydroxyvitamin D Influence Muscle Development during Puberty in Girls? - A 7-Year Longitudinal Study

Vitamin D is well known for its regulatory role in calcium and phosphate homeostasis, but its role in muscle mass and strength during growth remains inconclusive. We explored the association of serum 25-hydroxyvitamin D (25(OH)D) with muscle development in girls from 11 to 18-years old. Whole body lean tissue mass (LM_WB), appendicular lean mass (aLM), muscle cross-sectional area at the lower leg (mCSA), maximal voluntary contraction of elbow flexors (MVC_elbow) and knee extensors (MVC_knee) were assessed in 217 girls aged 10–13 years (at baseline), 215 in 2-year and 226 in 7.5-year follow-up. Serum concentration of 25(OH)D and intact parathyroid hormone (PTH) were analyzed retrospectively and girls were categorized according to their 25(OH)D levels (consistently insufficient 25(OH)D G_LL <50 nmol/l and consistently sufficient G_HH >50 nmol/l from baseline to 7-year follow-up). We found that 25(OH)D level declined until menarche (p<0.05) while LM_WB, aLM, mCSA, MVC_elbow and MVC_knee continued to increase (p<0.001 for all) post menarche. At pre-menarche, the G_LL (n = 34) had higher LM_WB and aLM than the G_HH (n = 21, p<0.05), while post-menarche the G_HH (n = 15) had a greater catch-up gain in LM_WB (p = 0.004), aLM (p = 0.001) and mCSA (p = 0.027) compared to the G_LL (n = 65) over the first 2-year period. At the age of 18, no differences in muscle mass/strength between the low (n = 151) and high (n = 77) levels of 25(OH)D groups were found. This finding was independent of vitamin D receptor genotype and other confounders. In conclusion, our results showed that levels of 25(OH)D have no significant negative influence on the development of muscle mass and strength during pubertal growth both with longitudinal and cross-sectional comparison. On the contrary, our results suggest that the temporary negative association between 25(OH)D and muscle mass arises as a consequence of fast growth prior to menarche, and this negative association is diminished through catch-up growth after menarche.     

Can Yeast (S. cerevisiae) Metabolic Volatiles Provide Polymorphic Signaling?

Chemical signaling between organisms is a ubiquitous and evolutionarily dynamic process that helps to ensure mate recognition, location of nutrients, avoidance of toxins, and social cooperation. Evolutionary changes in chemical communication systems progress through natural variation within the organism generating the signal as well as the responding individuals. A promising yet poorly understood system with which to probe the importance of this variation exists between D. melanogaster and S. cerevisiae. D. melanogaster relies on yeast for nutrients, while also serving as a vector for yeast cell dispersal. Both are outstanding genetic and genomic models, with Drosophila also serving as a preeminent model for sensory neurobiology. To help develop these two genetic models as an ecological model, we have tested if - and to what extent - S. cerevisiae is capable of producing polymorphic signaling through variation in metabolic volatiles. We have carried out a chemical phenotyping experiment for 14 diverse accessions within a common garden random block design. Leveraging genomic sequences for 11 of the accessions, we ensured a genetically broad sample and tested for phylogenetic signal arising from phenotypic dataset. Our results demonstrate that significant quantitative differences for volatile blends do exist among S. cerevisiae accessions. Of particular ecological relevance, the compounds driving the blend differences (acetoin, 2-phenyl ethanol and 3-methyl-1-butanol) are known ligands for D. melanogasters chemosensory receptors, and are related to sensory behaviors. Though unable to correlate the genetic and volatile measurements, our data point clear ways forward for behavioral assays aimed at understanding the implications of this variation.     

Clinical Characteristics Associated with Borrelia burgdorferi Sensu Lato Skin Culture Results in Patients with Erythema Migrans

Clinical characteristics associated with isolation of Borrelia burgdorferi sensu lato from skin have not been fully evaluated. To gain insight into predictors for a positive EM skin culture, we compared basic demographic, epidemiologic, and clinical data in 608 culture-proven and 501 culture-negative adult patients with solitary EM. A positive Borrelia spp. skin culture was associated with older age, a time interval of >2 days between tick bite and onset of the skin lesion, EM ≥5 cm in diameter, and location of the lesion on the extremities, whereas several other characteristics used as clinical case definition criteria for the diagnosis of EM (such as tick bite at the site of later EM, information on expansion of the skin lesion, central clearing) were not. A patient with a 15-cm EM lesion had almost 3-fold greater odds for a positive skin culture than patients with a 5-cm lesion. Patients with a free time interval between the tick bite and onset of EM had the same probability of a positive skin culture as those who did not recall a tick bite (OR=1.02); however, the two groups had >3-fold greater odds for EM positivity than patients who reported a tick bite with no interval between the bite and onset of the lesion. In conclusion, several yet not all clinical characteristics used in EM case definitions were associated with positive Borrelia spp. skin culture. The findings are limited to European patients with solitary EM caused predominantly by B. afzelii but may not be valid for other situations.     

The Effects of Catalyst Layer Deposition Methodology on Electrode Performance

The catalyst layer of the cathode is arguably the most critical component of low‐temperature fuel cells and carbon dioxide (CO₂) electrolysis cells because their performance is typically limited by slow oxygen (O₂) and CO₂ reduction kinetics. While significant efforts have focused on developing cathode catalysts with improved activity and stability, fewer efforts have focused on engineering the catalyst layer structure to maximize catalyst utilization and overall electrode and system performance. Here, we study the performance of cathodes for O₂ reduction and CO₂ reduction as a function of three common catalyst layer preparation methods: hand‐painting, air‐brushing, and screen‐printing. We employed ex‐situ X‐ray micro‐computed tomography (MicroCT) to visualize the catalyst layer structure and established data processing procedures to quantify catalyst uniformity. By coupling structural analysis with in‐situ electrochemical characterization, we directly correlate variation in catalyst layer morphology to electrode performance. MicroCT and SEM analyses indicate that, as expected, more uniform catalyst distribution and less particle agglomeration, lead to better performance. Most importantly, the analyses reported here allow for the observed differences over a large geometric volume as a function of preparation methods to be quantified and explained for the first time. Depositing catalyst layers via a fully‐automated air‐brushing method led to a 56% improvement in fuel cell performance and a significant reduction in electrode‐to‐electrode variability. Furthermore, air‐brushing catalyst layers for CO₂ reduction led to a 3‐fold increase in partial CO current density and enhanced product selectivity (94% CO) at similar cathode potential but a 10‐fold decrease in catalyst loading as compared to previous reports.