Principal factors that determine the extension of detection range in molecular beacon aptamer/conjugated polyelectrolyte bioassays.

A strategy to extend the detection range of weakly-binding targets is reported that takes advantage of fluorescence resonance energy transfer (FRET)-based bioassays based on molecular beacon aptamers (MBAs) and cationic conjugated polyelectrolytes (CPEs). In comparison to other aptamer-target pairs, the aptamer-based adenosine triphosphate (ATP) detection assays are limited by the relatively weak binding between the two partners. In response, a series of MBAs were designed that have different stem stabilities while keeping the constant ATP-specific aptamer sequence in the loop part. The MBAs are labeled with a fluorophore and a quencher at both termini. In the absence of ATP, the hairpin MBAs can be opened by CPEs via a combination of electrostatic and hydrophobic interactions, showing a FRET-sensitized fluorophore signal. In the presence of ATP, the aptamer forms a G-quadruplex and the FRET signal decreases due to tighter contact between the fluorophore and quencher in the ATP/MBA/CPE triplex structure. The FRET-sensitized signal is inversely proportional to [ATP]. The extension of the detection range is determined by the competition between opening of the ATP/MBA G-quadruplex by CPEs and the composite influence by ATP/aptamer binding and the stem interactions. With increasing stem stability, the weak binding of ATP and its aptamer is successfully compensated to show the resistance to disruption by CPEs, resulting in a substantially broadened detection range (from millimolar up to nanomolar concentrations) and a remarkably improved limit of detection. From a general perspective, this strategy has the potential to be extended to other chemical- and biological-assays with low target binding affinity

Role of Staphylococcal Superantigen in Atopic Dermatitis: Influence on Keratinocytes

Staphylococcus aureus may perform an crucial function in atopic dermatitis (AD), via the secretion of superantigens, including staphylococcal enterotoxins (SE) A or B, and toxic shock syndrome toxin-1 (TSST-1). Dysregulated cytokine production by keratinocytes (KCs) upon exposure to staphylococcal superantigens (SsAgs) may be principally involved in the pathophysiology of AD. We hypothesized that lesional KCs from AD may react differently to SsAgs compared to nonlesional skin or normal skin from nonatopics. We conducted a comparison of HLA-DR or CD1a expression in lesional skin as opposed to that in nonlesional or normal skin by immunohistochemistry (IHC). We also compared, using ELISA, the levels of IL-1α, IL-1β, and TNF-α secreted by cultured KCs from lesional, nonlesional, and normal skin, after the addition of SEA, SEB and TSST-1. IHC revealed that both HLA-DR and CD1a expression increased significantly in the epidermis of lesional skin versus nonlesional or normal skin in quite a similar manner. IL-1α, IL-1β, and TNF-α secretion was also significantly elevated in the cultured KCs from lesional skin after the addition of SsAgs. Our results indicated that KCs from lesional skin appear to react differently to SsAgs and increased proinflammatory cytokine production in response to SsAgs may contribute to the pathogenesis of AD

‘Evidence of an auxin signal pathway, microRNA167-ARF8-GH3, and its response to exogenous auxin in cultured rice cells’

MicroRNA167 (miR167) was shown to cleave auxin responsive factor 8 (ARF8) mRNA in cultured rice cells. MiR167 level was found to be controlled by the presence of auxin in the growth medium. When cells grew in auxin-free medium, miR167 level decreased, resulting in an increase in the level of ARF8 mRNA. Cells growing in the normal growth medium containing auxin showed a reversed trend. It was also shown that expression of OsGH3-2, an rice IAA-conjugating enzyme, was positively regulated by ARF8. Delivery of synthesized miR167 into cells led to decrease of both ARF8 mRNA and OsGH3-2 mRNA. This study provides an evidence in which the exogeneous auxin signal is transduced to OsGH3-2 through miR167 and ARF8 in sequence. This proposed auxin signal transduction pathway, auxin-miR167-ARF8-OsGH3-2, could be, in conjunction with the other microRNA-mediated auxin signals, an important one for responding to exogeneous auxin and for determining the cellular free auxin level which guides appropriate auxin responses

Persistent neuronal Ube3a expression in the suprachiasmatic nucleus of Angelman syndrome model mice

Mutations or deletions of the maternal allele of the UBE3A gene cause Angelman syndrome (AS), a severe neurodevelopmental disorder. The paternal UBE3A/Ube3a allele becomes epigenetically silenced in most neurons during postnatal development in humans and mice; hence, loss of the maternal allele largely eliminates neuronal expression of UBE3A protein. However, recent studies suggest that paternal Ube3a may escape silencing in certain neuron populations, allowing for persistent expression of paternal UBE3A protein. Here we extend evidence in AS model mice (Ube3am–/p+) of paternal UBE3A expression within the suprachiasmatic nucleus (SCN), the master circadian pacemaker. Paternal UBE3A-positive cells in the SCN show partial colocalization with the neuropeptide arginine vasopressin (AVP) and clock proteins (PER2 and BMAL1), supporting that paternal UBE3A expression in the SCN is often of neuronal origin. Paternal UBE3A also partially colocalizes with a marker of neural progenitors, SOX2, implying that relaxed or incomplete imprinting of paternal Ube3a reflects an overall immature molecular phenotype. Our findings highlight the complexity of Ube3a imprinting in the brain and illuminate a subpopulation of SCN neurons as a focal point for future studies aimed at understanding the mechanisms of Ube3a imprinting

Signal Transduction Mechanisms Underlying Group I mGluR-mediated Increase in Frequency and Amplitude of Spontaneous EPSCs in the Spinal Trigeminal Subnucleus Oralis of the Rat

Group I mGluRs (mGluR1 and 5) pre- and/or postsynaptically regulate synaptic transmission at glutamatergic synapses. By recording spontaneous EPSCs (sEPSCs) in the spinal trigeminal subnucleus oralis (Vo), we here investigated the regulation of glutamatergic transmission through the activation of group I mGluRs. Bath-applied DHPG (10 μM/5 min), activating the group I mGluRs, increased sEPSCs both in frequency and amplitude; particularly, the increased amplitude was long-lasting. The DHPG-induced increases of sEPSC frequency and amplitude were not NMDA receptor-dependent. The DHPG-induced increase in the frequency of sEPSCs, the presynaptic effect being further confirmed by the DHPG effect on paired-pulse ratio of trigeminal tract-evoked EPSCs, an index of presynaptic modulation, was significantly but partially reduced by blockades of voltage-dependent sodium channel, mGluR1 or mGluR5. Interestingly, PKC inhibition markedly enhanced the DHPG-induced increase of sEPSC frequency, which was mainly accomplished through mGluR1, indicating an inhibitory role of PKC. In contrast, the DHPG-induced increase of sEPSC amplitude was not affected by mGluR1 or mGluR5 antagonists although the long-lasting property of the increase was disappeared; however, the increase was completely inhibited by blocking both mGluR1 and mGluR5. Further study of signal transduction mechanisms revealed that PLC and CaMKII mediated the increases of sEPSC in both frequency and amplitude by DHPG, while IP3 receptor, NO and ERK only that of amplitude during DHPG application. Altogether, these results indicate that the activation of group I mGluRs and their signal transduction pathways differentially regulate glutamate release and synaptic responses in Vo, thereby contributing to the processing of somatosensory signals from orofacial region

Eco-Innovation Indices as Tools for Measuring Eco-Innovation

Measuring eco-innovation helps us understand the overall trends and raises awareness in society. Measuring eco-innovation at the national level and making comparisons across countries may allow us to benchmark performance and foster policy learning. This paper assesses two indices developed in two different regions: The ASEM Eco-Innovation Index (ASEI) by the ASEM SMEs Eco-Innovation Center, based in Republic of Korea; and the Eco-Innovation Scoreboard (Eco-IS) developed by the Eco-Innovation Observatory, based in the European Union. This paper aims to examine and compare the features of both and attempts to obtain insights on their strengths and weaknesses. Towards this aim, our paper assesses those scoreboards against four criteria stemming from innovation analysis: (1) relevance of areas and stakeholders covered; (2) ability to indicate changes; (3) directions towards common goals; and (4) ability to facilitate further changes. We conclude both are promising, despite data shortages, and have great potential to contribute towards the sustainable development goals (SDGs), particularly with regard to the SDGs on sustainable industrialization and sustainable consumption and production. In comparison, the ASEI covers more countries than the Eco-IS. However, the ASEI has limitations on measuring indicators due to limited data availability in Asian countries. The Eco-IS is closely linked with the regional and national policies for eco-innovation in Europe, while the ASEI’s impact appears more limited, as of now. In conclusion, the research results give insights into key areas, goals and applications of eco-innovation indices, and can help upgrading eco-innovation indices. This research helps interpret the scores of two indices better and facilitate application of the scores in the multiple ways. It is expected that this research contributes to developing and modifying a global eco-innovation index and enhancing the ability of these indices to facilitate eco-innovation strategies at national levels and across relevant actors

Body extract of tail amputated zebrafish promotes culturing of primary fin cells from glass catfish

The most spectacular regenerative events in vertebrates are epimorphic regeneration. In this study, interestingly, a whole-body extract 24 h after tail amputation enhanced primary cell growth and viability compared to that of a non-tail amputated body. Additionally, these effects of extract treatment in vitro were dose-dependent occurring at concentrations of 0.02, 0.05 and 0.1 mg/ml. This is the first in vitro study on the interaction between primary fin cells from glass catfish and tail amputated body extracts of zebrafish. These results provide an essential knowledge base for rational approaches to tissue and organ regeneration.Keywords: Cell growth, cell viability, extract, glass catfish, regeneration, zebrafishAfrican Journal of Biotechnology Vol. 12(12), pp. 1449-145