RESUMEN
Sensory axons degenerate following separation from their cell body, but partial injury to peripheral nerves may leave the integrity of damaged axons preserved. We show that an endogenous ligand for the natural killer (NK) cell receptor NKG2D, Retinoic Acid Early 1 (RAE1), is re-expressed in adult dorsal root ganglion neurons following peripheral nerve injury, triggering selective degeneration of injured axons. Infiltration of cytotoxic NK cells into the sciatic nerve by extravasation occurs within 3 days following crush injury. Using a combination of genetic cell ablation and cytokine-antibody complex stimulation, we show that NK cell function correlates with loss of sensation due to degeneration of injured afferents and reduced incidence of post-injury hypersensitivity. This neuro-immune mechanism of selective NK cell-mediated degeneration of damaged but intact sensory axons complements Wallerian degeneration and suggests the therapeutic potential of modulating NK cell function to resolve painful neuropathy through the clearance of partially damaged nerves.
Asunto(s)
Células Asesinas Naturales/fisiología , Proteínas Asociadas a Matriz Nuclear/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Traumatismos de los Nervios Periféricos/metabolismo , Animales , Axones , Ganglios Espinales/citología , Ganglios Espinales/metabolismo , Células Asesinas Naturales/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Subfamilia K de Receptores Similares a Lectina de Células NK/metabolismo , Regeneración Nerviosa , Neuronas/citología , Neuronas Aferentes/inmunología , Neuronas Aferentes/metabolismo , Proteínas Asociadas a Matriz Nuclear/fisiología , Proteínas de Transporte Nucleocitoplasmático/fisiología , Dolor , Traumatismos de los Nervios Periféricos/inmunología , Enfermedades del Sistema Nervioso Periférico , Nervio Ciático , Células Receptoras Sensoriales/metabolismoRESUMEN
Aiming at developing an affordable and easily implementable computational protocol for routine prediction of spectral properties of rigid molecular dyes, density functional theory, and time-dependent density functional theory were used in conjunction with a vibronic coupling scheme for band shape estimate. To predict the perceived color of molecules in solution, a model has been setup linking the UV-vis spectra predicted at ab initio level to the L*a*b* colorimetric parameters. The results show that a mixed protocol, implying the use of a global hybrid functional for the prediction of adiabatic energy differences and a range separated hybrid for the prediction of potential energy curvature, allows perceived colors to be quantitatively predicted, as demonstrated by the comparison of L*a*b* colorimetric parameters obtained from computed and experimental spectra. © 2017 Wiley Periodicals, Inc.
RESUMEN
Although axonal damage induces rapid changes in gene expression in primary sensory neurons, it remains unclear how this process is initiated. The transcription factor ATF3, one of the earliest genes responding to nerve injury, regulates expression of downstream genes that enable axon regeneration. By exploiting ATF3 reporter systems, we identify topoisomerase inhibitors as ATF3 inducers, including camptothecin. Camptothecin increases ATF3 expression and promotes neurite outgrowth in sensory neurons in vitro and enhances axonal regeneration after sciatic nerve crush in vivo. Given the action of topoisomerases in producing DNA breaks, we determine that they do occur immediately after nerve damage at the ATF3 gene locus in injured sensory neurons and are further increased after camptothecin exposure. Formation of DNA breaks in injured sensory neurons and enhancement of it pharmacologically may contribute to the initiation of those transcriptional changes required for peripheral nerve regeneration.
Asunto(s)
Factor de Transcripción Activador 3/metabolismo , Axones/metabolismo , Roturas del ADN/efectos de los fármacos , ADN-Topoisomerasas de Tipo I/metabolismo , Traumatismos de los Nervios Periféricos/metabolismo , Células Receptoras Sensoriales/metabolismo , Animales , ADN-Topoisomerasas de Tipo I/efectos de los fármacos , Expresión Génica/fisiología , Ratones Endogámicos C57BL , Regeneración Nerviosa/efectos de los fármacos , Regeneración Nerviosa/fisiología , Proyección Neuronal/fisiología , Nervio Ciático/metabolismoRESUMEN
Chronic neuropathic pain is a major morbidity of neural injury, yet its mechanisms are incompletely understood. Hypersensitivity to previously non-noxious stimuli (allodynia) is a common symptom. Here, we demonstrate that the onset of cold hypersensitivity precedes tactile allodynia in a model of partial nerve injury, and this temporal divergence was associated with major differences in global gene expression in innervating dorsal root ganglia. Transcripts whose expression change correlates with the onset of cold allodynia were nociceptor related, whereas those correlating with tactile hypersensitivity were immune cell centric. Ablation of TrpV1 lineage nociceptors resulted in mice that did not acquire cold allodynia but developed normal tactile hypersensitivity, whereas depletion of macrophages or T cells reduced neuropathic tactile allodynia but not cold hypersensitivity. We conclude that neuropathic pain incorporates reactive processes of sensory neurons and immune cells, each leading to distinct forms of hypersensitivity, potentially allowing drug development targeted to each pain type.
Asunto(s)
Conducta Animal , Hiperalgesia/fisiopatología , Neuralgia/fisiopatología , Transcriptoma , Animales , Frío , Hiperalgesia/etiología , Hiperalgesia/inmunología , Macrófagos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuralgia/complicaciones , Neuralgia/inmunología , Células Receptoras Sensoriales/metabolismo , Linfocitos T/inmunología , Canales Catiónicos TRPV/deficiencia , TactoRESUMEN
Neuropathic pain is a major, intractable clinical problem and its pathophysiology is not well understood. Although recent gene expression profiling studies have enabled the identification of novel targets for pain therapy, classical study designs provide unclear results owing to the differential expression of hundreds of genes across sham and nerve-injured groups, which can be difficult to validate, particularly with respect to the specificity of pain modulation. To circumvent this, we used two outbred lines of rats, which are genetically similar except for being genetically segregated as a result of selective breeding for differences in neuropathic pain hypersensitivity. SerpinA3N, a serine protease inhibitor, was upregulated in the dorsal root ganglia (DRG) after nerve injury, which was further validated for its mouse homolog. Mice lacking SerpinA3N developed more neuropathic mechanical allodynia than wild-type (WT) mice, and exogenous delivery of SerpinA3N attenuated mechanical allodynia in WT mice. T lymphocytes infiltrate the DRG after nerve injury and release leukocyte elastase (LE), which was inhibited by SerpinA3N derived from DRG neurons. Genetic loss of LE or exogenous application of a LE inhibitor (Sivelastat) in WT mice attenuated neuropathic mechanical allodynia. Overall, we reveal a novel and clinically relevant role for a member of the serpin superfamily and a leukocyte elastase and crosstalk between neurons and T cells in the modulation of neuropathic pain.
Asunto(s)
Proteínas de Fase Aguda/metabolismo , Inhibidores Enzimáticos/farmacología , Elastasa de Leucocito/antagonistas & inhibidores , Serpinas/metabolismo , Linfocitos T/citología , Animales , Separación Celular , Dependovirus/genética , Femenino , Ganglios Espinales/metabolismo , Hiperalgesia/fisiopatología , Hibridación in Situ , Masculino , Ratones , Ratones Transgénicos , Neuralgia , Neuronas/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Dolor/fisiopatología , Fenotipo , Reacción en Cadena de la Polimerasa , Ratas , Regulación hacia ArribaRESUMEN
Axon regeneration in the CNS requires reactivating injured neurons' intrinsic growth state and enabling growth in an inhibitory environment. Using an inbred mouse neuronal phenotypic screen, we find that CAST/Ei mouse adult dorsal root ganglion neurons extend axons more on CNS myelin than the other eight strains tested, especially when pre-injured. Injury-primed CAST/Ei neurons also regenerate markedly in the spinal cord and optic nerve more than those from C57BL/6 mice and show greater sprouting following ischemic stroke. Heritability estimates indicate that extended growth in CAST/Ei neurons on myelin is genetically determined, and two whole-genome expression screens yield the Activin transcript Inhba as most correlated with this ability. Inhibition of Activin signaling in CAST/Ei mice diminishes their CNS regenerative capacity, whereas its activation in C57BL/6 animals boosts regeneration. This screen demonstrates that mammalian CNS regeneration can occur and reveals a molecular pathway that contributes to this ability.
Asunto(s)
Axones/fisiología , Ganglios Espinales/fisiología , Regeneración Nerviosa/fisiología , Neuropatía Ciática/fisiopatología , Traumatismos de la Médula Espinal/fisiopatología , Animales , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Endogámicos NOD , Neuropatía Ciática/patología , Traumatismos de la Médula Espinal/patologíaRESUMEN
Human genetic studies have revealed an association between GTP cyclohydrolase 1 polymorphisms, which decrease tetrahydrobiopterin (BH4) levels, and reduced pain in patients. We now show that excessive BH4 is produced in mice by both axotomized sensory neurons and macrophages infiltrating damaged nerves and inflamed tissue. Constitutive BH4 overproduction in sensory neurons increases pain sensitivity, whereas blocking BH4 production only in these cells reduces nerve injury-induced hypersensitivity without affecting nociceptive pain. To minimize risk of side effects, we targeted sepiapterin reductase (SPR), whose blockade allows minimal BH4 production through the BH4 salvage pathways. Using a structure-based design, we developed a potent SPR inhibitor and show that it reduces pain hypersensitivity effectively with a concomitant decrease in BH4 levels in target tissues, acting both on sensory neurons and macrophages, with no development of tolerance or adverse effects. Finally, we demonstrate that sepiapterin accumulation is a sensitive biomarker for SPR inhibition in vivo.
Asunto(s)
Biopterinas/análogos & derivados , Regulación de la Expresión Génica/fisiología , Inflamación/metabolismo , Neuralgia/metabolismo , Oxidorreductasas de Alcohol/metabolismo , Animales , Antiinflamatorios no Esteroideos/uso terapéutico , Biopterinas/metabolismo , Presión Sanguínea/efectos de los fármacos , Péptido Relacionado con Gen de Calcitonina/metabolismo , Modelos Animales de Enfermedad , Inhibidores Enzimáticos/uso terapéutico , GTP Ciclohidrolasa/genética , Regulación de la Expresión Génica/efectos de los fármacos , Inflamación/inducido químicamente , Inflamación/tratamiento farmacológico , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Ratones , Ratones Transgénicos , Neuralgia/inducido químicamente , Neuralgia/tratamiento farmacológico , Dimensión del Dolor , Umbral del Dolor/efectos de los fármacos , Umbral del Dolor/fisiología , Tiempo de Reacción/efectos de los fármacos , Tiempo de Reacción/genética , Nervio Ciático/metabolismo , Células Receptoras Sensoriales/efectos de los fármacos , Células Receptoras Sensoriales/metabolismo , Sulfasalazina/uso terapéutico , Factores de TiempoRESUMEN
OBJECTIVE: Plasma kallikrein (PK) has been identified in vitreous fluid obtained from individuals with diabetic retinopathy and has been implicated in contributing to retinal vascular dysfunction. In this report, we examined the effects of PK on retinal vascular functions and thickness in diabetic rats. RESEARCH DESIGN AND METHODS: We investigated the effects of a selective PK inhibitor, ASP-440, and C1 inhibitor (C1-INH), the primary physiological inhibitor of PK, on retinal vascular permeability (RVP) and hemodynamics in rats with streptozotocin-induced diabetes. The effect of intravitreal PK injection on retinal thickness was examined by spectral domain optical coherence tomography. RESULTS: Systemic continuous administration of ASP-440 for 4 weeks initiated at the time of diabetes onset inhibited RVP by 42% (P = 0.013) and 83% (P < 0.001) at doses of 0.25 and 0.6 mg/kg per day, respectively. Administration of ASP-440 initiated 2 weeks after the onset of diabetes ameliorated both RVP and retinal blood flow abnormalities in diabetic rats measured at 4 weeks' diabetes duration. Intravitreal injection of C1-INH similarly decreased impaired RVP in rats with 2 weeks' diabetes duration. Intravitreal injection of PK increased both acute RVP and sustained focal RVP (24 h postinjection) to a greater extent in diabetic rats compared with nondiabetic control rats. Intravitreal injection of PK increased retinal thickness compared with baseline to a greater extent (P = 0.017) in diabetic rats (from 193 ± 10 µm to 223 ± 13 µm) compared with nondiabetic rats (from 182 ± 8 µm to 193 ± 9 µm). CONCLUSIONS: These results show that PK contributes to retinal vascular dysfunctions in diabetic rats and that the combination of diabetes and intravitreal injection of PK in rats induces retinal thickening.