Publications by authors named "Carla Ghelardini"

286 Publications

Adenosine A3 agonists reverse neuropathic pain via T cell-mediated production of IL-10.

J Clin Invest 2021 Feb 23. Epub 2021 Feb 23.

Department of Pharmacological and Physiological Science, Saint Louis University School of Medicine, St. Louis, United States of America.

The A3 adenosine receptor (A3AR) has emerged as a therapeutic target with A3AR agonists to tackle the global challenge of neuropathic pain; investigation into their mode of action is essential for ongoing clinical development. A3ARs on immune cells, and their activation during pathology, modulates cytokine release. Thus, immune cells as a cellular substrate for the pharmacological action of A3AR agonists is enticing but unknown. Studies herein discovered that RagKO mice lacking T- and B-cells are insensitive to the anti-allodynic effects of A3AR agonists versus wild-type (WT) mice. Similar findings were observed in interleukin-10 and interleukin-10 receptor knockout mice. Adoptive transfer of CD4+ T-cells (CD4+-T) from WT mice infiltrated the dorsal root ganglion (DRG) and restored A3AR agonist-mediated anti-allodynia in RagKO mice; CD4+-T from Adora3KO or Il10KO mice did not. Transfer of CD4+-T from WT, but not Il10KO, into Il10KO mice fully reinstated anti-allodynic effects of A3AR activation. Transfer of CD4+-T from WT, but not Il10KO, into Adora3KO mice fully reinstated anti-allodynic effects of A3AR activation. Notably, A3AR agonism reduced DRG neuron excitability when co-cultured with CD4+-T in an IL-10-dependent manner. A3AR actions on CD4+-T infiltrate in the DRG decreased phosphorylation of GluN2B-containing N-methyl-D-aspartate receptors at Tyr1472, a modification associated with regulating neuronal hypersensitivity. Our findings establish that activation of A3AR on CD4+-T cells to release of IL-10 is required and sufficient for A3AR agonists as therapeutics.
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http://dx.doi.org/10.1172/JCI139299DOI Listing
February 2021

Efficacy of Extract against Inflammatory Pain: In Vivo Studies in Mice.

Mar Drugs 2021 Jan 21;19(2). Epub 2021 Jan 21.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA)-Pharmacology and Toxicology Section, University of Florence, Viale Gaetano Pieraccini, 6, 50139 Florence, Italy.

(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) 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.
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http://dx.doi.org/10.3390/md19020048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7909763PMC
January 2021

Pain Modulation in WAG/Rij Epileptic Rats (A Genetic Model of Absence Epilepsy): Effects of Biological and Pharmacological Histone Deacetylase Inhibitors.

Front Pharmacol 2020 3;11:549191. Epub 2020 Dec 3.

Department of Pharmacy, University of Naples Federico II, Naples, Italy.

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 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- (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 ( < 0.001). BUT and BUT + VPA treatment reduced hypersensitivity ( < 0.01). VPA was significantly effective only after 1 month ( < 0.01). All the three receptors are involved in BUT + VPA effects ( < 0.001). BUT and BUT + VPA decreased the expression of NFkB and enhanced glutathione reductase ( < 0.01); protein oxidation (carbonylation) was reduced ( < 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.
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http://dx.doi.org/10.3389/fphar.2020.549191DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7745735PMC
December 2020

Extra virgin olive oil and related by-products (Olea europaea L.) as natural sources of phenolic compounds for abdominal pain relief in gastrointestinal disorders in rats.

Food Funct 2020 Dec 25;11(12):10423-10435. Epub 2020 Nov 25.

Department of Neuroscience, Psychology, Drug Research and Child Health - NEUROFARBA - Pharmacology and Toxicology Section, University of Florence, Viale Pieraccini 6, 50139, Florence, Italy.

Management of abdominal pain, a common symptom of IBDs and IBS, is still a clinical problem. Extra virgin olive oil (EVOO), a main component of the Mediterranean diet, shows positive effects on chronic inflammation in IBDs. In this study, the effect of the oral administration of EVOO (3 mL) and two olive milling by-products, DPA (300 mg kg) and DRF (300 mg kg), on preventing the development of abdominal pain in a DNBS-induced colitis model in rats was evaluated. The doses were chosen with the aim of simulating a plausible daily intake in humans. DPA and EVOO treatments significantly reduced the abdominal viscero-motor response to colon-rectal distension at 2 and 3 mL of balloon distension volume, both 7 and 14 days after the DNBS-injection. DRF showed efficacy in the reduction of visceral hypersensitivity only with 3 mL balloon inflation. In awake animals, DPA and DRF reduced pain perception (evaluated as abdominal withdrawal reflex) with all balloon distension volumes, while EVOO was effective only with higher distension volumes. Fourteen days after the DNBS-injection, all samples reduced the macroscopic intestinal damage (quantified as the macroscopic damage score) also showing, at the microscopic level, a reduction of the inflammatory infiltrate (quantified by hematoxylin and eosin analysis), fibrosis (highlighted by picrosirius red staining), the increase in mast cells and their degranulation (analyzed by triptase immunohistochemistry). This is the first report on the promotion of abdominal pain relief in a rat model obtained administering EVOO and two derived by-products. Our results suggest a protective role of phenol-rich EVOO and milling by-products, which may be proposed as food ingredients for novel functional foods.
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http://dx.doi.org/10.1039/d0fo02293dDOI Listing
December 2020

Therapeutic potential for coxibs-nitric oxide releasing hybrids in cystic fibrosis.

Eur J Med Chem 2021 Jan 31;210:112983. Epub 2020 Oct 31.

Department of Chemistry and Technologies of Drug, Sapienza University of Rome, Piazzale A. Moro 5, 00185, Rome, Italy. Electronic address:

This review discusses the rational for further studies of COX-2 inhibitors-NO releaser hybrids (NO-Coxibs) in the pharmacological treatment of the airway inflammation in Cystic Fibrosis (CF). Our research group developed several classes of NO-Coxibs for the pharmacological treatment of arthritis, and among them several compounds showed an outstanding in vivo efficacy and good pharmacokinetic properties. The good antiinflammatory properties displayed by these compounds during the previous screening could, by itself, suggest appropriate candidates for further testing in CF.
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http://dx.doi.org/10.1016/j.ejmech.2020.112983DOI Listing
January 2021

Eruca sativa Mill. seed extract promotes anti-obesity and hypoglycemic effects in mice fed with a high-fat diet.

Phytother Res 2020 Nov 3. Epub 2020 Nov 3.

Department of Pharmacy, University of Pisa, Pisa, Italy.

Obesity is currently considered a major source of morbidity, with dramatic complications on health status and life expectancy. Several studies demonstrated the positive effects of Brassicaceae vegetables on obesity and related diseases, partially attributing these beneficial properties to glucosinolates and their derivatives isothiocyanates. Recently, isothiocyanates have been described as a hydrogen sulfide (H S)-releasing moiety, suggesting that H S may be at least in part responsible for the beneficial effects of Brassicaceae. In this work, the metabolic effects of an extract obtained from Eruca sativa Mill. seeds (E.S., Brassicaceae), containing high levels of glucoerucin, were evaluated in an experimental model of obesity. Male balb/c mice were fed for 10 weeks with standard (Std) diet or high fat (HF) diet supplemented with E.S. E.S. significantly contained the body weight gain in this obesity model, improving also glucose homeostasis. Interestingly, lower values of white adipose tissue mass and a significant reduction of adipocytes size were also observed. Moreover, E.S. enhanced the adipocytes metabolism, improving the citrate synthase activity and reduced triglyceride levels in mice fed with HF diet. Taken together, these results suggest that E.S. is endowed with an interesting translational and nutraceutical value in the prevention of metabolic disorders, suggesting that H S could be a key player.
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http://dx.doi.org/10.1002/ptr.6941DOI Listing
November 2020

()-3-Furan-2-yl---tolyl-acrylamide and its Derivative DM489 Decrease Neuropathic Pain in Mice Predominantly by α7 Nicotinic Acetylcholine Receptor Potentiation.

ACS Chem Neurosci 2020 11 19;11(21):3603-3614. Epub 2020 Oct 19.

Illawarra Health and Medical Research Institute (IHMRI), University of Wollongong, Wollongong, NSW 2522, Australia.

The main objective of this study was to determine whether ()-3-furan-2-yl---tolyl-acrylamide (PAM-2) and its structural derivative DM489 produce anti-neuropathic pain activity using the streptozotocin (STZ)- and oxaliplatin-induced neuropathic pain animal models. To assess possible mechanisms of action, the pharmacological activity of these compounds was determined at α7 and α9α10 nicotinic acetylcholine receptors (nAChRs) and Ca2.2 channels expressed alone or coexpressed with G protein-coupled GABA receptors. The animal results indicated that a single dose of 3 mg/kg PAM-2 or DM489 decreases STZ-induced neuropathic pain in mice, and chemotherapy-induced neuropathic pain is decreased by PAM-2 (3 mg/kg) and DM489 (10 mg/kg). The observed anti-neuropathic pain activity was inhibited by the α7-selective antagonist methyllycaconitine. The coadministration of oxaliplatin with an inactive dose (1 mg/kg) of PAM-2 decreased the development of neuropathic pain after 14, but not 7, days of cotreatment. The electrophysiological results indicated that PAM-2 potentiates human (h) and rat (r) α7 nAChRs with 2-7 times higher potency than that for hCa2.2 channel inhibition and an even greater difference compared to that for rα9α10 nAChR inhibition. These results support the notion that α7 nAChR potentiation is likely the predominant molecular mechanism underlying the observed anti-nociceptive pain activity of these compounds.
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http://dx.doi.org/10.1021/acschemneuro.0c00476DOI Listing
November 2020

Intranasal Low-Dose Naltrexone Against Opioid Side Effects: A Preclinical Study.

Front Pharmacol 2020 18;11:576624. Epub 2020 Sep 18.

Department of Neuroscience, Psychology, Drug Research and Child Health - NEUROFARBA - Pharmacology and Toxicology Section, University of Florence, Florence, Italy.

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 morphine and 60 mg kg 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.
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http://dx.doi.org/10.3389/fphar.2020.576624DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7531600PMC
September 2020

The active second-generation proteasome inhibitor oprozomib reverts the oxaliplatin-induced neuropathy symptoms.

Biochem Pharmacol 2020 12 1;182:114255. Epub 2020 Oct 1.

Dept. of Pharmacy and Biotechnology, Alma Mater Studiorum - University of Bologna, Via Irnerio 48, 40126 Bologna, Italy.

Oxaliplatin-induced neuropathy (OXAIN) is a major adverse effect of this antineoplastic drug, widely used in the treatment of colorectal cancer. Although its molecular mechanisms remain poorly understood, recent evidence suggest that maladaptive neuroplasticity and oxidative stress may participate to the development of this neuropathy. Given the role played on protein remodeling by ubiquitin-proteasome system (UPS) in response to oxidative stress and in neuropathic pain, we investigated whether oxaliplatin might cause alterations in the UPS-mediated degradation pathway, in order to identify new pharmacological tools useful in OXAIN. In a rat model of OXAIN (2.4 mg kg i.p., daily for 10 days), a significant increase in chymotrypsin-(β5) like activity of the constitutive proteasome 26S was observed in the thalamus (TH) and somatosensory cortex (SSCx). In addition, the selective up-regulation of β5 and LMP7 (β5i) subunit gene expression was assessed in the SSCx. Furthermore, this study revealed that oprozomib, a selective β5 subunit proteasome inhibitor, is able to normalize the spinal prodynorphin gene expression upregulation induced by oxaliplatin, as well as to revert mechanical allodynia and thermal hyperalgesia observed in oxaliplatin-treated rats. These results underline the relevant role of UPS in the OXAIN and suggest new pharmacological targets to counteract this severe adverse effect. This preclinical study reveals the involvement of the proteasome in the oxaliplatin-induced neuropathy and adds useful information to better understand the molecular mechanism underlying this pain condition. Moreover, although further evidence is required, these findings suggest that oprozomib could be a therapeutic option to counteract chemotherapy-induced neuropathy.
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http://dx.doi.org/10.1016/j.bcp.2020.114255DOI Listing
December 2020

Faecal microbiota transplant from aged donor mice affects spatial learning and memory via modulating hippocampal synaptic plasticity- and neurotransmission-related proteins in young recipients.

Microbiome 2020 10 1;8(1):140. Epub 2020 Oct 1.

Dept. of Experimental and Clinical Medicine, University of Florence, 50134, Florence, Italy.

Background: The gut-brain axis and the intestinal microbiota are emerging as key players in health and disease. Shifts in intestinal microbiota composition affect a variety of systems; however, evidence of their direct impact on cognitive functions is still lacking. We tested whether faecal microbiota transplant (FMT) from aged donor mice into young adult recipients altered the hippocampus, an area of the central nervous system (CNS) known to be affected by the ageing process and related functions.

Results: Young adult mice were transplanted with the microbiota from either aged or age-matched donor mice. Following transplantation, characterization of the microbiotas and metabolomics profiles along with a battery of cognitive and behavioural tests were performed. Label-free quantitative proteomics was employed to monitor protein expression in the hippocampus of the recipients. We report that FMT from aged donors led to impaired spatial learning and memory in young adult recipients, whereas anxiety, explorative behaviour and locomotor activity remained unaffected. This was paralleled by altered expression of proteins involved in synaptic plasticity and neurotransmission in the hippocampus. Also, a strong reduction of bacteria associated with short-chain fatty acids (SCFAs) production (Lachnospiraceae, Faecalibaculum, and Ruminococcaceae) and disorders of the CNS (Prevotellaceae and Ruminococcaceae) was observed. Finally, the detrimental effect of FMT from aged donors on the CNS was confirmed by the observation that microglia cells of the hippocampus fimbria, acquired an ageing-like phenotype; on the contrary, gut permeability and levels of systemic and local (hippocampus) cytokines were not affected.

Conclusion: These results demonstrate that age-associated shifts of the microbiota have an impact on protein expression and key functions of the CNS. Furthermore, these results highlight the paramount importance of the gut-brain axis in ageing and provide a strong rationale to devise therapies aiming to restore a young-like microbiota to improve cognitive functions and the declining quality of life in the elderly. Video Abstract.
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http://dx.doi.org/10.1186/s40168-020-00914-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7532115PMC
October 2020

The endocannabinoid system dual-target ligand N-cycloheptyl-1,2-dihydro-5-bromo-1-(4-fluorobenzyl)-6-methyl-2-oxo-pyridine-3-carboxamide improves disease severity in a mouse model of multiple sclerosis.

Eur J Med Chem 2020 Dec 29;208:112858. Epub 2020 Sep 29.

Department of Pharmacy, University of Pisa, 56126, Pisa, Italy. Electronic address:

Multiple sclerosis is a chronic inflammatory demyelinating disorder of the central nervous system that eventually leads to progressive neurodegeneration and disability. Recent findings highlighted the emerging role of each target of the endocannabinoid system in controlling the symptoms and disease progression of multiple sclerosis. Therefore, multi-target modulators of the endocannabinoid system could provide a more effective pharmacological strategy as compared to the single target modulation. In this work, N-cycloheptyl-1,2-dihydro-5-bromo-1-(4-fluorobenzyl)-6-methyl-2-oxo-pyridine-3-carboxamide (B2) was identified as the most promising compound with dual agonism at cannabinoid receptors type-1 and cannabinoid receptors type-2 and good drug-like properties. In in vitro assays, B2 reduced glutamate release from rat synaptosomes through interaction with cannabinoid receptors type-1 and modulated the production of the pro- and anti-inflammatory cytokines (interleukins IL-1β and IL-6 and interleukin IL-10 respectively) via cannabinoid receptors type-2 activation. Furthermore, B2 demonstrated antinociceptive effects in an animal model of neuropathic pain and efficacy in an experimental autoimmune encephalomyelitis model of multiple sclerosis.
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http://dx.doi.org/10.1016/j.ejmech.2020.112858DOI Listing
December 2020

Toxicological Profile of the Pain-Relieving Antioxidant Compound Thioctic Acid in Its Racemic and Enantiomeric Forms.

Antioxidants (Basel) 2020 Aug 14;9(8). Epub 2020 Aug 14.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA)-Pharmacology and Toxicology Section, University of Florence, Viale Gaetano Pieraccini, 6, 50139 Florence, Italy.

Thioctic acid is a multipotent antioxidant compound existing as dextrorotatory (+), eutomer and naturally occurring and levorotatory (-). It has been proven to help fight many pathologies and is sold as racemate. In agreement with studies claiming a greater biopotency of the eutomer compared to the levorotatory compound, we recently preclinically and clinically showed that (+) thioctic acid is a pain-reliever as effective as double-dosed racemate. We investigated acute and subchronical toxicity of (+/-) thioctic acid, (-) thioctic acid, (+) thioctic acid and (+) salt thioctic acid on Sprague-Dawley rats. For acute toxicity, compounds were administered intraperitoneally (i.p.) with a single-injection at 125, 240, 360, 480 µmol/kg, then rodents were tested for motorial coordination and minimum lethal dose (LDmin). A subtoxic dose (360 µmol/kg) was administered i.p. for 15 days and we finally evaluated motorial impairment, glycemia, organ toxicity, and apoptosis state. Acutely administered, the highest doses of all thioctic acid compounds negatively affected motorial ability and (-) thioctic acid LDmin resulted higher than the others. Subchronic administrations caused overall body weight loss, motorial impairment, mass loss in some organs. (+/-) and (-) thioctic acid injections enhanced caspase-3 activity in some organs, (-) enantiomer-treated animals displayed more marked organ toxicity signs. Together with our previous study on the biologic role of enantiomers, these data suggest a therapeutic use of (+) enantiomer-based formulations, thus lowering dose and toxicity without affecting the positive effects brought by the drug.
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http://dx.doi.org/10.3390/antiox9080749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7464875PMC
August 2020

Deepening the Mechanisms of Visceral Pain Persistence: An Evaluation of the Gut-Spinal Cord Relationship.

Cells 2020 07 24;9(8). Epub 2020 Jul 24.

Department of Neuroscience, Psychology, Drug Research and Child Health, Neurofarba, Pharmacology and Toxicology Section, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

The management of visceral pain is a major clinical problem in patients affected by gastrointestinal disorders. The poor knowledge about pain chronicization mechanisms prompted us to study the functional and morphological alterations of the gut and nervous system in the animal model of persistent visceral pain caused by 2,4-dinitrobenzenesulfonic acid (DNBS). This agent, injected intrarectally, induced a colonic inflammation peaking on day 3 and remitting progressively from day 7. In concomitance with bowel inflammation, the animals developed visceral hypersensitivity, which persisted after colitis remission for up to three months. On day 14, the administration of pain-relieving drugs (injected intraperitoneally and intrathecally) revealed a mixed nociceptive, inflammatory and neuropathic pain originating from both the peripheral and central nervous system. At this time point, the colonic histological analysis highlighted a partial restitution of the , transmural collagen deposition, infiltration of mast cells and eosinophils, and upregulation of substance P (SP)-positive nerve fibers, which were surrounded by eosinophils and MHC-II-positive macrophages. A significant activation of microglia and astrocytes was observed in the dorsal and ventral horns of spinal cord. These results suggest that the persistence of visceral pain induced by colitis results from maladaptive plasticity of the enteric, peripheral and central nervous systems.
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http://dx.doi.org/10.3390/cells9081772DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7464824PMC
July 2020

Development of a stable oral pediatric solution of hydrochlorothiazide by the combined use of cyclodextrins and hydrophilic polymers.

Int J Pharm 2020 Sep 24;587:119692. Epub 2020 Jul 24.

Department of Chemistry, University of Florence, Florence, Italy. Electronic address:

Hydrochlorothiazide (HCT) is widely used in pediatrics for hypertension management. Due to the lack of pediatric commercial forms, community or hospital pharmacies generally prepare HCT extemporaneous pediatric suspensions by dispersing in water a portion of a crushed tablet or the drug powder; however, any dose or stability control is usually done on these preparations. Obtaining stable HCT solutions is very challenging, due to its low water-solubility and pH-dependent degradation. The aim of this work was to develop a stable 2 mg/mL-HCT oral pediatric solution without using co-solvents. Combined use of cyclodextrins (CD) and hydrophilic polymers was exploited to improve poor HCT solubility and stability. HPβCD and SBEβCD were selected, considering their safe toxicological profiles, while PVP resulted the best among the tested polymers. Low PVP concentrations (0.2-1.0%) improved the solubilizing efficiency of both CDs, allowing to reach the prefixed HCT concentration. Different CD-PVP concentrations were used to prepare several 2 mg/mL-HCT solutions in pH 5.5 buffer. The best stability was shown by solutions containing the highest SBEβCD concentration (25 mM), which allowed a 3-months stability at 4 °C. In vivo studies on rats showed that such formulation allowed a more pronounced and more reproducible diuretic effect than the corresponding HCT suspension.
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http://dx.doi.org/10.1016/j.ijpharm.2020.119692DOI Listing
September 2020

The Anti-Inflammatory and Pain-Relieving Effects of AR170, an Adenosine A Receptor Agonist, in a Rat Model of Colitis.

Cells 2020 06 21;9(6). Epub 2020 Jun 21.

School of Pharmacy, Medicinal Chemistry Unit, University of Camerino, 62032 Camerino (MC), Italy.

The pharmacological activation of A receptors has shown potential usefulness in the management of bowel inflammation. However, the role of these receptors in the control of visceral hypersensitivity in the presence of intestinal inflammation has not been investigated. The effects of AR170, a potent and selective A receptor agonist, and dexamethasone (DEX) were tested in rats with 2,4-dinitrobenzene sulfonic acid (DNBS)-induced colitis to assess their tissue inflammatory parameters. The animals received AR170, DEX, or a vehicle intraperitoneally for 6 days, starting 1 day before the induction of colitis. Visceral pain was assessed by recording the abdominal responses to colorectal distension in animals with colitis. Colitis was associated with a decrease in body weight and an increase in spleen weight. The macroscopic damage score and tissue tumor necrosis factor (TNF), interleukin 1β (IL-1β), and myeloperoxidase (MPO) levels were also enhanced. AR170, but not DEX, improved body weight. Both drugs counteracted the increase in spleen weight, ameliorated macroscopic colonic damage, and decreased TNF, IL-1β, and MPO tissue levels. The enhanced visceromotor response (VMR) in rats with colitis was decreased via AR170 administration. In rats with colitis, AR170 counteracted colonic inflammatory cell infiltration and decreased pro-inflammatory cytokine levels, thereby relieving visceral hypersensitivity.
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http://dx.doi.org/10.3390/cells9061509DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7348903PMC
June 2020

The Use of the Selective Imidazoline I Receptor Agonist Carbophenyline as a Strategy for Neuropathic Pain Relief: Preclinical Evaluation in a Mouse Model of Oxaliplatin-Induced Neurotoxicity.

Neurotherapeutics 2020 07;17(3):1005-1015

Dept. of Neuroscience, Psychology, Drug Research and Child Health - NEUROFARBA - Pharmacology and Toxicology Section, University of Florence, Viale Gaetano Pieraccini 6, 50139, Florence, Italy.

Anti-cancer therapy based on the repeated administration of oxaliplatin is limited by the development of a disabling neuropathic syndrome with detrimental effects on the patient's quality of life. The lack of effective pharmacological approaches calls for the identification of innovative therapeutic strategies based on new targets. We focused our attention on the imidazoline I receptor (I-R) and in particular on the selective I-R agonist 2-(1-([1,1'-biphenyl]-2-yl)propan-2-yl)-4,5-dihydro-1H-imidazole) (carbophenyline). The purpose of this work was the preclinical evaluation of the efficacy of carbophenyline on oxaliplatin-induced neuropathic pain in mice. Carbophenyline, acutely per os administered (0.1-10 mg kg), induced a dose-dependent anti-hyperalgesic effect that was completely blocked by the pre-treatment with the I-R antagonist 3 or the I/α receptor antagonist efaroxan, confirming the I-R-dependent mechanism. Conversely, pre-treatment with the I-R antagonist BU224 did not block the anti-nociceptive effect evoked by carbophenyline. Repeated oral administrations of carbophenyline (1 mg kg) for 14 days, starting from the first day of oxaliplatin injection, counteracted the development of neuropathic pain in all behavioral tests (cold plate, Von Frey, and paw pressure tests) carried out 24 h after the last carbophenyline treatment on days 7 and 14. In the dorsal horn of the spinal cord, carbophenyline significantly decreased the oxaliplatin-induced astrocyte activation detected by immunofluorescence staining by the specific labelling with GFAP antibody. In conclusion, carbophenyline showed anti-neuropathic properties both after acute and chronic treatment with preventive effect against oxaliplatin-induced astrocyte activation in the spinal cord. Therefore, I-R agonists emerge as a new class of candidates for the management of oxaliplatin-induced neuropathic pain.
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http://dx.doi.org/10.1007/s13311-020-00873-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7609613PMC
July 2020

Treatment of Non-Alcoholic Steatosis: Preclinical Study of a New Nutraceutical Multitarget Formulation.

Nutrients 2020 Jun 18;12(6). Epub 2020 Jun 18.

Department of Neuroscience, Psychology, Drug Research and Child Health-Neurofarba-Pharmacology and Toxicology Section, University of Florence, 50139 Florence, Italy.

Multifactorial pathogenesis of non-alcoholic steatohepatitis (NASH) disease, a wide-spread liver pathology associated with metabolic alterations triggered by hepatic steatosis, should be hit by multitarget therapeutics. We tested a multicomponent food supplement mixture (AP-NHm), whose components have anti-dislipidemic, antioxidant and anti-inflammatory effects, on in vitro and in vivo models of NASH. In vitro, hepatic cells cultures were treated for 24 h with 0.5 mM oleic acid (OA): in the co-treatment set cells were co-treated with AP-NH mixtures (AP-NHm, 1:3:10 ratio) and in the post-injury set AP-NHm was added for 48 h after OA damage. In vivo, C57BL/6 mice were fed with high-fat diet (HFD) for 12 weeks, inducing NASH at 7th week, and treated with AP-NHm at two dosages (1:3 ratio) in co-treatment or post-injury protocols, while a control group was fed with a standard diet. In in vitro co-treatment protocol, alterations of redox balance, proinflammatory cytokines release and glucose uptake were restored in a dose-dependent manner, at highest dosages also in post-injury regimen. In both regimens, pathologic dyslipidemias were also ameliorated by AP-NHm. In vivo, high-dose-AP-NHm-co-treated-HFD mice dose-dependently gained less body weight, were protected from dyslipidemia, and showed a lower liver weight. Dose-dependently, AP-NHm treatment lowered hepatic LDL, HDL, triglycerides levels and oxidative damage; co-treatment regimen was anti-inflammatory, reducing TNF-α and IL-8 levels. Hepatic lipidic infiltration significantly decreased in co-treated and post-injury-AP-NHm-HFD animals. The multitarget approach with AP-NHm was effective in preventing and reducing NASH-related pathologic features, warranting for the clinical development of this compound.
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http://dx.doi.org/10.3390/nu12061819DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7353335PMC
June 2020

Pomegranate Mesocarp against Colitis-Induced Visceral Pain in Rats: Effects of a Decoction and Its Fractions.

Int J Mol Sci 2020 Jun 17;21(12). Epub 2020 Jun 17.

Department of Neuroscience, Psychology, Drug Research and Child Health-NEUROFARBA-Pharmacology and Toxicology Section, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

The management of chronic visceral pain related to Inflammatory Bowel Diseases or Irritable Bowel Syndrome is still a clinical problem and new therapeutic strategies continue to be investigated. In the present study, the efficacy of a pomegranate decoction and of its polysaccharide and ellagitannin components in preventing the development of colitis-induced abdominal pain in rats was evaluated. After colitis induction by 2,4-dinitrobenzenesulfonic acid (DNBS), the pomegranate decoction (300 mg kg), polysaccharides (300 mg kg), and ellagitannins (45 mg kg) were orally administered for 14 days. Repeated treatment with decoction reduced visceral hypersensitivity in the colitic animals both at 7 and 14 days. Similar efficacy was shown by polysaccharides, but with lower potency. Ellagitannins administered at dose equivalent to decoction content showed higher efficacy in reducing the development of visceral pain. Macroscopic and microscopic evaluations performed on the colon 14 days after the damage showed that all three preparations reduced the overall amount of mast cells, the number of degranulated mast cells, and the density of collagen fibers in the mucosal stroma. Although ellagitannins seem to be responsible for most of the beneficial effects of pomegranate on DNBS-induced colitis, the polysaccharides support and enhance its effect. Therefore, pomegranate mesocarp preparations could represent a complementary approach to conventional therapies for promoting abdominal pain relief.
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http://dx.doi.org/10.3390/ijms21124304DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7353021PMC
June 2020

Coronaridine congeners decrease neuropathic pain in mice and inhibit α9α10 nicotinic acetylcholine receptors and Ca2.2 channels.

Neuropharmacology 2020 09 12;175:108194. Epub 2020 Jun 12.

Department of Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), Section of Pharmacology and Toxicology, University of Florence, 50139, Florence, Italy.

The primary aim of this study was to determine the anti-neuropathic activity of (±)-18-methoxycoronaridine [(±)-18-MC] and (+)-catharanthine in mice by using the oxaliplatin-induced neuropathic pain paradigm and cold plate test. The results showed that both coronaridine congeners induce anti-neuropathic pain activity at a dose of 72 mg/kg (per os), whereas a lower dose (36 mg/kg) of (+)-catharanthine decreased the progress of oxaliplatin-induced neuropathic pain. To determine the underlying molecular mechanism, electrophysiological recordings were performed on α9α10, α3β4, and α4β2 nAChRs as well as voltage-gated calcium (Ca2.2) channels modulated by G protein-coupled γ-aminobutyric acid type B receptors (GABARs). The results showed that (±)-18-MC and (+)-catharanthine competitively inhibit α9α10 nAChRs with potencies higher than that at α3β4 and α4β2 nAChRs and directly block Ca2.2 channels without activating GABARs. Considering the potency of the coronaridine congeners at Cav2.2 channels and α9α10 nAChRs, and the calculated brain concentration of (+)-catharanthine, it is plausible that the observed anti-neuropathic pain effects are mediated by peripheral and central mechanisms involving the inhibition of α9α10 nAChRs and/or Ca2.2 channels.
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http://dx.doi.org/10.1016/j.neuropharm.2020.108194DOI Listing
September 2020

Design, synthesis and biological evaluation of 7-substituted 4-phenyl-6H-imidazo[1,5-a]thieno[3,2-f] [1,4]diazepines as safe anxiolytic agents.

Eur J Med Chem 2020 Aug 18;200:112405. Epub 2020 May 18.

Dipartimento di Biotecnologie, Chimica e Farmacia, (Dipartimento d'Eccellenza 2018-2022), Università di Siena, Via Aldo Moro 2, 53100, Siena, Italy. Electronic address:

A series of 4-phenyl-6H-imidazo[1,5-a]thieno[3,2-f][1,4]diazepine-7-carboxylate esters were synthesized and tested as central benzodiazepine receptor (CBR) ligands by the ability to displace [H]flumazenil from rat cortical membranes. All the compounds showed high affinity with IC values ranging from 5.19 to 16.22 nM. In particular, compounds 12b (IC = 8.66 nM) and 12d (IC = 5.19 nM) appeared as the most effective ligands being their affinity values significantly lower than that of diazepam (IC = 18.52 nM). Compounds 12a-f were examined in vivo for their pharmacological effects in mice and five potential benzodiazepine (BDZ) actions were thus taken into consideration: anxiolytic, anticonvulsant, anti-amnesic, hypnotic, and locomotor activities. All the new synthesized compounds were able to induce a significant antianxiety effect and, among them, compound 12f protected pentylenetetrazole (PTZ)-induced convulsions in a dose-dependent manner reaching a 40% effect at 30 mg/kg. In addition, all the compounds were able to significantly prevent the memory impairment evoked by scopolamine, while none of them was able to interfere with pentobarbital-evoked sleep and influence motor coordination. Moreover, title compounds did not affect locomotor and exploratory activity at the same time and doses at which the anti-anxiety effect was observed. Finally, molecular docking simulations were carried out in order to assess the binding mode for compounds 12a-f. The obtained results demonstrated that these compounds bind the BDZ binding site in a similar fashion to flumazenil.
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http://dx.doi.org/10.1016/j.ejmech.2020.112405DOI Listing
August 2020

Intra-Articular Route for the System of Molecules 14G1862 from : Pain Relieving and Protective Effects in a Rat Model of Osteoarthritis.

Nutrients 2020 May 31;12(6). Epub 2020 May 31.

Department of Neuroscience, Psychology, Drug Research and Child Health-NEUROFARBA-Pharmacology and Toxicology Section, University of Florence, Viale Gaetano Pieraccini 6, 50139 Florence, Italy.

Current pharmacological therapies for the management of chronic articular diseases are far from being satisfactory, so new strategies need to be investigated. We tested the intra-articular pain relieving properties of a system of molecules from a characterized extract (14G1862) in a rat model of osteoarthritis induced by monoiodoacetate (MIA). 14G1862 (0.2-2 mg mL) was intra-articularly (i.a.) injected 7 days after MIA, behavioural and histological evaluations were performed 14, 30 and 60 days after treatments. Moreover, the effect of 14G1862 on nitrate production and iNOS expression in RAW 264.7 macrophages stimulated with LPS was assessed. , 14G1862 treatment attenuated LPS-induced NO production and iNOS expression in a comparable manner to celecoxib. , 14G1862 significantly reduced mechanical allodynia and hyperalgesia, spontaneous pain and motor alterations starting on day 14 up to day 60. The efficacy was higher or comparable to that evoked by triamcinolone acetonide (100 μg i.a.) used as reference drug. Histological evaluation highlighted the improvement of several morphological parameters in MIA + 14G1862-treated animals with particularly benefic effects on joint space and fibrin deposition. In conclusion, i.a. treatment with is a candidate to be a novel effective approach for osteoarthritis therapy.
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http://dx.doi.org/10.3390/nu12061618DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7352185PMC
May 2020

Novel formyl peptide receptor (FPR) agonists with pyridinone and pyrimidindione scaffolds that are potentially useful for the treatment of rheumatoid arthritis.

Bioorg Chem 2020 07 28;100:103880. Epub 2020 Apr 28.

NEUROFARBA, Sezione Farmaceutica e Nutraceutica, Università degli Studi di Firenze, Via Ugo Schiff 6, 50019 Sesto Fiorentino, Italy.

The resolution of inflammation is an active response involving the interaction of pro-resolving mediators with specific receptors, such as N-formyl peptide receptor 2 (FPR2). FPRs represent potentially important therapeutic targets for the treatment of some pathologies, including asthma and rheumatoid arthritis. Previously, we identified selective or mixed FPR agonists with a pyridazin-3(2H)-one scaffold, all containing a 4-bromophenylacetamide fragment at N-2. The most effective compounds in this series were EC3, a potent mixed FPR1/FPR2/FPR3 agonist, and EC10, which had a preference for FPR1. We report here a new series of pyridinone and pyrimidindione derivatives containing the 4-(bromophenyl)acetamide substituent that was essential for activity in the pyridazinone series. All new compounds were evaluated for FPR agonist activity in HL60 cells transfected with FPR1 or FPR2 and in human neutrophils. While most of the pyridinone derivatives had reasonable FPR agonist activity in the submicromolar/micromolar range, the pyrimidindione derivatives were less active. Compound 2a (N-(4-bromophenyl)-2-[3-cyano-5-(3-methoxyphenyl)-6-methyl-2-oxopyridin-1(2H)-yl]acetamide) was the most active pyridinone derivative and had a 10-fold preference for FPR2 (EC = 120 nM) versus FPR1 (EC = 1.6 μM). To assess their therapeutic activity, compounds 2a, EC3, and EC10 were evaluated in vivo using a rat model of rheumatoid arthritis. All three compounds increased the pain threshold and reduced pain hypersensitivity in the treated rats versus control rats, although 2a and EC10 were much more effective than EC3. Thus, these FPR agonists represent potential leads to develop for the treatment of inflammatory diseases such as rheumatoid arthritis.
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http://dx.doi.org/10.1016/j.bioorg.2020.103880DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7409366PMC
July 2020

Acute visceral pain relief mediated by A3AR agonists in rats: involvement of N-type voltage-gated calcium channels.

Pain 2020 Sep;161(9):2179-2190

Department of Neuroscience, Psychology, Drug Research and Child Health-Neurofarba-Division of Pharmacology and Toxicology, Florence, Italy.

Abstract: Pharmacological tools for chronic visceral pain management are still limited and inadequate. A3 adenosine receptor (A3AR) agonists are effective in different models of persistent pain. Recently, their activity has been related to the block of N-type voltage-gated Ca2+ channels (Cav2.2) in dorsal root ganglia (DRG) neurons. The present work aimed to evaluate the efficacy of A3AR agonists in reducing postinflammatory visceral hypersensitivity in both male and female rats. Colitis was induced by the intracolonic instillation of 2,4-dinitrobenzenesulfonic acid (DNBS; 30 mg in 0.25 mL 50% EtOH). Visceral hypersensitivity was assessed by measuring the visceromotor response and the abdominal withdrawal reflex to colorectal distension. The effects of A3AR agonists (MRS5980 and Cl-IB-MECA) were evaluated over time after DNBS injection and compared to that of the selective Cav2.2 blocker PD173212, and the clinically used drug linaclotide. A3AR agonists significantly reduced DNBS-evoked visceral pain both in the postinflammatory (14 and 21 days after DNBS injection) and persistence (28 and 35 days after DNBS) phases. Efficacy was comparable to effects induced by linaclotide. PD173212 fully reduced abdominal hypersensitivity to control values, highlighting the role of Cav2.2. The effects of MRS5980 and Cl-IB-MECA were completely abolished by the selective A3AR antagonist MRS1523. Furthermore, patch-clamp recordings showed that A3AR agonists inhibited Cav2.2 in dorsal root ganglia neurons isolated from either control or DNBS-treated rats. The effect on Ca2+ current was PD173212-sensitive and prevented by MRS1523. A3AR agonists are effective in relieving visceral hypersensitivity induced by DNBS, suggesting a potential therapeutic role against abdominal pain.
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http://dx.doi.org/10.1097/j.pain.0000000000001905DOI Listing
September 2020

Effects of the Combination of β-Hydroxy-β-Methyl Butyrate and R(+) Lipoic Acid in a Cellular Model of Sarcopenia.

Molecules 2020 04 30;25(9). Epub 2020 Apr 30.

Department of Neurosciences, Psychology, Drug Research and Child Health-Neurofarba-Pharmacology and Toxicology Section, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

Sarcopenia is a clinical problem associated with several pathological and non-pathological conditions. The aim of the present research is the evaluation of the pharmacological profile of the leucine metabolite β-hydroxy-β-methyl butyrate (HMB) associated with the natural R(+) stereoisomer of lipoic acid (R(+)LA) in a cellular model of muscle wasting. The C2C12 cell line is used as myoblasts or is differentiated in myotubes, sarcopenia is induced by dexamethasone (DEX). A Bonferroni significant difference procedure is used for a post hoc comparison. DEX toxicity (0.01-300 µM concentration range) is evaluated in myoblasts to measure cell viability and caspase 3 activation after 24 h and 48 h; cell incubation with 1 µM DEX for 48 h is chosen as optimal treatment for decreasing cell viability and increasing caspase 3 activity. R(+)LA or HMB significantly prevents DEX-induced cell mortality; the efficacy is improved when 100 µM R(+)LA is combined with 1 mM HMB. Regarding myoblasts, this combination significantly reduces DEX-evoked O production and protein oxidative damage. During the early phase of myotube formation, the mixture preserves the number of myogenin-positive cells, whereas it completely prevents the DEX-dependent damage in a later phase of myotube differentiation (7 days), as evaluated by cell diameter and percentage of multinucleated cells. R(+)LA in association with HMB is suggested for sarcopenia therapy.
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http://dx.doi.org/10.3390/molecules25092117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7249096PMC
April 2020

Phenyl(thio)phosphon(amid)ate Benzenesulfonamides as Potent and Selective Inhibitors of Human Carbonic Anhydrases II and VII Counteract Allodynia in a Mouse Model of Oxaliplatin-Induced Neuropathy.

J Med Chem 2020 05 14;63(10):5185-5200. Epub 2020 May 14.

Department of NEUROFARBA, Pharmaceutical and Nutraceutical Section, University of Florence, Via Ugo Schiff 6, 50019 Sesto Fiorentino, Italy.

Human carbonic anhydrase (CA; EC 4.2.1.1) isoforms II and VII are implicated in neuronal excitation, seizures, and neuropathic pain (NP). Their selective inhibition over off-target CAs is expected to produce an anti-NP action devoid of side effects due to promiscuous CA modulation. Here, a drug design strategy based on the observation of (dis)similarities between the target CA active sites was planned with benzenesulfonamide derivatives and, for the first time, a phosphorus-based linker. Potent and selective CA II/VII inhibitors were identified among the synthesized phenyl(thio)phosphon(amid)ates -. X-ray crystallography depicted the binding mode of phosphonic acid to both CAs II and VII. The most promising derivatives, after evaluation of their stability in acidic media, were tested in a mouse model of oxaliplatin-induced neuropathy. The most potent compound racemic mixture was subjected to HPLC enantioseparation, and the identification of the eutomer, the ()-enantiomer, allowed to halve the dose totally relieving allodynia in mice.
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http://dx.doi.org/10.1021/acs.jmedchem.9b02135DOI Listing
May 2020

β-Sitosterol Loaded Nanostructured Lipid Carrier: Physical and Oxidative Stability, In Vitro Simulated Digestion and Hypocholesterolemic Activity.

Pharmaceutics 2020 Apr 22;12(4). Epub 2020 Apr 22.

Department of Chemistry, University of Florence, via Schiff 6, Sesto Fiorentino, 50019 Florence, Italy.

The objective of the present study was to explore the potential of nanostructured lipid carriers (NLCs) for improving the oral delivery of β-sitosterol, a poorly water-soluble bioactive component with hypocholesterolemic activity. Two β-sitosterol formulations with different solid lipid compositions were prepared by melt emulsification, followed by the sonication technique, and the effect of storage conditions and simulated digestion on the physical, chemical and oxidative stability, bioaccessibility and release were extensively studied. Both NLC preparations remained relatively stable during the four weeks of storage at different conditions (4, 25 and 40 °C), with more superior stability at lower temperatures. The in vitro digestion experiment indicated a high physical stability after exposure to the simulated mouth and stomach stages and an improved overall β-sitosterol bioaccessibility at the end of the digestion. The NLCs presented an increased solubility and gradual release which could be justified by the remarkable affinity of β-sitosterol to the complex lipid mixture. An in vivo study demonstrated an improved reduction in the total cholesterol and low-density lipoprotein cholesterol plasma levels in mice compared with the drug suspension. These investigations evidenced the potential of the developed NLC formulations for the enhancement of solubility and in vivo performance of β-sitosterol.
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http://dx.doi.org/10.3390/pharmaceutics12040386DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7237988PMC
April 2020

Bacopa monnieri as augmentation therapy in the treatment of anhedonia, preclinical and clinical evaluation.

Phytother Res 2020 Sep 1;34(9):2331-2340. Epub 2020 Apr 1.

Department Neurofarba, Psychiatry Section, University of Florence, Florence, Italy.

Bacopa monnieri (L.) is widely used in Ayurvedic medicine as a neural tonic for improving intelligence and memory. Several studies highlighted its efficacy in neuropsychiatric diseases but there is no evidence regarding anhedonia. Aim of the present work was to preclinically and clinically test against anhedonia a standardized B. monnieri extract (20% bacosides). In a mouse model of a depressive-like syndrome induced by lipopolysaccharide (LPS), the daily administration of the extract (50-200 mg kg , p.o.) for 1 week, dose-dependently counteracted the immobility time in Porsolt and Tail suspension tests (p < .01). At the sucrose preference test (directly related to the ability for feeling pleasure) the extract treatment (100 and 200 mg kg ) counteracted the reduction of sucrose intake induced by LPS (p < .01). Moreover, B. monnieri significantly reduced cytokines, cortisol, and artemin LPS-dependent alterations in plasma while increased the brain-derived neurotrophic factor levels (p < .05). The efficacy of the same extract was tested in a clinical study in which 42 patients with significant degree of anhedonia (evaluated as Snaith-Hamilton Pleasure Scale [SHAPS] score ≥ 3) were enrolled. Patients were divided into two groups and treated with citalopram or citalopram associated with B. monnieri (300 mg bid) for 4 weeks. The Pears Sample T-test showed a significant improvement (p < .05) in relevant scales (Hamilton depression rating scale, SHAPS, and strength and difficulties questionnaire) in the extract-treated group in comparison to citalopram alone was recorded. These data suggest that B. monnieri extract may be effective for the management of anhedonia and therefore should be considered for future controlled trials.
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http://dx.doi.org/10.1002/ptr.6684DOI Listing
September 2020

Functional Selectivity and Antinociceptive Effects of a Novel KOPr Agonist.

Front Pharmacol 2020 5;11:188. Epub 2020 Mar 5.

Department of Pharmacy and Biotechnology, University of Bologna, Bologna, Italy.

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 functional selectivity and 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 cAMP ELISA and β-arrestin 2 recruitment assays. Antinociception and antihypersensitivity were assessed in mice 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.
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http://dx.doi.org/10.3389/fphar.2020.00188DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7066533PMC
March 2020

The Citrus Flavonoid Naringenin Protects the Myocardium from Ageing-Dependent Dysfunction: Potential Role of SIRT1.

Oxid Med Cell Longev 2020 25;2020:4650207. Epub 2020 Jan 25.

Department of Pharmacy, University of Pisa, Via Bonanno 6, 56126 Pisa, Italy.

Sirtuin 1 (SIRT1) enzyme plays a pivotal role in the regulation of many physiological functions. In particular, it is implicated in ageing-related diseases, such as cardiac hypertrophy, myocardial infarct, and endothelial dysfunction; moreover, its expression decreases with age. Therefore, an effective strategy to extend the lifespan and improve cardiovascular function is the enhancement of the expression/activity of SIRT1 with exogenous agents. The Citrus flavonoid naringenin (NAR) presents structural similarity with the natural SIRT1 activator resveratrol. In this study, we demonstrate through assays that NAR significantly activates SIRT1 enzyme and shows antisenescence effects. The binding mode of NAR into SIRT1 was detailed investigated through studies. Moreover, chronic administration (for six months) of NAR (100 mg/kg/day) to 6-month-old mice leads to an enhancement of SIRT1 expression and a marked reduction of reactive oxygen species production in myocardial tissue. Furthermore, at the end of the treatment, the plasma levels of two well-known markers of cardiovascular inflammation, TNF- and IL6, are significantly reduced in 12-month-old mice treated with NAR, as well as the cardiovascular risk (total cholesterol/HDL ratio) compared to control mice. Finally, the age-associated fibrotic remodeling, which is well detected through a Mallory trichrome staining in the vehicle-treated 12-month-old mice, is significantly reduced by the chronic treatment with NAR. Moreover, an improvement of myocardium functionality is highlighted by the enhancement of citrate synthase activity and stabilization of the mitochondrial membrane potential after NAR treatment. Taken together, these results suggest that a nutraceutical approach with NAR may have positive impacts on many critical hallmarks of myocardial senescence, contributing to improve the cardiac performance in aged subjects.
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http://dx.doi.org/10.1155/2020/4650207DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7003265PMC
September 2020

Researching New Therapeutic Approaches for Abdominal Visceral Pain Treatment: Preclinical Effects of an Assembled System of Molecules of Vegetal Origin.

Nutrients 2019 Dec 20;12(1). Epub 2019 Dec 20.

Department of Neuroscience, Psychology, Drug Research and Child Health-Neurofarba-Pharmacology and Toxicology Section, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

Abdominal pain is a frequent symptom of irritable bowel syndrome (IBS) and inflammatory bowel diseases (IBDs). Although the knowledge of these pathologies is progressing, new therapeutic strategies continue to be investigated. In the present study, the effect of a system of molecules of natural origin (a medical device according to EU Directive 93/42/EC, engineered starting from resins, polysaccharides and and polyphenols) was evaluated against the intestinal damage and visceral pain development in DNBS-induced colitis model in rats. The system (250 and 500 mg kg) was orally administered once daily, starting three days before the injection of 2,4-dinitrobenzenesulfonic acid (DNBS) and for 14 days thereafter. The viscero-motor response (VMR) to colon-rectal balloon distension (CRD) was used as measure of visceral sensitivity. The product significantly reduced the VMR of DNBS-treated animals. Its effect on pain threshold was better than dexamethasone and mesalazine, and not lower than amitriptyline and otilonium bromide. At microscopic and macroscopic level, the tested system was more effective in protecting the intestinal mucosa than dexamethasone and mesalazine, promoting the healing of tissue lesions. Therefore, we suggest that the described system of molecules of natural origin may represent a therapeutic option to manage painful bowel diseases.
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http://dx.doi.org/10.3390/nu12010022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7019336PMC
December 2019