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Growing evidence suggests that opioid analgesics modulate angiogenesis during pathophysiological processes. Vascular endothelial growth factor-A (VEGF-A) was recently proposed to be involved in pain development. To date, no anti-angiogenic drug is used for pain management. When administered in a bioavailable formulation, (i.e., ultramicronized) N-palmitoylethanolamine (PEA) delays the onset of morphine tolerance, improves morphine analgesic activity and reduces angiogenesis in in vivo models. This study aimed at investigating whether VEGF-A is involved in PEA-induced delay of morphine tolerance. The anti-VEGF-A monoclonal antibody bevacizumab was used as a reference drug. Preemptive and concomitant treatment with ultramicronized PEA delayed morphine tolerance and potentiated the analgesic effect of morphine, while counteracting morphine-induced increase of VEGF-A in the nervous system. Similar results were obtained when bevacizumab was administered together with morphine. Of note, bevacizumab showed an analgesic effect per se. In equianalgesic treatment regimens (obtained through increasing morphine doses and associating PEA), PEA resulted in lower expression of VEGF-A in dorsal root ganglia (DRG) and spinal cord compared to morphine alone. Similar results were observed for plasma levels of the soluble VEGF receptor 1 (sFLT-1). Moreover, in morphine-treated animals, two pain-related genes (i.e., Serpina3n and Eaat2) showed a more than 3-fold increase in their expression at spinal cord and DRG level, with the increase being significantly counteracted by PEA treatment. This study supports the hypothesis that the effects of PEA on morphine analgesia and tolerance may be mediated by the down-modulation of VEGF-A and sFLT-1 in the nervous system and plasma, respectively.

Background: Prolonged exposure to stress is a risk factor for the onset of several disorders. Modern life is burdened by a pervasive prevalence of stress, which represents a major societal challenge requiring new therapeutic strategies. In this context, botanical drug-based therapies can have a paramount importance.Methods: Here we studied the preventive effects of a repeated treatment (p.o. daily, 3 weeks) with a combination of Centella asiatica (200 mg/kg), Echinacea purpurea (20 mg/kg) and Zingiber officinale (150 mg/kg) standardized extracts, on the chronic social defeat stress (CSDS) deleterious outcomes. After 10 days of CSDS exposure, male mice’ performances were evaluated in paradigms relevant for social (social interaction test), emotional (tail suspension test), cognitive (novel object recognition) domains as well as for pain perception (cold plate and von Frey tests) and motor skills (rotarod). Mice were then sacrificed, the spinal cords, hippocampi and frontal cortices dissected and processed for RT-PCR analysis.Results: Extracts mix treatment prevented stress-induced social aversion, memory impairment, mechanical and thermal allodynia and reduced behavioural despair independently of stress exposure. The treatment stimulated hippocampal and cortical BDNF and TrkB mRNA levels and counteracted stress-induced alterations in pro- (TNF-α, IL-1β and IL-6) and anti-inflammatory (IL4, IL10) cytokines expression in the same areas. It also modulated expression of pain related genes (GFAP and Slc1a3) in the spinal cord.Conclusion: The treatment with the extracts mix obtained from C. asiatica, E. purpurea and Z. officinale may represent a promising strategy to promote resilience and prevent the deleterious effects induced by extended exposure to psychosocial stress.

Globally, waterborne viral infections significantly threaten public health. While current European Union regulations stipulate that drinking water must be devoid of harmful pathogens, they do not specifically address the presence of enteric viruses in water used for irrigation or food production. Traditional virus detection methods rely on molecular biology assays, requiring specialized personnel and laboratory facilities. Here, we describe an electrochemical sandwich enzyme-linked immunomagnetic assay (ELIME) for the detection of the hepatitis A virus (HAV) in water matrices. This method employed screen-printed electrodes as the sensing platform and utilized commercially available pre-activated magnetic beads to provide a robust foundation for the immunological reaction. The ELIME assay demonstrated exceptional analytical performance in only 185 min achieving a detection limit of 0.5 genomic copies per milliliter (g.c./mL) and exhibiting good reproducibility with a relative standard deviation (RSD) of 7% in HAV-spiked drinking and processing water samples. Compared with the real-time RT-qPCR method described in ISO 15216-1, the ELIME assay demonstrated higher sensitivity, although the overall linearity of the method was moderate. These analytical attributes highlight the potential of the ELIME assay as a rapid and viable alternative for HAV detection in water used for agriculture and food processing.

Psoriasis is a chronic immune-mediated inflammatory cutaneous disease characterized by elevated levels of inflammatory cytokines and adipokine Lipocalin-2 (LCN-2). Recently, natural plant-based products have been studied as new antipsoriatic compounds. We investigate the ability of a leaf extract of the marine plant Posidonia oceanica (POE) to inhibit psoriatic dermatitis in C57BL/6 mice treated with Imiquimod (IMQ). One group of mice was topically treated with IMQ (IMQ mice) for 5 days, and a second group received POE orally before each topical IMQ treatment (IMQ-POE mice). Psoriasis Area Severity Index (PASI) score, thickness, and temperature of the skin area treated with IMQ were measured in both groups. Upon sacrifice, the organs were weighed, and skin biopsies and blood samples were collected. Plasma and lesional skin protein expression of IL-17, IL-23, IFN-γ, IL-2, and TNF-α and plasma LCN-2 concentration were evaluated by ELISA. PASI score, thickness, and temperature of lesional skin were reduced in IMQ-POE mice, as were histological features of psoriatic dermatitis and expression of inflammatory cytokines and LCN-2 levels. This preliminary study aims to propose P. oceanica as a promising naturopathic anti-inflammatory treatment that could be introduced in Complementary Medicine for psoriasis.

Interleukin-6 (IL-6) plays a crucial role in autoimmunity and chronic inflammation. This study aims to develop a low-cost, simple-to-manufacture, and user-friendly label-free electrochemical point-of-care device for the rapid detection of IL-6 in patients with psoriasis. Precisely, a sandwich-based format immunosensor was developed using two primary antibodies (mAb-IL6 clone-5 and clone-7) and screen-printed electrodes modified with an inexpensive recycling electrochemical enhancing material, called biochar. mAb-IL6 clone-5 was used as a covalently immobilized capture bioreceptor on modified electrodes, and mAb-IL6 clone-7 was used to recognize the immunocomplex (Anti-IL6 clone-5 and IL-6) and form the sandwich. Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to conduct electrochemical characterization of the layer-by-layer assembly of the immunosensor, while square wave voltammetry (SWV) was used to perform the sensing. The developed immunosensor demonstrated robust analytical performance in buffer solution, with a wide linear range (LR) by varying from 2 to 250 pg/mL, a good limit of detection (LOD) of 0.78 pg/mL and reproducibility (RSD

Throughout adulthood neural stem cells divide in neurogenic niches–the dentate gyrus of the hippocampus and the subventricular zone–producing progenitor cells and new neurons. Stem cells self-renew, thus preserving their pool. Furthermore, the number of stem/progenitor cells in the neurogenic niches decreases with age. We have previously demonstrated that the cyclin-dependent kinase inhibitor p16Ink4a maintains, in aged mice, the pool of dentate gyrus stem cells by preventing their activation after a neurogenic stimulus such as exercise (running). We showed that, although p16Ink4a ablation by itself does not activate stem/progenitor cells, exercise strongly induced stem cell proliferation in p16Ink4a knockout dentate gyrus, but not in wild-type. As p16Ink4a regulates stem cell self-renewal during aging, we sought to profile the dentate gyrus transcriptome from p16Ink4a wild-type and knockout aged mice, either sedentary or running for 12 days. By pairwise comparisons of differentially expressed genes and by correlative analyses through the DESeq2 software, we identified genes regulated by p16Ink4a deletion, either without stimulus (running) added, or following running. The p16Ink4a knockout basic gene signature, i.e., in sedentary mice, involves upregulation of apoptotic, neuroinflammation- and synaptic activity-associated genes, suggesting a reactive cellular state. Conversely, another set of 106 genes we identified, whose differential expression specifically reflects the pattern of proliferative response of p16 knockout stem cells to running, are involved in processes that regulate stem cell activation, such as synaptic function, neurotransmitter metabolism, stem cell proliferation control, and reactive oxygen species level regulation. Moreover, we analyzed the regulation of these stem cell-specific genes after a second running stimulus. Surprisingly, the second running neither activated stem cell proliferation in the p16Ink4a knockout dentate gyrus nor changed the expression of these genes, confirming that they are correlated to the stem cell reactivity to stimulus, a process where they may play a role regulating stem cell activation.

Abstract Background Neuropathic pain is a clinically relevant adverse effect of several anticancer drugs that markedly impairs patients’ quality of life and frequently leads to dose reduction or therapy discontinuation. The poor knowledge about the mechanisms involved in neuropathy development and pain chronicization, and the lack of effective therapies, make treatment of chemotherapy-induced neuropathic pain an unmet medical need. In this context, the vascular endothelial growth factor A (VEGF-A) has emerged as a candidate neuropathy hallmark and its decrease has been related to pain relief. In the present study, we have investigated the role of VEGF-A and its receptors, VEGFR-1 and VEGFR-2, in pain signalling and in chemotherapy-induced neuropathy establishment as well as the therapeutic potential of receptor blockade in the management of pain. Methods Behavioural and electrophysiological analyses were performed in an in vivo murine model, by using selective receptor agonists, blocking monoclonal antibodies or siRNA-mediated silencing of VEGF-A and VEGFRs. Expression of VEGF-A and VEGFR-1 in astrocytes and neurons was detected by immunofluorescence staining and confocal microscopy analysis. Results In mice, the intrathecal infusion of VEGF-A (VEGF165 isoforms) induced a dose-dependent noxious hypersensitivity and this effect was mediated by VEGFR-1. Consistently, electrophysiological studies indicated that VEGF-A strongly stimulated the spinal nociceptive neurons activity through VEGFR-1. In the dorsal horn of the spinal cord of animals affected by oxaliplatin-induced neuropathy, VEGF-A expression was increased in astrocytes while VEGFR-1 was mainly detected in neurons, suggesting a VEGF-A/VEGFR-1-mediated astrocyte-neuron cross-talk in neuropathic pain pathophysiology. Accordingly, the selective knockdown of astrocytic VEGF-A by intraspinal injection of shRNAmir blocked the development of oxaliplatin-induced neuropathic hyperalgesia and allodynia. Interestingly, both intrathecal and systemic administration of the novel anti-VEGFR-1 monoclonal antibody D16F7, endowed with anti-angiogenic and antitumor properties, reverted oxaliplatin-induced neuropathic pain. Besides, D16F7 effectively relieved hypersensitivity induced by other neurotoxic chemotherapeutic agents, such as paclitaxel and vincristine. Conclusions These data strongly support the role of the VEGF-A/VEGFR-1 system in mediating chemotherapy-induced neuropathic pain at the central nervous system level. Thus, treatment with the anti-VEGFR-1 mAb D16F7, besides exerting antitumor activity, might result in the additional advantage of attenuating neuropathic pain when combined with neurotoxic anticancer agents.

Experimental evidence suggests that neuroinflammation is a key pathological event of many diseases affecting the nervous system. It has been well recognized that these devastating illnesses (e.g., Alzheimer’s, Parkinson’s, depression, and chronic pain) are multifactorial, involving many pathogenic mechanisms, reason why pharmacological treatments are unsatisfactory. The purpose of this study was to evaluate the efficacy of a vegetal mixture capable of offering a multiple approach required to manage the multifactoriality of neuroinflammation. A mixture composed of Zingiber officinale (150 mg kg−1), Echinacea purpurea (20 mg kg−1), and Centella asiatica (200 mg kg−1) was tested in a mouse model of systemic neuroinflammation induced by lipopolysaccharide (LPS, 1 mg kg−1). Repeated treatment with the vegetal mixture was able to completely counteract thermal and mechanical allodynia as reported by the Cold plate and von Frey tests, respectively, and to reduce the motor impairments as demonstrated by the Rota rod test. Moreover, the mixture was capable of neutralizing the memory loss in the Passive avoidance test and reducing depressive-like behavior in the Porsolt test, while no efficacy was shown in decreasing anhedonia as demonstrated by the Sucrose preference test. Finally, LPS stimulation caused a significant increase in the activation of glial cells, of the central complement proteins and of inflammatory cytokines in selected regions of the central nervous system (CNS), which were rebalanced in animals treated with the vegetal mixture. In conclusion, the vegetal mixture tested thwarted the plethora of symptoms evoked by LPS, thus being a potential candidate for future investigations in the context of neuroinflammation.

Obesity is a health concern worldwide, and its onset is multifactorial. In addition to metabolic syndrome, a high-fat diet induces many deleterious downstream effects, such as chronic systemic inflammation, a loss of gut barrier integrity, and gut microbial dysbiosis, with a reduction of many butyrate-producing bacteria. These conditions can be ameliorated by increasing legumes in the daily diet. White and kidney beans (Phaseolus vulgaris L.) and their non-nutritive bioactive component phaseolamin were demonstrated to mitigate several pathological features related to a metabolic syndrome-like condition. The aim of the present study was to investigate the molecular pathways involved in the protective effects on the intestinal and liver environment of a chronic oral treatment with P. vulgaris extract (PHAS) on a murine model of the high-fat diet. Results show that PHAS treatment has an anti-inflammatory effect on the liver, colon, and cecum. This protective effect was mediated by peroxisome proliferator-activated receptor (PPAR)-α and γ. Moreover, we also observed that repeated PHAS treatment was able to restore tight junctions’ expression and protective factors of colon and cecum integrity disrupted in HFD mice. This improvement was correlated with a significant increase of butyrate levels in serum and fecal samples compared to the HFD group. These data underline that prolonged treatment with PHAS significantly reduces some pathological features related to the metabolic syndrome-like condition, such as inflammation and intestinal barrier disruption; therefore, PHAS could be a valid tool to be associated with the therapeutic strategy.

Rheumatoid arthritis is an autoimmune disorder that causes chronic joint pain, swelling, and movement impairment, resulting from prolonged inflammation-induced cartilage and bone degradation. The pathogenesis of RA, which is still unclear, makes diagnosis and treatment difficult and calls for new therapeutic strategies to cure the disease. Recent research has identified FPRs as a promising druggable target, with AMC3, a novel agonist, showing preclinical efficacy in vitro and in vivo. In vitro, AMC3 (1–30 µM) exhibited significant antioxidant effects in IL-1β (10 ng/mL)-treated chondrocytes for 24 h. AMC3 displayed a protective effect by downregulating the mRNA expression of several pro-inflammatory and pro-algic genes (iNOS, COX-2, and VEGF-A), while upregulating genes essential for structural integrity (MMP-13, ADAMTS-4, and COLIAI). In vivo, AMC3 (10 mg kg−1) prevented hypersensitivity and restored postural balance in CFA-injected rats after 14 days. AMC3 attenuated joint alterations, reduced joint inflammatory infiltrate, pannus formation, and cartilage erosion. Chronic AMC3 administration reduced transcriptional changes of genes causing excitotoxicity and pain (EAATs and CCL2) and prevented morphological changes in astrocytes, including cell body hypertrophy, processes length, and thickness, caused by CFA in the spinal cord. This study demonstrates the usefulness of AMC3 and establishes the groundwork for further research.

One of the main issues in the cultural heritage field of restoration chemistry is the identification of greener and more effective methods for the wet cleaning of paper artefacts, which serve as witnesses to human history and custodians of cultural values. In this context, we propose a biocompatible method to perform wet cleaning on paper based on the use of 1 MHz ultrasound in combination with water-dispersed polyvinyl alcohol microbubbles (PVAMBs), followed by dabbing with PVA-based hydrogel. This method can be applied to both old and new papers. FTIR spectroscopy, X-ray diffraction, HPLC analysis, pH measurements and tensile tests were performed on paper samples, to assess the efficacy of the cleaning system. According to the results, ultrasound-activated PVAMB application allows for an efficient interaction with rough and porous cellulose paper profiles, promoting the removal of cellulose degradation byproducts, while the following hydrogel dabbing treatment guarantees the removal of cleaning materials residues. Moreover, the results also pointed out that after the treatment no thermal or mechanical damages had affected the paper. In conclusion, the readability of these kinds of artifacts can be improved without causing an alteration of their structural properties, while mitigating the risk of ink diffusion.

Tendinopathies represent about 45% of musculoskeletal lesions and they are a big burden in clinics characterized by activity-related pain, focal tendon tenderness and intra-tendinous imaging changes. Many approaches have been proposed for tendinopathies’ management (e.g., nonsteroidal anti-inflammatory drugs, corticosteroids, eccentric exercises, laser therapy), unfortunately with very little support of efficacy or serious side effects, thus making the identification of new treatments fundamental. The aim of the study was to test the protective and pain reliever effect of thymoquinone (TQ)-loaded formulations in a rat model of tendinopathy induced by carrageenan intra-tendon injection (20 µL of carrageenan 0.8% on day 1). Conventional (LP-TQ) and hyaluronic acid (HA)-coated TQ liposomes (HA-LP-TQ) were characterized and subjected to in vitro release and stability studies at 4 °C. Then, TQ and liposomes were peri-tendon injected (20 µL) on days 1, 3, 5, 7 and 10 to evaluate their antinociceptive profile using mechanical noxious and non-noxious stimuli (paw pressure and von Frey tests), spontaneous pain (incapacitance test) and motor alterations (Rota rod test). Liposomes containing 2 mg/mL of TQ and covered with HA (HA-LP-TQ2) reduced the development of spontaneous nociception and hypersensitivity for a long-lasting effect more than the other formulations. The anti-hypersensitivity effect matched with the histopathological evaluation. In conclusion, the use of TQ encapsulated in HA-LP liposomes is suggested as a new treatment for tendinopathies.

The distinction between glial painful and protective pathways is unclear and the possibility to finely modulate the system is lacking. Focusing on painful neuropathies, we studied the role of interleukin 1α (IL-1α), an alarmin belonging to the larger family of damage-associated molecular patterns endogenously secreted to restore homeostasis.The treatment of rat primary neurons with increasing doses of the neurotoxic anticancer drug oxaliplatin (0.3–100μM, 48 h) induced the release of IL-1α. The knockdown of the alarmin in neurons leads to their higher mortality when co-cultured with astrocytes. This toxicity was related to increased extracellular ATP and decreased release of transforming growth factor β1, mostly produced by astrocytes.In a rat model of neuropathy induced by oxaliplatin, the intrathecal treatment with IL-1α was able to reduce mechanical and thermal hypersensitivity both after acute injection (100 ng and 300 ng) and continuous infusion (100 and 300 ng/die−1). Ex vivo analysis on spinal purified astrocyte processes (gliosomes) and nerve terminals (synaptosomes) revealed the property of IL-1α to reduce the endogenous glutamate release induced by oxaliplatin. This protective effect paralleled with an increased number of GFAP-positive cells in the spinal cord, suggesting the ability of IL-1α to evoke a positive, conservative astrocyte phenotype.Endogenous IL-1α induced protective signals in the cross-talk between neurons and astrocytes. Exogenously administered in rats, IL-1α prevented neuropathic pain in the presence of spinal glutamate decrease and astrocyte activation.

Persistent pain can be managed with opioids, but their use is limited by the onset of tolerance. Ultramicronized N-palmitoylethanolamine (PEA) in vivo delays morphine tolerance with mechanisms that are still unclear. Since glial cells are involved in opioid tolerance and mast cells (MCs) are pivotal targets of PEA, we hypothesized that a potential mechanism by which PEA delays opioid tolerance might depend on the control of the crosstalk between these cells. Morphine treatment (30 μM, 30 min) significantly increased MC degranulation of RBL-2H3 cells, which was prevented by pre-treatment with PEA (100 μM, 18 h), as evaluated by β-hexosaminidase assay and histamine quantification. The impact of RBL-2H3 secretome on glial cells was studied. Six-hour incubation of astrocytes with control RBL-2H3-conditioned medium, and even more so co-incubation with morphine, enhanced CCL2, IL-1β, IL-6, Serpina3n, EAAT2 and GFAP mRNA levels. The response was significantly prevented by the secretome from PEA pre-treated RBL-2H3, except for GFAP, which was further upregulated, suggesting a selective modulation of glial signaling. In conclusion, ultramicronized PEA down-modulated both morphine-induced MC degranulation and the expression of inflammatory and pain-related genes from astrocytes challenged with RBL-2H3 medium, suggesting that PEA may delay morphine tolerance, regulating MC-astrocyte crosstalk.

Furosemide (FUR), an active pharmaceutical ingredient (API) belonging to a group of drugs known as loop diuretics, has widespread use, but, is characterized by a strong instability to light, which causes chemical transformations that could give a yellowing phenomenon and have a significant impact from a health and marketing point of view. Many studies have tried to explain this phenomenon under different experimental conditions, but no detailed explanation of the yellowing phenomenon has been provided. This work, unlike the others, provides an overall view and explanation of the behavior of FUR in relation to the yellowing phenomenon, both in the solution and in solid state, considering several aspects, such as light exposure, presence of oxygen, and moisture effects.

The racemic mixture dimiracetam negatively modulates NMDA-induced glutamate release in rat spinal cord synaptosomal preparations and is orally effective in models of neuropathic pain. In this study, we compared the effects of dimiracetam, its R- or S-enantiomers, and the R:S 3:1 non-racemic mixture (MP-101). In vitro, dimiracetam was more potent than its R- or S-enantiomers in reducing the NMDA-induced [3H]D-aspartate release in rat spinal cord synaptosomes. Similarly, acute oral administration of dimiracetam was more effective than a single enantiomer in the sodium monoiodoacetate (MIA) paradigm of painful osteoarthritis. Then, we compared the in vitro effects of a broad range of non-racemic enantiomeric mixtures on the NMDA-induced [3H]D-aspartate release. Dimiracetam was a more potent blocker than each isolated enantiomer but the R:S 3:1 non-racemic mixture (MP-101) was even more potent than dimiracetam, with an IC50 in the picomolar range. In the chronic oxaliplatin-induced neuropathic pain model, MP-101 showed a significantly improved anti-neuropathic profile, and its effect continued one week after treatment suspension. MP-101 also performed better than dimiracetam in animal models of cognition and depression. Based on the benign safety and tolerability profile previously observed with racemic dimiracetam, MP-101 appears to be a novel, promising clinical candidate for the prevention and treatment of several neuropathic and neurological disorders.

Conus regius is a marine venomous mollusk of the Conus genus that captures its prey by injecting a rich cocktail of bioactive disulfide bond rich peptides called conotoxins. These peptides selectively target a broad range of ion channels, membrane receptors, transporters, and enzymes, making them valuable pharmacological tools and potential drug leads. C. regius-derived conotoxins are particularly attractive due to their marked potency and selectivity against specific nicotinic acetylcholine receptor subtypes, whose signalling is involved in pain, cognitive disorders, drug addiction, and cancer. However, the species-specific differences in sensitivity and the low stability and bioavailability of these conotoxins limit their clinical development as novel therapeutic agents for these disorders. Here, we give an overview of the main pharmacological features of the C. regius-derived conotoxins described so far, focusing on the molecular mechanisms underlying their potential therapeutic effects. Additionally, we describe adoptable chemical engineering solutions to improve their pharmacological properties for future potential clinical translation.

The dentate gyrus of the hippocampus and the subventricular zone are neurogenic niches where neural stem and progenitor cells replicate throughout life to generate new neurons. The Btg1 gene maintains the stem cells of the neurogenic niches in quiescence. The deletion of Btg1 leads to an early transient increase of stem/progenitor cells division, followed, however, by a decrease during adulthood of their proliferative capability, accompanied by apoptosis. Since a physiological decrease of neurogenesis occurs during aging, the Btg1 knockout mouse may represent a model of neural aging. We have previously observed that the defective neurogenesis of the Btg1 knockout model is rescued by the powerful neurogenic stimulus of physical exercise (running). To identify genes responsible for stem and progenitor cells maintenance, we sought here to find genes underlying this premature neural aging, and whose deregulated expression could be rescued by running. Through RNA sequencing we analyzed the transcriptomic profiles of the dentate gyrus isolated from Btg1 wild-type or Btg1 knockout adult (2-month-old) mice submitted to physical exercise or sedentary. In Btg1 knockout mice, 545 genes were deregulated, relative to wild-type, while 2081 genes were deregulated by running. We identified 42 genes whose expression was not only down-regulated in the dentate gyrus of Btg1 knockout, but was also counter-regulated to control levels by running in Btg1 knockout mice, vs. sedentary. Among these 42 counter-regulated genes, alpha-synuclein (Snca), Fos, Arc and Npas4 showed significantly greater differential regulation. These genes control neural proliferation, apoptosis, plasticity and memory and are involved in aging. In particular, Snca expression decreases during aging. We tested, therefore, whether an Snca-expressing lentivirus, by rescuing the defective Snca levels in the dentate gyrus of Btg1 knockout mice, could also reverse the aging phenotype, in particular the defective neurogenesis. We found that the exogenous expression of Snca reversed the Btg1 knockout-dependent decrease of stem cell proliferation as well as the increase of progenitor cell apoptosis. This indicates that Snca has a functional role in the process of neural aging observed in this model, and also suggests that Snca acts as a positive regulator of stem cell maintenance.

A simple, eco-friendly, cost-effective and rapid microwave-assisted method has been developed to synthetize dendritic silver nanostructures, composed of silver nanoparticles (AgNPs), using white grape pomace aqueous extract (WGPE) as both reducing and capping agent. With this aim, WGPE and AgNO3 (1 mM) were mixed at different ratio, and microwave irradiated at 700 W, for 40 s. To understand the role of bioactive compounds involved in the green synthesis of AgNPs, preliminary chemical characterization, FT-IR analysis and 1H NMR metabolite profiling of WGPE were carried out. The effects of bioactive extract concentration and stability over time on AgNPs formation were also evaluated. WGPE-mediated silver nanostructures were then characterized by UV–vis, FTIR analyses, and scanning electron microscopy. Interestingly, the formation of dendritic nanostructures, originated from the self-assembly of Ag rounded nanoparticles (average diameter of 33 ± 6 nm), was observed and ascribed to the use of microwave power and the presence of organic components within the used WGPE, inducing an anisotropic crystal growth and promoting a diffusion-limited aggregation mechanism. The bio-dendritic synthetized nanostructures were also evaluated for potential applications in bio-sensing and agricultural fields. Cyclic voltammetry measurements in 0.5 M phosphate + 0.1 M KCl buffer, pH 7.4 showed that green AgNPs possess the electroactive properties typical of AgNPs produced using chemical protocol. The biological activity of synthetized AgNPs was evaluated by in-vitro antifungal activity against F. graminearum. Additionally, a phytotoxicity evaluation of synthetized green nanostructures was carried out on wheat seed germination. Results highlighted the potential of WGPE as green agent for bio-inspired nanomaterial synthesis, and of green Ag nanostructures, which can be used as antifungal agent and in biosensing applications. Keywords: Agro-food wastes, Grape pomace, Green synthesis, Dendritic silver nanostructures

This paper proposes a deep leaning technique for accurate detection and reliable classification of organic pollutants in water. The pollutants are detected by means of cyclic voltammetry characterizations made by using low-cost disposable screen-printed electrodes. The paper demonstrates the possibility of strongly improving the detection of such platforms by modifying them with nanomaterials. The classification is addressed by using a deep learning approach with convolutional neural networks. To this end, the results of the voltammetry analysis are transformed into equivalent RGB images by means of Gramian angular field transformations. The proposed technique is applied to the detection and classification of hydroquinone and benzoquinone, which are particularly challenging since these two pollutants have a similar electroactivity and thus the voltammetry curves exhibit overlapping peaks. The modification of electrodes by carbon nanotubes improves the sensitivity of a factor of about ×25, whereas the convolution neural network after Gramian transformation correctly classifies 100% of the experiments.

Biochar is a pyrolytic material with several environmental benefits such as reducing greenhouse gas emissions, sequestering atmospheric carbon and contrasting global warming. However, nowadays, it has moved to the forefront for its conductivity and electron transfer properties, finding applications in the fabrication of electrochemical platforms. In this field, researchers have focused on low-cost biomass capable of replacing more popular and expensive carbonaceous nanomaterials (i.e., graphene, nanotubes and quantum dots) in the realization of sensitive cost-effectiveness and eco-friendly electrochemical tools. This review discusses recent developments of biochar-modified screen-printed electrodes (SPEs). Special attention has been paid to biochar’s manufacturing processes, electron-donating capabilities and sensing applications. Examples of representative works are introduced to explain the distinct roles of biochar in several electro-bioanalytical strategies.

We have previously generated a mouse model (Ptch1+/−/Tis21KO), which displays high frequency spontaneous medulloblastoma, a pediatric tumor of the cerebellum. Early postnatal cerebellar granule cell precursors (GCPs) of this model show, in consequence of the deletion of Tis21, a defect of the Cxcl3-dependent migration. We asked whether this migration defect, which forces GCPs to remain in the proliferative area at the cerebellar surface, would be the only inducer of their high frequency transformation. In this report we show, by further bioinformatic analysis of our microarray data of Ptch1+/−/Tis21KO GCPs, that, in addition to the migration defect, they show activation of the PI3K/AKT/mTOR pathway, as the mRNA levels of several activators of this pathway (e.g., Lars, Rraga, Dgkq, Pdgfd) are up-regulated, while some inhibitors (e.g. Smg1) are down-regulated. No such change is observed in the Ptch1+/− or Tis21KO background alone, indicating a peculiar synergy between these two genotypes. Thus we investigated, by mRNA and protein analysis, the role of PI3K/AKT/mTOR signaling in MBs and in nodules from primary Ptch1+/−/Tis21KO MB allografted in the flanks of immunosuppressed mice. Activation of the PI3K/AKT/mTOR pathway is seen in full-blown Ptch1+/−/Tis21KO MBs, relative to Ptch1+/−/Tis21WT MBs. In Ptch1+/−/Tis21KO MBs we observe that the proliferation of neoplastic GCPs increases while apoptosis decreases, in parallel with hyper-phosphorylation of the mTOR target S6, and, to a lower extent, of AKT. In nodules derived from primary Ptch1+/−/Tis21KO MBs, treatment with MEN1611, a novel PI3K inhibitor, causes a dramatic reduction of tumor growth, inhibiting proliferation and, conversely, increasing apoptosis, also of tumor CD15+ stem cells, responsible for long-term relapses. Additionally, the phosphorylation of AKT, S6 and 4EBP1 was significantly inhibited, indicating inactivation of the PI3K/AKT/mTOR pathway. Thus, PI3K/AKT/mTOR pathway activation contributes to Ptch1+/−/Tis21KO MB development and to high frequency tumorigenesis, observed when the Tis21 gene is down-regulated. MEN1611 could provide a promising therapy for MB, especially for patient with down-regulation of Btg2 (human ortholog of the murine Tis21 gene), which is frequently deregulated in Shh-type MBs.

A novel flow injection microfluidic immunoassay system for continuous monitoring of saxitoxin, a lethal biotoxin, in seawater samples is presented in this article. The system consists of a preimmobilized G protein immunoaffinity column connected in line with a lab-on-chip setup. The detection of saxitoxin in seawater was carried out in two steps: an offline incubation step (competition reaction) performed between the analyte of interest (saxitoxin or Ag, as standard or seawater sample) and a tracer (an enzyme-conjugated antigen or Ag*) toward a specific polyclonal antibody. Then, the mixture was injected through a “loop” of a few μL using a six-way injection valve into a bioreactor, in line with the valve. The bioreactor consisted of a small glass column, manually filled with resin upon which G protein has been immobilized. When the mixture flowed through the bioreactor, all the antibody-antigen complex, formed during the competition step, is retained by the G protein. The tracer molecules that do not interact with the capture antibody and protein G are eluted out of the column, collected, and mixed with an enzymatic substrate directly within the microfluidic chip, via the use of two peristaltic pumps. When Ag* was present, a color change (absorbance variation, ΔAbs) of the solution is detected at a fixed wavelength (655 nm) by an optical chip docking system and registered by a computer. The amount of saxitoxin, present in the sample (or standard), that generates the variation of the intensity of the color, will be directly proportional to the concentration of the analyte in the analyzed solution. Indeed, the absorbance response increased proportionally to the enzymatic product and to the concentration of saxitoxin in the range of 3.5 × 10–7–2 × 10–5 ng ml−1 with a detection limit of 1 × 10–7 ng ml−1 (RSD% 15, S N−1 equal to 3). The immunoanalytical system has been characterized, optimized, and tested with seawater samples. This analytical approach, combined with the transportable and small-sized instrumentation, allows for easy in situ monitoring of marine water contaminations.

Peripheral neuropathies, characterized by altered nociceptive and muscular functions, are related to oxidative stress. Thioctic acid is a natural antioxidant existing as two optical isomers, but most clinically used as racemic mixture. The present study investigated the central nervous system’s changes which followed loose-ligation-derived compression of sciatic nerve, the putative neuroprotective role of thioctic acid and the pain-alleviating effect on low-back pain suffering patients. Loose ligation of the right sciatic nerve was performed in spontaneously hypertensive rats (SHR), a model of increased oxidative stress, and in normotensive Wistar-Kyoto rats (WKY). Animals with sciatic nerve ligation were left untreated or were treated intraperitoneally for 15 days with 250 μmol·kg−1·die−1 of (+/−)-thioctic acid; 125 μmol·kg−1·die−1 of (+/−)-thioctic acid; 125 μmol·kg−1·die−1 of (+)-thioctic acid lysine salt; 125 μmol·kg−1·die−1 of (−)-thioctic acid; 300 μmol·kg−1·die−1 pregabalin. Control SHR and WKY rats received the same amounts of vehicle. The clinical trial NESTIORADE (Sensory-Motor Neuropathies of the Sciatic Nerve: Comparative evaluation of the effect of racemic and dextro-rotatory forms of thioctic acid) examined 100 patients (49 males and 51 females aged 53 ± 11 years) dividing them into two equal-numbered groups, each treated daily for 60 days with 600 mg of (+/−)-thioctic acid or (+)-thioctic acid, respectively. The trial was registered prior to patient enrollment at EudraCT website (OSSC Number: 2011-000964-81). In the preclinical study, (+)-thioctic acid was more active than (+/−)- or (−)-enantiomers in relieving pain and protecting peripheral nerve as well as in reducing oxidative stress and astrogliosis in the spinal cord. Main findings of NESTIORADE clinical trial showed a greater influence on painful symptomatology, a quicker recovery and a better impact on quality of life of (+)-thioctic acid vs. (+/−)-thioctic acid. These data may have a pharmacological and pharmacoeconomical relevance and suggest that thioctic acid, above all (+)-enantiomer, could be considered for treatment of low-back pain involving neuropathy.

Water pollution is nowadays a global problem and the effective detection of pollutants is of fundamental importance. Herein, a facile, efficient, robust, and rapid (response time < 2 min) method for the determination of important quinone-based industrial pollutants such as hydroquinone and benzoquinone is reported. The recognition method is based on the use of screen-printed electrodes as sensing platforms, enhanced with carbon-based nanomaterials. The enhancement is achieved by modifying the working electrode of such platforms through highly sensitive membranes made of Single- or Multi-Walled Carbon Nanotubes (SWNTs and MWNTs) or by graphene nanoplatelets. The modified sensing platforms are first carefully morphologically and electrochemically characterized, whereupon they are tested in the detection of different pollutants (i.e., hydroquinone and benzoquinone) in water solution, by using both cyclic and square-wave voltammetry. In particular, the sensors based on film-deposited nanomaterials show good sensitivity with a limit of detection in the nanomolar range (0.04 and 0.07 μM for SWNT- and MWNT-modified SPEs, respectively) and a linear working range of 10 to 1000 ppb under optimal conditions. The results highlight the improved performance of these novel sensing platforms and the large-scale applicability of this method for other analytes (i.e., toxins, pollutants).

Screen-printed electrodes are the most recent generation of low-cost, mass-produced, sensitive and portable devices for the measurement of analytes of interest. The responses of these platforms, in terms of current intensity and reproducibility, are strongly influenced by factors such as printing procedures, type of ink, substrates, etc. In this paper, an improved inverse-designed screen-printed electrode (IDSPE) is proposed. The electrochemical performance is compared with that obtained using classical screen printed electrodes (SEPs), showing enhanced sensitivity and signal-to-noise ratio (background current minimization 32 ± 3nA and 0.64 ± 0.01nA, for SPE and IDSPE, respectively). A full comparison between inverse and classical screen-printed electrodes is carried out using various electroactive species (potassium ferricyanide, ascorbic acid, hexaammineruthenium(III) chloride and NADH) and two different electrochemical techniques (cyclic and square-wave voltammetry). In tests conducted with potassium ferricyanide the sensitivity of the IDSPE shows a nearly four-fold improvement, and a limit of detection three times better than the values obtained employing the classical SPE. The reproducibility (RSD%) in tests conducted with ascorbic acid is 6% and 12% for IDSPE and SPE, respectively. Moreover, surface modification of both screen-printed electrodes (SPE and IDSPE) with biochar obtained from recycled brewers waste (Bio-SPEs and Bio-IDSPEs), further improves their electrochemical performance, in terms, for example, of the heterogeneous electron transfer constant (0.0024 and 0.0018 for Bio-SPE and Bio-IDSPE, respectively).

Epigenetic mechanisms are involved in epilepsy and chronic pain development. About that, we studied the effects of the natural histone deacetylase (HDAC) inhibitor sodium butyrate (BUT) in comparison with valproic acid (VPA) in a validated genetic model of generalized absence epilepsy and epileptogenesis. WAG/Rij rats were treated with BUT (30 mg/kg), VPA (300 mg/kg), and their combination (BUT + VPA) daily per os for 6 months. Rats were subjected at Randall–Selitto, von Frey, hot plate, and tail flick tests after 1, 3, and 6 months of treatment to evaluate hypersensitivity to noxious and non-noxiuous stimuli. Moreover, PPAR-γ (G3335 1 mg/kg), GABA-B (CGP35348 80 mg/kg), and opioid (naloxone 1 mg/kg) receptor antagonists were administrated to investigate the possible mechanisms involved in analgesic activity. The expression of NFkB, glutathione reductase, and protein oxidation (carbonylation) was also evaluated by Western blot analysis. WAG/Rij rats showed an altered pain threshold throughout the study (p < 0.001). BUT and BUT + VPA treatment reduced hypersensitivity (p < 0.01). VPA was significantly effective only after 1 month (p < 0.01). All the three receptors are involved in BUT + VPA effects (p < 0.001). BUT and BUT + VPA decreased the expression of NFkB and enhanced glutathione reductase (p < 0.01); protein oxidation (carbonylation) was reduced (p < 0.01). No effect was reported with VPA. In conclusion BUT, alone or in coadministration with VPA, is a valuable candidate for managing the epilepsy-related persistent pain.

Opioids are broad spectrum analgesics that are an integral part of the therapeutic armamentarium to combat pain in the clinical practice. Unfortunately, together with analgesia, a number of adverse effects can occur such as nausea, vomiting, constipation, gastrointestinal alterations and cognitive impairments. Naltrexone is a competitive antagonist of opioid receptors commonly used to treat opioid addiction; its oral use against agonists side effects is limited by the decrease of opioids-therapeutic efficacy and own adverse effects. The intranasal delivery of naltrexone could offer a quick and effective achievement of CNS based on extracellular mechanisms including perineural and perivascular transport. The aim of the study was to test the efficacy of intranasal low-dose naltrexone in reducing intraperitoneal morphine and oxycodone side effects in rodents. In mice, 1 μg naltrexone intranasally administered 30 min before opioids reduced cognitive impairments and motor alteration induced by 10 mg kg−1 morphine and 60 mg kg−1 oxycodone in the Passive avoidance and Rota rod tests, respectively. Moreover, naltrexone rebalanced opioid-induced reduction of the intestinal transit and latency of feces expulsion as well as food intake inhibition. Importantly, 1 μg naltrexone instillation did not block analgesia as demonstrated by the Hot plate test. In rats, intranasal naltrexone counteracted the opioid-induced pica phenomenon related to emesis and increased water and palatable food intake. The effects were comparable to that achieved by metoclopramide used as reference drug. Treatments did not influence body weight. Lastly, the safety of the intranasal delivery has been checked by hematoxylin–eosin staining that did not show histological alterations of the nasal cavity. In conclusion, intranasal low-dose naltrexone counteracted morphine and oxycodone induced gastrointestinal and CNS side effects without impairing opioid analgesia. It is a candidate to be a valid clinical strategy deserving deep analysis.

In the adult mammalian brain new neurons are continuously generated throughout life in two niches, the dentate gyrus of the hippocampus and the subventricular zone. This process, called adult neurogenesis, starts from stem cells, which are activated and enter the cell cycle. The proliferative capability of stem cells progressively decreases during aging. The population of stem cells is generally quiescent, and it is not clear whether the potential for stem cells to expand is limited, or whether they can expand and then return to quiescence, remaining available for further activation. Certain conditions may deregulate stem cells quiescence and self-renewal. In fact we discuss the possibility of activation of stem cells by neurogenic stimuli as a function of the intensity of the stimulus (i.e., whether this is physiological or pathological), and of the deregulation of the system (i.e., whether the model is aged or carrying genetic mutations in the gene network controlling quiescence). It appears that when the system is aged and/or carrying mutations of quiescence-maintaining genes, preservation of the quiescent state of stem cells is more critical and stem cells can be activated by a neurogenic stimulus which is ineffective in normal conditions. Moreover, when a neurogenic stimulus is in itself a cause of brain damage (e.g., kainic acid treatment) the activation of stem cells occurs bypassing any inhibitory control. Plausibly, with strong neurogenic stimuli, such as kainic acid injected into the dentate gyrus, the self-renewal capacity of stem cells may undergo rapid exhaustion. However, the self-renewal capability of stem cells persists when normal stimuli are elicited in the presence of a mutation of one of the quiescence-maintaining genes, such as p16Ink4a, p21Cip1 or Btg1. In this case, stem cells become promptly activated by a neurogenic stimulus even during aging. This indicates that stem cells retain a high proliferative capability and plasticity, and suggests that stem cells are protected against the response to stimulus and are resilient to exhaustion. It will be interesting to assess at which functional degree of deregulation of the quiescence-maintaining system, stem cells will remain responsive to repeated neurogenic stimuli without undergoing exhaustion of their pool.

Kappa opioid receptor (KOPr) agonists represent alternative analgesics for their low abuse potential, although relevant adverse effects have limited their clinical use. Functionally selective KOPr agonists may activate, in a pathway-specific manner, G protein-mediated signaling, that produces antinociception, over β-arrestin 2-dependent induction of p38MAPK, which preferentially contributes to adverse effects. Thus, functionally selective KOPr agonists biased toward G protein-coupled intracellular signaling over β-arrestin-2-mediated pathways may be considered candidate therapeutics possibly devoid of many of the typical adverse effects elicited by classic KOPr agonists. Nonetheless, the potential utility of functionally selective agonists at opioid receptors is still highly debated; therefore, further studies are necessary to fully understand whether it will be possible to develop more effective and safer analgesics by exploiting functional selectivity at KOPr. In the present study we investigated in vitro functional selectivity and in vivo antinociceptive effects of LOR17, a novel KOPr selective peptidic agonist that we synthesized. LOR17-mediated effects on adenylyl cyclase inhibition, ERK1/2, p38MAPK phosphorylation, and astrocyte cell proliferation were studied in HEK-293 cells expressing hKOPr, U87-MG glioblastoma cells, and primary human astrocytes; biased agonism was investigated via cAMP ELISA and β-arrestin 2 recruitment assays. Antinociception and antihypersensitivity were assessed in mice via warm-water tail-withdrawal test, intraperitoneal acid-induced writhing, and a model of oxaliplatin-induced neuropathic cold hypersensitivity. Effects of LOR17 on locomotor activity, exploratory activity, and forced-swim behavior were also assayed. We found that LOR17 is a selective, G protein biased KOPr agonist that inhibits adenylyl cyclase and activates early-phase ERK1/2 phosphorylation. Conversely to classic KOPr agonists as U50,488, LOR17 neither induces p38MAPK phosphorylation nor increases KOPr-dependent, p38MAPK-mediated cell proliferation in astrocytes. Moreover, LOR17 counteracts, in a concentration-dependent manner, U50,488-induced p38MAPK phosphorylation and astrocyte cell proliferation. Both U50,488 and LOR17 display potent antinociception in models of acute nociception, whereas LOR17 counteracts oxaliplatin-induced thermal hypersensitivity better than U50,488, and it is effective after single or repeated s.c. administration. LOR17 administered at a dose that fully alleviated oxaliplatin-induced thermal hypersensitivity did not alter motor coordination, locomotor and exploratory activities nor induced pro-depressant-like behavior. LOR17, therefore, may emerge as a novel KOPr agonist displaying functional selectivity toward G protein signaling and eliciting antinociceptive/antihypersensitivity effects in different animal models, including oxaliplatin-induced neuropathy.

About 30% of medulloblastomas (MBs), a tumor of the cerebellum, arise from cerebellar granule cell precursors (GCPs) undergoing transformation following activation of the Sonic hedgehog (Shh) pathway. To study this process, we generated a new MB model by crossing Patched1 heterozygous (Ptch1+/−) mice, which develop spontaneous Shh-type MBs, with mice lacking B-cell translocation gene 1 (Btg1), a regulator of cerebellar development. In MBs developing in Ptch1+/− mice, deletion of Btg1 does not alter tumor and lesion frequencies, nor affect the proliferation of neoplastic precursor cells. However, in both tumors and lesions arising in Ptch1+/− mice, ablation of Btg1 increases by about 25% the apoptotic neoplastic precursor cells, as judged by positivity to activated caspase-3. Moreover, although Btg1 ablation in early postnatal GCPs, developing in the external granule cell layer, leads to a significant increase of proliferation, and decrease of differentiation, relative to wild-type, no synergy occurs with the Ptch1+/− mutation. However, Btg1 deletion greatly increases apoptosis in postnatal GCPs, with strong synergy between Btg1-null and Ptch1+/− mutations. That pronounced increase of apoptosis observed in Ptch1+/−/Btg1 knockout young or neoplastic GCPs may be responsible for the lack of effect of Btg1 ablation on tumorigenesis. This increased apoptosis may be a consequence of increased expression of protein arginine methyltransferase 1 (Prmt1) protein that we observe in Btg1 knockout/Ptch1+/− MBs. In fact, apoptotic genes, such as BAD, are targets of Prmt1. Moreover, in Btg1-null MBs, we observed a two-fold increase of cells positive to CD15, which labels tumor stem cells, raising the possibility of activation of quiescent tumor cells, known for their role in long-term resistance to treatment and relapses. Thus, Btg1 appears to play a role in cerebellar tumorigenesis by regulating the balance between apoptosis and proliferation during MB development, also influencing the number of tumor stem cells.

Most therapies used in patients affected by inflammatory bowel diseases are ineffective in preventing the development of chronic visceral hypersensitivity, mainly due to inflammation-induced enteric neuroplasticity. Glucosinolates, secondary metabolites mainly of Brassicaceae with anti-inflammatory and neuroprotective properties, are effective in treating both neuropathic and arthritis pain through H2S release and Kv7 potassium channel activation. The aim of this work was to investigate the protective and anti-hyperalgesic efficacy of a defatted seed meal from Eruca sativa Mill. (Brassicaceae), rich in glucosinolates, in a rat model of colitis induced by 2,4-dinitrobenzene sulfonic acid (DNBS). The mechanisms of action were also investigated. Visceral pain was assessed by measuring the abdominal response to colorectal distension. Fifteen days after colitis induction, the acute administration of E. sativa defatted seed meal (0.1–1 g kg−1 p.o.) dose-dependently relieved pain. This effect was hampered by co-administering an H2S scavenger or a selective Kv7 blocker. Administering E. sativa (1 g kg−1) for 14 days, starting after DNBS injection, contributed to counteracting visceral pain persistence in the post-inflammatory phase of colitis by promoting colon healing from the damage and reducing enteric gliosis. E. sativa defatted seed meal might be employed as a nutraceutical tool for supporting abdominal pain relief in patients.

Chronic pain management requires increasing doses of opioids, the milestone of painkillers, which may result in the onset of tolerance with exacerbated side effects. Maintaining stable analgesia with low doses of opioids is thus imperative. N-palmitoylethanolamine (PEA) is an endogenous lipid compound endowed with pain-relieving as well as anti-inflammatory properties. The ultramicronized formulation of PEA was recently demonstrated to be able to modulate morphine’s effects, delaying tolerance and improving efficacy. To evaluate the possible application to other opioids, in this study, we analysed the capacity of ultramicronized PEA to regulate analgesia and tolerance induced by oxycodone and tramadol. Pre-emptive and continuative treatment with ultramicronized PEA (30 mg kg−1, daily, per os) delayed the onset of opioid tolerance and enhanced opioid analgesia when it was acutely administered in association with tramadol (20 mg kg−1, daily, subcutaneously) or oxycodone (0.5 mg kg−1, daily, subcutaneously). Moreover, PEA exerted antinociceptive effects on tolerant rats, suggesting the use of PEA together with opioids for stable, long-lasting analgesia. To that purpose, the oxycodone dose needed to be increased from 0.3 mg kg−1 (day 1) up to 1 mg kg−1 (day 31) in the oxycodone + vehicle group; the tramadol dose was progressively enhanced from 15 mg kg−1 to 50 mg kg−1 in 31 days in the tramadol + vehicle group. Acute oral co-treatment with PEA (120 mg kg−1) achieved the same analgesia without increasing the dose of both opioids. The behavioural effects of PEA on opioid chronic treatment paralleled a decrease in astrocyte activation in the dorsal horn of the spinal cord (a marker of the development of opioid tolerance) and with a modulation of mRNA expression of IL-6 and serpin-A3. In conclusion, pre- and co-administration of ultramicronized PEA delayed the development of tramadol tolerance, potentiating either oxycodone or tramadol analgesia and allowing a long-lasting analgesic effect with a low opioid dose regimen. The use of PEA is suggested for clinical purposes to support the opioid-based management of persistent pain.

The use of carbon nanomaterials (CNMs) in sensors and biosensor realization is one of the hottest topics today in analytical chemistry. In this work, a comparative in-depth study, exploiting different nanomaterial (MWNT-CO2H, -NH2, -OH and GNP) modified screen-printed electrodes (SPEs), is reported. In particular, the sensitivity, the heterogeneous electron transfer constant (k0), and the peak-to-peak separation (ΔE) have been calculated and analyzed. After which, an electrochemical amperometric sensor capable of determining uric acid (UA), based on the nano-modified platforms previously characterized, is presented. The disposable UA biosensor, fabricated modifying working electrode (WE) with Prussian Blue (PB), carbon nanotubes, and uricase enzyme, showed remarkable analytical performances toward UA with high sensitivity (CO2H 418 μA μM−1 cm−2 and bare SPE-based biosensor, 33 μA μM−1 cm−2), low detection limits (CO2H 0.5 nM and bare SPE-based biosensors, 280 nM), and good repeatability (CO2H and bare SPE-based biosensors, 5% and 10%, respectively). Moreover, the reproducibility (RSD%) of these platforms in tests conducted for UA determination in buffer and urine samples results are equal to 6% and 15%, respectively. These results demonstrate that the nanoengineered electrode exhibited good selectivity and sensitivity toward UA even in the presence of interfering species, thus paving the way for its application in other bio-fluids such as simple point-of-care (POC) devices.

Acute inflammation is particularly relevant in the pathogenesis of visceral hypersensitivity associated with inflammatory bowel diseases. Glia within the enteric nervous system, as well as within the central nervous system, contributes to neuroplasticity during inflammation, but whether enteric glia has the potential to modify visceral sensitivity following colitis is still unknown. This work aimed to investigate the occurrence of changes in the neuron–glial networks controlling visceral perception along the gut–brain axis during colitis, and to assess the effects of peripheral glial manipulation. Enteric glia activity was altered by the poison fluorocitrate (FC; 10 µmol kg−1 i.p.) before inducing colitis in animals (2,4-dinitrobenzenesulfonic acid, DNBS; 30 mg in 0.25 mL EtOH 50%), and visceral sensitivity, colon damage, and glia activation along the pain pathway were studied. FC injection significantly reduced the visceral hyperalgesia, the histological damage, and the immune activation caused by DNBS. Intestinal inflammation is associated with a parallel overexpression of TRPV1 and S100β along the gut–brain axis (colonic myenteric plexuses, dorsal root ganglion, and periaqueductal grey area). This effect was prevented by FC. Peripheral glia activity modulation emerges as a promising strategy for counteracting visceral pain induced by colitis.

A3 adenosine receptor (A3AR) agonists have emerged as potent relievers of neuropathic pain by a T cell-mediated production of IL-10. The H4 histamine receptor (H4R), also implicated in pain modulation, is expressed on T cells playing a preeminent role in its activation and release of IL-10. To improve the therapeutic opportunities, this study aimed to verify the hypothesis of a possible cross-talk between A3AR and H4R in the resolution of neuropathic pain. In the mouse model of Chronic Constriction Injury (CCI), the acute intraperitoneal co-administration of the A3AR agonist IB-MECA (0.5 mg/kg) and the H4R agonist VUF 8430 (10 mg/kg), were additive in counteracting mechano-allodynia increasing IL-10 plasma levels. In H4R−/− mice, IB-MECA activity was reduced, lower pain relief and lower modulation of plasma IL-1β, TNF-α, IL-6 and IL-10 were shown. The complete anti-allodynia effect of IB-MECA in H4R−/− mice was restored after intravenous administration of CD4+ T cells obtained from naïve wild type mice. In conclusion, a role of the histaminergic system in the mechanism of A3AR-mediated neuropathic pain relief was suggested highlighting the driving force evoked by CD4+ T cells throughout IL-10 up-regulation.

Persistent pain affecting patients with inflammatory bowel diseases (IBDs) is still very difficult to treat. Carbonic anhydrase (CA) represents an intriguing pharmacological target considering the anti-hyperalgesic efficacy displayed by CA inhibitors in both inflammatory and neuropathic pain models. The aim of this work was to evaluate the effect of inhibiting CA IV, particularly when expressed in the gut, on visceral pain associated with colitis induced by 2,4-di-nitrobenzene sulfonic acid (DNBS) in rats. Visceral sensitivity was assessed by measuring animals’ abdominal responses to colorectal distension. Repeated treatment with the selective CA IV inhibitors AB-118 and NIK-67 effectively counteracted the development of visceral pain induced by DNBS. In addition to pain relief, AB-118 showed a protective effect against colon damage. By contrast, the anti-hyperalgesic activity of NIK-67 was independent of colon healing, suggesting a direct protective effect of NIK-67 on visceral sensitivity. The enzymatic activity and the expression of CA IV resulted significantly increased after DNBS injection. NIK-67 normalised CA IV activity in DNBS animals, while AB-118 was partially effective. None of these compounds influenced CA IV expression through the colon. Although further investigations are needed to study the underlying mechanisms, CA IV inhibitors are promising candidates in the search for therapies to relieve visceral pain in IBDs.

The new configurationally stable, unsymmetrical 2,12-diaza[6]helicene was synthesized as a racemate and the enantiomers were separated in an enantiopure state by semi-preparative HPLC on chiral stationary phase. Under selected alkylation conditions it was possible to obtain both the enantiopure 2-N-mono- and di-N-ethyl quaternary iodides. Metathesis with bis(trifluoromethanesulfonyl)imide anion gave low-melting salts which were tested as inherently chiral additives to achiral ionic liquids for the electrochemical enantiodiscrimination of chiral organic probes in voltammetric experiments. Remarkable differences in the oxidation potentials of the enantiomers of two probes, a chiral ferrocenyl amine and an aminoacid, were achieved; the differences increase with increasing additive concentration and number of alkylated nitrogen atoms.

Osteoarthritis is the most widespread joint-affecting disease. The management of persistent pain remains inadequate and demands new therapeutic strategies. In this study, we explored the pain relieving and protective properties of a single intra-articular (i.a.) injection of khellin loaded in nanovesicles (K-Ves) based on ascorbyl decanoate plus phosphatidylcholine in a rat model of osteoarthritis (OA) induced by monosodium iodoacetate (MIA) treatment. The developed nanovesicles (approximately 136 nm) had a narrow size distribution (PdI 0.26), a good recovery (about 80%) and a worthy encapsulation efficiency (about 70%) with a ζ-potential of about −40 mV. The stability of K-Ves was assessed in simulated synovial fluid. Seven days after the articular damage with MIA, both K-Ves and a suspension of khellin (K, 50 μL) were i.a. injected. K-Ves significantly counteracted MIA-induced hypersensitivity to mechanical noxious (paw pressure test) and non-noxious stimuli (von Frey test) and significantly reduced the postural unbalance related to spontaneous pain (incapacitance test) and the motor alterations (beam balance test) 7 and 14 days after the i.a. injection. K was partially active only on day 7 after the treatment. The histology emphasized the improvement of several morphological factors in MIA plus K-Ves-treated animals. In conclusion, K-Ves could be successfully used for the local treatment of osteoarthritis.

In this work, an innovative coating strategy that is able to prolong the shelf-life of fresh-cut kiwifruit was proposed, and the effectiveness of the procedure was evaluated for a period of ten days under cold storage (4 °C). Alginate (2% m/v) functionalized with green extracts from hop (Humulus lupulus L.) cones (HE; 0.5 and 1%, v/v) was used as a coating material in order to assess the best performing strategy, leading to the most stable product. At the concentrations used to formulate the edible coatings, no contribution related to hop bitterness on the final product was recorded. The results were compared to control samples (without edible coating and coated only with alginate at 2% m/v). The plant extract was characterized by its main chemical traits and by 1H NMR profiling, revealing the presence of antioxidant and antimicrobial bioactive compounds (i.e., alpha and beta hop acids, xanthohumol). Furthermore, the characteristics of the samples during cold storage were evaluated by physico-chemical (i.e., weight loss, soluble solid content, titratable acidity, pH, color attributes) and nutraceutical (i.e., total polyphenol, ascorbic acid content, total carotenoids, chlorophylls) traits. The results showed that the incorporation of hop extracts into the edible coatings tested was able to preserve the quality and nutraceutical traits of fresh-cut kiwifruit during cold storage, thus prolonging their shelf life and marketability.

Abstract Neurotoxicity is a major side effect of platinum derivatives both during and after treatment. In the absence of effective pharmacological compounds, the opportunity to identify safe adjuvant treatments among medicinal plants seems appropriate. Astragali radix is an adaptogenic herbal product recently analyzed in platinum-treated cancer patients. With the aim of evaluating the anti-neuropathic profile of Astragali radix, a previously characterized aqueous (Aqu) and two hydroalcoholic (20%HA and 50%HA) extracts were tested in a rat model of oxaliplatin-induced neuropathy. Repeated administrations significantly reduced oxaliplatin-dependent hypersensitivity with 50%HA, the most effective, fully preventing mechanical and thermal hypersensitivity. Ex vivo, 50%HA reduced morphometric and molecular alterations induced by oxaliplatin in peripheral nerve and dorsal-root-ganglia. In the spinal cord and in brain areas, 50%HA significantly decreased activation of microglia and astrocytes. Furthermore, 50%HA prevented the nephro- and hepato-toxicity induced by the anticancer drug. The protective effect of 50%HA did not alter oxaliplatin-induced apoptosis in colon tumors of Pirc rats, an Apc-driven model of colon carcinogenesis. The hydroalcoholic extract (50%HA) of Astragali radix relieves pain and promotes the rescue mechanisms that protect nervous tissue from the damages triggering chronic pain. A safe profile strongly suggests the usefulness of this natural product in oxaliplatin-induced neuropathy.

In the neurogenic niches—the dentate gyrus of the hippocampus and the subventricular zone (SVZ) adjacent to lateral ventricles—stem cells continue to divide during adulthood, generating progenitor cells and new neurons, and to self-renew, thus maintaining the stem cell pool. During aging, the numbers of stem/progenitor cells in the neurogenic niches are reduced. The preservation of the neurogenic pool is committed to a number of antiproliferative genes, with the role of maintaining the quiescence of neural cells. The cyclin-dependent kinase inhibitor p16Ink4a, whose expression increases with age, controls the expansion of SVZ aging stem cells, since in mice its deficiency prevents the decline of neurogenesis in SVZ. No change of neurogenesis is however observed in the p16Ink4a-null dentate gyrus. Here, we hypothesized that p16Ink4a plays a role as a regulator of the self-renewal of the stem cell pool also in the dentate gyrus, and to test this possibility we stimulated the dentate gyrus neural cells of p16Ink4a-null aging mice with physical exercise, a powerful neurogenic activator. We observed that running highly induced the generation of new stem cells in the p16Ink4a-null dentate gyrus, forcing them to exit from quiescence. Stem cells, notably, are not induced to proliferate by running in wild-type (WT) mice. Moreover, p16Ink4a-null progenitor cells were increased by running significantly above the number observed in WT mice. The new stem and progenitor cells generated new neurons, and continued to actively proliferate in p16Ink4a-null mice longer than in the WT after cessation of exercise. Thus, p16Ink4a prevents aging dentate gyrus stem cells from being activated by exercise. Therefore, p16Ink4a may play a role in the maintenance of dentate gyrus stem cells after stimulus, by keeping a reserve of their self-renewal capacity during aging.

Folic acid (FA) is the synthetic surrogate of the essential B vitamin folate, alternatively named folacin, pteroylglutamic acid or vitamin B9. FA is an electroactive compound that helps our body to create and keep our cells healthy: it acts as the main character in a variety of synthetic biological reactions such as the synthesis of purines, pyrimidine (thus being indirectly implied in DNA synthesis), fixing and methylation of DNA. Therefore, physiological folate deficiency may be responsible for severe degenerative conditions, including neural tube defects in developing embryos and megaloblastic anaemia at any age. Moreover, being a water-soluble molecule, it is constantly lost and has to be reintegrated daily; for this reason, FA supplements and food fortification are, nowadays, extremely diffused and well-established practices. Consequently, accurate, reliable and precise analytical techniques are needed to exactly determine FA concentration in various media. Thus, the aim of this review is to report on research papers of the past 5 years (2016–2020) dealing with rapid and low-cost electrochemical determination of FA in food or biological fluid samples.

Configurationally stable 5-aza[6]helicene (1) was envisaged as a promising scaffold for non-conventional ionic liquids (IL)s. It was prepared, purified, and separated into enantiomers by preparative HPLC on a chiral stationary phase. Enantiomerically pure quaternary salts of 1 with appropriate counterions were prepared and fully characterized. N-octyl-5-aza[6]helicenium bis triflimidate (2) was tested in very small quantities as a selector in achiral IL media to perform preliminary electrochemical enantiodifferentiation experiments on the antipodes of two different chiral probes. The new organic salt exhibited outstanding enantioselection performance with respect to these probes, thus opening the way to applications in the enantioselective electroanalysis of relevant bioactive molecules.

Posidonia oceanica (L.) Delile is traditionally used for its beneficial properties. Recently, promising antioxidant and anti-inflammatory biological properties emerged through studying the in vitro activity of the ethanolic leaves extract (POE). The present study aims to investigate the anti-inflammatory and analgesic role of POE in mice. Inflammatory pain was modeled in CD-1 mice by the intraplantar injection of carrageenan, interleukin IL-1β and formalin. Pain threshold was measured by von Frey and paw pressure tests. Nociceptive pain was studied by the hot-plate test. POE (10–100 mg kg−1) was administered per os. The paw soft tissue of carrageenan-treated animals was analyzed to measure anti-inflammatory and antioxidant effects. POE exerted a dose-dependent, acute anti-inflammatory effect able to counteract carrageenan-induced pain and paw oedema. Similar anti-hyperalgesic and anti-allodynic results were obtained when inflammation was induced by IL-1β. In the formalin test, the pre-treatment with POE significantly reduced the nocifensive behavior. Moreover, POE was able to evoke an analgesic effect in naïve animals. Ex vivo, POE reduced the myeloperoxidase activity as well as TNF-α and IL-1β levels; further antioxidant properties were highlighted as a reduction in NO concentration. POE is the candidate for a new valid strategy against inflammation and pain.