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Acid-sensing ion channels (ASICs), which act as proton-gating sodium channels, have garnered attention as pharmacological targets. ASIC1a isoform, notably prevalent in the central nervous system, plays an important role in synaptic plasticity, anxiety, neurodegeneration, etc. In the peripheral nervous system, ASIC1a shares prominence with ASIC3, the latter well established for its involvement in pain signaling, mechanical sensitivity, and inflammatory hyperalgesia. However, the precise contributions of ASIC1a in peripheral functions necessitate thorough investigation. To dissect the specific roles of ASICs, peptide ligands capable of modulating these channels serve as indispensable tools. Employing molecular modeling, we designed the peptide targeting ASIC1a channel from the sea anemone peptide Ugr9-1, originally targeting ASIC3. This peptide (A23K) retained an inhibitory effect on ASIC3 (IC 50 9.39 µM) and exhibited an additional inhibitory effect on ASIC1a (IC 50 6.72 µM) in electrophysiological experiments. A crucial interaction between the Lys23 residue of the A23K peptide and the Asp355 residue in the thumb domain of the ASIC1a channel predicted by molecular modeling was confirmed by site-directed mutagenesis of the channel. However, A23K peptide revealed a significant decrease in or loss of analgesic properties when compared to the wild-type Ugr9-1. In summary, using A23K, we show that negative modulation of the ASIC1a channel in the peripheral nervous system can compromise the efficacy of an analgesic drug. These results provide a compelling illustration of the complex balance required when developing peripheral pain treatments targeting ASICs.

Transient receptor potential vanilloid subtype 3 (TRPV3) is an ion channel implicated in skin physiology and itch. TRPV3 inhibitors can present a novel strategy for combating debilitating itch conditions, and medicinal plants are a natural pool of such compounds. Here, we report the isolation of a TRPV3-inhibiting compound from Andrographis paniculata , a medicinal plant with anti-inflammatory properties whose bioactive components are poorly characterized in terms of molecular targets. Using 1 H and 13 C NMR and high-resolution mass spectrometry, the compound was identified as a labdane-type diterpenoid, 14-deoxy-11,12-didehydroandrographolide (ddA). The activity of the compound was evaluated by fluorescent calcium assay and manual whole-cell patch-clamp technique. ddA inhibited human TRPV3 in stably expressing CHO and HaCaT keratinocytes, acting selectively among other TRP channels implicated in itch and inflammation and not showing toxicity to HaCaT cells. Antipruritic effects of the compound were evaluated in scratching behavior models on ICR mice. ddA suppressed itch induced by the TRPV3 activator carvacrol. Additionally, ddA potently suppressed histamine-induced itch with efficacy comparable to loratadine, a clinically used antihistamine drug. These results suggest the potential of ddA as a possible safe and efficacious alternative for antipruritic therapy.

Tafalgin (Taf) is a tetrapeptide opioid used in clinical practice in Russia as an analgesic drug for subcutaneous administration as a solution (4 mg/mL; concentration of 9 mM). We found that the acid-sensing ion channels (ASICs) are another molecular target for this molecule. ASICs are proton-gated sodium channels that mediate nociception in the peripheral nervous system and contribute to fear and learning in the central nervous system. Using electrophysiological methods, we demonstrated that Taf could increase the integral current through heterologically expressed ASIC with half-maximal effective concentration values of 0.09 mM and 0.3 mM for rat and human ASIC3, respectively, and 1 mM for ASIC1a. The molecular mechanism of Taf action was shown to be binding to the channel in the resting state and slowing down the rate of desensitization. Taf did not compete for binding sites with both protons and ASIC3 antagonists, such as APETx2 and amiloride (Ami). Moreover, Taf and Ami together caused an unusual synergistic effect, which was manifested itself as the development of a pronounced second desensitizing component. Thus, the ability of Taf to act as a positive allosteric modulator of these channels could potentially cause promiscuous effects in clinical practice. This fact must be considered in patients' treatment.

The neuropeptide nocistatin (NS) is expressed by the nervous system cells and neutrophils as a part of a precursor protein and can undergo stepwise limited proteolysis. Previously, it was shown that rat NS (rNS) is able to activate acid-sensing ion channels (ASICs) and that this effect correlates with the acidic nature of NS. Here, we investigated changes in the properties of rNS in the course of its proteolytic degradation by comparing the effects of the full-size rNS and its two cleavage fragments on the rat isoform 3 ASICs (ASIC3) expressed in X. laevis oocytes and pain perception in mice. The rNS acted as both positive and negative modulator by lowering the steady-state desensitization of ASIC3 at pH 6.8-7.0 and reducing the channel's response to stimuli at pH 6.0-6.9, respectively. The truncated rNSΔ21 peptide lacking 21 amino acid residues from the N-terminus retained the positive modulatory activity, while the C-terminal pentapeptide (rNSΔ30) acted only as a negative ASIC3 modulator. The effects of the studied peptides were confirmed in animal tests: rNS and rNSΔ21 induced a pain-related behavior, whereas rNSΔ30 showed the analgesic effect. Therefore, we have shown that the mode of rNS action changes during its stepwise degradation, from an algesic molecule through a pain enhancer to a pain reliever (rNSΔ30 pentapeptide), which can be considered as a promising drug candidate.

Efficient DNA sample preparation from fungi with the rigid cell walls is still critical for successful polymerase chain reaction (PCR), one of the basic platforms in molecular diagnostics of fungi, especially in medical mycology. Common methods that involve different chaotropes to yield DNA samples have found a limited application for fungi. Here we describe a novel procedure for efficient production of permeable fungal cell envelopes with DNA inside as suitable templates for PCR. This procedure is facile, relies on boiling of fungal cells in aqueous solutions of selected chaotropic agents and additives and enables to remove RNA and proteins from PCR template samples. The use of chaotropic solutions containing 7 M urea, 1% sodium dodecyl sulfate (SDS), up to100 mM ammonia and/or 25 mM sodium citrate was the best option to yield highly purified DNA-containing cell envelopes from all fungal strains under study, including clinical Candida and Cryptococcusisolates. After treatment with the selected chaotropic mixtures, the fungal cell walls had undergone loosening and were no longer a barrier to release DNA in PCR as evident from electron microscopy examinations and successful target gene amplifications. Overall, the developed simple, fast, and low-cost approach to produce PCR-suitable templates in the form of DNA encased by permeable cell walls can find application in molecular diagnostics., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Acid-sensing ion channels (ASICs) are proton-gated ion channels that mediate nociception in the peripheral nervous system and contribute to fear and learning in the central nervous system. Sevanol was reported previously as a naturally-occurring ASIC inhibitor from thyme with favorable analgesic and anti-inflammatory activity. Using electrophysiological methods, we found that in the high micromolar range, the compound effectively inhibited homomeric ASIC1a and, in sub- and low-micromolar ranges, positively modulated the currents of α1β2γ2 GABA A receptors. Next, we tested the compound in anxiety-related behavior models using a targeted delivery into the hippocampus with parallel electroencephalographic measurements. In the open field, 6 µM sevanol reduced both locomotor and θ-rhythmic activity similar to GABA, suggesting a primary action on the GABAergic system. At 300 μM, sevanol markedly suppressed passive avoidance behavior, implying alterations in conditioned fear memory. The observed effects could be linked to distinct mechanisms involving GABA A R and ASIC1a. These results elaborate the preclinical profile of sevanol as a candidate for drug development and support the role of ASIC channels in fear-related functions of the hippocampus.

Chemoselective O-alkylation of 1-aryl-3-polyfluoroalkylpyrazol-5-oles under basic conditions resulted in a series of 5-alkoxypyrazoles (26 derivatives). They showed an acceptable ADME profile (in silico) and can be considered as drug-like. In experiments in vivo (CD-1 mice), it was found that the obtained compounds do not have toxic properties at a dose of more than 150 mg/kg (for most compounds at a dose of >300 mg/kg, and for lead compounds - >600 mg/kg). 22 Compounds from this series demonstrated from moderate to high analgesic effects (28-104 % at 1 h and 37-109 % at 2 h after administration) in vivo in the hot plate test (SD rats, 15 mg/kg, intraperitoneal (ip)). The lead compound was 4-([1-phenyl-3-(trifluoromethyl)pyrazol-5-yl]oxy)butan-1-ol, which not only increased the latent period in the hot plate test by 103 % at both measurement points but also showed a pronounced analgesic effect under conditions of capsaicin-induced nociception (CD-1 mice, 15 mg/kg, ip). According to molecular modeling, all synthesized compounds can interact with the TRPV1 ion channel. This biological target was confirmed in in vitro experiments on Chinese hamster ovary cells expressing rTRPV1. 5-Alkoxypyrazoles were partial agonists of the TRPV1 ion channel in various degree, and the most active was the same pyrazole as in in vivo tests., (© 2023 Wiley-VCH GmbH.)

Acid-sensing ion channels (ASICs) have been known as sensors of a local pH change within both physiological and pathological conditions. ASIC-targeting peptide toxins could be potent molecular tools for ASIC-manipulating in vitro, and for pathology treatment in animal test studies. Two sea anemone toxins, native Hmg 1b-2 and recombinant Hmg 1b-4, both related to APETx-like peptides, inhibited the transient current component of human ASIC3-Δ20 expressed in Xenopus laevis oocytes, but only Hmg 1b-2 inhibited the rat ASIC3 transient current. The Hmg 1b-4 action on rASIC3 as a potentiator was confirmed once again. Both peptides are non-toxic molecules for rodents. In open field and elevated plus maze tests, Hmg 1b-2 had more of an excitatory effect and Hmg 1b-4 had more of an anxiolytic effect on mouse behavior. The analgesic activity of peptides was similar and comparable to diclofenac activity in an acid-induced muscle pain model. In models of acute local inflammation induced by λ-carrageenan or complete Freund's adjuvant, Hmg 1b-4 had more pronounced and statistically significant anti-inflammatory effects than Hmg 1b-2. It exceeded the effect of diclofenac and, at a dose of 0.1 mg/kg, reduced the volume of the paw almost to the initial volume. Our data highlight the importance of a comprehensive study of novel ASIC-targeting ligands, and in particular, peptide toxins, and present the slightly different biological activity of the two similar toxins.

Purinergic P2X7 receptors (P2X7) have now been proven to play an important role and represent an important therapeutic target in many pathological conditions including neurodegeneration. Here, we investigated the impact of peptides on purinergic signaling in Neuro-2a cells through the P2X7 subtype in in vitro models. We have found that a number of recombinant peptides, analogs of sea anemone Kunitz-type peptides, are able to influence the action of high concentrations of ATP and thereby reduce the toxic effects of ATP. The influx of calcium, as well as the fluorescent dye YO-PRO-1, was significantly suppressed by the studied peptides. Immunofluorescence experiments confirmed that the peptides reduce the P2X7 expression level in neuronal Neuro-2a cells. Two selected active peptides, HCRG1 and HCGS1.10, were found to specifically interact with the extracellular domain of P2X7 and formed stable complexes with the receptor in surface plasmon resonance experiments. The molecular docking approach allowed us to establish the putative binding sites of the most active HCRG1 peptide on the extracellular domain of the P2X7 homotrimer and propose a mechanism for regulating its function. Thus, our work demonstrates the ability of the Kunitz-type peptides to prevent neuronal death by affecting signaling through the P2X7 receptor.

4-Arylhydrazinylidene-5-(polyfluoroalkyl)pyrazol-3-ones (4-AHPs) were found to be obtained by the regiospecific cyclization of 2-arylhydrazinylidene-3-(polyfluoroalkyl)-3-oxoesters with hydrazines, by the azo coupling of 4-nonsubstituted pyrazol-5-oles with aryldiazonium chlorides or by the firstly discovered acid-promoted self-condensation of 2-arylhydrazinylidene-3-oxoesters. All the 4-AHPs had an acceptable ADME profile. Varying the substituents in 4-AHPs promoted the switching or combining of their biological activity. The polyfluoroalkyl residue in 4-AHPs led to the appearance of an anticarboxylesterase action in the micromolar range. An NH-fragment and/or methyl group instead of the polyfluoroalkyl one in the 4-AHPs promoted antioxidant properties in the ABTS, FRAP and ORAC tests, as well as anti-cancer activity against HeLa that was at the Doxorubicin level coupled with lower cytotoxicity against normal human fibroblasts. Some Ph-N-substituted 4-AHPs could inhibit the growth of N. gonorrhoeae bacteria at MIC 0.9 μg/mL. The possibility of using 4-AHPs for cell visualization was shown. Most of the 4-AHPs exhibited a pronounced analgesic effect in a hot plate test in vivo at and above the diclofenac and metamizole levels except for the ones with two chlorine atoms in the aryl group. The methylsulfonyl residue was proved to raise the anti-inflammatory effect also. A mechanism of the antinociceptive action of the 4-AHPs through blocking the TRPV1 receptor was proposed and confirmed using in vitro experiment and molecular docking.

A novel peptide AnmTX Sco 9a-1 with the β-hairpin fold was isolated from the swimming sea anemone Stomphia coccinea (Actinostolidae family). The peptide consists of 28 amino acid residues, including modified hydroxyproline residue, and its measured molecular mass is 2960 Da. The peptide was not toxic on mice; however, it stimulated their exploratory motivation and active search behavior, and demonstrated an anti-anxiety effect. AnmTX Sco 9a-1 at doses of 0.1 and 1 mg/kg reduced the volume of edema during 24 h better than the nonsteroidal anti-inflammatory drug, Diclofenac, at dose of 1 mg/kg in a model of acute local λ-carrageenan-induced inflammation. ELISA analysis of the animal's blood showed that peptide at a dose of 1 mg/kg reduced the content of tumor necrosis factor-α (TNF-α), a pro-inflammatory mediator responsible in the edema development, up to the level of TNF-α in the intact group. Besides, AnmTX Sco 9a-1 demonstrated a significant analgesic effect on acute pain sensitivity in the carrageenan-induced thermal hyperalgesia model at doses of 0.1 and 1 mg/kg. Activity of AnmTX Sco 9a-1 was shown not to be associated with modulation of nociceptive ASIC channels.

The nicotinic acetylcholine receptors (nAChRs) are prototypical ligand-gated ion channels, provide cholinergic signaling, and are modulated by various venom toxins and drugs in addition to neurotransmitters. Here, four APETx-like toxins, including two new toxins, named Hmg 1b-2 Met ox and Hmg 1b-5, were isolated from the sea anemone Heteractis magnifica and characterized as novel nAChR ligands and acid-sensing ion channel (ASIC) modulators. All peptides competed with radiolabeled α-bungarotoxin for binding to Torpedo californica muscle-type and human α7 nAChRs. Hmg 1b-2 potentiated acetylcholine-elicited current in human α7 receptors expressed in Xenopus laevis oocytes. Moreover, the multigene family coding APETx-like peptides library from H. magnifica was described and in silico surface electrostatic potentials of novel peptides were analyzed. To explain the 100% identity of some peptide isoforms between H. magnifica and H. crispa , 18S rRNA, COI, and ITS analysis were performed. It has been shown that the sea anemones previously identified by morphology as H. crispa belong to the species H. magnifica .

Plant lignans exhibit a wide range of biological activities, which makes them the research objects of potential use as therapeutic agents. They provide diverse naturally-occurring pharmacophores and are available for production by chemical synthesis. A large amount of accumulated data indicates that lignans of different structural groups are apt to demonstrate both anti-inflammatory and antioxidant effects, in many cases, simultaneously. In this review, we summarize the comprehensive knowledge about lignan use as a bioactive agent in disorders associated with oxidative stress and inflammation, pharmacological effects in vitro and in vivo, molecular mechanisms underlying these effects, and chemical synthesis approaches. This article provides an up-to-date overview of the current data in this area, available in PubMed, Scopus, and Web of Science databases, screened from 2000 to 2022.

CO 2 inhalation is currently the most common method of euthanasia for laboratory rats and mice, and it is often used for further terminal blood sampling for clinical biochemical assays. Lately, this method has been criticized due to animal welfare issues associated with some processes that develop after CO 2 inhalation. The stress reaction and the value of the clinical laboratory parameters significantly depend on the used anesthetics, method, and the site of blood sampling. Especially in small rodents, an acute terminal state followed by a cascade of metabolic reactions that can affect the studied biochemical profile may develop and cause unnecessary suffering of animals. The aim of this study was to compare the stability of biochemical parameters of outbred Sprague Dawley rats and CD-1 mice serum collected after CO 2 inhalation or the intramuscular injection of tiletamine-zolazepam-xylazine (TZX). The serum content of total protein and albumin, cholesterol, triglycerides, aspartate aminotransferase (AST), alanine aminotr ansferase (ALT), alkaline phosphatase (ALP), total bilirubin, and creatinine was decreased by the injection of TZX in comparison with CO 2 inhalation. In addition, the levels of calcium, phosphates, chlorides and potassium were lowered by TZX vs. CO 2 administration, while the level of sodium increased. Finally, the level of the majority of serum clinical biochemical parameters in rats and mice tend to be overestimated after CO 2 inhalation, which may lead to masking the possible effect of anti-inflammatory drugs in animal tests. Injection anesthesia for small rodents with TZX is a more feasible method for terminal blood sampling, which also reduces the suffering of animals.

Human neuroblastoma SH-SY5Y is a prominent neurobiological tool used for studying neuropathophysiological processes. We investigated acid-sensing (ASIC) and transient receptor potential vanilloid-1 (TRPV1) and ankyrin-1 (TRPA1) ion channels present in untreated and differentiated neuroblastoma SH-SY5Y to propose a new means for their study in neuronal-like cells. Using a quantitative real-time PCR and a whole-cell patch-clamp technique, ion channel expression profiles, functionality, and the pharmacological actions of their ligands were characterized. A low-level expression of ASIC1a and ASIC2 was detected in untreated cells. The treatment with 10 μM of retinoic acid (RA) for 6 days resulted in neuronal differentiation that was accompanied by a remarkable increase in ASIC1a expression, while ASIC2 expression remained almost unaltered. In response to acid stimuli, differentiated cells showed prominent ASIC-like currents. Detailed kinetic and pharmacological characterization suggests that homomeric ASIC1a is a dominant isoform among the present ASIC channels. RA-treatment also reduced the expression of TRPV1 and TRPA1, and minor electrophysiological responses to their agonists were found in untreated cells. Neuroblastoma SH-SY5Y treated with RA can serve as a model system to study the effects of different ligands on native human ASIC1a in neuronal-like cells. This approach can improve the characterization of modulators for the development of new neuroprotective and analgesic drugs.

Wireless Sensor Networks (WSNs) have gained great significance from researchers and industry due to their wide applications. Energy and resource conservation challenges are facing the WSNs. Nevertheless, clustering techniques offer many solutions to address the WSN issues, such as energy efficiency, service redundancy, routing delay, scalability, and making WSNs more efficient. Unfortunately, the WSNs are still immature, and suffering in several aspects. This paper aims to solve some of the downsides in existing routing protocols for WSNs; a Lightweight and Efficient Dynamic Cluster Head Election routing protocol (LEDCHE-WSN) is proposed. The proposed routing algorithm comprises two integrated methods, electing the optimum cluster head, and organizing the re-clustering process dynamically. Furthermore, the proposed protocol improves on others present in the literature by combining the random and periodic electing method in the same round, and the random method starts first at the beginning of each round/cycle. Moreover, both random and periodic electing methods are preceded by checking the remaining power to skip the dead nodes and continue in the same way periodically with the rest of the nodes in the round. Additionally, the proposed protocol is distinguished by deleting dead nodes from the network topology list during the re-clustering process to address the black holes and routing delay problems. Finally, the proposed algorithm's mathematical modeling and analysis are introduced. The experimental results reveal the proposed protocol outperforms the LEACH protocol by approximately 32% and the FBCFP protocol by 8%, in terms of power consumption and network lifetime. In terms of Mean Package Delay, LEDCHE-WSN improves the LEACH protocol by 42% and the FBCFP protocol by 15%, and regarding Loss Ratio, it improves the LEACH protocol by approximately 46% and FBCFP protocol by 25%.

In the common chickweed Stellaria media , two antimicrobial peptides (AMPs), SmAMP1.1a and SmAMP1.2a, have been shown to be proteolytically released as products of the expression of a single gene, proSmAMP1 . In this study, the gene proSmAMP1 was introduced into two potato varieties, Zhukovsky ranny and Udacha. These early-maturing varieties were shown to be susceptible to early blight caused by Alternaria spp. Most transgenic lines of either variety having strong expression of the target gene demonstrated high levels of resistance to Alternaria spp. during three years of cultivation, but did not otherwise differ from the initial varieties. Disease severity index (DSI) was introduced as a complex measure of plant susceptibility to early blight, taking into account the diameter of lesions caused by the Alternaria spp., the fungus sporulation intensity and its incubation period duration. Across all transgenic lines, the DSI inversely correlated both with the target gene expression and the copy number in the plant genome. Our results are promising for improving the resistance of potato and other crops to early blight by expression of AMPs from wild plants.

Although TRPV1 ion channel has been attracting researchers' attention for many years, its functions in animal organisms, the principles of regulation, and the involvement in pathological processes have not yet been fully clarified. Mutagenesis experiments and structural studies have identified the structural features of the channel and binding sites for its numerous ligands; however, these studies are far from conclusion. This review summarizes recent achievements in the TRPV1 research with special focus on structural and functional studies of the channel and on its ligands, which are extremely diverse in their nature and interaction specificity to TRPV1. Particular attention was given to the effects of numerous endogenous agonists and antagonists that can fine-tune the channel sensitivity to its usual activators, such as capsaicin, heat, acids, or their combination. In addition to the pain sensing not covered in this review, the TRPV1 channel was found to be involved in the regulation of many important physiological and pathological processes and, therefore, can be considered as a promising therapeutic target in the treatment of various diseases, such as pneumonia, ischemia, diabetes, epilepsy, schizophrenia, psoriasis, etc.

Lynx1 is a GPI-tethered protein colocalized with nicotinic acetylcholine receptors (nAChRs) in the brain areas important for learning and memory. Previously, we demonstrated that at low micromolar concentrations the water-soluble Lynx1 variant lacking GPI-anchor (ws-Lynx1) acts on α7-nAChRs as a positive allosteric modulator. We hypothesized that ws-Lynx1 could be used for improvement of cognitive processes dependent on nAChRs. Here we showed that 2 µM ws-Lynx1 increased the acetylcholine-evoked current at α7-nAChRs in the rat primary visual cortex L1 interneurons. At higher concentrations ws-Lynx1 inhibits α7-nAChRs expressed in Xenopus laevis oocytes with IC 50  ~ 50 µM. In mice, ws-Lynx1 penetrated the blood-brain barrier upon intranasal administration and accumulated in the cortex, hippocampus, and cerebellum. Chronic ws-Lynx1 treatment prevented the olfactory memory and motor learning impairment induced by the α7-nAChRs inhibitor methyllycaconitine (MLA). Enhanced long-term potentiation and increased paired-pulse facilitation ratio were observed in the hippocampal slices incubated with ws-Lynx1 and in the slices from ws-Lynx1-treated mice. Long-term potentiation blockade observed in MLA-treated mice was abolished by ws-Lynx1 co-administration. To understand the mechanism of ws-Lynx1 action, we studied the interaction of ws-Lynx1 and MLA at α7-nAChRs, measured the basal concentrations of endogenous Lynx1 and the α7 nAChR subunit and their association in the mouse brain. Our findings suggest that endogenous Lynx1 limits α7-nAChRs activation in the adult brain. Ws-Lynx1 partially displaces Lynx1 causing positive modulation of α7-nAChRs and enhancement of synaptic plasticity. Ws-Lynx1 and similar compounds may constitute useful hits for treatment of cognitive deficits associated with the cholinergic system dysfunction., (© 2020 International Society for Neurochemistry.)

To the present, different efficient but expensive, multistage, and time-consuming technologies have been developed to deliver ribonucleic acids (RNA) into eukaryotic cells. Here, we report a simple and feasible solution to design RNA nanocarriers based on nucleic acid condensation by bi- and trivalent metal ions during thermal cycling. Efficient RNA conversion to nanoparticles with small size (10-50 nm) suitable for transfection was achieved using cations Ni 2+ , Co 2+ or Cu 2+ alone or in combination with Ca 2+ at the specially selected concentrations (2.0 mM-3.5 mM), low ionic strength, and narrow pH range (8.0-8.5). Other ions - Mn 2+ , Zn 2+ , Tb 3+ , or Gd 3+ - caused RNA-cleaving effect that was abolished in the presence of Ni 2+ , Co 2+ , Zn 2+ , or Cu 2+ . Naked RNA-metal ion nanoparticles were extremely unstable in phosphate buffer and sensitive to serum ribonucleases (RNases), and this problem was solved by treatment with polyarginines-16 and 8. Polyarginine-stabilized nanoparticles, containing malachite green (MG) aptamer RNA and metal cations, crossed the cell membrane, dissociated in the cytoplasm, and preserved the functionality of transported RNA, as judged from efficient transfection of human embryonic kidney 293 cells. The technology, involving RNA condensation by metal cations, can be used as a cheap alternative to produce nanoscale carriers to deliver various RNAs into cells in vitro and in vivo .Communicated by Ramaswamy H. Sarma.

Among acid-sensing ion channels (ASICs), ASIC1a and ASIC3 subunits are the most widespread and prevalent in physiological and pathophysiological conditions. They participate in synaptic plasticity, learning and memory, as well as the perception of inflammatory and neurological pain, making these channels attractive pharmacological targets. Sevanol, a natural lignan isolated from Thymus armeniacus , inhibits the activity of ASIC1a and ASIC3 isoforms, and has a significant analgesic and anti-inflammatory effect. In this work, we described the efficient chemical synthesis scheme of sevanol and its analogues, which allows us to analyze the structure-activity relationships of the different parts of this molecule. We found that the inhibitory activity of sevanol and its analogues on ASIC1a and ASIC3 channels depends on the number and availability of the carboxyl groups of the molecule. At the structural level, we predicted the presence of a sevanol binding site based on the presence of molecular docking in the central vestibule of the ASIC1a channel. We predicted that this site could also be occupied in part by the FRRF-amide peptide, and the competition assay of sevanol with this peptide confirmed this prediction. The intravenous (i.v.), intranasal (i.n.) and, especially, oral (p.o.) administration of synthetic sevanol in animal models produced significant analgesic and anti-inflammatory effects. Both non-invasive methods of sevanol administration (i.n. and p.o.) showed greater efficacy than the invasive (i.v.) method, thus opening new horizons for medicinal uses of sevanol.

Acid-sensing ion channels (ASICs) are of the most sensitive molecular sensors of extracellular pH change in mammals. Six isoforms of these channels are widely represented in membranes of neuronal and non-neuronal cells, where these molecules are involved in different important regulatory functions, such as synaptic plasticity, learning, memory, and nociception, as well as in various pathological states. Structural and functional studies of both wild-type and mutant ASICs are essential for human care and medicine for the efficient treatment of socially significant diseases and ensure a comfortable standard of life. Ligands of ASICs serve as indispensable tools for these studies. Such bioactive compounds can be synthesized artificially. However, to date, the search for such molecules has been most effective amongst natural sources, such as animal venoms or plants and microbial extracts. In this review, we provide a detailed and comprehensive structural and functional description of natural compounds acting on ASICs, as well as the latest information on structural aspects of their interaction with the channels. Many of the examples provided in the review demonstrate the undoubted fundamental and practical successes of using natural toxins. Without toxins, it would not be possible to obtain data on the mechanisms of ASICs' functioning, provide detailed study of their pharmacological properties, or assess the contribution of the channels to development of different pathologies. The selectivity to different isoforms and variety in the channel modulation mode allow for the appraisal of prospective candidates for the development of new drugs., (Copyright © 2020 Osmakov, Khasanov, Andreev, Lyukmanova and Kozlov.)

The ion channel TRPV1, which is one of the most important integrators of pain and inflammatory stimuli, is considered a promising therapeutic target in the treatment of pain conditions. In this work, we performed a comparative study of the analgesic effect in the "hot plate" test of recombinant analogues of Kunitz-type peptides from the sea anemone Heteractis crispa venom: APHC1-modulator of TRPV1 and HCRG21-a full blocker of TRPV1. As a result of biological tests, it was shown that the full blocker HCRG21, despite the higher value of 50% effective concentration of TRPV1 inhibition, had an equal analgesic ability with the APHC1 upon intramuscular administration and retained it for 13 h of observation. The analgesic effect of APHC1 at a dose of 0.1 mg/kg when administered intramuscularly developed very quickly in 5 min but lasted 3 h. The differences in the pharmacodynamic profile of the peptides are in good agreement with different mechanisms of binding to TRPV1.

In a recent computational study, we revealed some mechanistic aspects of TRPV1 (transient receptor potential channel 1) thermal activation and gating and proposed a set of probable functionally important residues - "hot spots" that have not been characterized experimentally yet. In this work, we analyzed TRPV1 point mutants G643A, I679A + A680G, and K688G/P combining molecular modeling, biochemistry, and electrophysiology. The substitution G643A reduced maximal conductivity that resulted in a normal response to moderate stimuli, but a relatively weak response to more intensive activation. I679A + A680G channel was severely toxic for oocytes most probably due to abnormally increased basal activity of the channel ("always open" gates). The replacement K688G presumably facilitated movements of TRP domain and disturbed its coupling to the pore, thus leading to spontaneous activation and enhanced desensitization of the channel. Finally, mutation K688P was suggested to impair TRP domain directed movement, and the mutated channel showed ~100-fold less sensitivity to the capsaicin, enhanced desensitization and weaker activation by the heat. Our results provide a better understanding of TRPV1 thermal and capsaicin-induced activation and gating. These observations provide a structural basis for understanding some aspects of TRPV1 channel functioning and depict potentially pathogenic mutations., (Copyright © 2020. Published by Elsevier B.V.)

Global industry is undergoing major transformations with the genesis of a new paradigm known as the Internet of Things (IoT) with its underlying technologies. Many company leaders are investing more effort and money in transforming their services to capitalize on the benefits provided by the IoT. Thereby, the decision makers in public waste management do not want to be outdone, and it is challenging to provide an efficient and real-time waste management system. This paper proposes a solution (hardware, software, and communications) that aims to optimize waste management and include a citizen in the process. The system follows an IoT-based approach where the discarded waste from the smart bin is continuously monitored by sensors that inform the filling level of each compartment, in real-time. These data are stored and processed in an IoT middleware providing information for collection with optimized routes and generating important statistical data for monitoring the waste collection accurately in terms of resource management and the provided services for the community. Citizens can easily access information about the public waste bins through the Web or a mobile application. The creation of the real prototype of the smart container, the development of the waste management application and a real-scale experiment use case for evaluation, demonstration, and validation show that the proposed system can efficiently change the way people deal with their garbage and optimize economic and material resources.

Currently, five peptide modulators of acid-sensing ion channels (ASICs) attributed to structural class 1b of sea anemone toxins have been described. The APETx2 toxin is the first and most potent ASIC3 inhibitor, so its homologs from sea anemones are known as the APETx-like peptides. We have discovered that two APETx-like peptides from the sea anemone Heteractis crispa , Hcr 1b-3 and Hcr 1b-4, demonstrate different effects on rASIC1a and rASIC3 currents. While Hcr 1b-3 inhibits both investigated ASIC subtypes with IC 50 4.95 ± 0.19 μM for rASIC1a and 17 ± 5.8 μM for rASIC3, Hcr 1b-4 has been found to be the first potentiator of ASIC3, simultaneously inhibiting rASIC1a at similar concentrations: EC 50 1.53 ± 0.07 μM and IC 50 1.25 ± 0.04 μM. The closest homologs, APETx2, Hcr 1b-1, and Hcr 1b-2, previously demonstrated the ability to inhibit hASIC3 with IC 50 63 nM, 5.5, and 15.9 μM, respectively, while Hcr 1b-2 also inhibited rASIC1a with IC 50 4.8 ± 0.3 μM. Computer modeling allowed us to describe the peculiarities of Hcr 1b-2 and Hcr 1b-4 interfaces with the rASIC1a channel and the stabilization of the expanded acidic pocket resulting from peptides binding which traps the rASIC1a channel in the closed state.

The generation of terahertz (THz) radiation during the propagation of subpicosecond pulses in liquid media is investigated using a theoretical model considering the relative contribution of Kerr and plasma nonlinearity. The dependences of the THz emission generation efficiency on the contribution of plasma nonlinearity with a fixed third-order nonlinearity value revealed the existence of weak and strong ionization modes. It is shown that the transition between these modes is determined by the ratio of plasma to Kerr nonlinearity coefficients and the pump energy. In the strong ionization mode and with the fixed contribution of plasma nonlinearity, the optical-to-THz conversion efficiency decreases with increasing Kerr nonlinearity due to the redistribution of the pump energy for the third-order effects. These results contribute to estimating the potential of liquid media as highly efficient THz sources.

With the integration of Wireless Sensor Networks and the Internet of Things, the smart grid is being projected as a solution for the challenges regarding electricity supply in the future. However, security and privacy issues in the consumption and trading of electricity data pose serious challenges in the adoption of the smart grid. To address these challenges, blockchain technology is being researched for applicability in the smart grid. In this paper, important application areas of blockchain in the smart grid are discussed. One use case of each area is discussed in detail, suggesting a suitable blockchain architecture, a sample block structure and the potential blockchain technicalities employed in it. The blockchain can be used for peer-to-peer energy trading, where a credit-based payment scheme can enhance the energy trading process. Efficient data aggregation schemes based on the blockchain technology can be used to overcome the challenges related to privacy and security in the grid. Energy distribution systems can also use blockchain to remotely control energy flow to a particular area by monitoring the usage statistics of that area. Further, blockchain-based frameworks can also help in the diagnosis and maintenance of smart grid equipment. We also discuss several commercial implementations of blockchain in the smart grid. Finally, various challenges to be addressed for integrating these two technologies are discussed.

By irradiating a water jet with double pulses, we demonstrate 4-fold higher THz wave generation than for a single pump pulse. The dependence of the enhanced THz signal on the temporal delay between two collinear pulses reveals the optimal time for launching signal pulse is near 2-4 ps, which corresponds to the time needed to create the complete pre-ionization state when sufficient electron density is already induced, and there is no plasma reflection of the pump pulse radiation. The increase in THz waves generation efficiency corresponds to the case of water jet excitation by the pulses with an optimal duration for a certain jet thickness, which is determined by the spatial pulse size. Using a theoretical model of the interaction of a high-intensity sub-picosecond pulse with an isotropic medium, we held a numerical simulation, which well describes the experimental results when using 3 ps value of population relaxation time. Thus, in this work, double pump method allows not only to increase the energy of the generated THz waves, but also to determine the characteristic excited state lifetime of liquid water. The optical-to-terahertz conversion efficiency in case of double pulse excitation of water column is of the order of 0.5⋅10 -3 , which exceeds the typical values for THz waves generation during two-color filamentation in air and comparable with the achievable values due to the optical rectification in some crystals.

Acid-sensing ion channels (ASICs), which are present in almost all types of neurons, play an important role in physiological and pathological processes. The ASIC1a subtype is the most sensitive channel to the medium's acidification, and it plays an important role in the excitation of neurons in the central nervous system. Ligands of the ASIC1a channel are of great interest, both fundamentally and pharmaceutically. Using a two-electrode voltage-clamp electrophysiological approach, we characterized lindoldhamine (a bisbenzylisoquinoline alkaloid extracted from the leaves of Laurus nobilis L.) as a novel inhibitor of the ASIC1a channel. Lindoldhamine significantly inhibited the ASIC1a channel's response to physiologically-relevant stimuli of pH 6.5-6.85 with IC 50 range 150-9 μM, but produced only partial inhibition of that response to more acidic stimuli. In mice, the intravenous administration of lindoldhamine at a dose of 1 mg/kg significantly reversed complete Freund's adjuvant-induced thermal hyperalgesia and inflammation; however, this administration did not affect the pain response to an intraperitoneal injection of acetic acid (which correlated well with the function of ASIC1a in the peripheral nervous system). Thus, we describe lindoldhamine as a novel antagonist of the ASIC1a channel that could provide new approaches to drug design and structural studies regarding the determinants of ASIC1a activation.

Acid-sensing ion channel (ASIC) channels belong to the family of ligand-gated ion channels known as acid-sensing (proton-gated) ion channels. Only a few activators of ASICs are known. These are exogenous and endogenous molecules that cause a persistent, slowly desensitized current, different from an acid-induced current. Here we describe a novel endogenous agonist of ASICs-peptide nocistatin produced by neuronal cells and neutrophils as a part of prepronociceptin precursor protein. The rat nocistatin evoked currents in X. laevis oocytes expressing rat ASIC1a, ASIC1b, ASIC2a, and ASIC3 that were very similar in kinetic parameters to the proton-gated response. Detailed characterization of nocistatin action on rASIC1a revealed a proton-like dose-dependence of activation, which was accompanied by a dose-dependent decrease in the sensitivity of the channel to the protons. The toxin mambalgin-2, antagonist of ASIC1a, inhibited nocistatin-induced current, therefore the close similarity of mechanisms for ASIC1a activation by peptide and protons could be suggested. Thus, nocistatin is the first endogenous direct agonist of ASICs. This data could give a key to understanding ASICs activation regulation in the nervous system and also could be used to develop new drugs to treat pathological processes associated with ASICs activation, such as neurodegeneration, inflammation, and pain.

Acid-sensing ion channels (ASICs) are proton-gated sodium-selective channels that are expressed in the peripheral and central nervous systems. ASIC1a is one of the most intensively studied isoforms due to its importance and wide representation in organisms, but it is still largely unexplored as a target for therapy. In this study, we demonstrated response of the ASIC1a to acidification in the presence of the daurisoline (DAU) ligand. DAU alone did not activate the channel, but in combination with protons, it produced the second peak component of the ASIC1a current. This second peak differs from the sustained component (which is induced by RF-amide peptides), as the second (DAU-induced) peak is completely desensitized, with the same kinetics as the main peak. The co-application of DAU and mambalgin-2 indicated that their binding sites do not overlap. Additionally, we found an asymmetry in the pH activation curve of the channel, which was well-described by a mathematical model based on the multiplied probabilities of protons binding with a pool of high-cooperative sites and a single proton binding with a non-cooperative site. In this model, DAU targeted the pool of high-cooperative sites and, when applied with protons, acted as an inhibitor of ASIC1a activation. Moreover, DAU's occupation of the same binding site most probably reverses the channel from steady-state desensitization in the pH 6.9-7.3 range. DAU features disclose new opportunities in studies of ASIC structure and function.

Polar liquids are strong absorbers of electromagnetic waves in the terahertz range, therefore, historically such liquids have not been considered as good candidates for terahertz sources. However, flowing liquid medium has explicit advantages, such as a higher damage threshold compared to solid-state sources and more efficient ionization process compared to gases. Here we report systematic study of efficient generation of terahertz radiation in flat liquid jets under sub-picosecond single-color optical excitation. We demonstrate how medium parameters such as molecular density, ionization energy and linear absorption contribute to the terahertz emission from the flat liquid jets. Our simulation and experimental measurements reveal that the terahertz energy has quasi-quadratic dependence on the optical excitation pulse energy. Moreover, the optimal pump pulse duration, which depends on the thickness of the jet is theoretically predicted and experimentally confirmed. The obtained optical-to-terahertz energy conversion efficiency is more than 0.05%. It is comparable to the commonly used optical rectification in most of electro-optical crystals and two-color air filamentation. These results, significantly advancing prior research, can be successfully applied to create a new alternative source of terahertz radiation.

The values of the nonlinear refractive index coefficient for various materials in the terahertz frequency range exceed the ones in both visible and NIR ranges by several orders of magnitude. This allows to create nonlinear switches, modulators, systems requiring lower control energies in the terahertz frequency range. We report the direct measurement of the nonlinear refractive index coefficient of liquid water by using the Z-scan method with broadband pulsed THz beam. Our experimental result shows that nonlinear refractive index coefficient in water is positive and can be as large as 7×10 -10 cm 2 /W in the THz frequency range, which exceeds the values for the visible and NIR ranges by 6 orders of magnitude. To estimate n 2 , we use the theoretical model that takes into account ionic vibrational contribution to the third-order susceptibility. We show that the origins of the nonlinearity observed are the anharmonicity of molecular vibrations.

Our previous studies demonstrated the formation of structurally diverse DNA-containing microparticles (DNA MPs) in PCR with Mg-pyrophosphate (MgPPi) as the structure-forming component. These DNA MPs were referred to major structural types: microdisks (2D MPs) with nanometer thickness and 3D MPs with sophisticated morphology and constructed from intersecting disks and their segments. Little is known about factors that influence both the morphology and size of DNA MPs, and the present study was aimed at fulfilling this gap. We showed that the addition of Mn 2+ cations to PCR mixtures caused the profound changes in MPs morphology, depending on DNA polymerase used (KlenTaq or Taq). Asymmetric PCR with 20-fold decrease in the concentration of one of two primers facilitated the predominant formation of microdisks with unusual structure. The addition of 1 mM Na-pyrophosphate to PCR mixtures with synthesized DNA and subsequent thermal cycling (10-15 cycles) were optimal to produce microdisks or nanometer 3D particles. Using electron microscopy, we studied also the structure of inorganic micro- and nanoparticles from MgPPi, formed during multiple heating and cooling cycles of a mixture of Mg 2+ and Na-pyrophosphate in various regimes. Also, we found the conditions to yield planar (Mg·Mn)PPi nanocrystals (diameter ~100 nm and thickness ~10 nm) which efficiently adsorbed exogenous DNA. These inorganic nanoparticles are promising for DNA delivery in transfection studies. Mechanisms to be involved in structural modifications of MPs and perspectives of their practical application are discussed.

TRPA1 is a cation channel located on the plasma membrane of many types of human and animal cells, including skin sensory neurons and epithelial cells of the intestine, lungs, urinary bladder, etc. TRPA1 is the major chemosensor that also responds to thermal and mechanical stimuli. Substances that activate TRPA1, e.g., allyl isothiocyanates (pungent components of mustard, horseradish, and wasabi), cinnamaldehyde from cinnamon, organosulfur compounds from garlic and onion, tear gas, acrolein and crotonaldehyde from cigarette smoke, etc., cause burning, mechanical and thermal hypersensitivity, cough, eye irritation, sneezing, mucus secretion, and neurogenic inflammation. An increased activity of TRPA1 leads to the emergence of chronic pruritus and allergic dermatitis and is associated with episodic pain syndrome, a hereditary disease characterized by episodes of debilitating pain triggered by stress. TRPA1 is now considered as one of the targets for developing new anti-inflammatory and analgesic drugs. This review summarizes information on the structure, function, and physiological role of this channel, as well as describes known TRPA1 ligands and their significance as therapeutic agents in the treatment of inflammation-associated pain.

Acid-sensing ion channel 3 (ASIC3) makes an important contribution to the development and maintenance of inflammatory and acid-induced pain. We compared different ASIC3 inhibitors (peptides from sea anemones (APETx2 and Ugr9-1) and nonpeptide molecules (sevanol and diclofenac)) in anti-inflammatory action and analgesic effects. All tested compounds had distinct effects on pH-induced ASIC3 current. APETx2 inhibited only transient current, whereas Ugr9-1 and sevanol decreased transient and sustained components of the current. The effect on mice was evaluated after administering an intramuscular injection in the acetic acid writhing pain model and the complete Freund's adjuvant-induced thermal hyperalgesia/inflammation test. The bell-shaped dependence of the analgesic effect was observed for APETx2 in the acetic acid-induced writhing test, as well as for sevanol and peptide Ugr9-1 in the thermal hyperalgesia test. This dependence could be evidence of the nonspecific action of compounds in high doses. Compounds reducing both components of ASIC3 current produced more significant pain relief than APETx2, which is an effective inhibitor of a transient current only. Therefore, the comparison of the efficacy of ASIC3 inhibitors revealed the importance of ASIC3-sustained currents' inhibition for promotion of acidosis-related pain relief.

APHC3 is an analgesic polypeptide that was found in the sea anemone (Heteractis crispa), and contains 56 amino acid residues. This polypeptide is of interest for the development of medications for diseases, associated with inflammatory or neuropathological processes, as well as its use as an analgesic. This work presents an innovative biotechnological method for APHC3 production. We have constructed a recombinant plasmid intended for biosynthesizing the fusion protein consisting of a chitin-binding domain, DnaB mini-intein from Synechocystis sp. capable of undergoing pH-dependent self-cleavage, and the target peptide. In the process of biosynthesis the fusion protein aggregates and forms the inclusion bodies that are welcomed since APHC3 is a cytotoxic peptide. The target peptide recovery process developed by us involves 3 chromatographic steps. The method developed by us enables to produce 940 mg of the recombinant APHC3 from 100 g of the inclusion bodies. The method is straightforward to implement and scale up. The recombinant APHC3 activity and effectiveness as an analgesic was proved by animal testing., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Background and Purpose: Acid-sensing ion channels (ASICs) play an important role in synaptic plasticity and learning, as well as in nociception and mechanosensation. ASICs are involved in pain and in neurological and psychiatric diseases, but their therapeutic potential is limited by the lack of ligands activating them at physiological pH., Experimental Approach: We extracted, purified and determined the structure of a bisbenzylisoquinoline alkaloid, lindoldhamine, (LIN) from laurel leaves. Its effect on ASIC3 channels were characterized, using two-electrode voltage-clamp electrophysiological recordings from Xenopus laevis oocytes., Key Results: At pH 7.4 or higher, LIN activated a sustained, proton-independent, current through rat and human ASIC3 channels, but not rat ASIC1a or ASIC2a channels. LIN also potentiated proton-induced transient currents and promoted recovery from desensitization in human, but not rat, ASIC3 channels., Conclusions and Implications: We describe a novel ASIC subtype-specific agonist LIN, which induced proton-independent activation of human and rat ASIC3 channels at physiological pH. LIN also acts as a positive allosteric modulator of human, but not rat, ASIC3 channels. This unique, species-selective, ligand of ASIC3, opens new avenues in studies of ASIC structure and function, as well as providing new approaches to drug design., (© 2017 The British Pharmacological Society.)

Sea anemones (Actiniaria) are intensely popular objects of study in venomics. Order Actiniaria includes more than 1,000 species, thus presenting almost unlimited opportunities for the discovery of novel biologically active molecules. The venoms of cold-water sea anemones are studied far less than the venoms of tropical sea anemones. In this work, we analysed the molecular venom composition of the cold-water sea anemone Cnidopus japonicus. Two sets of NGS data from two species revealed molecules belonging to a variety of structural classes, including neurotoxins, toxin-like molecules, linear polypeptides (Cys-free), enzymes, and cytolytics. High-throughput proteomic analyses identified 27 compounds that were present in the venoms. Some of the toxin-like polypeptides exhibited novel Cys frameworks. To characterise their function in the venom, we heterologously expressed 3 polypeptides with unusual Cys frameworks (designated CjTL7, CjTL8, and AnmTx Cj 1c-1) in E. coli. Toxicity tests revealed that the CjTL8 polypeptide displays strong crustacean-specific toxicity, while AnmTx Cj 1c-1 is toxic to both crustaceans and insects. Thus, an improved NGS data analysis algorithm assisted in the identification of toxins with unusual Cys frameworks showing no homology according to BLAST. Our study shows the advantage of combining omics analysis with functional tests for active polypeptide discovery.

Acid-sensing ion channels (ASICs) ASIC3 expressed mainly in peripheral sensory neurons play an important role in pain perception and inflammation development. In response to acidic stimuli, they can generate a unique biphasic current. At physiological pH 7.4, human ASIC3 isoform (hASIC3) is desensitized and able to generate only a sustained current. We found endogenous isoquinoline alkaloids (EIAs), which restore hASIC3 from desensitization and recover the transient component of the current. Similarly, rat ASIC3 isoform (rASIC3) can also be restored from desensitization (at pH < 7.0) by EIAs with the same potency. At physiological pH and above, EIAs at high concentrations were able to effectively activate hASIC3 and rASIC3. Thus, we found first endogenous agonists of ASIC3 channels that could both activate and prevent or reverse desensitization of the channel. The decrease of EIA levels could be suggested as a novel therapeutic strategy for treatment of pain and inflammation.

TRPV1 (vanilloid) receptors are activated by different types of stimuli including capsaicin, acidification and heat. Various ligands demonstrate stimulus-dependent action on TRPV1. In the present work we studied the action of polypeptides isolated from sea anemone Heteractis crispa (APHC1, APHC2 and APHC3) on rat TRPV1 receptors stably expressed in CHO cells using electrophysiological recordings, fluorescent Ca2+ measurements and molecular modeling. The APHCs potentiated TRPV1 responses to low (3-300 nM) concentrations of capsaicin but inhibited responses to high (>3.0 μM) concentrations. The activity-dependent action was also found for TRPV1 responses to 2APB and acidification. Thus the action mode of APHCs is bimodal and depended on the activation stimuli strength-potentiation of low-amplitude responses and no effect/inhibition of high-amplitude responses. The double-gate model of TRPV1 activation suggests that APHC-polypeptides may stabilize an intermediate state during the receptor activation. Molecular modeling revealed putative binding site at the outer loops of TRPV1. Binding to this site can directly affect activation by protons and can be allosterically coupled with capsaicin site. The results are important for further investigations of both TRPV1 and its ligands for potential therapeutic use.

A novel bioactive peptide named τ-AnmTx Ueq 12-1 (short name Ueq 12-1) was isolated and characterized from the sea anemone Urticina eques. Ueq 12-1 is unique among the variety of known sea anemone peptides in terms of its primary and spatial structure. It consists of 45 amino acids including 10 cysteine residues with an unusual distribution and represents a new group of sea anemone peptides. The 3D structure of Ueq 12-1, determined by NMR spectroscopy, represents a new disulfide-stabilized fold partly similar to the defensin-like fold. Ueq 12-1 showed the dual activity of both a moderate antibacterial activity against Gram-positive bacteria and a potentiating activity on the transient receptor potential ankyrin 1 (TRPA1). Ueq 12-1 is a unique peptide potentiator of the TRPA1 receptor that produces analgesic and anti-inflammatory effects in vivo . The antinociceptive properties allow us to consider Ueq 12-1 as a potential analgesic drug lead with antibacterial properties.

The transient receptor potential ankyrin-repeat 1 (TRPA1) is an important player in pain and inflammatory pathways. It is a promising target for novel drug development for the treatment of a number of pathological states. A novel peptide producing a significant potentiating effect on allyl isothiocyanate- and diclofenac-induced currents of TRPA1 was isolated from the venom of sea anemone Metridium senile. It is a 35-amino acid peptide cross-linked by two disulfide bridges named τ-AnmTX Ms 9a-1 (short name Ms 9a-1) according to a structure similar to other sea anemone peptides belonging to structural group 9a. The structures of the two genes encoding the different precursor proteins of Ms 9a-1 were determined. Peptide Ms 9a-1 acted as a positive modulator of TRPA1 in vitro but did not cause pain or thermal hyperalgesia when injected into the hind paw of mice. Intravenous injection of Ms 9a-1 (0.3 mg/kg) produced a significant decrease in the nociceptive and inflammatory response to allyl isothiocyanate (the agonist of TRPA1) and reversed CFA (Complete Freund's Adjuvant)-induced inflammation and thermal hyperalgesia. Taken together these data support the hypothesis that Ms 9a-1 potentiates the response of TRPA1 to endogenous agonists followed by persistent functional loss of TRPA1-expressing neurons. We can conclude that TRPA1 potentiating may be useful as a therapeutic approach as Ms 9a-1 produces significant analgesic and anti-inflammatory effects in mice models of pain., (© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.)

In this paper, we demonstrate experimentally for the first time (to our knowledge) the generation of spectral supercontinuum (SC) of more than two octaves with high intensity in a water jet. The spectrum of the generated SC extends from 350 to 1400 nm, with intensities up to 10 11   W/cm 2 , and its generation efficiency is more than 50%. For the pump intensity 3.0×10 12   W/cm 2 in the spectral range from 400 to 800 nm, the spectrum is nearly flat (less than 40% deviation), which is useful for many applications.

Peptide Ugr9-1 from the venom of sea anemone Urticina grebelnyi selectively inhibits the ASIC3 channel and significantly reverses inflammatory and acid-induced pain in vivo. A close homolog peptide Ugr 9-2 does not have these features. To find the pharmacophore residues and explore structure-activity relationships of Ugr 9-1, we performed site-directed mutagenesis of Ugr 9-2 and replaced several positions by the corresponding residues from Ugr 9-1. Mutant peptides Ugr 9-2 T9F and Ugr 9-2 Y12H were able to inhibit currents of the ASIC3 channels 2.2 times and 1.3 times weaker than Ugr 9-1, respectively. Detailed analysis of the spatial models of Ugr 9-1, Ugr 9-2 and both mutant peptides revealed the presence of the basic-aromatic clusters on opposite sides of the molecule, each of which is responsible for the activity. Additionally, Ugr9-1 mutant with truncated N- and C-termini retained similar with the Ugr9-1 action in vitro and was equally potent in vivo model of thermal hypersensitivity. All together, these results are important for studying the structure-activity relationships of ligand-receptor interaction and for the future development of peptide drugs from animal toxins., (Copyright © 2015 Elsevier Ltd. All rights reserved.)