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1.
Exp Eye Res ; 222: 109168, 2022 09.
Artículo en Inglés | MEDLINE | ID: mdl-35777472

RESUMEN

Dry eye disease (DED), a multifactorial disease of the tears and ocular system, causes loss of tear film homeostasis with damage to the ocular surface. This study aimed to assess whether a peculiar matrix based on sodium hyaluronate (HA), xanthan gum (XNT), glycine (GLY) and betaine (BET) as osmoprotectants, could be involved in biological responses. Wound healing assay on human corneal epithelial (HCE) cells in monolayer showed a synergistic effect of the combination of HA + XNT (**p ≤ 0.01) together with an efficient extracellular matrix remodeling of the formulation in SkinEthic™ HCE 3D-model sought by integrin beta-1 (ITGß1) expression and morphological analysis by hematoxylin and eosin (H&E), compared to a reference marketed product. The synergistic effect of HA + XNT + GLY + BET showed an antioxidant effect on HCE cells (***p ≤ 0.001). Real-time PCR analysis showed that the combination of GLY + BET seemed to ameliorate the effect exhibited by the single osmoprotectants in reducing tumor necrosis factor-alpha (TNFα, #p ≤ 0.05), interleukin-1 beta (IL1ß, ####p ≤ 0.0001) and cyclooxygenases-2 (COX2, ####p ≤ 0.0001) genes in SIRC cells under hyperosmotic stress. Furthermore, pretreatment with XNT, alone and in combination (##p ≤ 0.01), reduced COX2 expression in human non-small cell lung cancer cells (A549). Finally, the formulation was well-tolerated following q.i.d. ocular administration in rabbits during a 28-day study. Due to the synergistic effect of its components, the matrix proved able to repair the ocular surface restoring cell homeostasis and to protect the ocular surface from pro-inflammatory pathways activation and oxidative damage, thus behaving as a reactive oxygen species (ROS) scavenger.


Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Síndromes de Ojo Seco , Neoplasias Pulmonares , Animales , Carcinoma de Pulmón de Células no Pequeñas/complicaciones , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Ciclooxigenasa 2 , Síndromes de Ojo Seco/metabolismo , Humanos , Neoplasias Pulmonares/complicaciones , Neoplasias Pulmonares/metabolismo , Conejos , Lágrimas/metabolismo
2.
Int J Mol Sci ; 22(9)2021 Apr 22.
Artículo en Inglés | MEDLINE | ID: mdl-33922399

RESUMEN

Eye drop formulations allowing topical treatment of retinal pathologies have long been sought as alternatives to intravitreal administration. This study aimed to assess whether a novel nanostructured microemulsions system (NaMESys) could be usefully employed to deliver sorafenib to the retina following topical instillation. NaMESys carrying 0.3% sorafenib (NaMESys-SOR) proved to be cytocompatible in vitro on rabbit corneal cells, and well-tolerated following b.i.d. ocular administration to rabbits during a 3-month study. In rats subject to retinal ischemia-reperfusion, NaMESys-SOR significantly inhibited retinal expression of tumor necrosis factor-alpha (TNFα, 20.7%) and inducible nitric oxide synthase (iNos, 87.3%) mRNAs in comparison to controls. Similarly, in streptozotocin-induced diabetic rats, NaMESys-SOR inhibited retinal expression of nuclear factor kappa B (NFκB), TNFα, insulin like growth factor 1 (IGF1), IGF1 receptor (IGF1R), vascular endothelial growth factor receptor 1 (VEGFR1) and 2 (VEGFR2) mRNAs by three-fold on average compared to controls. Furthermore, a reduction in TNFα, VEGFR1 and VEGFR2 protein expression was observed by western blot. Moreover, in mice subject to laser-induced choroidal neovascularization, NaMESys-SOR significantly inhibited neovascular lesions by 54%. In conclusion, NaMESys-SOR was shown to be a well-tolerated ophthalmic formulation able to deliver effective amounts of sorafenib to the retina, reducing proinflammatory and pro-angiogenic mediators in reliable models of proliferative retinopathies. These findings warrant further investigations on the full therapeutic potential of NaMESys-SOR eye drops, aiming to address unmet needs in the pharmacotherapy of retinal neovascular diseases.


Asunto(s)
Neovascularización Coroidal/tratamiento farmacológico , Diabetes Mellitus Experimental/complicaciones , Retinopatía Diabética/tratamiento farmacológico , Nanoestructuras/administración & dosificación , Enfermedades de la Retina/tratamiento farmacológico , Neovascularización Retiniana/tratamiento farmacológico , Sorafenib/farmacología , Administración Oftálmica , Animales , Retinopatía Diabética/etiología , Retinopatía Diabética/patología , Modelos Animales de Enfermedad , Emulsiones , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Nanoestructuras/química , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/farmacología , Conejos , Ratas , Ratas Sprague-Dawley , Enfermedades de la Retina/patología , Sorafenib/administración & dosificación
3.
Mol Pain ; 13: 1744806917697009, 2017 01.
Artículo en Inglés | MEDLINE | ID: mdl-28326943

RESUMEN

Background L-acetylcarnitine, a drug marketed for the treatment of chronic pain, causes analgesia by epigenetically up-regulating type-2 metabotropic glutamate (mGlu2) receptors in the spinal cord. Because the epigenetic mechanisms are typically long-lasting, we hypothesized that analgesia could outlast the duration of L-acetylcarnitine treatment in models of inflammatory and neuropathic pain. Results A seven-day treatment with L-acetylcarnitine (100 mg/kg, once a day, i.p.) produced an antiallodynic effect in the complete Freund adjuvant mouse model of chronic inflammatory pain. L-Acetylcarnitine-induced analgesia persisted for at least 14 days after drug withdrawal. In contrast, the analgesic effect of pregabalin, amitryptiline, ceftriaxone, and N-acetylcysteine disappeared seven days after drug withdrawal. L-acetylcarnitine treatment enhanced mGlu2/3 receptor protein levels in the dorsal region of the spinal cord. This effect also persisted for two weeks after drug withdrawal and was associated with increased levels of acetylated histone H3 bound to the Grm2 gene promoter in the dorsal root ganglia. A long-lasting analgesic effect of L-acetylcarnitine was also observed in mice subjected to chronic constriction injury of the sciatic nerve. In these animals, a 14-day treatment with pregabalin, amitryptiline, tramadol, or L-acetylcarnitine produced a significant antiallodynic effect, with pregabalin displaying the greatest efficacy. In mice treated with pregabalin, tramadol or L-acetylcarnitine the analgesic effect was still visible 15 days after the end of drug treatment. However, only in mice treated with L-acetylcarnitine analgesia persisted 37 days after drug withdrawal. This effect was associated with an increase in mGlu2/3 receptor protein levels in the dorsal horns of the spinal cord. Conclusions Our findings suggest that L-acetylcarnitine has the unique property to cause a long-lasting analgesic effect that might reduce relapses in patients suffering from chronic pain.


Asunto(s)
Acetilcarnitina/farmacología , Acetilcarnitina/uso terapéutico , Epigénesis Genética/efectos de los fármacos , Inflamación/tratamiento farmacológico , Neuralgia/tratamiento farmacológico , Amitriptilina/uso terapéutico , Analgésicos/farmacología , Analgésicos/uso terapéutico , Animales , Enfermedad Crónica , Modelos Animales de Enfermedad , Adyuvante de Freund/efectos adversos , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/etiología , Inflamación/inducido químicamente , Masculino , Ratones , Ratones Endogámicos C57BL , Manejo del Dolor , Pregabalina/uso terapéutico , Receptores de Glutamato Metabotrópico/metabolismo , Factores de Tiempo , Tramadol/uso terapéutico
4.
Pharmacol Res ; 67(1): 1-9, 2013 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-23073075

RESUMEN

Fingolimod (FTY720), a novel drug approved for the treatment of relapsing-remitting multiple sclerosis, activates different sphingosine-1-phosphate receptor (S1PR) subtypes. Its primary mechanism of action is to reduce the egress of T lymphocytes from secondary lymphoid organs, thus restraining neuroinflammation and autoimmunity. However, recent evidence suggests that the action of FTY720 involves S1PRs expressed by cells resident in the CNS, including neurons. Here, we examined the effect of FTY720, its active metabolite, FTY720-P, and sphingosine-1-phosphate (S1P) on neuronal viability using a classical in vitro model of excitotoxic neuronal death. Mixed cultures of mouse cortical cells were challenged with toxic concentrations of N-methyl-d-aspartate (NMDA) for 10 min, and neuronal death was assessed 20 h later. FTY720, FTY720-P, and S1P were all neuroprotective when applied 18-20 h prior to the NMDA pulse. Neuroprotection was attenuated by pertussis toxin, and inhibited by the selective type-1 S1PR (S1P1R) antagonist, W146, and by inhibitors of the mitogen associated protein kinase (MAPK) and the phosphatidylinositol-3-kinase (PtdIns-3-K) pathways. Both FTY720 and FTY720-P retained their protective activity in pure cultures of mouse or rat cortical neurons. These data offer the first direct demonstration that FTY720 and its active metabolite protect neurons against excitotoxic death.


Asunto(s)
Inmunosupresores/farmacología , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Organofosfatos/farmacología , Glicoles de Propileno/farmacología , Esfingosina/análogos & derivados , Animales , Astrocitos/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Células Cultivadas , Femenino , Clorhidrato de Fingolimod , Lisofosfolípidos/farmacología , Ratones , Ratas , Esfingosina/farmacología
5.
Mol Pain ; 8: 77, 2012 Oct 23.
Artículo en Inglés | MEDLINE | ID: mdl-23088864

RESUMEN

BACKGROUND: Pharmacological activation of type-2 metabotropic glutamate receptors (mGlu2 receptors) causes analgesia in experimental models of inflammatory and neuropathic pain. Presynaptic mGlu2 receptors are activated by the glutamate released from astrocytes by means of the cystine/glutamate antiporter (System x(c)(-) or Sx(c)(-)). We examined the analgesic activity of the Sx(c)(-) activator, N-acetyl-cysteine (NAC), in mice developing inflammatory or neuropathic pain. RESULTS: A single injection of NAC (100 mg/kg, i.p.) reduced nocifensive behavior in the second phase of the formalin test. NAC-induced analgesia was abrogated by the Sxc- inhibitor, sulphasalazine (8 mg/kg, i.p.) or by the mGlu2/3 receptor antagonist, LY341495 (1 mg/kg, i.p.). NAC still caused analgesia in mGlu3(-/-) mice, but was inactive in mGlu2(-/-) mice. In wild-type mice, NAC retained the analgesic activity in the formalin test when injected daily for 7 days, indicating the lack of tolerance. Both single and repeated injections of NAC also caused analgesia in the complete Freund's adjuvant (CFA) model of chronic inflammatory pain, and, again, analgesia was abolished by LY341495. Data obtained in mice developing neuropathic pain in response to chronic constriction injury (CCI) of the sciatic nerve were divergent. In this model, a single injection of NAC caused analgesia that was reversed by LY341495, whereas repeated injections of NAC were ineffective. Thus, tolerance to NAC-induced analgesia developed in the CCI model, but not in models of inflammatory pain. The CFA and CCI models differed with respect to the expression levels of xCT (the catalytic subunit of Sx(c)(-)) and activator of G-protein signaling type-3 (AGS3) in the dorsal portion of the lumbar spinal cord. CFA-treated mice showed no change in either protein, whereas CCI mice showed an ipislateral reduction in xCT levels and a bilateral increase in AGS3 levels in the spinal cord. CONCLUSIONS: These data demonstrate that pharmacological activation of Sxc- causes analgesia by reinforcing the endogenous activation of mGlu2 receptors. NAC has an excellent profile of safety and tolerability when clinically used as a mucolytic agent or in the management of acetaminophen overdose. Thus, our data encourage the use of NAC for the experimental treatment of inflammatory pain in humans.


Asunto(s)
Acetilcisteína/uso terapéutico , Analgésicos/uso terapéutico , Dolor/tratamiento farmacológico , Receptores de Glutamato Metabotrópico/metabolismo , Sistema de Transporte de Aminoácidos y+/genética , Sistema de Transporte de Aminoácidos y+/metabolismo , Animales , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Glutamato Metabotrópico/genética
6.
Pharmaceutics ; 13(11)2021 Nov 18.
Artículo en Inglés | MEDLINE | ID: mdl-34834371

RESUMEN

Uveal melanoma is the second most common melanoma and the most common intraocular malignant tumour of the eye. Among various treatments currently studied, Sorafenib was also proposed as a promising drug, often administered with other compounds in order to avoid resistance mechanisms. Despite its promising cellular activities, the use of Sorafenib by oral administration is limited by its severe side effects and the difficulty to reach the target. The encapsulation into drug delivery systems represents an interesting strategy to overcome these limits. In this study, different lipid nanoparticulate formulations were prepared and compared in order to select the most suitable for the encapsulation of Sorafenib. In particular, two solid lipids (Softisan or Suppocire) at different concentrations were used to produce solid lipid nanoparticles, demonstrating that higher amounts were able to achieve smaller particle sizes, higher homogeneity, and longer physical stability. The selected formulations, which demonstrated to be biocompatible on Statens Seruminstitut Rabbit Cornea cells, were modified to improve their mucoadhesion, evaluating the effect of two monovalent cationic lipids with two lipophilic chains. Sorafenib encapsulation allowed obtaining a sustained and prolonged drug release, thus confirming the potential use of the developed strategy to topically administer Sorafenib in the treatment of uveal melanoma.

7.
Sci Rep ; 8(1): 13361, 2018 09 06.
Artículo en Inglés | MEDLINE | ID: mdl-30190524

RESUMEN

In cerebellar Purkinje cells (PCs) type-1 metabotropic glutamate (mGlu1) receptors play a key role in motor learning and drive the refinement of synaptic innervation during postnatal development. The cognate mGlu5 receptor is absent in mature PCs and shows low expression levels in the adult cerebellar cortex. Here we found that mGlu5 receptors were heavily expressed by PCs in the early postnatal life, when mGlu1α receptors were barely detectable. The developmental decline of mGlu5 receptors coincided with the appearance of mGlu1α receptors in PCs, and both processes were associated with specular changes in CpG methylation in the corresponding gene promoters. It was the mGlu1 receptor that drove the elimination of mGlu5 receptors from PCs, as shown by data obtained with conditional mGlu1α receptor knockout mice and with targeted pharmacological treatments during critical developmental time windows. The suppressing activity of mGlu1 receptors on mGlu5 receptor was maintained in mature PCs, suggesting that expression of mGlu1α and mGlu5 receptors is mutually exclusive in PCs. These findings add complexity to the the finely tuned mechanisms that regulate PC biology during development and in the adult life and lay the groundwork for an in-depth analysis of the role played by mGlu5 receptors in PC maturation.


Asunto(s)
Regulación hacia Abajo , Epigénesis Genética , Células de Purkinje/metabolismo , Receptores AMPA/metabolismo , Receptores de Ácido Kaínico/biosíntesis , Sinapsis/metabolismo , Animales , Islas de CpG , Metilación de ADN , Masculino , Ratones , Ratones Noqueados , Células de Purkinje/citología , Receptores AMPA/genética , Receptores de Ácido Kaínico/genética , Sinapsis/genética
8.
Sci Rep ; 5: 17799, 2015 Dec 08.
Artículo en Inglés | MEDLINE | ID: mdl-26643205

RESUMEN

The kynurenine pathway of tryptophan metabolism has been implicated in the pathophysiology of psychiatric disorders, including schizophrenia. We report here that the kynurenine metabolite, xanturenic acid (XA), interacts with, and activates mGlu2 and mGlu3 metabotropic glutamate receptors in heterologous expression systems. However, the molecular nature of this interaction is unknown, and our data cannot exclude that XA acts primarily on other targets, such as the vesicular glutamate transporter, in the CNS. Systemic administration of XA in mice produced antipsychotic-like effects in the MK-801-induced model of hyperactivity. This effect required the presence of mGlu2 receptors and was abrogated by the preferential mGlu2/3 receptor antagonist, LY341495. Because the mGlu2 receptor is a potential drug target in the treatment of schizophrenia, we decided to measure serum levels of XA and other kynurenine metabolites in patients affected by schizophrenia. Serum XA levels were largely reduced in a large cohort of patients affected by schizophrenia, and, in patients with first-episode schizophrenia, levels remained low after 12 months of antipsychotic medication. As opposed to other kynurenine metabolites, XA levels were also significantly reduced in first-degree relatives of patients affected by schizophrenia. We suggest that lowered serum XA levels might represent a novel trait marker for schizophrenia.


Asunto(s)
Carácter Cuantitativo Heredable , Receptores de Glutamato Metabotrópico/agonistas , Esquizofrenia/diagnóstico , Esquizofrenia/metabolismo , Xanturenatos/metabolismo , Adulto , Anciano , Animales , Biomarcadores , Encéfalo/metabolismo , Estudios de Casos y Controles , Femenino , Células HEK293 , Humanos , Quinurenina/metabolismo , Masculino , Metabolómica/métodos , Ratones , Persona de Mediana Edad , Unión Proteica , Esquizofrenia/sangre , Transducción de Señal , Membranas Sinápticas/metabolismo , Xanturenatos/sangre , Adulto Joven
9.
Neuropharmacology ; 81: 237-43, 2014 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-24565643

RESUMEN

Cinnabarinic acid (CA) is an endogenous metabolite of the kynurenine pathway which acts as an orthosteric agonist of type-4 metabotropic glutamate receptor (mGlu4). We now report that systemic administration of CA (0.1-10 mg/kg, i.p.) was highly protective against experimental autoimmune encephalomyelitis (EAE) induced by the myelin oligodendrocyte glycoprotein (MOG35-55) peptide, which models multiple sclerosis in mice. Full protection against EAE required daily injections of CA since the time of immunization, similarly to what reported for the mGlu4 enhancer N-phenyl-7-(hydroxyimino)cyclopropa[b]chromen-1acarboxamide (PHCCC). CA treatment boosted an immune response dominated by regulatory T (Treg) cells at the expenses of Th17 cells. In addition, exogenous CA enhanced endogenous CA formation in lymphocytes, suggesting the occurrence of a positive feedback loop sustaining immune tolerance. To examine whether activation of mGlu4 could account for the protective activity of CA against EAE, we used mGlu4 knockout mice. As expected, these mice displayed a more severe form of EAE in response to immunization. CA was still protective against EAE in mGlu4-deficient mice, although its action was significantly reduced both at high and low CA doses. This suggests that the action of CA against neuroinflammation involves multiple mechanisms including the activation of mGlu4. These data further suggest that CA is one possible bridge between activation of the kynurenine pathway and immune tolerance aimed at restraining neuroinflammation.


Asunto(s)
Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Agonistas de Aminoácidos Excitadores/uso terapéutico , Oxazinas/uso terapéutico , Receptores de Glutamato Metabotrópico/metabolismo , Animales , Benzopiranos/uso terapéutico , Sistema Nervioso Central/patología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/inducido químicamente , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteína Básica de Mielina/metabolismo , Glicoproteína Mielina-Oligodendrócito/toxicidad , Miosina Tipo II/metabolismo , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/genética , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/metabolismo , Fragmentos de Péptidos/toxicidad , Proteínas de Dominio T Box/genética , Proteínas de Dominio T Box/metabolismo , Células Th17/efectos de los fármacos , Factores de Tiempo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
10.
Mol Brain ; 6: 48, 2013 Nov 19.
Artículo en Inglés | MEDLINE | ID: mdl-24252411

RESUMEN

BACKGROUND: Spinocerebellar ataxia type 1 (SCA1) is a genetic disorder characterized by severe ataxia associated with progressive loss of cerebellar Purkinje cells. The mGlu1 metabotropic glutamate receptor plays a key role in mechanisms of activity-dependent synaptic plasticity in the cerebellum, and its dysfunction is linked to the pathophysiology of motor symptoms associated with SCA1. We used SCA1 heterozygous transgenic mice (Q154/Q2) as a model for testing the hypothesis that drugs that enhance mGlu1 receptor function may be good candidates for the medical treatment of SCA1. RESULTS: Symptomatic 30-week old SCA1 mice showed reduced mGlu1 receptor mRNA and protein levels in the cerebellum. Interestingly, these mice also showed an intense expression of mGlu5 receptors in cerebellar Purkinje cells, which normally lack these receptors. Systemic treatment of SCA1 mice with the mGlu1 receptor positive allosteric modulator (PAM), Ro0711401 (10 mg/kg, s.c.), caused a prolonged improvement of motor performance on the rotarod and the paw-print tests. A single injection of Ro0711401 improved motor symptoms for several days, and no tolerance developed to the drug. In contrast, the mGlu5 receptor PAM, VU0360172 (10 mg/kg, s.c.), caused only a short-lasting improvement of motor symptoms, whereas the mGlu1 receptor antagonist, JNJ16259685 (2.5 mg/kg, i.p.), further impaired motor performance in SCA1 mice. The prolonged symptomatic benefit caused by Ro0711401 outlasted the time of drug clearance from the cerebellum, and was associated with neuroadaptive changes in the cerebellum, such as a striking reduction of the ectopically expressed mGlu5 receptors in Purkinje cells, increases in levels of total and Ser880-phosphorylated GluA2 subunit of AMPA receptors, and changes in the length of spines in the distal dendrites of Purkinje cells. CONCLUSIONS: These data demonstrate that pharmacological enhancement of mGlu1 receptors causes a robust and sustained motor improvement in SCA1 mice, and lay the groundwork for the development of mGlu1 receptor PAMs as novel "cerebellum-specific", effective, and safe symptomatic drugs for the treatment of SCA1 in humans.


Asunto(s)
Amidas/farmacología , Receptores de Glutamato Metabotrópico/metabolismo , Ataxias Espinocerebelosas/metabolismo , Ataxias Espinocerebelosas/patología , Animales , Cerebelo/efectos de los fármacos , Cerebelo/metabolismo , Cerebelo/patología , Cromatografía Líquida de Alta Presión , Modelos Animales de Enfermedad , Regulación de la Expresión Génica/efectos de los fármacos , Espectrometría de Masas , Ratones , Actividad Motora/efectos de los fármacos , Niacinamida/análogos & derivados , Niacinamida/farmacología , Células de Purkinje/efectos de los fármacos , Células de Purkinje/metabolismo , Células de Purkinje/patología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptor del Glutamato Metabotropico 5/genética , Receptor del Glutamato Metabotropico 5/metabolismo , Receptores de Glutamato Metabotrópico/genética , Ataxias Espinocerebelosas/genética , Ataxias Espinocerebelosas/fisiopatología
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