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The mammalian brain stores and distinguishes among episodic memories, i.e. memories formed during the personal experience, through a mechanism of pattern separation computed in the hippocampal dentate gyrus. Decision-making for food-related behaviors, such as the choice and intake of food, might be affected in obese subjects by alterations in the retrieval of episodic memories. Adult neurogenesis in the dentate gyrus regulates the pattern separation. Several molecular factors affect adult neurogenesis and exert a critical role in the development and plasticity of newborn neurons. Orexin-A/hypocretin-1 and downstream endocannabinoid 2-arachidonoylglycerol signaling are altered in obese mice. Here, we show that excessive orexin-A/2-arachidonoylglycerol/cannabinoid receptor type-1 signaling leads to the dysfunction of adult hippocampal neurogenesis and the subsequent inhibition of plasticity and impairment of pattern separation. By inhibiting orexin-A action at orexin-1 receptors we rescued both plasticity and pattern separation impairment in obese mice, thus providing a molecular and functional mechanism to explain alterations in episodic memory in obesity., The authors show that adult hippocampal neurogenesis is altered in the dentate gyrus of obese mice with subsequent inhibition of long-term potentiation and impairment of pattern separation. Inhibition of orexin-A action at orexin-1 receptors rescued both impairments in obese mice.

Traumatic brain injury (TBI) is one of the main causes of death in young people for which currently no efficacious treatment exists. Recently, we have reported that mice with mild-TBI with a specific injury in the insula showed elevated levels of a little investigated N-acyl amino acid, N-oleoylglycine (OlGly). N-acyl amino acids have recently experienced an increased interest because of their important biological activities. They belong to the endocannabinoidome family of lipids with structural similarities with the endocannabinoids (eCBs). The aim of this study was to test the neuroprotective and antihyperalgesic actions of OlGly in a model of mouse mild-TBI (mTBI) and its effect on levels of eCBs and N-acylethanolamines at the end of treatment. Following mTBI, mice were administered a daily injection of OlGly (10-50-100 mg/kg i.p.) for 14 days. Treatment with OlGly normalized motor impairment and behavior in the light/dark box test, ameliorated TBI-induced thermal hyperalgesia and mechanical allodynia, and normalized aggressiveness and depression. Moreover, levels of eCBs and some N-acylethanolamines underwent significant changes 60 days after TBI, especially in the prefrontal cortex and hypothalamus, and OlGly reversed some of these changes. In conclusion, our findings reveal that OlGly ameliorates the behavioral alterations associated with mTBI in mice, while concomitantly modulating eCB and eCB-like mediator tone.

Chronic pain is a persistent, complex condition that contributes to impaired mood, anxiety and emotional problems. Osteoarthritis (OA) is one of the major causes of chronic pain in adults and elderly people. A substantial body of evidence demonstrate that hippocampal neural circuits, especially monoamine dopamine and serotonin levels, contributes to negative affect and avoidance motivation experienced during pain. Current pharmacological strategies for OA patients are unsatisfying and the endocannabinoid system modulation might represent an alternative for the treatment of OA-related pain. In the present study, we used a rat model of osteoarthritis induced by intra-articular injection of sodium monoiodoacetate to assess, 28 days post-induction, the contribution of endocannabinoid system on the possible alteration in pain perception and affective behavior, in LTP and monoamine levels in the lateral entorhinal cortex-dentate gyrus pathway. The results show that OA-related chronic pain induces working memory impairment and depressive-like behavior appearance, diminishes LTP, decreases dopamine levels and increases serotonin levels in the rat dentate gyrus. URB597 administration (i.p., 1 mg/kg) reduces hyperalgesia and mechanical allodynia, improves recognition memory and depressive-live behavior, restores LTP and normalizes monoamine levels in the hippocampus. The effect was observed 60-120 min post-treatment and was blocked by AM251, which proves the action of URB597 via the CB

Following injury, the endocannabinoid system is activated in the brain suggesting a strategic role in the self-repair mechanisms. Indeed endocannabinoid system manipulation ameliorates traumatic brain injury (TBI) symptoms. Cannabidiol (CBD), together with △9-tetrahydrocannabinol (THC), is the main phytocannabinoid extracted from the plant Cannabis sativa, and it plays anti-inflammatory, antioxidant, neuroprotective, anticonvulsant, hypnotic, and antiemetic effects and has proven to be useful in neuropsychiatric, neurodegenerative, post-traumatic stress, and ischemic disorders. Unlike THC, CBD is not psychoactive and enhances the beneficial and reduces the side effects of THC. CBD has negligible action on cannabinoid receptors and modulates the endocannabinoid system throughout the inhibition of endocannabinoid degradation and reuptake. It also stimulates serotonin 1A (5-HT1A), adenosine 2A (A2A), transient receptor potential vanilloid subtype 1 (TRPV1), and nuclear peroxisome proliferator-activated receptor γ (PPARγ). We collect in this chapter all the preclinical and clinical evidence on the beneficial effects of CBD in the TBI considering it important for two main reasons: the lack of effective therapy for the TBI and the good tolerability of the CBD.

Osteoarthritis is a very disabling disease that can be treated with both non-pharmacological and pharmacological approaches. In the last years, pharmaceutical-grade chondroitin sulfate (CS) and glucosamine emerged as symptomatic slow-acting molecules, effective in pain reduction and improved function in patients affected by osteoarthritis. CS is a sulfated glycosaminoglycan that is currently produced mainly by extraction from animal tissues, and it is commercialized as a pharmaceutical-grade ingredient and/or food supplement. However, public concern on animal product derivatives has prompted the search for alternative non-extractive production routes. Thus, different approaches were established to obtain animal-free natural identical CS. On the other hand, the unsulfated chondroitin, which can be obtained via biotechnological processes, demonstrated promising anti-inflammatory properties in vitro, in chondrocytes isolated from osteoarthritic patients. Therefore, the aim of this study was to explore the potential of chondroitin, with respect to the better-known CS, in an in vivo mouse model of knee osteoarthritis. Results indicate that the treatment with biotechnological chondroitin (BC), similarly to CS, significantly reduced the severity of mechanical allodynia in an MIA-induced osteoarthritic mouse model. Decreased cartilage damage and a reduction of inflammation- and pain-related biochemical markers were also observed. Overall, our data support a beneficial activity of biotechnological unsulfated chondroitin in the osteoarthritis model tested, thus suggesting BC as a potential functional ingredient in pharmaceuticals and nutraceuticals with the advantage of avoiding animal tissue extraction.

Neuropathic pain has long-term consequences in terms of affective and cognitive disturbances suggesting the involvement of supraspinal mechanisms. In the present study, we used the spared nerve injury (SNI) model to characterize the development of sensory and aversive components of neuropathic pain, and to determine their electrophysiological impact across PFC and limbic regions. Moreover, we evaluated the regulation of several genes involved in immune response and inflammation triggered by the SNI.We showed that SNI led to sensorial hypersensitivity (cold and mechanical stimuli) and depressive-like behavior lasting 12 months after nerve injury. Interestingly, changes in non-emotional cognitive tasks (novel object recognition and Y maze) showed in 1-month SNI mice, were not evident normal in 12 months-SNI animals. In vivo electrophysiology revealed an impaired the Long Term Potentiation (LTP) at prefrontal cortex (PFC)- nucleus accumbens core (NAcore) pathway in both 1 and 12 months SNI. On the other hand, a reduced neural activity was recorded in the lateral entorhinal cortex (LEC)-dentate gyrus (DG) pathway in 1 month-, but not in 12 months- SNI mice. Finally, we observed the upregulation of specific genes involved in involved in immune response in the hippocampus of 1 month-, but not 12 months-SNI mice, suggesting a neuroinflammatory response which may contribute to the SNI phenotype.These data suggest that distinct brain circuits may drive the psychiatric components of neuropathic pain and pave the way for better investigate the long-term consequences of peripheral nerve injury in which most of the available drugs are to date unsatisfactory.

The dorsal striatum, apart from controlling voluntary movement, displays a recently demonstrated pain inhibition. It is connected to the descending pain modulatory system and in particular to the rostral ventromedial medulla through the medullary dorsal reticular nucleus. Diseases of the basal ganglia, such as Parkinson's disease, in addition to being characterized by motor disorders, are associated with pain and hyperactivation of the excitatory transmission. A way to counteract glutamatergic hyperactivation is through the activation of group III metabotropic glutamate receptors (mGluRs), which are located on presynaptic terminals inhibiting neurotransmitter release. So far the mGluRs of group III have been the least investigated, owing to a lack of selective tools. More recently, selective ligands for each mGluR of group III, in particular positive and negative allosteric modulators, have been developed and the role of each subtype is starting to emerge. The neuroprotective potential of group III mGluRs in pathological conditions, such as those characterized by elevate glutamate, has been recently shown. In the dorsal striatum, mGluR7 and mGluR8 are located at glutamatergic corticostriatal terminals and their stimulation inhibits pain in pathological conditions such as neuropathic pain. The two receptors in the dorsal striatum have instead a different role in pain control in normal conditions. This review will discuss recent results focusing on the contribution of mGluR7 and mGluR8 in the dorsal striatal control of pain. The role of mGluR4, whose antiparkinsonian activity is widely reported, will also be addressed.

Chronic pain is associated with cognitive deficits. Palmitoylethanolamide (PEA) has been shown to ameliorate pain and pain-related cognitive impairments by restoring glutamatergic synapses functioning in the spared nerve injury (SNI) of the sciatic nerve in mice. SNI reduced mechanical and thermal threshold, spatial memory and LTP at the lateral entorhinal cortex (LEC)-dentate gyrus (DG) pathway. It decreased also postsynaptic density, volume and dendrite arborization of DG and increased the expression of metabotropic glutamate receptor 1 and 7 (mGluR1 and mGluR7), of the GluR1, GluR1s845 and GluR1s831 subunits of AMPA receptor and the levels of glutamate in the DG. The level of the endocannabinoid 2-arachidonoylglycerol (2-AG) was instead increased in the LEC. Chronic treatment with PEA, starting from when neuropathic pain was fully developed, was able to reverse mechanical allodynia and thermal hyperalgesia, memory deficit and LTP in SNI wild type, but not in PPAR alpha null, mice. PEA also restored the level of glutamate and the expression of phosphorylated GluR1 subunits, postsynaptic density and neurogenesis. Altogether, these results suggest that neuropathic pain negatively affects cognitive behavior and related LTP, glutamatergic synapse and synaptogenesis in the DG. In these conditions PEA treatment alleviates pain and cognitive impairment by restoring LIT and synaptic maladaptative changes in the LEC-DG pathway. These outcomes open new perspectives for the use of the N-acylethanolamines, such as PEA, for the treatment of neuropathic pain and its central behavioural sequelae.

Due to the widespread use of non-steroidal anti-inflammatory drugs (NSAIDs), the incidence of NSAID-associated adverse events has increased exponentially over the past decades. Ketoprofen (ketoprofen acid, KA) is a widely used NSAID and, like with other NSAIDs, its use can be associated with adverse effects that especially involve the gastrointestinal tract and the kidney. The salification of KA with L-lysine has led to the synthesis of ketoprofen lysine salt (KLS), which is characterized by higher solubility and a more rapid gastrointestinal absorption compared to KA. Previous studies have reported that KLS has also an increased gastric tolerance in vitro, and this is due to the inhibition of lipid peroxidation and reactive oxygen species scavenging effects of L-lysine. Here, we report in vivo tolerability/toxicity studies that were conducted prior seeking KLS marketing authorization, in which we compared KLS and KA safety profile, focusing in particular on the evaluation of the gastrointestinal and renal tolerability of the drugs administered orally to dogs. Our results demonstrate that KLS has an increased in vivo gastrointestinal tolerability compared to KA and show, for the first time, that KLS has also increased in vivo renal tolerability compared to KA, thus supporting the concept that L-lysine may counteract NSAID-induced oxidative stress-mediated gastrointestinal and renal injury.

The 2-amino-4-(3-hydroxy-5-methylisoxazol-4-yl)-butyric acid, homo-AMPA, an analog of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and 2-aminoadipic acid, has shown no activity towards ionotropic and metabotropic glutamate 1, 2, 3, 4, 5, and 7 receptors (mGluR1-7), agonist activity at mGluR6 while the activity at mGluR8 was never investigated. The effect of homo-AMPA on pain control has been never investigated. In this study we evaluated the effect of intra-ventrolateral periaqueductal grey (VL PAG) microinjections of homo-AMPA on pain responses and the activity of pain-responding neurons of the rostral ventromedial medulla (RVM), the "pronociceptive" ON cells, and the "antinociceptive" OFF cells. The study was performed in control and diabetic neuropathic mice. Homo-AMPA decreased mechanical allodynia in diabetic neuropathic mice. Homo-AMPA increased also the latency to tail-flick, decreased the ongoing activity, the pain stimulus-evoked burst of firing, and the duration of the burst of the ON cells in both, control and neuropathic mice. Homo-AMPA also increased the ongoing activity, decreased and delayed the pause of the OFF cells in control mice. Unlike the retina, we did not find the transcript and protein for mGluR6 in the VL PAG. Alpha-methyl-serine-O-phosphate, a group III mGluRs antagonist, blocked the anti-allodynic effect of homo-AMPA. Considering the absence of both, mGluR6 in VL-PAG and homo-AMPA activity at mGluR4 and mGluR7 at the dose used, mGluR8 could be the target on which homo-AMPA produces the observed effects. The target of homo-AMPA capable of evoking analgesia at a very low dose and in conditions of diabetic neuropathy deserves further consideration.

The bioactive form of vitamin D, 1,25-dihydroxyvitamin D (1,25D3), exerts immunomodulatory actions resulting in neuroprotective effects potentially useful against neurodegenerative and autoimmune diseases. In fact, vitamin D deficiency status has been correlated with painful manifestations associated with different pathological conditions. In this study, we have investigated the effects of vitamin D deficiency on microglia cells, as they represent the main immune cells responsible for early defense at central nervous system (CNS), including chronic pain states. For this purpose, we have employed a model of low vitamin D intake during gestation to evaluate possible changes in primary microglia cells obtained from postnatal day(P)2-3 pups. Afterwards, pain measurement and microglia morphological analysis in the spinal cord level and in brain regions involved in the integration of pain perception were performed in the parents subjected to vitamin D restriction. In cultured microglia, we detected a reactive—activated and proliferative—phenotype associated with intracellular reactive oxygen species (ROS) generation. Oxidative stress was closely correlated with the extent of DNA damage and increased β-galactosidase (B-gal) activity. Interestingly, the incubation with 25D3 or 1,25D3 or palmitoylethanolamide, an endogenous ligand of peroxisome proliferator-activated-receptor-alpha (PPAR-α), reduced most of these effects. Morphological analysis of ex-vivo microglia obtained from vitamin-D-deficient adult mice revealed an increased number of activated microglia in the spinal cord, while in the brain microglia appeared in a dystrophic phenotype. Remarkably, activated (spinal) or dystrophic (brain) microglia were detected in a prominent manner in females. Our data indicate that vitamin D deficiency produces profound modifications in microglia, suggesting a possible role of these cells in the sensorial dysfunctions associated with hypovitaminosis D.

Chemotherapy-induced neuropathic pain is a clinically relevant adverse effect of several anticancer drugs leading to dose reduction or therapy discontinuation. The lack of knowledge about the mechanisms of neuropathy development and pain chronicization makes chemotherapy-induced neuropathic pain treatment an unmet medical need. In this context, the vascular endothelial growth factor A (VEGF-A) has emerged as a neurotoxicity biomarker in a model of chemotherapy-induced neuropathy, and its decrease has been related to pain relief. Aim of this study was to clarify the VEGF-A-dependent pain signaling in the CNS for individuating new targeted therapeutic approaches. In mice, the intrathecal infusion of VEGF-A induced a dose-dependent noxious hypersensitivity mediated by the VEGF receptor 1 (VEGFR-1) as demonstrated by pharmacological and genetic tools. In electrophysiological study, VEGF-A stimulated the spinal nociceptive neurons activity through VEGFR-1. In the dorsal horn of the spinal cord, VEGF-A increased in astrocytes of animals affected by neuropathy suggesting this cell population as a source of the potent pain mediator. Accordingly, the selective knockdown of astrocytic VEGF-A, by shRNAmir, blocked the development of oxaliplatin-induced neuropathic pain. Besides, the anti-VEGFR-1 mAb D16F7 (previously described as anticancer) effectively relieved neuropathic pain induced by chemotherapeutic agents. In conclusion, astrocyte-released VEGF-A is a new player in the complex neuron-glia network that oversees physiological and pathological pain and D16F7 mAb rises as a potent pain killer strategy.

Traumatic brain injury (TBI) is one of the main causes of death in young people for which currently no efficacious treatment exists. Recently, we have reported that mice with mild-TBI with a specific injury in the insula showed elevated levels of a little investigated

Recent evidence points to the gut microbiota as a regulator of brain and behavior, although it remains to be determined if gut bacteria play a role in chronic pain. The endocannabinoid system is implicated in in- flammation and chronic pain processing at both the gut and central nervous system (CNS) levels. In the present study, we used low Vitamin D dietary intake in mice and evaluated possible changes in gut microbiota, pain processing and endocannabinoid system signaling. Vitamin D deficiency induced a lower microbial diversity characterized by an increase in Firmicutes and a decrease in Verrucomicrobia and Bacteroidetes. Concurrently, vitamin D deficient mice showed tactile allodynia associated with neuronal hyperexcitability and alterations of endocannabinoid system members (endogenous mediators and their receptors) at the spinal cord level. Changes in endocannabinoid (anandamide and 2-ara- chidonoylglycerol) levels were also observed in the duodenum and colon. Remarkably, the anti-inflammatory anandamide congener, palmitoylethanolamide, counteracted both the pain behaviour and spinal biochemical changes in vitamin D deficient mice, whilst increasing the levels of Akkermansia, Eubacterium and Enterobacteriaceae, as compared with vehicle-treated mice. Finally, induction of spared nerve injury in normal or vitamin D deficient mice was not accompanied by changes in gut microbiota composition. Our data suggest the existence of a link between Vitamin D deficiency – with related changes in gut bacterial composition – and altered nociception, possibly via molecular mechanisms involving the endocannabinoid and related mediator signaling systems.

Traumatic brain injury (TBI) represents an important public health problem and is followed by neuroinflammation and neurological dysfunctions. It has been suggested that brain trauma is often associated to deep behavioral alterations and chronic pain-like syndrome. Despite inducing minimal brain damage, mild TBI (mTBI) leads to persistent behavioral changes, including anxiety, depression, social interaction impairment, and aggressiveness. The clinical management of these symptoms is still unsatisfactory and new pharmacological treatments are needed, especially for the aggressiveness and depression. In a mouse model of mTBI, we investigated the effect of 2-Pentadecyl-2-Oxazoline (PEA-OXA), a natural compound, that is a secondary metabolite, found in green and roasted coffee beans, on both the pain perception, and neuropsychiatric dysfunctions. We found that the compound acts as a α2 adrenergic antagonist and this mechanism is here described for the first time. Mild TBI mice, starting from 14-d post-trauma, developed anxious and aggressive behavior, whilst depressive-like behavior and impaired social interactions were observed from the 60th d onward. PEA-OXA normalized all the behavioral changes investigated. We also investigated the memory impairments through Morris Water Maze (MWM) test. Both sham and mTBI mice treated with PEA-OXA showed amelioration in the reversal task of the MWM. Nevertheless, the main symptom of the long-term mTBI is represented by the depressive-like behavior, which was completely reversed by PEA-OXA repeated administration. In humans, mTBI-induced depression precedes the appearance of dementias and is characterized by a massive deficit of GABAergic transmission in the cortices. We found that PEA-OXA normalized the GABA changes in the prefrontal cortex. In order to prove the α2-mediated effect of the PEA-OXA we have performed open field test in naïve animals by microinjecting into the medial prefrontal cortex the dexomedetomidine, a selective α2 agonist with or without PEA-OXA co-injection. We found that PEA-OXA antagonized the α2 agonist effect on the locomotor activity. Moreover, PEA-OXA microinjection into the medial prefrontal cortex induced an enhancement of dopamine release. Collectively, these data suggest that this natural compound, through its multi-target activity is able to: i) ameliorate behavioral alterations (i.e. depression), ii) selectively normalize cortical GABA levels, iii) rescue the impaired neuronal activity in the prefrontal cortex.

Focusing on the importance of the free phenolic hydroxyl moiety, a family of 23 alkylresorcinol-based compounds were developed and evaluated for their cannabinoid receptor binding properties. The non-symmetrical hexylresorcinol derivative 29 turned out to be a CB2-selective competitive antagonist/inverse agonist endowed with good potency. Both the olivetol- and 5-(2-methyloctan-2-yl)resorcinol-based derivatives 23 and 24 exhibited a significant antinociceptive activity. Interestingly, compound 24 proved to be able to activate both cannabinoid and TRPV1 receptors. Even if cannabinoid receptor subtype selectivity remained a goal only partially achieved, results confirm the validity of the alkylresorcinol nucleus as skeleton for the identification of potent cannabinoid receptor modulators.

Neuropathic pain is a pathological condition induced by a lesion or disease affecting the somatosensory system, with symptoms like allodynia and hyperalgesia. It has a multifaceted pathogenesis as it implicates several molecular signaling pathways involving peripheral and central nervous systems. Affective and cognitive dysfunctions have been reported as comorbidities of neuropathic pain states, supporting the notion that pain and mood disorders share some common pathogenetic mechanisms. The understanding of these pathophysiological mechanisms requires the development of animal models mimicking, as far as possible, clinical neuropathic pain symptoms. Among them, the Spared Nerve Injury (SNI) model has been largely characterized in terms of behavioral and functional alterations. This model is associated with changes in neuronal firing activity at spinal and supraspinal levels, and induces late neuropsychiatric disorders (such as anxious-like and depressive-like behaviors, and cognitive impairments) comparable to an advanced phase of neuropathy. The goal of this review is to summarize current findings in preclinical research, employing the SNI model as a tool for identifying pathophysiological mechanisms of neuropathic pain and testing pharmacological agent.

Aging is a physiological condition involving progressive degenerative modifications and loss of functionality in all organ systems. In general, aged patients are frail and sensitive to anesthesia and surgery. Clinical practice indicates that being more sensitive to anesthetic drugs and more susceptible to the side effects, elderly requires usually lower doses to reach the clinical anesthesia. This is mainly associated with the significant changes in pharmacokinetics (and pharmacodynamics) occurring with advancing age. However, how the dose–response relationship is affected by age should be better defined. This chapter focuses on the main changes that physiologically occur with age, how these changes affect organs and systems, and their impact on anesthetic care. The agents used in general anesthesia are discussed in detail with particular emphasis on aspects concerning the dose, safety, and efficacy of their use in the elderly. Moreover, we highlight some recent preclinical evidence on the use of general anesthetics in aged animals.

The study investigated the role of the metabotropic glutamate receptor subtype 7 (mGluR7) in pain signalling in the dorsal striatum of sham and neuropathic rats. Supraspinal circuitries involved in the dorsal striatum control of pain were also explored. In the sham rats, microinjection of N,N'-bis(diphenylmethyl)-1,2-ethanediamine (AMN082), a selective mGluR7 positive allosteric modulator, into the dorsal striatum, facilitated pain, increased the activity of the ON cells and inhibited the activity of the OFF cells in the rostral ventromedial medulla, and decreased glutamate levels in the dorsal striatum. Conversely, AMN082 inhibited pain and the activity of the ON cells while increased the activity of the OFF cells in rats with spared nerve injury (SNI) of the sciatic nerve. AMN082 also decreased glutamate levels in the dorsal striatum of SNI rats. The effect of AMN082 on mechanical allodynia and glutamate release was blocked by 6-(2,4-dimethylphenyl)-2-ethyl-6,7-dihydro-4(5H)-benzoxazolone (ADX71743), a selective mGluR7 negative allosteric modulator. Moreover, in the sham rats, AMN082 increased the activity of total nociceptive convergent neurons in the dorsal reticular nucleus while in the SNI rats, such activity was decreased. The administration of lidocaine into the subthalamic nucleus abolished the effect of AMN082 on the total nociceptive convergent neurons in the sham rats but not in the SNI rats. Thus, the dual effect of mGluR7 in facilitating or inhibiting pain responses may be due to the recruitment of different pathways of the basal ganglia, the indirect or direct pathway, in physiological or pathological conditions, respectively.

// Flavio Fazioli 1, * , Gianluca Colella 2, * , Roberta Miceli 3 , Mariano Giuseppe Di Salvatore 1 , Michele Gallo 1 , Serena Boccella 4 , Annarosaria De Chiara 5 , Carlo Ruosi 2, * and Filomena de Nigris 6, * 1 Division of Musculoskeletal Oncology Surgery, National Cancer Institute G. Pascale, Naples, Italy 2 Department of Human Health, Federico II University of Naples, Naples, Italy 3 S.C. Cellular Biology and Biotherapy, National Cancer Institute G. Pascale, Naples, Italy 4 Department of Experimental Medicine, University of Campania “Luigi Vanvitelli”, Naples Italy 5 Division of Pathology, National Cancer Institute G. Pascale Foundation, Naples, Italy 6 Department of Biochemistry, Biophysics and General Pathology, University of Campania “Luigi Vanvitelli”, Naples Italy * These authors have contributed equally to this work Correspondence to: Filomena de Nigris, email: Filomena.denigris@unina2.it Keywords: recurrences, sarcoma, angiogenesis, stem cells, AKT Received: March 31, 2017 Accepted: May 29, 2017 Published: June 28, 2017 ABSTRACT Giant cell tumors of bone (GCTB) are rare sarcomas with a high rate of unpredictable local relapse. Studies suggest that surgical methods affect recurrence, supporting the idea that local disease develops from re-growth of residual cancer cells. To identify early prognostic markers of individual risk of recurrence, we evaluated the effect of post-surgery fluids from a cohort of GCTB patients on growth of primary and established sarcoma cell lines, and mice xenograph. Post-surgery fluids increased cell growth and enhanced expression of CD44 ++ , the principal receptor for the extracellular matrix component hyaluronan and the mesenchymal stem marker CD117 + . Cancer cells became highly invasive and tumorigenic, acquiring stemness properties, and activated AKT/mTOR pathway. Prolonged stimulation with post-surgery fluids down-regulated the mesenchymal gene TWIST1 and Vimentin protein, and transdifferentiated cells into tubule-like structures positive to the endothelial markers VE-Cadherin and CD31 + . In mice, post-surgery fluids gave rise to larger and more vascularized tumors than control, while in patients AKT/mTOR pathway activation was associated with recurrence by logistic regression (Kaplan-Meier; P

Inflammation plays a key role in the pathogenesis of several chronic diseases. The urokinase plasminogen activator receptor (uPAR) exerts a plethora of functions in both physiological and pathological processes, including inflammation. In this study, we evaluated the anti-inflammatory effect of a novel peptide ligand of uPAR, UPARANT, in different animal models of inflammation. Rats and mice were divided in different groups (n = 5) for single or repeated administration of vehicle (9% DMSO in 0.9% NaCl), UPARANT (6, 12 and 24 mg/kg) or dexamethasone (2 mg/kg). Animals were subjected to carrageenan-induced paw oedema or zymosan-induced peritonitis. UPARANT effects were tested on: (1) the carrageenan-induced paw oedema volume, (2) the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and the nitrite/nitrate (NOx) levels in the paw exudates, (3) cells recruitment into the peritoneal cavity after zymosan injection and (4) NOx levels in the peritoneal lavage. UPARANT (12 and 24 mg/kg) reduced inflammation in both experimental paradigms. Analysis of pro-inflammatory enzymes revealed that administration of UPARANT reduced iNOS, COX2 and NO over-production. Our study provides a solid evidence that UPARANT reduces the severity of inflammation in diverse animal models, thus representing a novel anti-inflammatory drug with potential advantages with respect to the typical steroidal agents.

It is a common opinion that metabotropic glutamate receptor subtype 6 (mGluR6) is expressed exclusively in the retina, and in particular in the dendrites of ON-bipolar cells. Glutamate released in darkness from photoreceptors activates mGluR6, which is negatively associated with a membrane non-selective cation channel, the transient receptor potential melanoma-related 1, TRPM1, resulting in cell hyperpolarization. The evidence that mGluR6 is expressed not only in the retina but also in other tissues and cell populations has accumulated over time. The expression of mGluR6 has been identified in microglia, bone marrow stromal and prostate cancer cells, B lymphocytes, melanocytes and keratinocytes and non-neural tissues such as testis, kidney, cornea, conjunctiva, and eyelid. The receptor also appears to be expressed in brain areas, such as the hypothalamus, cortex, hippocampus, nucleus of tractus solitarius, superior colliculus, axons of the corpus callosum and accessory olfactory bulb. The pharmacological activation of mGluR6 in the hippocampus produced an anxiolytic-like effect and in the periaqueductal gray analgesic potential. This review aims to collect all the evidence on the expression and functioning of mGluR6 outside the retina that has been accumulated over the years for a broader view of the potential of the receptor whose retinal confinement appears understimated.

Neuropathic pain afflicts millions of individuals and represents a major health problem for which there is limited effective and safe therapy. Emerging literature links altered sphingolipid metabolism to nociceptive processing. However, the neuropharmacology of sphingolipid signaling in the central nervous system in the context of chronic pain remains largely unexplored and controversial. We now provide evidence that sphingosine-1-phosphate (S1P) generated in the dorsal horn of the spinal cord in response to nerve injury drives neuropathic pain by selectively activating the S1P receptor subtype 1 (S1PR1) in astrocytes. Accordingly, genetic and pharmacological inhibition of S1PR1 with multiple antagonists in distinct chemical classes, but not agonists, attenuated and even reversed neuropathic pain in rodents of both sexes and in two models of traumatic nerve injury. These S1PR1 antagonists retained their ability to inhibit neuropathic pain during sustained drug administration, and their effects were independent of endogenous opioid circuits. Moreover, mice with astrocyte-specific knockout of S1pr1 did not develop neuropathic pain following nerve injury, thereby identifying astrocytes as the primary cellular substrate of S1PR1 activity. On a molecular level, the beneficial reductions in neuropathic pain resulting from S1PR1 inhibition were driven by interleukin 10 (IL-10), a potent neuroprotective and anti-inflammatory cytokine. Collectively, our results provide fundamental neurobiological insights that identify the cellular and molecular mechanisms engaged by the S1PR1 axis in neuropathic pain and establish S1PR1 as a target for therapeutic intervention with S1PR1 antagonists as a class of nonnarcotic analgesics.

Neurological dysfunctions are the most impactful and persistent consequences of traumatic brain injury (TBI). Indeed, previous reports suggest that an association between TBI and chronic pain syndromes, as well anxio-depressive behaviors, tends to be more common in patients with mild forms of TBI. At present, no effective treatment options are available for these symptoms. In the present study, we used a weight drop mild TBI mouse model to investigate the effect of a commercially available 10% Cannabidiol (CBD) oil on both the sensorial and neuropsychiatric dysfunctions associated with mild TBI through behavioral and biomolecular approaches. TBI mice developed chronic pain associated with anxious and aggressive behavior, followed by a late depressive-like behavior and impaired social interaction. Such behaviors were related with specific changes in neurotransmitters release at cortical levels. CBD oral treatment restored the behavioral alterations and partially normalized the cortical biochemical changes. In conclusion, our data show some of the brain modifications probably responsible for the behavioral phenotype associated with TBI and suggest the CBD as a pharmacological tool to improve neurological dysfunctions caused by the trauma.

Non-small-cell lung cancer (NSCLC) accounts for 85%–90% of all cases of lung cancer that is the most deadly type of cancer. Despite advances in chemotherapy and radiotherapy, severe side effects and frequent drug resistance limit the success of the treatments, and the identification of new therapeutic options still represents a crucial challenge. Here, we provide the evidence for the therapeutic potential of an aptamer-microRNA (miR) complex (AmiC) composed by an aptamer (GL21.T), able to bind and antagonize the oncogenic receptor Axl, and the miR-137, downregulated in lung cancer and involved in cell survival and proliferation. We found that, when applied to Axl-expressing NSCLC cancer cells, the complex is effectively internalized, increasing miR cellular levels and downregulating miR targets. Most importantly, the complex combines the inhibitory function of the GL21.T aptamer and miR-137, leading to a negative impact on NSCLC migration and growth. The described AmiC thus represents a promising tool for the development of new therapeutic approaches for NSCLC. Keywords: aptamers, miRNAs, NSCLC, targeted delivery